Method for adjusting reward and derogatory

Abstract

The present application relates to a method of treating depression, anorexia or greedy comprising modulating the activity of ACC neurons.

Description

Technical Field

[0001] The present application relates to methods for treating depression, anorexia and bulimia. The present application also relates to a method for regulating reward devaluation. Background of the Invention

[0003] Depression is a kind of mental illness, and it affects more than 300 million people in the whole world, and has brought huge global burden.During depressive episode, patient experiences depressed mood (feeling sad, irritable, empty) or loses the joy or interest of activity.Also there are several other symptoms, and it may comprise the feeling of inattention, excessive guilt or low self-worth, despair to the future, think of death or suicide, sleep interruption and feeling particularly tired or energy is low.Depression may also be relevant with appetite, causes for example bulimia or anorexia, and it may cause obesity or excessive weight consumption, and affects physical health.

[0004] Among dozens of symptoms, lack of happiness is the one most directly associated with suicidal thoughts in patients with depression. However, the specific neural mechanisms underlying this lack of happiness are not fully understood, and for many patients with depression, there are no effective and adequate therapies that improve all symptoms. Therefore, further exploration of the mechanisms and targets or potential treatments for depression is needed. SUMMARY OF THE INVENTION

[0006] In this application, new targets for treating depression, anorexia and bulimia are studied. Based on the studies, the inventors provide methods for treating depression, anorexia and bulimia.

[0007] In a first aspect, a method for treating depression or anorexia is provided. In some embodiments, depression or anorexia is treated by alleviating reward devaluation. In some embodiments, the method comprises inhibiting, destroying, interrupting or blocking ACC neurons in a subject in need thereof. In some embodiments, ACC neurons are ACC pyramidal neurons. In some embodiments, the neurons are basolateral amygdala (BLA)-projecting ACC neurons. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are destroyed. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are destroyed. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons are destroyed. In some embodiments, all signals from the ACC are blocked or interrupted. In some embodiments, all signals from the ACC pyramidal neurons are blocked or interrupted. In some embodiments, all signals from the BLA-projecting ACC neurons are blocked or interrupted. In some embodiments, the activation of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC is inhibited. In some embodiments, the activation of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons is inhibited. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the activation of BLA-projecting ACC neurons is inhibited. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are hyperpolarized. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are hyperpolarized. In some embodiments, all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons are hyperpolarized.

[0008] In some embodiments, a method for inhibiting, destroying, interrupting or blocking ACC neurons comprises biological, chemical and / or physical methods. In some embodiments, a method for inhibiting, destroying, interrupting or blocking ACC neurons uses chemical genetic means, optogenetic means, electrical stimulation of the brain, pharmacological modulation, or a combination thereof.

[0009] In some embodiments, the method for destroying ACC neurons comprises surgically removing ACC neurons. In some embodiments, the method for destroying ACC neurons comprises introducing an apoptotic protein into ACC neurons. In some embodiments, the method for destroying ACC neurons comprises introducing the coding sequence of an apoptotic protein into ACC neurons. In some embodiments, the apoptotic protein is caspase 3. In some embodiments, the method for destroying ACC neurons comprises introducing a toxin receptor protein and its ligand into ACC neurons. In some embodiments, the method for destroying ACC neurons comprises introducing the coding sequence of a toxin receptor protein and its ligand into ACC neurons. In some embodiments, the toxin receptor protein is DTR (diphtheria toxin receptor). In some embodiments, the toxin receptor protein is expressed in the ACC neurons of the subject, and the toxin is administered to the subject.

[0010] In some embodiments, the method for interrupting or blocking ACC neuron signals comprises introducing an inhibitor of a neurotransmitter synthesized or secreted by ACC neurons into ACC neurons. In some embodiments, the method for interrupting or blocking ACC neuron signals comprises introducing an antagonist of a neurotransmitter synthesized or secreted by ACC neurons into ACC neurons. In some embodiments, the method for interrupting or blocking ACC neuron signals comprises introducing a neutralizer of a neurotransmitter synthesized or secreted by ACC neurons into ACC neurons. In some embodiments, the method for interrupting or blocking ACC neuron signals comprises introducing TeNT (tetanus toxin) into ACC neurons. In some embodiments, the method for interrupting or blocking ACC neuron signals comprises inhibiting the release of neurotransmitters. In some embodiments, the method for interrupting or blocking ACC neuron signals comprises inhibiting the synthesis of neurotransmitters.

[0011] In some embodiments, the method for inhibiting ACC neurons comprises electrical brain stimulation. In some embodiments, the electrical brain stimulation is high frequency stimulation or deep brain stimulation. In some embodiments, the electrical frequency used in the electrical brain stimulation exceeds 50 Hz. In some embodiments, the electrical brain stimulation is performed via a brain-computer interface.

[0012] In some embodiments, the method for inhibiting ACC neurons comprises introducing into the ACC neurons one or more compounds selected from the group consisting of:

[0013] 1) Gi / o coupled receptors;

[0014] 2) ligands or agonists of Gi / o coupled receptors;

[0015] 3) Gi / o coupled receptors and their ligands or agonists;

[0016] 4) anion channel proteins;

[0017] 5) ligands or agonists of anion channel proteins;

[0018] 6) Anion channel proteins and their ligands or agonists;

[0019] 7) Inhibitors or antagonists of cation channel proteins;

[0020] 8) Inhibitory neurotransmitters;

[0021] 9) apoptosis proteins; and

[0022] 10) Toxin receptors and their ligands.

[0023] In some embodiments, the Gi / o coupled receptor is a DREADD (designer receptor activated only by designer drugs). In some embodiments, the DREADD is a variant of the muscarinic acetylcholine receptor. In some embodiments, the DREADD is hM4Di or hM4Dnrxn. In some embodiments, the DREADD is a variant of the kappa opioid receptor. In some embodiments, the DREADD is a KORDi. In some embodiments, the ligand or agonist of the DREADD is selected from the group consisting of clozapine, CNO (clozapine-N-oxide), C21 (compound 21), perlapine, DCZ (desclozapine), JHU37152, JHU37160, and salvinorin B. In some embodiments, the DREADD is a variant of the muscarinic acetylcholine receptor, and the ligand or agonist is clozapine, CNO, C21, perlapine, DCZ, JHU37152, or JHU37160. In some embodiments, the DREADD is a variant of the muscarinic acetylcholine receptor, and the ligand or agonist of the DREADD is salvinorin B. In some embodiments, the DREADD is expressed in ACC neurons of the subject, and the ligand of the DREADD is administered to the subject.

[0024] In some embodiments, the Gi / o-coupled receptor is a Gi / o-coupled light-sensitive protein. In some embodiments, the Gi / o-coupled light-sensitive protein is an opsin. In some embodiments, the Gi / o-coupled light-sensitive protein is selected from the group consisting of Lamplight (Lamprey Parapinopsin), rod opsin, cone opsin, μ-opioid receptor-rod opsin chimera, and Opto-MOR. In some embodiments, the Gi / o-coupled light-sensitive protein is expressed in ACC neurons of a subject, and light of certain wavelengths is used to stimulate the Gi / o-coupled light-sensitive protein.

[0025] In some embodiments, the anion channel protein is a ligand-gated ion channel protein (LGIC) or a variant thereof. In some embodiments, the LGIC is a chloride ion channel protein. In some embodiments, the LGIC is a GABAA receptor. In some embodiments, the LGIC is a glycine receptor. In some embodiments, the LGIC and its ligand are any one selected from the following groups: GlyR-M and ivermectin, GluCl and ivermectin, PSAM-GlyR and PSEM89S, and GABAA and zolpidem. In some embodiments, the LGIC is expressed in ACC neurons of a subject, and a ligand of the LGIC is administered to the subject.

[0026] In some embodiments, the anion channel protein is a light-gated anion channel protein. In some embodiments, the light-gated anion channel protein is selected from the group consisting of eNpHR3.0, Arch, eBR, iC1C2, ChloC, ACRs, GtACR1, and GtACR2. In some embodiments, the light-gated anion channel protein is expressed in ACC neurons of a subject, and light of certain wavelengths acts directly or indirectly on the light-gated anion channel protein and causes the opening of the light-gated anion channel.

[0027] In some embodiments, the one or more compounds are selected from the group consisting of muscimol, clozapine-N-oxide, nalfurafine, salvinorin B, allatostatin, 8-chloro-11-[4-(1,1-dideuteroethyl)piperazin-1-yl]-5H-dibenzo[b,e][1,4]diazepine, NMDAR antagonists, AMPAR antagonists, GABA and its analogs, GABA receptor agonists. In a second aspect, the present application provides a method for treating obesity or bulimia in a subject. In some embodiments, the method comprises promoting or accelerating reward devaluation.

[0028] In some embodiments, the method includes enhancing the signal of the ACC neurons of the subject. In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA projection ACC neurons. In some embodiments, the signal of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC is enhanced. In some embodiments, the signal of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons is enhanced. In some embodiments, the signal of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA projection ACC neurons is enhanced. In some embodiments, the method for enhancing signaling of ACC neurons comprises promoting the release or synthesis of neurotransmitters secreted from ACC neurons.

[0029] In some embodiments, the method includes activating the subject's ACC neurons. In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA projection ACC neurons. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are activated. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are activated. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA projection ACC neurons are activated.

[0030] In some embodiments, the method for activating ACC neurons includes biological, chemical and / or physical methods. In some embodiments, the method for activating ACC neurons uses chemical genetics, optogenetics, electrical stimulation, pharmacological modulation, or a combination thereof.

[0031] In some embodiments, the method for activating ACC neurons comprises introducing into the ACC neurons one or more compounds selected from the group consisting of:

[0032] 1) Gs-coupled receptors;

[0033] 2) Gs-coupled receptors and ligands or agonists of Gs-coupled receptors;

[0034] 3) Gq-coupled receptors;

[0035] 4) Gq-coupled receptors and ligands or agonists of Gq-coupled receptors;

[0036] 5) Cation channel proteins;

[0037] 6) Cation channel proteins and ligands or agonists of cation channel proteins;

[0038] 7) inhibitors or antagonists of anion channel proteins; and

[0039] 8) Excitatory neurotransmitters.

[0040] In some embodiments, the Gs-coupled receptor or Gq-coupled receptor is a DREADD. In some embodiments, the DREADD is selected from the group consisting of hM3Dq, Rq(R165L), or hM3Ds, and the ligand or agonist is clozapine, CNO (clozapine-N-oxide), C21 (Compound 21), perlapine, DCZ (desclozapine), JHU37152, and JHU37160. In some embodiments, the DREADD is expressed in ACC neurons, and a ligand or agonist of the DREADD is administered to a subject.

[0041] In some embodiments, the Gs-coupled receptor or Gq-coupled receptor is a Gs-coupled or Gq-coupled light-sensitive protein. In some embodiments, the Gs-coupled or Gq-coupled light-sensitive protein is an opsin. In some embodiments, the opsin is selected from the group consisting of cOpn5, hOPN5, and JellyOp (an opsin from Charybdea rastonii). In some embodiments, the Gs-coupled light-sensitive protein or Gq-coupled light-sensitive protein is expressed in ACC neurons of a subject, and light of certain wavelengths is used to stimulate the Gs-coupled light-sensitive protein or Gq-coupled light-sensitive protein.

[0042] In some embodiments, the cation channel protein is a LGIC protein. In some embodiments, the LGIC protein is selected from the group consisting of a 5-HT3 receptor, an acid-sensing ion channel (ASIC) protein, an epithelial sodium channel (ENaC), an ionotropic glutamate receptor, an IP3 receptor, a nicotinic acetylcholine receptor, a P2X receptor, a ryanodine receptor, and a zinc-activated channel protein (ZAC). In some embodiments, the LGIC is expressed in ACC neurons, and a ligand or agonist of the LGIC is administered to a subject.

[0043] In some embodiments, the cation channel protein is a light-gated cation channel protein. In some embodiments, the light-gated cation channel protein is selected from the group consisting of ChR2, CheRiff, Chronos, and variants thereof. Examples of ChR2 variants include, but are not limited to, ChR2(H134R), ChETA, ReaChR, bReaChES, Chrimson, ChrimsonR, C1C2, C1V1, C1V1(t), C1V1(t / t), oChIEF, ChRmine, ChRmine2.0, ChRger1, ChRger2, and ChRger3. Examples of Chronos include, but are not limited to, CsChR, CoChR, VChR1, and CheRiff. In some embodiments, the light-gated cation channel protein is expressed in the ACC neurons of the subject, and light of certain wavelengths is used to stimulate the light-gated cation channel protein, resulting in the opening of the light-gated cation channel.

[0044] In some embodiments, the one or more compounds are selected from the group consisting of glutamate and its analogs, NMDAR agonists, AMPAR agonists, and agonists for other glutamate receptors.

[0045] In a third aspect, the present application also provides a method for alleviating reward devaluation in a subject, which comprises inhibiting, destroying, interrupting or blocking ACC neurons.

[0046] In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA projection ACC neurons. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are inhibited, destroyed, interrupted, or blocked. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are inhibited, destroyed, interrupted, or blocked. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA projection ACC neurons are inhibited, destroyed, interrupted, or blocked. In some embodiments, all or some (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are hyperpolarized. In some embodiments, all or some (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are hyperpolarized. In some embodiments, all or some (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons are hyperpolarized.

[0047] In some embodiments, the method comprises inhibiting the signaling or activation of ACC neurons. In some embodiments, the method comprises removing ACC neurons. In some embodiments, the method comprises promoting or inducing apoptosis of ACC neurons. In some embodiments, the method is a biological, chemical and / or physical method. In some embodiments, the method comprises chemical genetic measures, optogenetic measures, electrical brain stimulation, pharmacological regulation measures, or a combination thereof. In some embodiments, the method comprises introducing one or more compounds selected from the following groups into ACC neurons:

[0048] 1) Gi / o coupled receptors;

[0049] 2) ligands or agonists of Gi / o coupled receptors;

[0050] 3) Gi / o coupled receptors and Gi / o coupled receptor ligands or agonists;

[0051] 4) anion channel proteins;

[0052] 5) ligands or agonists of anion channel proteins;

[0053] 6) Anion channel proteins and ligands or agonists of anion channel proteins;

[0054] 7) Inhibitors or antagonists of cation channel proteins;

[0055] 8) Inhibitory neurotransmitters;

[0056] 9) Inhibitors of neurotransmitters synthesized or secreted by ACC neurons;

[0057] 10) apoptosis proteins;

[0058] 11) Toxin receptors;

[0059] 12) Ligands of toxin receptors.

[0060] In some embodiments, methods for reducing reward devaluation can be used to treat depression or anorexia in a subject.

[0061] In some embodiments, the method for inhibiting, destroying, disrupting or blocking ACC neurons is the same as the method for treating depression or anorexia in the first aspect.

[0062] In some embodiments, the toxin receptor protein is DTR (diphtheria toxin receptor). In some embodiments, the toxin receptor protein is expressed in the ACC neurons of the subject, and the toxin is administered to the subject. In some embodiments, DREADD is expressed in the ACC neurons of the subject, and the ligand of DREADD is administered to the subject, wherein the DREADD is a Gi / o coupled DREADD. In some embodiments, Gi / o coupled light-sensitive protein is expressed in the ACC neurons of the subject, and light of certain wavelengths is used to stimulate the Gi / o coupled light-sensitive protein. In some embodiments, LGIC is expressed in the ACC neurons of the subject, and the ligand of LGIC is administered to the subject. In some embodiments, light-gated anion channel protein is expressed in the ACC neurons of the subject, and light of certain wavelengths acts directly or indirectly on the light-gated anion channel protein and causes the opening of the light-gated anion channel.

[0063] In some embodiments, the method of inhibiting ACC neurons comprises electrical brain stimulation. In some embodiments, the electrical brain stimulation is high frequency stimulation or deep brain stimulation. In some embodiments, the electrical frequency used in the electrical brain stimulation exceeds 50 Hz. In some embodiments, the electrical brain stimulation is performed via a brain-computer interface.

[0064] In a fourth aspect, the present application also provides a method for increasing the devaluation of rewards in a subject, comprising activating ACC neurons or enhancing the activity of ACC neurons.

[0065] In some embodiments, ACC neurons are ACC pyramidal neurons. In some embodiments, ACC neurons are BLA projection ACC neurons. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are activated, or their activity is enhanced. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are activated, or their activity is enhanced. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA projection ACC neurons are activated, or their activity is enhanced.

[0066] In some embodiments, the method for increasing the devaluation of a subject's reward comprises activation of ACC neurons, or enhancing or prolonging the activity of ACC neurons. In some embodiments, the method is a biological, chemical, and / or physical method. In some embodiments, the method comprises chemical genetic measures, optogenetic measures, electrical brain stimulation, pharmacological modulation measures, or a combination thereof. In some embodiments, the method comprises introducing one or more compounds selected from the group consisting of:

[0067] 1) Gs-coupled receptors;

[0068] 2) Gs-coupled receptors and ligands or agonists of Gs-coupled receptors;

[0069] 3) Gq-coupled receptors;

[0070] 4) Gq-coupled receptors and ligands or agonists of Gq-coupled receptors;

[0071] 5) Cation channel proteins;

[0072] 6) Cation channel proteins and ligands or agonists of cation channel proteins;

[0073] 7) inhibitors or antagonists of anion channel proteins; and

[0074] 8) Excitatory neurotransmitters.

[0075] In some embodiments, a Gs-coupled or Gq-coupled DREADD is expressed in ACC neurons, and a ligand or agonist of the DREADD is administered to a subject. In some embodiments, a Gs-coupled light-sensitive protein or a Gq-coupled light-sensitive protein is expressed in ACC neurons of a subject, and light of a certain wavelength is used to stimulate the Gs-coupled light-sensitive protein or the Gq-coupled light-sensitive protein. In some embodiments, a LGIC is expressed in ACC neurons, and a ligand or agonist of the LGIC is administered to a subject, and the LGIC is a cation channel protein. In some embodiments, a light-gated cation channel protein is expressed in ACC neurons of a subject, and light of a certain wavelength is used to stimulate the light-gated cation channel protein, resulting in the opening of the light-gated cation channel.

[0076] In some embodiments of all of the above aspects, a Gi / O-coupled receptor, anion channel protein, apoptotic protein, toxin receptor, Gs-coupled receptor, Gq-coupled receptor, or cation channel protein is introduced into ACC neurons by introducing a nucleic acid encoding a Gi / O-coupled receptor, anion channel protein, apoptotic protein, toxin receptor, Gs-coupled receptor, Gq-coupled receptor, or cation channel protein into ACC neurons. In some embodiments, the nucleic acid is inserted into the genome of the ACC neurons by genetic engineering methods. In some embodiments, the genetic engineering methods utilize genome editing tools or recombinase systems. In some embodiments, the genome editing tools are selected from the group consisting of a CRISPR system, ZFNs, TALENs, and homing endonucleases. In some embodiments, the genetic engineering methods utilize a recombinase system, wherein the recombinase system comprises: 1) a nucleic acid encoding a Gi / O-coupled receptor, anion channel protein, apoptotic protein, toxin receptor, Gs-coupled receptor, Gq-coupled receptor, or cation channel protein, flanked by recombinase recognition sites at the 3' and 5' ends, and 2) a recombinase or a nucleic acid encoding the recombinase corresponding to the recombinase recognition sites. In some embodiments, the nucleic acid flanked by recombinase recognition sites at the 3' and 5' ends of 1) and the recombinase or nucleic acid encoding the recombinase of 2) are introduced into the neuronal cell body and axon terminal of the ACC neuron, respectively. In some embodiments, the nucleic acids of 1) and 2) are contained in different vectors. In some embodiments, the vector introduced into the axon terminal travels retrogradely in the neuron. In some embodiments, the vector introduced into the axon terminal is an AAV2 retroviral vector. In some embodiments, the recombinase system is a Cre / loxP system.

[0077] Also provided is a use of a compound in the preparation of a medicament for treating depression or anorexia in a subject by introducing the drug into ACC neurons, and a compound or a pharmaceutical composition comprising the compound for treating depression or anorexia in a subject by introducing the drug into ACC neurons, wherein the compound is selected from the group consisting of:

[0078] 1) Gi / o coupled receptor or nucleic acid encoding Gi / o coupled receptor;

[0079] 2) ligands or agonists of Gi / o coupled receptors;

[0080] 3) Gi / o coupled receptors (or nucleic acids encoding Gi / o coupled receptors) and ligands or agonists of Gi / o coupled receptors;

[0081] 4) anion channel protein or a nucleic acid encoding an anion channel protein;

[0082] 5) ligands or agonists of anion channel proteins;

[0083] 6) anion channel protein (or nucleic acid encoding anion channel protein) and ligands or agonists of anion channel protein;

[0084] 7) Inhibitors or antagonists of cation channel proteins;

[0085] 8) Inhibitory neurotransmitters;

[0086] 9) Inhibitors of neurotransmitters synthesized or secreted by ACC neurons;

[0087] 10) apoptotic protein or nucleic acid encoding apoptotic protein;

[0088] 11) Toxin receptor or nucleic acid encoding toxin receptor;

[0089] 12) Ligands of toxin receptors.

[0090] In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons.

[0091] Also provided is a use of a compound in the preparation of a medicament for treating obesity or bulimia in a subject by introducing the drug into ACC neurons, and a compound or a pharmaceutical composition comprising the compound for treating obesity or bulimia in a subject by introducing the drug into ACC neurons, wherein the compound is selected from the group consisting of:

[0092] 1) Gs-coupled receptor or nucleic acid encoding a Gs-coupled receptor;

[0093] 2) Gs-coupled receptors (or nucleic acids encoding Gs-coupled receptors) and ligands or agonists of Gs-coupled receptors;

[0094] 3) a Gq-coupled receptor or a nucleic acid encoding a Gq-coupled receptor;

[0095] 4) Gq-coupled receptors (or nucleic acids encoding Gq-coupled receptors) and ligands or agonists of Gq-coupled receptors;

[0096] 5) a cation channel protein or a nucleic acid encoding a cation channel protein;

[0097] 6) cation channel proteins (or nucleic acids encoding cation channel proteins) and ligands or agonists of cation channel proteins;

[0098] 7) inhibitors or antagonists of anion channel proteins; and

[0099] 8) Excitatory neurotransmitters.

[0100] In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons.

[0101] Also provided is a use of a compound in the preparation of a medicament for modulating reward devaluation in a subject by introducing the drug into ACC neurons, and a compound or a pharmaceutical composition comprising the compound for modulating reward devaluation in a subject by introducing the drug into ACC neurons, wherein the compound is selected from the group consisting of:

[0102] 1) G protein-coupled receptor or nucleic acid encoding a G protein-coupled receptor;

[0103] 2) G protein-coupled receptor ligands or agonists;

[0104] 3) G protein-coupled receptors (or nucleic acids encoding G protein-coupled receptors) and ligands or agonists of G protein-coupled receptors;

[0105] 4) ion channel protein or nucleic acid encoding ion channel protein;

[0106] 5) ligands or agonists of ion channel proteins;

[0107] 6) ion channel proteins (or nucleic acids encoding ion channel proteins) and ligands or agonists of ion channel proteins;

[0108] 7) Inhibitors or antagonists of ion channel proteins;

[0109] 8) neurotransmitters or neurotransmitter analogs;

[0110] 9) Inhibitors of neurotransmitters synthesized or secreted by ACC neurons;

[0111] 10) an apoptotic protein or a nucleic acid encoding an apoptotic protein; and

[0112] 11) Toxin receptors or nucleic acids encoding toxin receptors and their ligands.

[0113] In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons.

[0114] Also provided is a use of a compound in the preparation of a medicament for alleviating reward devaluation in a subject by introducing the drug into ACC neurons, and a compound or a pharmaceutical composition comprising the compound for alleviating reward devaluation in a subject by introducing the drug into ACC neurons, wherein the compound is selected from the group consisting of:

[0115] 1) Gi / o coupled receptor or nucleic acid encoding Gi / o coupled receptor;

[0116] 2) ligands or agonists of Gi / o coupled receptors;

[0117] 3) Gi / o coupled receptors (or nucleic acids encoding Gi / o coupled receptors) and Gi / o coupled receptor ligands or agonists;

[0118] 4) anion channel protein or a nucleic acid encoding an anion channel protein;

[0119] 5) ligands or agonists of anion channel proteins;

[0120] 6) anion channel protein (or nucleic acid encoding anion channel protein) and ligands or agonists of anion channel protein;

[0121] 7) Inhibitors or antagonists of cation channel proteins;

[0122] 8) Inhibitory neurotransmitters;

[0123] 9) Inhibitors of neurotransmitters synthesized or secreted by ACC neurons;

[0124] 10) apoptotic protein or nucleic acid encoding apoptotic protein;

[0125] 11) a toxin receptor or a nucleic acid encoding a toxin receptor; and

[0126] 12) Ligands of toxin receptors.

[0127] In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons.

[0128] Also provided is a use of a compound in the preparation of a medicament for increasing reward devaluation in a subject by introducing the drug into ACC neurons, and a compound or a pharmaceutical composition comprising the compound for increasing reward devaluation in a subject by introducing the drug into ACC neurons, wherein the compound is selected from the group consisting of:

[0129] 1) Gs-coupled receptor or nucleic acid encoding a Gs-coupled receptor;

[0130] 2) Gs-coupled receptors (or nucleic acids encoding Gs-coupled receptors) and ligands or agonists of Gs-coupled receptors;

[0131] 3) a Gq-coupled receptor or a nucleic acid encoding a Gq-coupled receptor;

[0132] 4) Gq-coupled receptors (or nucleic acids encoding Gq-coupled receptors) and ligands or agonists of Gq-coupled receptors;

[0133] 5) a cation channel protein or a nucleic acid encoding a cation channel protein;

[0134] 6) cation channel proteins (or nucleic acids encoding cation channel proteins) and ligands or agonists of cation channel proteins;

[0135] 7) inhibitors or antagonists of anion channel proteins; and

[0136] 8) Excitatory neurotransmitters.

[0137] In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA-projecting ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons. In some embodiments, the drug is introduced into all or a portion (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons.

[0138] On the other hand, the present application also provides a device, equipment, facility, instrument or apparatus for treating depression or anorexia in a subject, which comprises an electrode, through which electrical stimulation is passed or by which ACC neurons are inhibited, destroyed, interrupted or blocked. In some embodiments, ACC neurons are ACC pyramidal neurons. In some embodiments, the neurons are basolateral amygdala (BLA) projection ACC neurons. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are destroyed. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are destroyed. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons are destroyed. In some embodiments, all signals from the ACC are blocked or interrupted. In some embodiments, all signals from the ACC pyramidal neurons are blocked or interrupted. In some embodiments, all signals from the BLA-projecting ACC neurons are blocked or interrupted. In some embodiments, the activation of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC is inhibited. In some embodiments, the activation of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons is inhibited. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the activation of BLA-projecting ACC neurons is inhibited. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC are hyperpolarized. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are hyperpolarized. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA-projecting ACC neurons are hyperpolarized. In some embodiments, the electrical stimulation is high frequency stimulation or deep brain stimulation. In some embodiments, the electrical frequency of the electrical stimulation exceeds 50 Hz.

[0139] On the other hand, the present application also provides a device, equipment, facility, instrument or apparatus for treating obesity or bulimia in a subject, which comprises an electrode, through which electrical stimulation is passed or from which the signal of the ACC neurons of the subject is activated or enhanced. In some embodiments, the ACC neurons are ACC pyramidal neurons. In some embodiments, the ACC neurons are BLA projection ACC neurons. In some embodiments, the signal of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the neurons in the ACC is enhanced. In some embodiments, the signal of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons is enhanced. In some embodiments, the signal of all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) BLA projection ACC neurons is enhanced. In some embodiments, the electrode enhances the signal of ACC neurons by promoting the release or synthesis of neurotransmitters secreted from all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) ACC neurons. In some embodiments, all or part (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) ACC neurons in the ACC are activated. In some embodiments, all or some (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the ACC pyramidal neurons are activated. In some embodiments, all or some (e.g., 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) of the BLA projecting ACC neurons are activated. In some embodiments, the electrical stimulation is deep brain stimulation.

[0140] In addition, the present application also provides a target, namely BLA projection neurons as a new target for diagnosing depression. Based on the study of the new target, a method for diagnosing depression or determining the risk of depression in a subject is provided, which includes detecting the degree of activation of BLA projection neurons, wherein a higher degree of activation compared to the average degree detected in a normal population indicates depression or the risk of depression. In some embodiments, detecting the degree of activation includes calculating the number of activated BLA projection neurons and / or measuring the activation duration of BLA projection neurons. In some embodiments, a higher degree of activation means a prolonged average activation time of BLA projection neurons and / or more activated BLA projection neurons. BRIEF DESCRIPTION OF THE DRAWINGS

[0141] Figure 1 a-1h, Chronic restraint stress (CRS)-induced depression is associated with behavioral hypersensitivity to reward devaluation.

[0142] ( Figure 1 a and Figure 1 b) Effects of CRS and ketamine (Ket) treatment (10 mg / kg; intraperitoneal injection one hour before behavioral testing) on ​​the immobility time in the forced swim test ( Figure 1 a) and reward preference in the sucrose preference test (SPT; Figure 1 b). Ctrl represents a group of mice that had not received prior CRS treatment. Each point represents data from a single mouse.

[0143] ( Figure 1 c) Effect of CRS on fluid (water or 1% sucrose) intake during SPT.

[0144] ( Figure 1 d) Fitting curves of the effect of CRS on 1% sucrose intake at different time points. The blue dashed line represents the CRS data normalized to the initial value of control mice.

[0145] ( Figure 1 e) Water intake in control, CRS, and CRS+ketamine mice after 48 h of water deprivation.

[0146] ( Figure 1 f) Figure 1 e Fitting curve of water uptake rate of the animal groups shown.

[0147] ( Figure 1 g) Experimental procedures for the reward devaluation test. Note that only water was provided during the three recovery days.

[0148] ( Figure 1 h) Effects of CRS with or without ketamine on reward devaluation. Box and whisker plot: boxes depict the upper and lower quartiles of the data, whiskers depict the range of the data, and dots depict individual data.

[0149] Figure 2 a-2s, ACC pyramidal neurons control reward devaluation.

[0150] ( Figure 2 a) Schematic diagram showing the method for expressing caspase 3 in ACC pyramidal neurons to induce cell ablation.

[0151] ( Figure 2 b) Representative images of ACC after caspase 3-induced ablation of pyramidal neurons. Red, NeuN staining of neurons; blue, DAPI counterstaining of nuclei.

[0152] ( Figure 2 c) Reward values ​​of mGFP-expressing control mice and caspase 3-expressing mice before and after selective satiety in the reward devaluation test (n=12 mice in the caspase 3 group and n=18 mice in the mGFP control group).

[0153] ( Figure 2 d) Representative micrographs of hM4Di-mcherry expression (red) in ACC pyramidal neurons.

[0154] ( Figure 2 e) Reward values ​​of mGFP-expressing control mice and hM4Di-expressing mice in a reward devaluation test performed 1 hour after CNO injection (n=10 mice per group).

[0155] ( Figure 2 f) Schematic diagram of the water intake test (30 min duration) in which lick events were monitored using a lickometer.

[0156] ( Figure 2 g) Water intake of mGFP-expressing control mice and caspase 3-expressing mice throughout a four-day water intake assay, preceded by 48 hours of water deprivation (n=12 mice for caspase 3 group and n=18 mice for mGFP control group).

[0157] ( Figure 2 h) Fitted curves (1 minute bin width) of water uptake rates for control mice and mice expressing caspase 3. The nested insets show the mean cumulative water uptake for the two animal groups on day 2 of the experiment.

[0158] ( Figure 2 i) Water intake in control mice and mice expressing hM3Dq in the water intake test: Mice were deprived of water for 24 hours before the first day of the experiment, and CNO (2 mg / kg, intraperitoneal injection) was administered 1 hour before the water intake test on the fourth day (n = 9 mice in the hM3Dq group and n = 10 mice in the mGFP control group).

[0159] ( Figure 2 j) Fitted curves of water uptake rates for control and hM3Dq-expressing mice during CNO (day 4). Nested subplots show the average cumulative water uptake curves for the two animal groups during CNO.

[0160] ( Figure 2 k) Timeline of kainic acid (KA)-induced ACC damage and high-fat diet-induced obesity.

[0161] ( Figure 2 l and Figure 2 m) Food intake of ACC-ablated mice and non-ablated control mice during 3 weeks of high-fat diet feeding ( Figure 21 ) and weight gain ( Figure 2 m) (KA group n = 9 mice and saline control group n = 7 mice).

[0162] ( Figure 2 n) Left panel shows typical ACC-ablated mice and non-ablated control littermates after 3 weeks of high-fat diet feeding. Right panel, body weight of ACC-ablated mice and non-ablated control mice during 3 weeks of high-fat diet feeding (n=9 mice in KA group and n=7 mice in saline control group).

[0163] ( Figure 2 o) Timeline of AAV vector injections and behavioral experiments used to investigate the effects of chemogenetic inhibition of ACC pyramidal neurons on depression-like behaviors.

[0164] ( Figure 2 p) Reward values ​​of CRS-inhibited mice expressing mGFP or hM4Di before and after selective satiety in the reward devaluation test (n=10 mice in the hM4Di group and n=9 mice in the mGFP control group).

[0165] ( Figure 2 q and Figure 2 r) Effect of chemogenetic inhibition of ACC pyramidal neurons on the immobility time during the forced swim test in CRS-inhibited mice ( Figure 2 q) and sucrose preference ( Figure 2 Effects of r). Normal saline was injected into hM4Di-expressing mice, or CNO was injected into mGFP-expressing mice as a control. CNO was injected one hour before the experiment (n = 8 mice in the CRS-hM4Di group and n = 12 mice in the CRS-mGFP control group).

[0166] ( Figure 2 s) Sucrose preference test in healthy (CRS-untreated) mice expressing mGFP, hM3Dq, or hM4Di in ACC pyramidal neurons. The test was performed 1 hour after CNO injection and lasted for 3 hours (n = 7 mice in the hM4Di group, n = 7 mice in the hM3Dq group, and n = 6 mice in the mGFP control group).

[0167] Figure 3 a-3m, Reward devaluation is shown to attenuate reward inhibition via ACC pyramidal neurons.

[0168] ( Figure 3 a and Figure 3b) Schematic diagram showing the fiber photometry approach for Ca2+ signals from ACC pyramidal neurons ( Figure 3 a) and methods of delivering liquid stimulation or electrical footshock (0.7 mA, 1 s; Figure 3 b) Liquid stimuli were delivered intraorally via a buccal fistula (water, 1% or 10% sucrose solution or 5 mM quinine solution).

[0169] ( Figure 3 c and Figure 3 d) Average Ca2+ responses of ACC pyramidal neurons to different stimuli over time ( Figure 3 c) and peak response value ( Figure 3 d) (n=6 mice in the electric shock group, n=9 mice in the quinine group, and n=14 mice in the water, 1% sucrose, and 10% sucrose groups).

[0170] ( Figure 3 e) Fiber photometry of Ca2+ signals in response to repeated intraoral water delivery (100 trials divided into five blocks of 20 trials).

[0171] ( Figure 3 f) Heatmap (left) and average trace (right) showing Ca2+ responses of ACC pyramidal neuron populations from representative mice.

[0172] ( Figure 3 g) Quantification of Ca2+ responses from different blocks of water delivery. The minimum Z score was defined as the minimum of the mean responses in (F) (n=12 mice for control GCaMP6m group and n=6 mice for CRS GCaMP6m group).

[0173] ( Figure 3 h) Difference in the minimum Z score (Δmin Z score) of devalued and non-devalued rewards after selective satiation (n = 9 mice per group).

[0174] ( Figure 3 i) Ca from individual ACC pyramidal neurons 2+ Schematic diagram of AAV vector injection and GRIN lens implantation for imaging.

[0175] ( Figure 3 j) Raw fluorescence traces of three neurons. Blue droplets indicate the delivery of sucrose reward.

[0176] ( Figure 3 k) Ca2+ response dynamics of all responding neurons in the five bins across the entire imaging period. Cells were sorted based on the strength of their response to reward delivery.

[0177] ( Figure 3 l and Figure 3 m) Percentage (l) and mean minimum Z score (m) of neurons inhibited by reward delivery throughout the imaging period.

[0178] Figure 4 a-4o, BLA-projecting ACC neurons form functional subpopulations in reward devaluation and depressive behaviors.

[0179] ( Figure 4 a) Axonal projections of ACC pyramidal neurons throughout the mouse brain.

[0180] ( Figure 4 b- Figure 4 d) ACC pyramidal neurons are classified into at least five subtypes based on the axonal morphology of individually reconstructed neurons. ( Figure 4 b) 3D view of 95 reconstructed individual ACC pyramidal neurons. ( Figure 4 c) Horizontal view of the five types of ACC pyramidal neurons classified according to their axonal targets. Figure 4 d) Heatmap of data for normalized axonal projection length for each neuron across 86 representative target brain regions. Each column represents a brain region and each row represents a single neuron.

[0181] ( Figure 4 e) Distinct collateral patterns of different types of ACC pyramidal neurons identified after infusion of rAAV2-retro-Cre vector into separate downstream brain regions (types 1-5: VTA, BLA, NAc, MS, and OFC, respectively) and subsequent infusion of AAV-DIO mGFP into the ACC.

[0182] ( Figure 4 f and Figure 4 g) Ablation of BLA-projecting ACC neurons ( Figure 4 f) and ablation of the other four types of ACC projection neurons ( Figure 4 g) Effect on water intake. Data from mice expressing caspase 3 are plotted as solid lines, and data from their corresponding mGFP-expressing control mice are plotted as dashed lines (ACC-BLA group: n = 10 caspase 3 mice and 7 mGFP control mice; ACC-VTA group: n = 4 caspase 3 mice and 4 mGFP control mice; ACC NAc group: n = 7 caspase 3 mice and 7 mGFP control mice; ACC-MS group: n = 7 caspase 3 mice and 6 mGFP control mice; ACC-OFC group: n = 5 caspase 3 mice and 5 mGFP control mice).

[0183] ( Figure 4h) Reward values ​​of ACC-BLA caspase 3-lesioned mice and ACC-BLA mGFP control mice during a reward devaluation test (n=7 mice for caspase 3 group and n=7 mice for mGFP control group).

[0184] ( Figure 4 i and Figure 4 j) Effects of single-dose chemogenetic activation of BLA-projecting ACC neurons on water intake in a reward devaluation test ( Figure 4 i) and the effects of multiple doses of chemogenetic activation on reward value (insensitive procedure; Figure 4 j; Figure 4 In i, n = 12 mice in the hM3Dq group and n = 10 mice in the mGFP control group; Figure 4 j (n = 9 mice in hM3Dq group and n = 6 mice in mPFP control group).

[0185] ( Figure 4 k and Figure 4 l) Before ketamine treatment (blue) or 1 day after treatment (rose red) ( Figure 4 l; 10 mg / kg IP), in water-deprived healthy mice ( Figure 4 Quantification of Ca2+ responses of BLA-projecting ACC neurons to water reward delivery in CRS-inhibited and water-deprived mice ( Figure 4 k Healthy ACC-BLA GCaMP6m group n = 8 mice and Figure 4 k CRS ACC-BLA GCaMP6m group n = 7 mice).

[0186] ( Figure 4 m) Timeline of DTR-mediated injury of BLA-projecting ACC neurons.

[0187] ( Figure 4 n and Figure 4 o) Effect of DTR-mediated ablation of BLA-projecting ACC neurons on forced swimming immobility time ( mGFP-expressing control mice and DTR-expressing mice (ACC-BLA DTR group n = 11 mice and ACC-BLA mGFP control group n = 8 mice) after DT administration. Figure 4 n) and sucrose preference ( Figure 4 o)’s impact.

[0188] Figure 5 Compound 21 (C21) was shown to inhibit ACC neurons via hM4Di, which resulted in enhanced sucrose preference.

[0189] Figure 6 a-6c shows overexpression of c-fos in depression model mice. ( Figure 6a) Shows the experimental process that generated this data. Immunohistochemical staining for the immediate early gene c-fos was performed after forced swimming challenge (FST). Figure 6 b) c-fos staining in the ACC of control and depression model mice. Blue is DAPI; rose indicates c-fos-positive cells. ( Figure 6 c) Density of c-fos-positive cells in the ACC of control and depression model mice.

[0190] Figure 7 a-7f, Behavioral tests of CRS mice and the antidepressant effect of ketamine.

[0191] ( Figure 7 a) Experimental timeline of chronic restraint stress (CRS) and behavioral assays.

[0192] ( Figure 7 b and Figure 7 c) Effects of CRS and ketamine treatment on forced swim test ( Figure 7 b) Immobility time and sucrose preference ( Figure 7 c) (n = 12 mice in the control group and n = 27 mice in the CRS group in the forced swim test, and n = 10 mice in the control group and n = 10 mice in the CRS group in the sucrose preference test).

[0193] ( Figure 7 d) Mean intake rates of different rewards in water-deprived, non-CRS-treated mice (water group n = 6 mice, 1% sucrose group n = 7 mice, and 10% sucrose group n = 8 mice).

[0194] ( Figure 7 e) Schematic diagram and timeline of the sensitive reward devaluation experiment, in which mice received two rewards and water (red rectangles) during a 3-day recovery phase.

[0195] ( Figure 7 f) If Figure 7 e shows reward devaluation in control and CRS mice in the sensitive reward devaluation test (n = 9 mice in the control group and n = 14 mice in the CRS group).

[0196] Figure 8 a-8d, chemical genetic manipulation of ACC neuronal activity and animal behavior.

[0197] ( Figure 8 a) Raw traces (top) and group data (bottom) of whole-cell patch-clamp recordings show that CNO (5 M) significantly inhibits action potential firing in ACC pyramidal neurons expressing hM4Di (n = 8 cells; left) but increases firing activity in ACC pyramidal neurons expressing hM3Dq (n = 7 cells; right) in slice preparations.

[0198] ( Figure 8 b) Chemoregulation of ACC pyramidal neurons significantly increased water reward intake during the 3-day experimental period (n=9 mice in the hM4Di group and n=11 mice in the mGFP control group).

[0199] ( Figure 8 c) Chemogenetic inhibition of ACC pyramidal neurons significantly slowed the decay of water reward intake rate during day 3, whereas no significant difference was found during the day 2 test without CNO application.

[0200] ( Figure 8 d) Water intake rate curves of control mice and hM3Dq mice without CNO treatment during the experimental period on day 3.

[0201] Figure 9 a-9e, Neurochemical lesion of ACC neurons induces excessive consumption of water reward.

[0202] ( Figure 9 a and Figure 9 b) saline control ( Figure 9 a) or kainic acid (KA; Figure 9 b) NeuN staining (red) of ACC neurons after infusion into the ACC. Blue indicates DAPI counterstaining of cell nuclei.

[0203] ( Figure 9 c) Extent of KA-induced damage (color represents the average degree of KA damage in the ACC from 7 mice).

[0204] ( Figure 9 d) Licking duration of control mice (n=5) and KA-lesioned mice (n=7) in the water intake test across 5 daily testing sessions.

[0205] ( Figure 9 e) Cumulative licking duration curves of control mice (n=5) and KA-lesioned mice (n=7) during day 3.

[0206] Figure 10 a-10f, Responses of ACC pyramidal neurons to water and high-fat food rewards.

[0207] ( Figure 10 a) Images show GCaMP6m expression in ACC pyramidal neurons (green). Blue indicates DAPI counterstaining of cell nuclei.

[0208] ( Figure 10 b) Schematic diagram of the fiber photometry system used to record Ca2+ signals from a population of ACC pyramidal neurons. DM, dichroic mirror; ND, neutral density filter.

[0209] ( Figure 10 c) Ca2+ signals in ACC pyramidal neurons during free water access in mice. Data aligned to the start of a lick (left panel) or the end of a lick (right panel). (Top panel) Heatmap of 21 licking bouts from the example mouse. (Middle panel) Average response of the example mouse during free water access. Gray trace represents the average licking rate during water access. (Bottom panel) Average response of 12 mice.

[0210] ( Figure 10 d)( Figure 10 Statistics of the group responses in b).

[0211] ( Figure 10 e) Schematic diagram showing fiber photometry of Ca2+ signaling in response to delivery of high-fat food pellets (~12 mg / pellet).

[0212] ( Figure 10 f) Responses of ACC pyramidal neurons after consumption of high-fat food pellets (n=8 mice).

[0213] Figure 11 a-11e, The inhibitory strength of ACC pyramidal neurons is related to the reward value.

[0214] ( Figure 11 a to Figure 11 c) Response water ( Figure 11 a), 1% sucrose ( Figure 11 b) or 10% sucrose ( Figure 11 c) Ca of ACC pyramidal neurons 2+ The upper panel shows a heatmap of the average responses to different rewards from 14 recording sites across 7 mice. The lower panel shows the average responses from all recording sites.

[0215] ( Figure 11 d and Figure 11 e) The inhibitory strength of ACC pyramidal neurons in response to different rewards, as measured by the minimum Z score ( Figure 11 d) and the area under zero Z score ( Figure 11 e) Measured (n=14 recording sites from 7 mice).

[0216] Figure 12 a-12f, Aversive stimuli activate ACC pyramidal neurons.

[0217] ( Figure 12 a) Schematic diagram of fiber photometry of footshock and Ca2+ signals from ACC pyramidal neurons.

[0218] ( Figure 12b) Trial-by-trial responses and mean Ca of ACC pyramidal neurons to footshock from a representative mouse. 2+ Heatmap image of the signal.

[0219] ( Figure 12 c) Responses of ACC pyramidal neurons to footshock from the entire experimental group (n=6 mice).

[0220] ( Figure 12 d) Schematic diagram of the auditory-induced fear conditioning test and fiber photometry.

[0221] ( Figure 12 e) Responses of ACC pyramidal neurons to fear-associated cues (7.5 kHz tone) and velocity curves from a representative experimental mouse.

[0222] ( Figure 12 f) Average responses and average speed of ACC pyramidal neurons to fear-related cues (7500 Hz sound) of all mice tested (n=12 mice).

[0223] Figure 13 a-13d, Responses of ACC pyramidal neurons to reward devaluation.

[0224] ( Figure 13 a) Quantification of the Ca2+ response from different blocks of water delivery as measured by the area under the zero Z-score across consecutive blocks.

[0225] ( Figure 13 b) Timeline of reward devaluation test and fiber photometry recording.

[0226] ( Figure 13 c) Schematic diagram showing the random intraoral delivery of one of two liquid rewards via a buccal fistula, with simultaneous Ca regulation of ACC pyramidal neurons. 2+ The signal was measured by fiber photometry.

[0227] ( Figure 13 d) Data from representative mice show that after selective satiation with pear juice, reward suppression (Δmin Z-score) changes for both rewards.

[0228] Figure 14 a-14l, In vivo imaging of ACC pyramidal neurons using a miniaturized microscope and an implanted GRIN lens.

[0229] ( Figure 14 a) Representative images of GRIN lens tracks in a coronal section containing the ACC. The midline is on the left.

[0230] ( Figure 14 b) Representative images of ACC pyramidal neurons showing GCaMP6m expression during in vivo imaging.

[0231] ( Figure 14 c and Figure 14 d) Responses of two ACC pyramidal neurons, one of which was inhibited by sucrose reward delivery ( Figure 14 c), while the other is activated by sucrose reward delivery ( Figure 14 d). (Top) Heat map of Ca2+ signals in response to reward delivery across 20 trials; (bottom) average response across 20 trials.

[0232] ( Figure 14 e) Raw traces of Ca2+ signals from eight ACC pyramidal neurons. Gray lines indicate the timing of footshock.

[0233] ( Figure 14 f and Figure 14 l) Responses of two ACC pyramidal neurons, one of which was excited by foot shock ( Figure 14 g), while the other is inhibited by foot shock ( Figure 14 h). (Left panel) Average response of 10 trials. (Right panel) Ca response to foot shock across trials 2+ Heatmap display of the signal.

[0234] ( Figure 14 h) Heat map image of the average Ca2+ signal of all imaged neurons in response to footshock.

[0235] ( Figure 14 i) (Left panel) Average response of all excitatory (rose red) or inhibitory (blue) neurons to footshock. (Right panel) Percentage of ACC pyramidal neurons that are excited, inhibited, or unresponsive to footshock.

[0236] ( Figure 14 j) Percentage of reward-activated, reward-inhibited, or reward-unresponsive neurons across the reward delivery bins (total number of imaged neurons was 228).

[0237] ( Figure 14 k) Percentage of neurons excited during reward delivery among all recorded neurons throughout the entire process.

[0238] ( Figure 14 l) Average maximum Z score of all excited neurons across the entire process.

[0239] Figure 15 , coronal and sagittal views of different types of ACC pyramidal neurons.

[0240] Figure 16 a-16f, Characterization of ACC axonal projections and soma distribution.

[0241] ( Figure 16 a) Axonal distribution of mGFP-labeled AM, ZI, or HDB-projecting ACC neurons, demonstrating distinct collateral patterns.

[0242] ( Figure 16 b- Figure 16 f) rAAV2-retro-H2B-GFP vector was infused into the ventral tegmental area (VTA; Figure 16 b), basolateral amygdala (BLA; Figure 16 c), nucleus accumbens (NAc; Figure 16 d), medial septal nucleus (MS; Figure 16 e), orbitofrontal cortex (OFC; Figure 16 f), Retrogradely labeled soma (green) has a unique distribution pattern in the ACC. Red indicates tdTomato expression in GABAergic neurons in Vgat-Cre Ai14 mice, indicating that no projection neurons are GABAergic.

[0243] DS, dorsal striatum; NAc, nucleus accumbens; MS, medial septal nucleus; CLA, claustrum; VA / AM, ventral anterior thalamic nucleus / anteromedial thalamic nucleus; V2MM, secondary visual cortex; LPMR, posterolateral thalamic nucleus-medial part of the thalamus; MD, dorsomedial thalamic nucleus; POA, preoptic area; ZI, zona incerta; BLA, basolateral amygdala; PH, posterior hypothalamic area; VTA, ventral tegmental area; SC, superior colliculus; PAG, periaqueductal gray; DRN, dorsal raphe nucleus; MRN, median raphe nucleus; Etc, extorhinal cortex; and DTg, dorsal tegmental nucleus.

[0244] Figure 17 a-17f, Effects of ablation of other downstream specific ACC neurons in the water uptake test.

[0245] By infusing AAV-DIO-Casp3 into the ACC and rAAV2-retro-Cre into multiple downstream centers [( Figure 17 a) Lateral posteromedial thalamus (LPMR), ( Figure 17 b) Dorsal raphe nucleus (DRN), ( Figure 17 c) the anteromedial / ventromedial thalamus (AM / VM), ( Figure 17 d) The horizontal edge of the Broca's diagonal band (HDB), ( Figure 17 e) The medial area of ​​the secondary visual cortex (V2MM) and ( Figure 17 f) claustrum (CLA)], projection-specific lesion of ACC neurons was performed.

[0246] Figure 18a-18e, Chemogenetic activation of BLA-projecting ACC neurons produces depressive-like behaviors.

[0247] ( Figure 18 a) Schematic diagram showing the method for expressing hM3Dq-mCherry in BLA-projecting ACC neurons of C57BL / 6N WT mice.

[0248] ( Figure 18 b) An image showing the expression profile of hM3Dq-mCherry in ACC.

[0249] ( Figure 18 c) Schematic diagram of AAV vector injection and chemogenetic activation of BLA projection neurons for multiple behavioral tests. hM3Dq-mCherry was expressed in BLA projection neurons, and cells were activated with a single dose of CNO (2 mg / kg; IP) before tests of water intake, sucrose preference, and forced swim immobility time, and with five low doses (0.5 mg / kg CNO for 2.5 days) before tests of reward devaluation (n = 9 mice for the hM3Dq-mCherry group and n = 10 mice for the mGFP control group).

[0250] ( Figure 18 d and Figure 18 e) Single-dose chemogenetic activation of BLA-projecting ACC neurons affects sucrose preference ( Figure 18 d) and forced swimming immobility time ( Figure 18 e) Effects of PD-L1 and PD-L2 expression on the mGFP-positive mice (n=9 mice in the hM3Dq-mCherry group and n=10 mice in the mGFP control group).

[0251] Figure 19 a-19d, Response maps of BLA-projecting ACC neurons to water reward delivery.

[0252] ( Figure 19 a) Trial-by-trial heat map images (100 trials total) showing Ca2+ expression in response to water delivery. 2+ Fiber photometry of the signal (left panel). The right panel shows the response averaged over 20 experimental blocks (color indicates the response from the first to the fifth block).

[0253] ( Figure 19 b) Quantification of Ca2+ responses in different blocks by inhibition area (under zero Z-score on the right side of (a)) (n=8 mice).

[0254] ( Figure 19 c) Heatmaps of representative CRS-inhibited mice and mean responses to water reward delivery in different blocks 1 day before and after injection of 10 mg / kg (ip) ketamine.

[0255] ( Figure 19 d) Quantification of Ca2+ responses by inhibition area in different groups of CRS-inhibited mice before and after ketamine injection (n=7 mice).

[0256] Figure 20 a-20f, Whole-cell recordings of BLA-projecting ACC neurons show increased neuronal excitability in CRS mice.

[0257] ( Figure 20 a) Schematic diagram showing the method for labeling BLA-projecting ACC neurons and CaMK2 immunostaining. rAAV2-retro-H2B-EYFP was injected into the BLA. Following H2B-EYFP expression, CaMK2 staining was performed in coronal brain sections containing the ACC.

[0258] ( Figure 20 b) CaMK2 immunoreactive signal (red) within BLA-projecting ACC neurons (green). The image on the right shows a magnified view of the dotted area in the left image.

[0259] ( Figure 20 c) Quantification of the proportion of CaMK2-immunopositive neurons in BLA-projecting ACC neurons (EYFP+&tdTomato+ / EYFP+; a total of 1497 EYFP cells from 16 brain sections from 2 mice).

[0260] ( Figure 20 d) Schematic showing the method for labeling BLA-projecting ACC neurons with H2B-EYFP and CRs prior to slice recordings.

[0261] ( Figure 20 e) Example traces of patch clamp recordings showing evoked firing of action potentials by BLA-projecting ACC neurons from control mice (left) and CRS-challenged mice (right). Both neurons were held at -65 mV and injected with 100 pA.

[0262] ( Figure 20 f) Action potential firing rate as a function of injected current in BLA-projecting ACC neurons from control mice (n=9 cells from 2 mice) or CRS-challenged mice (n=12 cells from 2 mice).

[0263] Figure 21 a-21d, Methods for ablating BLA-projecting ACC neurons.

[0264] ( Figure 21a) Schematic diagram showing the AAV vector injection method and determination of ablation efficiency. rAAC2-retro-Cre was injected into the BLA, and rAAV-DIO DTR (for lesion) or rAAV-DIO-mGFP (for control) was injected into the ACC. rAAV2-retro-mScarlet was also injected into the BLAs to retrogradely label BLA projection neurons.

[0265] ( Figure 21 b) Timeline of AAV vector injection and histochemical determination.

[0266] ( Figure 21 c) Representative images showing the expression profiles of mScarlet in BLA-projecting ACC neurons in control and DTR mice after DT administration.

[0267] ( Figure 21 d) Quantification of the average number of mScarlet-labeled BLA-projecting ACC cells in 50 μm-thick sections in the ACC-BLA mGFP control group (22 ACC regions from 11 mice) and the ACC-BLA DTR ablation group (16 ACC regions from 8 mice).

[0268] In the above figures, data are mean ± SEM. ns indicates not significant. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001. Detailed Description of the Invention

[0270] In the present application, a direct relationship between depression and reward devaluation is disclosed. The following examples show that inhibiting or destroying ACC neurons enhances reward intake and alleviates depression-related symptoms, while activating ACC neurons accelerates reward devaluation and contributes to bulimia and obesity. In some embodiments, obesity is obesity induced by excessive consumption. And among the neurons of the ACC, ACC pyramidal neurons, especially basolateral amygdala (BLA)-projecting ACC neurons, have the highest correlation with reward devaluation. Therefore, the present application provides a method for treating depression, anorexia, obesity and bulimia, which includes regulating the activity of ACC neurons, regulating the signals of ACC neurons or affecting the survival state of ACC neurons. The present application also provides a method for diagnosing depression, which includes detecting the degree of activation of ACC neurons. In some embodiments, ACC neurons are ACC pyramidal neurons. In some embodiments, ACC neurons are BLA-projecting ACC neurons.

[0271] Any compound or measure that can be used to modulate the activity of ACC neurons, modulate the signaling of ACC neurons, or affect the survival state of ACC neurons can be used alone or in combination in the technical solutions of this application. Here, the measures include any applicable physical, chemical, or biological measures or their combinations. Exemplary measures include chemogenetics, optogenetics, electroencephalography, pharmacological modulation, and surgery. The term "compound" as used herein encompasses any applicable synthetic or naturally derived chemical substance, including small molecule compounds and macromolecular compounds, such as proteins and nucleic acids. In this method, the compound is used by introducing the compound into ACC neurons. In this application, "introducing" a compound into ACC neurons means contacting the compound with the ACC neurons. Any physical, biological, or chemical method can be used to introduce the compound. For example, when the compound is a protein, methods for introducing the compound into ACC neurons include directly contacting the protein with the ACC neurons and expressing the protein in contact with the ACC neurons. In other words, as long as the protein is in contact with the ACC neurons in some manner, the protein can also be introduced into the ACC neurons by introducing a nucleic acid encoding the protein and expressing the protein into the ACC neurons or other cells near the ACC neurons.

[0272] The compounds used in this application include but are not limited to:

[0273] 1) G protein-coupled receptors,

[0274] 2) G protein-coupled receptor ligands or agonists;

[0275] 4) ion channel proteins;

[0276] 5) ligands or agonists of ion channel proteins;

[0277] 7) Inhibitors or antagonists of ion channel proteins;

[0278] 8) neurotransmitters or neurotransmitter analogs;

[0279] 9) Inhibitors of neurotransmitters synthesized or secreted by ACC neurons;

[0280] 10) apoptotic proteins; and

[0281] 11) Toxin receptors and their ligands.

[0282] definition

[0283] As used herein, the term "reward devaluation" refers to a normal physiological phenomenon induced by sensory-specific satiety. For example, repeated intake of a certain food can lead to a decrease in the pleasure and desire to eat it, while other foods may become more appealing. If reward devaluation fails, the subject is likely to continue consuming excessive amounts of food, leading to bulimia and possibly obesity. Conversely, when reward devaluation occurs too quickly, the subject may not consume enough energy to sustain themselves, leading to weight loss and poor health, which may be associated with anorexia and depression.

[0284] The term "mitigating reward devaluation" encompasses reducing, moderating, or alleviating the level or extent of reward devaluation, or slowing the development of reward devaluation, while the term "exacerbating reward devaluation" refers to increasing, intensifying, exacerbating, or heightening the level or extent of reward devaluation, or accelerating the development of reward devaluation.

[0285] As used herein, the term "depression" (also referred to as major depressive disorder or clinical depression) is a common but serious mood disorder. It can cause severe symptoms that affect how a subject feels, thinks, and handles daily activities, such as sleeping, eating, or working. In some embodiments, depression is an independent disease. In some embodiments, depression is a symptom of another disease. There are different types of depression, and some of them develop due to specific circumstances, including major depressive disorder, dysthymia, perinatal depression, seasonal affective disorder, and depression with psychotic symptoms. In the present application, when treating a subject for depression, the subject is experiencing a depressive disorder, condition, disease, or symptom.

[0286] As used herein, the term "overconsumption" refers to a state in which reward intake (e.g., food intake) exceeds individual needs. When food overconsumption occurs, too much nutrition and / or energy is provided to the subject, leading to overweight and obesity. Overconsumption is also a symptom of bulimia.

[0287] As used herein, the term "anhedonia" refers to the inability to experience pleasure. This is a common symptom of depression and other mental health disorders. In the following example, sucrose preference was tested in an animal model of depression to measure the state of anhedonia.

[0288] The term "anterior cingulate cortex," or "ACC," refers to the front portion of the cingulate cortex (or limbic lobe), a horseshoe-shaped structure near the center of the brain. The ACC is a part of the cerebral cortex beneath the frontal and parietal lobes and surrounds the head of the corpus callosum.

[0289] The term "pyramidal neuron" or "pyramidal cell" refers to a multipolar neuron found in brain regions including the cerebral cortex, hippocampus, and amygdala. Pyramidal neurons are the main excitatory units of the prefrontal cortex and corticospinal tract of mammals, and CaMKII (calcium / calmodulin-dependent protein kinase type II) is generally considered to be a molecular marker of pyramidal neurons in the cortex and hippocampus. And "ACC pyramidal neurons" refer to pyramidal neurons in the ACC.

[0290] The term "neurotransmitter" refers to a chemical substance that neurons use to communicate with each other and their target tissues during synaptic transmission. Naturally, if a neurotransmitter stimulates the target cell to take action, it is an excitatory neurotransmitter, acting at an excitatory synapse. On the other hand, if it inhibits the target cell, it is an inhibitory neurotransmitter, acting at an inhibitory synapse. Although, as used herein, "excitatory neurotransmitters" or "inhibitory neurotransmitters" can be artificially synthesized and introduced directly into the target neuron.

[0291] The term "basolateral amygdala-projecting ACC neuron" or "BLA-projecting ACC neuron" is an anatomically defined type of neuron. "Basolateral amygdala-projecting ACC neuron" or "BLA-projecting ACC neuron" refers to a neuron whose cell body is located in the ACC and projects axons to the BLA. This type of neuron can be specifically targeted by using, for example, retrotransduction-engineered viruses such as CAV-2 (canine adenovirus 2) or rAAV2 retroviruses.

[0292] As used herein, the terms "chemical genetics method," "chemical genetics approach," and "chemical genetics means" are used interchangeably and refer to a method of treating a subject by introducing a protein into a target cell of the subject and contacting the introduced protein with a ligand that activates or inhibits the introduced protein, wherein activation or inhibition of the introduced protein modulates the activity of the target cell. In some embodiments, the target cell is activated by activating the introduced protein. In some embodiments, the target cell is inhibited by activating the introduced protein. In some embodiments, the target cell is activated by inhibiting the introduced protein. In some embodiments, the target cell is inhibited by inhibiting the introduced protein. In some embodiments, the ligand only activates or inhibits the target cell in the subject and does not activate or inhibit other cells. In some embodiments, the ligand only activates or inhibits the introduced protein in the subject and does not activate or inhibit other proteins. The ligand of the introduced protein can be any chemical substance, such as a small molecule or a peptide. The introduced protein can be an engineered protein or a synthetic protein, and can also be wild type or mutant. In some embodiments, the introduced protein is a G protein-coupled receptor, an ion channel protein, or a variant thereof. Illustrative examples of introduced proteins include, but are not limited to, DREADDs and ligand-gated ion channel (LGIC) proteins. In some embodiments, to use a chemical genetics approach in a subject (e.g., in treating a disease), a DREADD or LGIC is located on the membrane of a target cell in the subject, and a ligand for the DREADD or LGIC is administered to the subject to activate or inhibit the DREADD or LGIC.

[0293] As used herein, the terms "optogenetic method," "optogenetic measure," or "optogenetic means" refer to a method of treating a subject by introducing a light-sensitive protein into the subject's target cells and modulating the activity of the target cells by light of a specific wavelength. In some embodiments, light of a specific wavelength activates the target cells via the light-sensitive protein. In some embodiments, light of a specific wavelength inhibits the target cells via the light-sensitive protein. In some embodiments, light of a specific wavelength activates the target cells via the light-sensitive protein, while another light of another specific wavelength inhibits the target cells via the light-sensitive protein. Illustrative examples of light-sensitive proteins include G protein-coupled light-sensitive proteins and light-gated ion channel proteins. The protein can be an engineered protein or a synthetic protein, and can also be wild type or mutant. In some embodiments, the use of a chemical genetics approach in a subject (e.g., in treating a disease) is to express a G protein-coupled light-sensitive protein or a light-gated ion channel protein in the target cells, and then activate or inhibit the G protein-coupled light-sensitive protein or light-gated ion channel protein by light of a specific wavelength. In some embodiments, the light-sensitive protein or light-gated ion channel protein is located on the membrane of the subject's target cells.

[0294] As used herein, "target neurons" or "target cells" refer to ACC neurons. In some embodiments, "target neurons" or "target cells" are ACC pyramidal neurons. In some embodiments, "target neurons" or "target cells" are BLA-projecting ACC neurons.

[0295] Tetanus neurotoxin, or TeNT, is a potent toxin produced by Clostridium tetani that inhibits neurotransmission. Expression of TENT on neurons can result in long-term inhibition of synaptic release.

[0296] The term "muscimol," also known as agarin or pantherine, is one of the main psychoactive components of the Amanita muscaria plant. Muscimol is a potent and selective stereoselective agonist of the GABAA receptor.

[0297] As an apoptotic protein, activated caspase 3 causes cell apoptosis. In some embodiments, the caspase 3 used in this application is an engineered caspase 3 precursor protein that lacks the cleavage site of the upstream caspase and encodes the cleavage site of the heterologous enzyme tobacco etch virus protease (TEVp). In some embodiments, the caspase 3 used in this application is pro-taCasp3, which is activated by TEVp and converted into taCasp3 that induces apoptosis. In some embodiments, target neurons are destroyed by introducing taCapase3 and TEVp.

[0298] The term "DTR" or "diphtheria toxin receptor" refers to the membrane-anchored form of heparin-binding EGF-like growth factor (HB-EGF precursor). HB-EGF precursor derived from toxin-sensitive animals, such as humans and monkeys, binds to DT-B (one of the two DT components) and functions as a toxin receptor. In some embodiments, DTR is introduced into target neurons in a subject who is insensitive to diphtheria toxin, and the target neurons are destroyed by administering diphtheria toxin to the subject.

[0299] G protein-coupled receptors

[0300] G protein-coupled receptors (GPCRs) are a diverse family of protein receptors that mediate cellular responses to external stimuli, such as Gi / o coupled receptors, Gs coupled receptors, and Gq coupled receptors. G proteins can be excitatory (Gs, G q / 11 , G 12 / 13) or inhibitory (e.g., Gi / o). Here, GPCR, Gi / o coupled receptor, Gs coupled receptor or Gq coupled receptor can be wild-type proteins or their variants. Illustrative examples of GPCR variants include ligand-activated GPCRs, such as DREADD (designer receptors specifically activated by designer drugs) and light-sensitive GPCRs. GPCRs also encompass GPCR variants that are no longer coupled to G proteins due to mutations in G proteins, as long as they still maintain the same stimulation to cells. For example, Rq (R165L) is a variant of a Gq coupled receptor, but is not coupled to a G protein, however, since it still gives neuronal stimulation, as the wild-type Gq coupled receptor does, Rq (R165L) is still included in the Gq coupled receptors.

[0301] The term "Gi / o-coupled receptor," also known as a seven-transmembrane domain receptor, refers to the portion of a GPCR that couples to a Gi / o protein. Gi / o-coupled receptors are a large group of cell surface receptors that detect extracellular molecules and activate cellular responses. Gi proteins inhibit the cAMP-dependent pathway primarily by inhibiting adenylate cyclase activity, reducing cAMP production from ATP, which in turn leads to decreased activity of cAMP-dependent protein kinase. Therefore, the ultimate effect of Gi is to inhibit cAMP-dependent protein kinase. Activation of Gi / o-coupled receptors in neurons inhibits neuronal activation.

[0302] The term "Gs-coupled receptor" refers to a portion of a GPCR that is coupled to a Gs protein. When a ligand activates a Gs-coupled receptor, the activated receptor acts as a guanine nucleotide exchange factor to promote the release of GDP from Gα and the binding of GTP to Gα, thereby driving the separation of GTP-bound Gα from Gβγ. In particular, the GTP-bound activated Gsα binds to adenylate cyclase to produce the second messenger cAMP, which in turn activates cAMP-dependent protein kinase (also known as protein kinase A or PKA). Activation of Gs-coupled receptors in neurons leads to neuronal activation.

[0303] The term "Gq-coupled receptor" refers to the portion of a GPCR that is coupled to a Gs protein. Activation of a Gq-coupled receptor stimulates the enzyme phospholipase Cβ (PLCβ), which leads to the hydrolysis of the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate. This results in the formation of two classic second messengers, inositol 1,4,5-triphosphate (which releases Ca from intracellular stores) and 2+ ) and diacylglycerol (which activates protein kinase C). Activation of Gq-coupled receptors in neurons leads to neuronal activation.

[0304] Ligand-activated GPCR

[0305] The term "ligand-activated GPCR" encompasses all GPCRs that can be activated by chemical substances. The activity of target cells can be modulated by introducing a ligand-activated GPCR into neurons and / or by introducing a ligand for a ligand-activated GPCR into target neurons. In particular, when the ligand does not activate endogenous receptors other than the introduced GPCR, specific regulation of target cells is possible.

[0306] Illustrative examples of ligand-activated GPCRs include, but are not limited to, serotonin receptors, muscarinic acetylcholine receptors, adenosine receptors, adhesion-like GPCRs, adrenergic receptors, angiotensin receptors, apelin receptors, bile acid receptors, bombesin receptors, bradykinin receptors, calcitonin receptors, calcium-sensing receptors, cannabinoid receptors, chemokine receptors, cholecystokinin receptors, frizzled-like GPCRs, complement peptide receptors, corticotropin-releasing factor receptors, dopamine receptors, endothelin receptors, estrogen (G protein-coupled) receptors, formyl peptide receptors, free fatty acid receptors, GABAB receptors, galanin receptors, ghrelin receptors, glucagon receptor family, glycoprotein hormone receptors, gonadotropin-releasing hormone receptors, GPR18, GPR55, and GPR119, histamine receptors, hydroxycarboxylic acid receptors, kisspeptin receptors, receptor), leukotriene receptors, lysophospholipid (LPA) receptors, sphingosine 1-phosphate (S1P) receptors, melanin-concentrating hormone receptors, melanocortin receptors, melatonin receptors, metabotropic glutamate receptors, motilin receptors, neuromedin U receptors, neuropeptide FF / neuropeptide AF receptors, neuropeptide S receptors, neuropeptide W / neuropeptide B receptors, neuropeptide Y receptors, neurotensin receptors, opioid receptors, orexin receptors, ketoglutarate receptors, P2Y receptors, parathyroid hormone receptors, peptide P518 receptors, platelet-activating factor receptors, prokineticin receptors, prolactin-releasing peptide receptors, prostanoid receptors, protease-activated receptors, relaxin family peptide receptors, somatostatin receptors, succinate receptors, tachykinin receptors, thyrotropin-releasing hormone receptors, trace amine receptors, urotensin receptors, vasopressin and oxytocin receptors, and VIP and P ACAP receptors.

[0307] As used herein, the term "DREADD" or "designer receptor activated only by designer drugs" refers to a class of artificially engineered GPCRs used in the field of chemical genetics or chemical genetics methods, and selectively activated by specific ligands. DREADDs can be generated from any GPCR, including the above-mentioned GPCRs. A DREADD ligand is a designer drug that specifically activates a DREADD. Illustrative examples of DREADDs include inhibitory DREADDs such as hM4Di, hM4Dnrxn, and KORDi, and excitatory DREADDs such as hM3Dq, Rq(R165L), and hM3Ds.

[0308] hM3Dq is a variant of the human M3 muscarinic (hM3) receptor. It can be activated by the inert clozapine metabolite clozapine-N-oxide (CNO), participating in the Gq signaling pathway. Gq signaling releases intracellular calcium stores and enhances neuronal excitation. As a result, hM3Dq-expressing neurons treated with CNO have a significantly increased firing rate. In addition to CNO, other DREADD ligands, such as compound 21, DCZ, and clozapine, can also activate hM3Dq.

[0309] Rq(R165L) is a variant of the human M3 muscarinic receptor that couples to β-arrestin to activate G protein-independent noncanonical GPCR signaling.

[0310] hM3Ds is a Gs-coupled DREADD generated by hM3Dq. In the presence of DREADD ligands (e.g., CNO, C21, DCZ), hM3Ds increases cAMP production but does not increase IP3 or intracellular Ca2+.

[0311] hM4Di is a variant of the human M4 muscarinic (hM4) receptor. It can be activated by the inert clozapine metabolite clozapine-N-oxide (CNO) and participates in the Gi signaling pathway. Gi signaling in neurons opens potassium channels, which leads to the influx of potassium ions, reducing the resting membrane potential of neurons and their ability to depolarize. Therefore, neurons expressing hM4Di treated with CNO have a dramatically reduced firing rate. In addition to CNO, other DREADD ligands, such as compound 21, DCZ, and clozapine, can also activate hM4Di.

[0312] hM4Dnrxn is an axon-selective variant of hM4Di. hM4Dnrxn was constructed by adding the intracellular amino acid sequence of neurexin-1 (aa1425-1479) to the C-terminus of hM4D, which also contains a C-terminal hemagglutinin (HA) tag.

[0313] KORDi is a variant of the Gi-coupled human kappa-opioid receptor, and salvinorin B (SALB) is an inert ligand of KORDi.

[0314] Non-limiting examples of ligand-activated GPCRs suitable for use as described herein include: CHRMI; GNRHR; GPR73; GPR45; PTHRI; CHRM2; GNRHR2; GPR73; GPR63; PTHR2; CHRM3; HRHI; GPRIO; GPR83; SCTR; CHRM4; HRH2; F2R; PGR15; ADCYAPIRI; CHRM5; HRH3; F2RL1; PGR15L; VIPRI; ADORA1; HRH4; F2RL2; GPR103; VIPR2; ADORA2A; FSHR 93; F2RL3; GPR103L; BAil; ADORA2B; LHCGR; P2RY1; GRCA; BAI2; ADORA3; TSHR; P2RY2; PGRI; BAB; P2RY12; GPR54; P2RY4; HGPCRII; CD97; GPR105; LTB4R; P2RY6; SALPR; EMRI ; GPR86; LTB4R2; P2RY11; MASI; EMR2; GPR87; MRGXI; LGR7; GPR90; EMR3; ADRAIA; MRGX2; LGR8; P2Y5; PGR16; ADRAIB; 275; LEC2; ADRA2A; MRGD; HTRIB; FKSG79; LEC3; ADRA2B; MrgAl; HTRID; PGR2; CELSRI; ADRA2C; MrgA2; 21; MrgA4; HTR2A; CMKLRI; GPR64; ADRB3; MrgA5; HTR2B; EBI2; PGR17; ADMR; MrgA6; HTR2C; GPCR150; DJ287G l4; C3AR1; MrgA7; HTR4; GPRI; KIAA0758; C5R1; MrgA8; HTR5A; GPR15; PGR18; GPR77; MrgA9; HTR5B; GPR17; P GR19; AGTRI; MrgAIO; HTR6; GPR18; PGR20; AGTR2; MrgAll; HTR7; GPR19; TEM5; AGTRLI; MrgA12; SSTRI; GPR2 0;KIAA1828;BRS3;MrgA13;SSTR2;GPR22;PGR21;GRPR31;MrgA14;SSTR3;GPR25;ETL;NMBR;MrgA15;SSTR4;<h2 style=";text-align:left;direction:ltr">GPR30;FLJ14454;BDKRBI;MrgA16;SSTR5;GPR31;GPR56;BDKRB2;MrgA19;G2A;GPR32;OAI;C NRI;MrgBl;GPR4;GPR33;PGR22;CNR2;MrgB2;GPR65;GPR34;PGR23;CCRI;MrgB3;GPR68;GPR 35;PGR24;CCR2;MrgB4;30EDGI;GPR39;PGR25;CCR3;MrgB5;EDG2;GPR40;PGR26;CCR4;MrgB 6;EDG3;GPR44;PGR27;CCR541;MrgB8;EDG4;GPR55;VLGRI;CCR6;MrgBIO;EDG5;GPR61;CCR7 43;MrgBll;EDG6;GPR62;CCR8;MrgB13;EDG7;GPR75;CCR9;GPR24;EDG8;GPR80;GPR2;SLT;TACRI;G PR82;CASR;CCRLI;MCIR;TACR2;GPR84;GABBRl;CCRL2;MC2R;TACR3;GPR88;GPR51;CCBP2;MC3R;TR HR;GPR91;GPRC5B;CMKBRlLl;MC4R;TRHR2;GPR92;GPRC5C;CMKBR1L2;MC5R;GPR57;GPRlOl;GPRC5D ;C 19;GRM2;IL8RB;GPR66;TAR2;HUMNPIIY20;GRM3;GPR9;NMU2R;TAR3;MRG;GRM4;CXCR4;NPFFlR;TAR4;MRGE;GRM5;BLRl;GPR74;GPR102;MRGF;GRM6;CXCR6;GPR7;TA7;MRGG;GRM7;CCKAR;GPR8;TA8;OP N3;GRM8;CCKBR;NPYlR;TAIO;OPN4;GPRC6A;CYSLTl;NPY2R;TAll;PGR4;PGR28;CYSLT2;PPYRl;TA1 2;PGR5;DRDl;NPY5R;TA14;PGR6;DRD2;NPY6R;TA15;PGR7;DRD3;NTSRl;GPR14;PGR8;DRD4;NTSR2;A VPRlA;PGRlO;FZDl;DRD5;OPRDl;AVPRlB;PGRll;FZD2;FY;OPRKl;AVPR2;PGR12;FZD3;TG1019;OPRMl;OXTR;PGR13;FZD4;HM74;O PRL1;GPR48;PGR14;FZD5;GPR81;OPNlLW;GPR49;RDCl;FZD6;EDNRA;OPNlMW;LGR6;RE2; FZD7; ; CALCRL; FPR-RS3; PTGER1; GPR21; CRHR1; FPR-RS4; PTGER2; GPR52; CRHR2; GALR1; PTGER3; GPR26; GIPR; TM7SF1; GALR2; PTGER4; LPlR; TM7SF1L2; GHSR; PTGIR; GPR37Ll; GLP2R; TM7SF3; GPR38; TBXA2R; GPR41; GHRHR; T PRA40; GPR43; hM1Dq, hM2Di, hM5Dq, hM4Di, hM4Dnrxn, KORDi, hM3Dq, Rq(R165L) and hM3Ds. ;

[0315] In the present application, any suitable ligand of a GPCR can also be introduced into neurons alone or together with a GPCR to modulate neuronal activity, for example, allatostatin for AlstR; CNO, clozapine, perampelamine, olanzapine, alosetron, fluperampelamine, and N4′-alkyl-substituted CNO analogs for hM4Di, hM3Dq, and GsD; and salvinorin B for KORD.

[0316] Photosensitive GPCR

[0317] Photosensitive GPCRs are GPCRs that are activated or inhibited by certain light signals. Illustrative examples of photosensitive GPCRs include opsin proteins and variants thereof. Opsin proteins use light-absorbing chromophores to receive light signals. Through the cis-trans isomerization of the chromophore, light energy is converted into chemical free energy, which in turn is used for conformational changes in the protein to activate the G protein. In some embodiments, the photosensitive GPCR is derived from the opsin of the opsin protein. Exemplary photosensitive GPCRs include Lamplight (lamprey lateral opsin), rod opsin, cone opsin, μ opioid receptor-rod opsin chimera, Opto-MOR, cOpn5, hOPN5, and JellyOp (opsin from Charybdea rastonii).

[0318] Ion channel proteins

[0319] As used herein, " ion channel " or " ion channel protein " is a protein molecule that spans the cell membrane, which allows ions to pass from one side of the membrane to the other side. In some embodiments, ion channel proteins include voltage-gated ion channels, ligand-gated ion channels, light-gated ion channels, mechanically sensitive ion channels and thermosensitive ion channels (e.g., TRPV4). In some embodiments, ion channel proteins include cation (e.g., calcium, potassium, sodium, proton) channel proteins, anion (e.g., chloride) channel proteins and non-selective ion channel proteins.

[0320] Here, when referring to an ion channel, a cation channel, or an anion channel, wild type or variants thereof are encompassed.

[0321] Ligand-gated ion channel proteins

[0322] Ligand-gated ion channel (LGIC) proteins refer to a large group of transmembrane proteins that allow or inhibit the passage of ions when they interact with specific chemicals (ligands). The binding of ligands to ion channel proteins directly results in the opening or closing of the channel. Ligand binding causes a dramatic change in the permeability of the channel to one or more specific ions; when the channel is inactive, effectively no ions can pass through the channel, but when ligands are bound, up to 10 ions per second can be allowed to pass through the channel. 7 ions or even more ions pass through.

[0323] In some embodiments, one or more subunits of the LGIC or variant thereof have been engineered to specifically bind a heterologous ligand, an exogenous ligand, and / or a synthetic ligand. In some embodiments, the specific ligand does not activate an endogenous receptor, but rather activates the engineered LGIC.

[0324] Illustrative examples of LGICs suitable for use in particular embodiments include, but are not limited to, 5-HT3 receptors, acid-sensing ion channels (ASICs), epithelial sodium channels (ENaCs), GABAA receptors, glycine receptors, ionotropic glutamate receptors, IP3 receptors, nicotinic acetylcholine receptors, P2X receptors, ryanodine receptors, zinc-activated channels (ZACs), and variants thereof.

[0325] Non-limiting examples of LGICs and their specific ligands used in the methods of the present application include those described in Frazier et al., Journal of Biological Chemistry, 2012; Magnus et al., Science, 2011; Lynag and Lynch, and Journal of Biological Chemistry, 2010. In some embodiments, the specific ligands of LGICs used in the methods of the present application include: ivermectin, moxidectin, selamectin, doramectin, emamectin, eprinomectin, and abamectin for GluClα, GluClβ, or GlyR-M; 22S PSAM-5HT3HC, used for PSAM GlyR PSEM 89S , PSEM for PSA nAChR 9S, capsaicin for TRPV1, and zolpidem for GABBA. In some embodiments, a LGIC suitable for use in the methods of the present application further comprises: HTR3A; HTR3B; HTR3C; HTR3D; HTR3E; ASIC1; ASIC2; ASIC3; SCNN1A; SCNN1B; SCNN1D; SCNN1G; GABRA1; GABRA2; GABRA3; GABRA4; GABRA5; GABRA6; GABRB1; GABRB2; GABRB3; GABRG1; GABRG2; GABRG3; GABRD; GABRE; GABRQ; GABRP; GABRR1; GABRR2; GABRR3; GLRA1; GLRA2; GLRA3; GLRA4; GLRB; GRIA1; GRIA2; GRIA3; GRIA 4; GRID1; GRID2; GRIK1; GRIK2; GRIK3; GRIK4; GRIK5; GRIN1; GRIN2A; GRIN2B; GRIN2C; GRIN2D; GRIN3A; GRIN3B; ITPR1; ITPR2; ITPR3; CHRNA1; CHRNA2; CHRNA3; CHRNA4; CHRNA5; CHRNA6; CHRNA7; CHRNA9; CHRNA10; CHRNB1; CHRNB2; CHRNB3; CHRNB4; CHRNG; CHRND; CHRNE; P2RX1; P2RX2; P2RX3; P2RX4; P2RX5; P2RX6; P2RX7; RYR1; RYR2; RYR3; and ZACN.

[0326] Light-gated ion channel proteins

[0327] The ion channel of light-gating is a group of ion channel proteins, and its opening or closing is the response to light. The ion channel of light-gating used herein can be naturally occurring or artificially synthesized. The example illustrative example of the ion channel of light-gating includes channel rhodopsin and its variant. As used herein, " ion channel protein of light-gating " also includes those based on any other type of ion channel protein engineering, such as ligand-gated ion channel (for example, nicotinic acetylcholine receptor) and voltage-gated ion channel. In addition, some ligands of ion channel of ligand-gating (for example, capsaicin and some cooling compounds, including menthol and icilin (icilin)) include potential receptor sites of light-labile blocking groups. Light-labile blocking groups and such receptor binding can cause the light-sensitive ligand of ion channel of ligand-gating, wherein light is by releasing active ligand as the indirect trigger of ion channel of ligand-gating.

[0328] Illustrative examples of light-gated ion channel proteins include light-gated anion channel proteins (e.g., eNpHR3.0, Arch, eBR, iC1C2, ChloC, ACRs, GtACR1, GtACR2) and light-gated cation channel proteins (e.g., ChR2 and variants thereof, such as ChR2(H134R), ChETA, ReaChR, bReaChES, Chrimson, ChrimsonR, C1C2, C1V1, C1V1(t), C1V1(t / t), oChIEF, ChRmine, ChRmine2.0, ChRger1, ChRger2, ChRger3, CheRiff and Chronos and variants thereof, such as CsChR, CoChR, VChR1, CheRiff).

[0329] In some embodiments, light-gated ion channel proteins are activated by light of a specific wavelength. In some embodiments, light-gated ion channel proteins are inhibited by light of a specific wavelength. In some embodiments, light-gated ion channel proteins are activated by light of a specific wavelength and inhibited by light of another wavelength. For example, combinations of light-gated ion channel proteins and specific wavelengths that inhibit neuronal activation include: Arch and 566 nm, eBR and 560 nm, iC1C2 and 475 nm, ChloC and 470 nm, etc.

[0330] Introducing the compound into target neurons

[0331] In some aspects of the present application, in a method for treating depression or anorexia, one or more compounds are introduced into target neurons to inhibit the activation or signaling of target neurons or to destroy neurons. In some aspects of the present application, in a method for treating obesity or bulimia, one or more compounds are introduced into target neurons to activate target neurons, or to enhance the activation of target neurons. In some aspects of the present application, in a method for alleviating or exacerbating reward devaluation, one or more compounds are introduced into target neurons to modulate the activity of target neurons.

[0332] Any suitable means for introducing a compound into a subject's neurons that can be used herein include physical methods, biological methods, chemical methods, or combinations thereof. In some embodiments, a compound can be introduced into a subject's target neurons by any method of administration. As used herein, the term "administration" or "administering" can refer to administration that is: oral, topical, intravenous, subcutaneous, transdermal, transdermal, intramuscular, intraarticular, parenteral, intraarteriole, intradermal, intraventricular, intraosseous, intraocular, intracranial, intraperitoneal, intralesional, intranasal, intracardial, intraarticular, intracavernous sinus, intrathecal, intravitreal, intracerebral ventricular, intratympanic, intracochlear, rectal, vaginal, by inhalation, by catheter, stent, or via an implanted reservoir or other device, actively or passively (e.g., by diffusion) administering the composition to the perivascular space and adventitia.

[0333] In some embodiments, the compound is a protein, and the protein is introduced by introducing a nucleic acid encoding the protein into the target neurons to express the protein. In some embodiments, the protein is a G-coupled receptor, an ion channel protein, an apoptotic protein, a toxin receptor or a toxic protein. In some embodiments, the expression of the protein or the insertion of the nucleic acid encoding the protein into the genome is controlled by one or more regulatory elements, such as promoters, enhancers, untranslated regions (UTRs), Kozak sequences, polyadenylation signals, additional restriction enzyme sites, multiple cloning sites, endogenous ribosome entry sites (IRES), recombinase recognition sites (e.g., LoxP, FRT and Att sites) or their recombinases, guide RNA target sites, stop codons, transcription termination signals, and polynucleotides encoding self-cleavage polypeptides, epitope tags, etc.

[0334] As is well known in the art, methods that can be used herein to accurately introduce proteins into neurons. Specifically, the combination of a nucleic acid encoding a protein and one or more compounds that contribute to the recombinant nucleic acid into the neuronal genome is divided into at least two parts, which are injected into the neuronal cell body and axon terminals respectively; and the part injected into the neuronal cell body is anterograde in the neuron, while the part injected into the axon terminals is retrograde in the neuron. Vectors and modifications can be used to help the combination enter the neuron and travel in the desired direction. In some embodiments, a recombinase system (e.g., Cre / loxP system) is used to specifically introduce a protein into the target neuron. In some embodiments, the nucleic acid encoding the protein is flanked by a recombinase recognition site and introduced into the target neuron from the neuronal cell body, and the recombinase (or nucleic acid encoding the recombinase) is introduced into the target neuron from the axon terminals. In some embodiments, the nucleic acid encoding the protein and the recombinase recognition site are anterograde in the target neuron, and the recombinase (or nucleic acid encoding the recombinase) is retrograde in the target neuron. In some embodiments, the nucleic acid encoding the protein and the recombinase recognition site are introduced into the target neuron from the axon terminal, and the recombinase (or nucleic acid encoding the recombinase) is introduced into the target neuron from the neuronal cell body. In some embodiments, the nucleic acid encoding the protein and the recombinase recognition site are retrograde in the target neuron, and the recombinase or recombinase (or nucleic acid encoding the recombinase) is anterograde in the target neuron. For example, in some embodiments, the target neuron is a BLA-projecting ACC neuron, the nucleic acid encoding the protein and the recombinase recognition site are injected on both sides of the BLA, and the recombinase (or nucleic acid encoding the recombinase) is injected on both sides of the ACC. In some embodiments, the target neuron is a BLA-projecting ACC neuron, the nucleic acid encoding the protein and the recombinase recognition site are injected on both sides of the ACC, and the recombinase (or nucleic acid encoding the recombinase) is injected on both sides of the BLA. Any vector or carrier that can carry a compound (e.g., a protein or nucleic acid) to anterograde or retrograde in neurons is applicable herein. In some embodiments, the vector or carrier is a viral vector. In some embodiments, exemplary viral vectors include those derived from adenovirus, retrovirus (e.g., lentivirus), parvovirus (e.g., adeno-associated virus (AAV), AAV-adenovirus chimeric virus, and adenovirus), herpes simplex virus-1 (HSV-1), rabies virus (RV), and pseudorabies virus (PRV). In some embodiments, the virus used to carry the compound for retrograde transport in neurons is a retrograde labeled adeno-associated virus (AAV2-retro). In some embodiments, the protein is expressed in target neurons of a subject, and its ligand is administered to the subject. Example

[0335] Example 1. Methods for testing reward devaluation and depressive states

[0336] Guided by the clinical theory that reward devaluation contributes to anhedonia in human MDD patients, the inventors first investigated whether the process of reward devaluation might be altered in depressive states. The inventors used chronic restraint stress (CRS) to induce depressive-like behavior in mice, which was assessed as an increase in immobility time in the forced swim test (an indicator of behavioral despair) and a decrease in sucrose preference in the sucrose preference test (an indicator of anhedonia) ( Figure 1 a, b and Figure 7 The rapid-acting antidepressant ketamine reversed two indicators of depression in CRS-treated mice one hour after drug administration ( Figure 1 a,b).

[0337] To explore potential changes in reward processing, the inventors conducted a more detailed analysis of the effect of sucrose preference on reward intake and consumption rate over time. In the sucrose preference test, CRS mice consumed significantly less sucrose than non-CRS control mice, while no difference in water intake was detected ( Figure 1 c). Monitoring licking rates using a lick meter showed that although both control and CRS mice initially exhibited high sucrose intake rates that declined over time, the intake rate in CRS mice declined significantly faster than that in control mice ( Figure 1 d).

[0338] Given that reward value is dynamically adjusted according to the animal's physiological needs, the inventors subsequently deprived mice of water to make water a reward. Compared with control mice, CRS mice consumed significantly less water and showed a faster decline in water intake rate, and both processes were reversed by ketamine treatment of CRS mice ( Figure 1 e, f). When mice with prior free access to water were given a choice of different reward values ​​(water, 1% sucrose, and 10% sucrose), they exhibited a higher intake rate for the higher value reward ( Figure 7 d), suggesting that reward intake rate can serve as an informative indicator of reward value. Thus, the faster decline in reward intake rate observed in CRS mice clearly corresponds to the accelerated reward devaluation in both the sucrose preference test and the water intake test.

[0339] The inventors then directly compared the performance of CRS mice with that of untreated (hereinafter referred to as "healthy") control mice in a sensory-specific satiety-based reward devaluation test ( Figure 1 g and Figure 7e). During the baseline test phase, the inventors identified the reward option (sucrose or pear juice) that the individual mice slightly preferred. On the day of the reward devaluation test, the mice were given free access to the preferred reward for 30 minutes (selective satiety) and then given the two choices two hours later.

[0340] By adjusting the reward reaccumulation schedule to control the level of reward devaluation, the inventors revealed the influence of the animals' depressive state on their sensitivity to reward devaluation. When mice had free access to all rewards (sucrose, pear juice, and water) during the accumulation period following the pre-test reward, both healthy control mice and CRS mice reduced their preference for the most recently consumed reward ( Figure 7 e, f). Therefore, the inventors call it the "sensitive behavior paradigm". Without reward reaccumulation ("insensitive behavior paradigm"), CRS mice, but not healthy control mice, showed reward devaluation ( Figure 1 g, h). In addition, ketamine reversed the reward devaluation deficit in CRS mice ( Figure 1 h). These results indicate that CRS-induced depression increases the sensitivity of animals to reward devaluation. Therefore, a method for testing reward devaluation and depression states was established, and changes in reward intake rate (e.g., expressed as sucrose / water preference) and immobility time reflect the reward devaluation rate and degree of depression in mice.

[0341] Example 2. Modulation of reward devaluation by ACC pyramidal neurons

[0342] The inventors of this application demonstrated that the activity of ACC neurons underlies reward devaluation and, by modulating the activity of ACC neurons, they achieved for the first time specific control of reward intake rate and reward devaluation in healthy mice and CRS mice. Specifically, the modulation methods included ablation, inhibition, and stimulation of ACC neurons.

[0343] To genetically ablate ACC pyramidal neurons marked by the expression of calcium / calmodulin-dependent protein kinase II (CaMK2), the inventors bilaterally infused the AAV-FLEX-taCapase3-TEVp vector into the ACC of CaMK2-Cre mice (JAX-005359, purchased from Jackson Laboratory) to induce caspase 3 expression and thereby lead to programmed cell death of ACC pyramidal neurons ( Figure 2 a, b). The inventors then examined whether such ablation of ACC pyramidal neurons would affect reward devaluation in healthy mice. Using a sensitive reward devaluation paradigm, the inventors found that reward devaluation was significantly attenuated in mice expressing caspase 3 compared to control mice expressing mGFP ( Figure 2 c), thus indicating that the observed reward devaluation requires the presence of ACC pyramidal neurons.

[0344] To inhibit the activity of ACC pyramidal neurons, we specifically expressed a designer drug-activated inhibitory receptor (DREADD), hM4Di, in the ACC of CaMK2-Cre mice by infusing the AAV-DIO-hM4Di vector, which allowed us to use the otherwise inert ligand clozapine-N-oxide (CNO; Figure 8 a) Reversible silencing of ACC pyramidal neuron activity. This chemogenetic approach abolished reward devaluation in mice expressing hM4Di but not in mGFP controls ( Figure 2 d, e) Reward devaluation in mice. These results support that reward devaluation can be controlled at the behavioral level by manipulating the activity of ACC pyramidal neurons in healthy mice.

[0345] Dysfunction of reward devaluation has been shown to affect reward intake and is clinically associated with anorexia and obesity. Therefore, the inventors investigated whether manipulation of ACC neurons could alter reward intake in healthy mice. The inventors found that genetic ablation of ACC pyramidal neurons resulted in significantly higher water intake and slower water intake decay ( Figure 2 fh). The inventors detected similar effects of chemogenetic inhibition of ACC pyramidal neurons on the day of CNO administration ( Figure 8 b, c).

[0346] In contrast, chemogenetic activation of ACC pyramidal neurons inhibited reward intake. The inventors expressed hM3Dq, an excitatory DREADD, in ACC pyramidal neurons to enable CNO-mediated reversible activation of these neurons ( Figure 8 a). Following chemogenetic activation of ACC pyramidal neurons, mice ingested significantly less water reward and displayed a significantly faster decline in reward intake rate compared to control mice expressing mGFP ( Figure 2 i, j, and Figure 8 d) These observations support that ACC pyramidal neurons negatively control reward intake. The results suggest that ablation or inhibition of ACC pyramidal neurons can alleviate anorexia, whereas activation of ACC pyramidal neurons reduces obesity by reducing appetite.

[0347] Based on the results from our chemical genetic manipulation of ACC pyramidal neurons, the inventors found that inducing neurochemical damage to ACC neurons leads to long-term reward overconsumption and ultimately to alarming obesity. Infusion of the excitotoxin kainic acid (KA) into the bilateral ACC resulted in widespread loss of ACC neurons and resulted in significantly higher water intake in the water intake test ( Figure 9 When fed a high-fat diet, the ACC-lesioned mice consumed significantly more food than saline control mice, continued to gain significantly more weight, and became extremely obese after three weeks ( Figure 2 kn). Therefore, disrupting ACC neurons can lead to acute and long-term changes in reward intake. Because this result is based on a widespread and random loss of ACC neurons (inadvertent loss of excitatory neurons), this suggests that inhibiting or disrupting the entire ACC enhances reward intake, while activating the entire ACC accelerates reward devaluation.

[0348] The inventors next tested how changing the activity of ACC pyramidal neurons affects depression-like behavior. In CRS mice, ACC neurons are prone to overexcitation. After forced swimming (FST) stimulation, CRS mice had significantly more immediate early gene c-fos positive neurons in the ACC region compared to normal mice (control), ( Figure 6 , which revealed the relationship between overexcited ACC neurons and depression. Chemical genetic inhibition of ACC pyramidal neurons significantly reduced reward devaluation ( Figure 2 o, p). In addition, such inhibition significantly reduced immobility time in the forced swim test and significantly increased sucrose preference ( Figure 2 q, r), the results showed improvement in behavioral despair and anhedonia. Here, compound 21 (C21), another inert ligand of hM4Di, was also tested. The results were the same ( Figure 5 In contrast, chemogenetic activation of ACC pyramidal neurons in healthy mice expressing hM3Dq significantly reduced sucrose preference ( Figure 2 Together, these results suggest that inhibition of ACC pyramidal neurons reduces depression-like behaviors in CRS mice, whereas activation of these cells induces depression-like behaviors in healthy mice.

[0349] In summary, any means of activating or inhibiting (or even partially or completely destroying) the ACC (especially pyramidal neurons in the ACC) can modulate reward devaluation and therefore help alleviate symptoms or diseases affected or induced by changes in reward devaluation (including but not limited to depression, depressive disorders, anorexia and obesity).

[0350] Example 3. Encoding reward devaluation by weakening reward inhibition

[0351] The strong behavioral effects of manipulating ACC pyramidal neuron activity raise the question of how these neurons represent reward signals in healthy and depressed states. Previous studies have revealed a variety of response patterns to rewarding and aversive stimuli in the ACC, which may reflect the presence of pyramidal neurons and several types of interneurons. The inventors of the present application focused on examining the dynamic response profiles of ACC pyramidal neurons (which are the main neurons in the cortex) at the neuronal population level and the single cell level.

[0352] The inventors first performed fiber photometry of Ca2+ dynamics ( Figure 3a), this method offers the advantage of reporting population neuronal activity from genetically identified cell types in behaving animals. The inventors of the present application infused the adeno-associated virus (AAV) vector AAV-DIO-GCaMP6m into the ACC of CaMK2-Cre mice to express the genetically encoded Ca2+ indicator GCaMP6m in pyramidal neurons ( Figure 10 a). In healthy mice actively seeking water rewards and high-fat food pellets, reward depletion leads to a decrease in the timing of Ca2+ signaling ( Figure 10 bf). When the inventors of the present application delivered rewards of different values ​​(water, 1% sucrose, and 10% sucrose) into the oral cavity of water-deprived mice through a buccal fistula, they observed that rewards of relatively higher values ​​triggered stronger inhibition of ACC pyramidal neurons ( Figure 3 bd and Figure 11 ae).

[0353] In contrast to rewarding stimuli, aversive stimuli activate ACC pyramidal neurons at a population level. Both oral delivery of the bitter agent quinine and aversive foot shocks elicit excitatory responses ( Figure 3 c and Figure 12 ac). Thus, these experiments with rewarding and aversive stimuli indicate that the strength of inhibition and excitation of ACC pyramidal neurons correlates with reward and aversive value, respectively ( Figure 3 d). Following auditory fear conditioning, an auditory tone was paired with a footshock, and the footshock-conditioned tone elicited a strong activation of these neurons, accompanied by a decrease in movement (a measure of fear) ( Figure 12 These results support that ACC pyramidal neurons negatively encode reward valence, rather than sensory salience or action vigor.

[0354] The inventors conducted two sets of recordings to investigate how ACC pyramidal neurons encode reward devaluation. In the first set of recording experiments, the inventors tracked Ca2+ dynamics during repeated oral delivery of water rewards for 100 trials ( Figure 3 e). This passive reward acquisition process minimizes the potential effects of physical effort associated with active reward seeking. In water-deprived mice, ACC pyramidal neurons were strongly inhibited by reward intake during the initial phase of the water delivery task; the strength of inhibition gradually decreased in response to repeated reward delivery, reaching a point of essentially zero inhibition after approximately 80 delivery trials ( Figure 3 f, g, and Figure 13 a). Consistent with the behavioral hypersensitivity of CRS mice to reward devaluation ( Figure 1 h) The reward inhibition strength of ACC pyramidal neurons in the CRS group was weaker than that in the healthy control group, and the decay rate was significantly faster ( Figure 3 g and Figure 13 a).

[0355] In a second set of recordings, the inventors combined a reward devaluation assay with fiber photometry via intraoral delivery of a liquid reward to test whether the attenuation of reward inhibition differed between previously consumed ("devalued") rewards and non-devalued rewards. The inventors found that selective satiety resulted in a significantly greater reduction in reward inhibition for devalued rewards compared to non-devalued rewards ( Figure 3 h and Figure 13 bd). Therefore, the results of these two fiber photometry experiments indicate that reward inhibition of ACC pyramidal neurons becomes gradually weakened after reward consumption. In addition, the smaller amplitude and faster decay of reward inhibition of ACC pyramidal neurons indicate hypersensitivity to reward devaluation in CRS mice.

[0356] Given that fiber photometry only reports group activity, and that individual ACC neurons are known to display a variety of response patterns, the inventors of the present application also investigated the response patterns of individual cells. Using a head-mounted miniature microscope and an implanted gradient index (GRIN) lens, the inventors of the present application performed Ca2+ imaging of individual ACC pyramidal neurons from healthy mice ( Figure 3 i and Figure 14 Similar to the fiber photometry results in the water delivery task, many ACC pyramidal neurons were strongly inhibited by reward delivery and strongly stimulated by footshock ( Figure 3 j and Figure 14 Importantly, both the number and the average strength of inhibited neurons were significantly reduced across the test blocks, each consisting of 20 water delivery trials ( Figure 3 km). A small number of ACC pyramidal neurons showed excitatory responses to reward; however, no differences were detected in the proportion of activated cells or the magnitude of the excitatory responses across sessions ( Figure 14 k, l). These imaging and fiber photometry results collectively support that reward devaluation is encoded by a reduction in reward inhibition.

[0357] The results of this example indicate that regulating the inhibitory state of the entire ACC or the population activity of ACC pyramidal neurons, or the number of ACC pyramidal neurons that can be inhibited by reward, can modulate reward devaluation and treat symptoms and diseases associated with reward devaluation.

[0358] Example 4. Regulation of reward devaluation and depression by BLA-projecting ACC neurons

[0359] ACC pyramidal neurons project axon terminals to many downstream brain regions. The inventors investigated which ACC projections may play a role in controlling reward intake and devaluation. By examining the axonal projection profiles of ACC pyramidal neurons based on mGFP expression, the inventors demonstrated that these neurons centrally innervate several cortical regions, the hypothalamus, the thalamus, and the brainstem ( Figure 4 a). By analyzing the axonal projection map of sparsely labeled single ACC pyramidal neurons at the whole-brain level ( Figure 4 b), the inventors divided the reconstructed neurons into five types, each type showing some different projection targets ( Figure 4 c, d; Figure 15 ).

[0360] The inventors verified the type-specific projection maps at the population level by injecting retrograde rAAV2-retro-Cre vectors into different representative downstream regions and delivering Cre-dependent AAV-DIO mGFP into the ACC of C57BL / 6N mice (purchased from Weitong Lihua, Beijing, China). Figure 4 e and Figure 16 a). The inventors note that type I to V pyramidal neurons are characterized by strong projections to the ventral tegmental area (VTA), BLA, nucleus accumbens (NAc), medial septum (MS), and OFC, respectively; however, some examined brain regions [e.g., dorsal striatum (DS)] receive input from multiple cell types ( Figure 4 d, e). Within the ACC, the cell bodies of the five neuronal types exhibit distinct but overlapping distributions within subregions and layers ( Figure 16 bf). These findings suggest the clear utility of monitoring projection areas (i.e., rather than ACC subregions) to investigate the function of each ACC pyramidal neuron type.

[0361] The use of a retrograde labeling approach allowed us to examine the effects of ablation of specific subpopulations of ACC pyramidal neurons on reward intake. The inventors injected the AAV-FLEX-taCapase3-TEVp vector bilaterally into the ACC and simultaneously injected the rAAV2-retro-Cre vector into one of 11 downstream regions (including the five representative target regions of the aforementioned ACC subpopulations) to induce projection-specific damage to ACC neurons. The inventors found that targeting neurons projecting to the BLA more significantly enhanced water reward intake ( ) than targeting neurons projecting to the other four downstream brain regions defined by the inventors. Figure 4 f, g, and Figure 17 Note that ablation of BLA-projecting ACC neurons abolishes reward devaluation in healthy mice ( Figure 4 h). These results suggest that the neuronal projection pathway from the ACC to the BLA normally inhibits reward intake and promotes reward devaluation.

[0362] Findings from projection-specific ablation experiments prompted us to investigate how chemogenetic activation of BLA-projecting ACC neurons might affect reward-related behaviors. Figure 18 ac), the inventors applied CNO and tested its effect in a reward intake test. Similar to activation of whole ACC pyramidal neurons, chemogenetic activation of BLA-projecting ACC neurons resulted in significantly lower water reward intake, weaker sucrose preference, and longer immobility time in the forced swim test in mice expressing hM3Dq compared to control mice expressing mGFP ( Figure 4 i and Figure 18 d, e). Simulating the long-term effects of CRS, multiple administrations of CNO over three days resulted in significantly stronger reward devaluation in mice expressing hM3Dq than in mice expressing mGFP ( Figure 4 j) Thus, activation of BLA-projecting ACC neurons increases reward devaluation and produces depression-like behaviors.

[0363] The inventors then performed physiological recordings to investigate whether CRS induces changes in the activity of BLA-projecting ACC neurons. Fiber photometry showed that BLA-projecting ACC neurons in healthy mice exhibited a reduction in reward inhibition after an accumulation of reward trials ( Figure 4 k and Figure 19 a, b). In CRS mice, BLA-projecting ACC neurons exhibited weaker and faster decay in reward inhibition during repeated reward delivery ( Figure 4 k). In addition, ketamine increased reward inhibition and slowed the decline of reward inhibition in mice deprived of water for two days ( Figure 4 l and Figure 19 c, d). Therefore, behavioral hypersensitivity to reward devaluation manifests as a stronger attenuation of reward inhibition in BLA-projecting ACC neurons. Given that CRS treatment reduces reward inhibition in BLA-projecting ACC neurons, the inventors examined whether these neurons in CRS mice differ in their intrinsic physiological properties compared to healthy mice. The inventors labeled BLA-projecting ACC pyramidal neurons ( Figure 20 Targeted whole-cell recordings in brain slices revealed that the excitability of BLA-projecting ACC neurons was significantly higher in CRS mice than in control mice ( Figure 20df), indicating the hyperexcitability of these neurons in depression. Thus, the inventors have actually constructed a depression model by selectively activating BLA-projecting ACC neurons in mice. Furthermore, through the chemogenetic activation method described in this example, the degree of inhibition of BLA-projecting ACC neurons can be controlled, and thus, a specific depression state can be set in the model mice.

[0364] Finally, the inventors examined whether ablation of BLA-projecting ACC neurons would improve depressive behavior induced by the CRS depression model. To enable temporally controlled neuronal ablation, the inventors expressed diphtheria toxin receptor (DTR) in BLA-projecting ACC neurons by infusing rAAV2-retro-Cre into the BLA and then infusing AAV-DIO-DTR-EGFP into the ACC of C57BL / 6N wild-type mice ( Figure 21 a). After model induction, DT administration eliminated the vast majority of BLA-projecting ACC neurons in DTR-expressing mice but not in mGFP-expressing control mice ( Figure 21 Behavioral tests of sucrose preference and forced swimming showed that ablation of BLA-projecting ACC neurons significantly reduced behavioral despair and anhedonia ( Figure 4 n, o), thus supporting the idea that modulation of specific ACC circuits could represent a beneficial intervention for treating reward devaluation-related symptoms and disorders, such as depression.

Claims

1. A method for treating depression or anorexia in a subject, comprising inhibiting, destroying, interrupting or blocking ACC (anterior cingulate cortex) neurons in the subject.

2. The method of claim 1, wherein the ACC neurons are ACC pyramidal neurons.

3. The method of claim 2, wherein the ACC neurons are basolateral amygdala (BLA)-projecting ACC neurons.

4. The method of any one of claims 1 to 3, comprising introducing one or more compounds or one or more nucleic acids encoding said compounds into said ACC neurons, wherein said compounds are selected from the group consisting of: 1) Gi / o coupled receptors; 2) ligands or agonists of Gi / o coupled receptors; 3) anion channel proteins; 4) ligands or agonists of anion channel proteins; 5) Inhibitors or antagonists of cation channel proteins; 6) Inhibitory neurotransmitters; 7) inhibitors of neurotransmitters synthesized or secreted by the ACC neurons; 8) apoptosis proteins; 9) Toxin receptors; as well as 10) ligands of toxin receptors, Preferably, the nucleic acid is introduced into the ACC neurons by genetic engineering methods; and Preferably, the genetic engineering method uses genome editing tools or recombinase systems.

5. The method of claim 4, wherein: The Gi / o coupled receptor is a DREADD (Designer Receptor Activated Only by Designer Drugs) or a Gi / o coupled light-sensitive protein.

6. The method of claim 5, wherein the DREADD is selected from the group consisting of hM4Di, hM4D nrxn and KORDi.

7. The method of claim 6, wherein the DREADD ligand or agonist is selected from the group consisting of clozapine, CNO (clozapine-N-oxide), C21 (Compound 21), perampelar, DCZ (desclozapine), JHU37152, JHU3760, and salvinorin B.

8. The method of claim 5, wherein the Gi / o coupled light-sensitive protein is selected from the group consisting of Lamplight (lamprey lateral opsin), rod opsin, cone opsin, μ opioid receptor (MOR)-rod opsin chimera, and Opto-MOR.

9. The method of claim 4, wherein the anion channel protein is a ligand-gated ion channel (LGIC) protein or a light-gated anion channel protein.

10. The method of claim 9, wherein the LGIC is selected from the group consisting of a GABAA receptor and a glycine receptor.

11. The method of claim 9, wherein the light-gated anion channel protein is selected from the group consisting of eNpHR3.0, Arch, eBR, iC1C2, ChloC, ACRs, GtACR1, and GtACR2.

12. The method of claim 4, wherein the one or more compounds are selected from the group consisting of TeNT (tetanus toxin), muscimol, clozapine-N-oxide, nalfurafine, salvinorin B, pleuronothin, 8-chloro-11-[4-(1,1-dideuteroethyl)piperazin-1-yl]-5H-dibenzo[b,e][1,4]diazepine, NMDAR antagonists, AMPAR antagonists, GABA and its analogs, GABA receptor agonists, caspase 3, and a combination of DTR or its ligands.

13. The method of any one of claims 1-3, wherein the method comprises surgical removal of the ACC neurons.

14. A method of treating obesity or bulimia in a subject, comprising activating ACC neurons in the subject.

15. The method of claim 14, wherein the ACC neurons are ACC pyramidal neurons.

16. The method of any one of claims 14-15, wherein the ACC neurons are BLA-projecting ACC neurons.

17. The method of any one of claims 14-16, comprising introducing into the subject one or more compounds that activate the ACC neurons.

18. The method of claim 17, wherein the one or more compounds are selected from the group consisting of: 1) Gs-coupled receptors; 2) ligands or agonists of Gs-coupled receptors; 3) Gq-coupled receptors; 4) ligands or agonists of Gq-coupled receptors; 5) Cation channel proteins; 6) Ligands or agonists of cation channel proteins; 7) inhibitors or antagonists of anion channel proteins; and 8) Excitatory neurotransmitters; Preferably, the Gs-coupled receptor, Gq-coupled receptor or cation channel protein is introduced into the ACC neuron by introducing a nucleic acid encoding the Gi / o-coupled receptor, anion channel protein, apoptosis protein or toxin receptor into the ACC neuron; preferably, the nucleic acid is introduced into the ACC neuron genome by a genetic engineering method; and preferably, the genetic engineering method uses a genome editing tool or a recombinase system.

19. The method of claim 18, wherein the Gs-coupled receptor or the Gq-coupled receptor is a DREADD or a Gs-coupled or Gq-coupled light-sensitive protein.

20. The method of claim 19, wherein the DREADD is selected from the group consisting of hM3Dq, Rq(R165L), and hM3Ds.

21. The method of claim 20, wherein the ligand or agonist of the Gs-coupled receptor or the Gq-coupled receptor is selected from the group consisting of clozapine, CNO (clozapine-N-oxide), C21 (Compound 21), perampin, DCZ (desclozapine), JHU37152, and JHU37160.

22. The method of claim 19, wherein the Gs-coupled or Gq-coupled light-sensitive protein is an opsin.

23. The method of claim 22, wherein the opsin is selected from the group consisting of cOpn5, hOPN5, and JellyOp (opsin from Charybdea rastonii).

24. The method of claim 19, wherein the cation channel protein is a LGIC protein or a light-gated cation channel protein.

25. The method of claim 24, wherein the LGIC protein is selected from the group consisting of a 5-HT3 receptor, an acid-sensing ion channel (ASIC) protein, an epithelial sodium channel (ENaC), an ionotropic glutamate receptor, an IP3 receptor, a nicotinic acetylcholine receptor, a P2X receptor, a ryanodine receptor, and a zinc-activated channel protein (ZAC).

26. The method of claim 19, wherein the light-gated cation channel protein is selected from the group consisting of ChR2, CheRiff, Chronos, and variants thereof.

27. The method of claim 19, wherein the one or more compounds are selected from the group consisting of an NMDAR agonist, an AMPAR agonist, and an agonist of a glutamate receptor.

28. A method of reducing reward devaluation in a subject comprising inhibiting, destroying, disrupting or blocking ACC neurons.

29. The method of claim 28, wherein the ACC neurons are ACC pyramidal neurons.

30. The method of claim 29, wherein the ACC neurons are BLA-projecting ACC neurons.

31. The method of any one of claims 28-30, wherein the ACC neurons are inhibited, destroyed, interrupted or blocked by one or more measures selected from the group consisting of chemogenetic measures, optogenetic measures, electrical brain stimulation and pharmacological modulation measures.

32. A method of increasing reward devaluation in a subject, comprising activating ACC neurons.

33. The method of claim 32, wherein the ACC neuron is an ACC pyramidal neuron.

34. The method of claim 33, wherein the ACC neurons are BLA-projecting ACC neurons.

35. The method of any one of claims 32-34, wherein the ACC neurons are activated by one or more measures selected from the group consisting of chemogenetic measures, optogenetic measures, electrical brain stimulation, and pharmacological modulation measures.

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