Natriuretic peptide signal channel preparation and application thereof

Abstract

The invention discloses a preparation of a natriuretic peptide signal channel. The preparation maintains the metabolite content of natriuretic peptide in vivo at a normal level. The invention also discloses an application of a medicine taking the brain natriuretic peptide or the related receptor thereof as an action target spot in preparation of a medicine for treating Alzheimer's disease or social disorder related diseases. According to the preparation of the natriuretic peptide signal channel, diseases such as Alzheimer's disease and social disorder are treated or relieved through specific action targets of the BNP or related receptors thereof, a new treatment target and a nerve regulation and control treatment strategy are provided for treating the diseases such as AD, and the preparation has a wide application prospect.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technologies, and particularly to a preparation of the natriuretic peptide signaling pathway and its applications. Background Art

[0002] Natriuretic Peptides (NPs) are a class of polypeptide hormones with various biological activities, mainly including atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP). These polypeptides play important roles in the cardiovascular system, such as regulating blood pressure, promoting diuresis and sodium excretion. In recent years, studies have found that natriuretic peptides not only play important roles in the cardiovascular system, but may also play roles in the nervous system and behavioral regulation. For example, natriuretic peptides act through their receptors (such as NPRA, NPRB, and NPRC) in the central nervous system to regulate the release of neurotransmitters and the excitability of neurons. In addition, natriuretic peptides may also affect social behavior and social disorders by regulating the stress response and emotional state. In the study of social disorders, the mechanism of action of the natriuretic peptide signaling pathway is not fully understood, but existing studies have shown that natriuretic peptides may affect an individual's social behavior and emotional response by regulating the levels of neurotransmitters and neuronal activities in the brain. Therefore, the development of preparations containing the natriuretic peptide signaling pathway may provide new ideas and methods for the treatment of social disorders.

[0003] Social disorders can give rise to a variety of related diseases and problems, including social anxiety disorder, depression, substance abuse, and suicide risk. Social anxiety disorder is manifested by fear and anxiety in social situations, causing considerable distress and functional impairment in at least some aspects of daily life. Prolonged social disorders may lead to depression, manifested as low mood, decreased interest, appetite and sleep disorders, etc. Some patients with social disorders may use alcohol or other drugs to reduce fear and depression in social activities, resulting in substance abuse problems. Severe social disorders and depression may increase the risk of suicide or attempted suicide. Currently, the treatment methods for social disorders mainly include psychotherapy, drug treatment, exposure therapy, and relaxation techniques. Cognitive behavioral therapy (CBT) is the most commonly used psychotherapy method, which reduces anxiety and fear by changing the patient's thinking patterns and behaviors. In terms of drug treatment, selective serotonin reuptake inhibitors (SSRI) and serotonin and norepinephrine reuptake inhibitors (SNRI) are commonly used drugs. Exposure therapy helps patients adapt to and cope with anxiety-provoking situations by gradually exposing them to such situations. Relaxation techniques include learning stress reduction skills such as deep breathing, meditation, and yoga, etc. The neural circuit mechanism of social disorders involves multiple brain regions and neural networks, including the hippocampus, basal forebrain, and default mode network (DMN). Studies have found that abnormal activation of the hippocampus is related to defects in social recognition ability. The neural circuit in the basal forebrain plays an important role in regulating social behavior, especially somatostatin (SST)-positive inhibitory neurons. The default mode network (DMN) plays a key role in social function and is related to a variety of neuropsychiatric diseases.

[0004] Alzheimer's disease (AD) is a neurodegenerative disease of the nervous system that mainly occurs in the elderly or pre-elderly. The clinical manifestations mainly include memory impairment, cognitive decline, language disorders, social disorders, and loss of daily living ability. With the aging of society, the number of demented elderly with Alzheimer's disease is increasing, bringing a heavy burden to the families of the diseased elderly. However, as an old neurodegenerative disease that has been studied for more than 100 years, scientists have not yet found the exact pathogenic mechanism, let alone an effective drug for the treatment of AD.

[0005] Currently, there is an urgent need to develop drugs that can treat and relieve diseases such as Alzheimer's disease and social disorders. Summary of the Invention

[0006] The present invention aims to solve the technical problem that there is currently no effective drug for the treatment of Alzheimer's disease, and provides a preparation of the natriuretic peptide signaling pathway, which treats or relieves diseases such as Alzheimer's disease and social disorders through specific action targets of brain natriuretic peptide (BNP) or its related receptors.

[0007] To solve the above technical problems, the present invention is achieved through the following technical solutions:

[0008] In one aspect of the present invention, a preparation of the natriuretic peptide signaling pathway is provided, and this preparation maintains the metabolite content of natriuretic peptide in the body at a normal level.

[0009] The preparation includes small molecule compounds, polypeptides, proteins, or nucleic acid drugs.

[0010] The preparation acts on nerve cells in the ventral hippocampal CA1 region of the brain.

[0011] The natriuretic peptide is brain natriuretic peptide (BNP).

[0012] In another aspect of the present invention, there is also provided the use of a drug targeting brain natriuretic peptide or its related receptor in the preparation of a drug for treating Alzheimer's disease or diseases related to social disorders.

[0013] In another aspect of the present invention, there is also provided the use of a drug for regulating the natriuretic peptide signaling pathway in the preparation of a drug for treating Alzheimer's disease or diseases related to social disorders.

[0014] The drug is a brain-targeted drug.

[0015] Preferably, the drug targets nerve cells in the ventral hippocampal CA1 region of the brain.

[0016] In another aspect of the present invention, there is also provided the use of the above preparation in the preparation of a drug for treating Alzheimer's disease or diseases related to social disorders.

[0017] In another aspect of the present invention, there is also provided a neuromodulatory substance for treating Alzheimer's disease or diseases related to social disorders, and the neuromodulatory substance is a substance for regulating the natriuretic peptide signaling pathway in the ventral hippocampal CA1 region of the brain.

[0018] The preparation of the natriuretic peptide signaling pathway of the present invention utilizes the characteristic that BNP is significantly elevated in AD patients, and through experiments, it is found and confirmed that administering excessive BNP at the ventral hippocampal CA1 in the mouse brain will significantly inhibit the social memory of the mouse. It is concluded that drugs for treating mental diseases such as AD can be developed through specific action targets of BNP or its related receptor, providing new treatment targets and neuromodulatory treatment strategies for treating or alleviating diseases such as Alzheimer's disease and social disorders, and having very broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] The present invention will be further described in detail below in conjunction with the drawings and specific embodiments.

[0020] Figure 1It is the graph of the results of the animal behavior experiment of administering BNP through catheter implantation 30 minutes before the behavior in Example 1 of the present invention;

[0021] Figure 2 It is the graph of the results that knocking out the NPR3 receptor in Example 2 of the present invention can inhibit the social memory of mice;

[0022] Figure 3 It is the graph of the results that the social memory inhibited by BNP in Example 3 of the present invention cannot be re - evoked by light activation. Detailed implementation manners

[0023] Brain natriuretic peptide (BNP) is significantly elevated in AD patients, and the content of natriuretic peptide metabolites in the bodies of patients with diseases such as AD is also significantly increased. To explore whether the excessive BNP causes symptoms such as memory impairment and social impairment in AD patients, or whether the memory impairment and other symptoms in AD patients cause the excessive BNP, the present invention designs experiments. Catheters are implanted in the ventral hippocampal CA1 region of the mouse brain to administer excessive brain natriuretic peptide (BNP). Through experiments, it is found and confirmed that administering excessive BNP will significantly inhibit the social memory of mice. And the present invention also conducts experiments by knocking out BNP - related receptors. The results show that when the natriuretic peptide receptor NPR1 receives excessive self - BNP after knocking out a class of NPR3 receptors, it will also inhibit the social memory of mice. The experimental results provide new ideas for the treatment and alleviation of diseases such as Alzheimer's disease and social disorders, and it is expected to treat mental diseases such as AD through the specific targeting of BNP and its related receptors.

[0024] Administering brain natriuretic peptide (BNP) through catheter implantation 30 minutes before the behavior in Example 1 will significantly inhibit the social memory of mice

[0025] 1. Materials and methods

[0026] 1.1 Animals

[0027] Animal care and experimental procedures were approved by the Animal Ethics Committee of Shanghai Fudan University School of Medicine. The behavioral assays in all examples were performed on awake, freely moving C57BL / 6J mice, NPR3 - FLOX gene - knockout and wild - type littermate control mice, NPR3 gene - knockout and wild - type littermate control mice (all 2 - 3 months old, C57BL / 6J genetic background).

[0028] 1.2 Social memory test model

[0029] The social memory test model used in this invention is mainly the three-chamber experiment. The boxes for the three-chamber experiment are rectangular instruments made of acrylic plates (Shanghai Xinruan Technology Information Co., Ltd.). The interior of the box is divided into three boxes of the same size (length 19 cm, width 45 cm, height 20 cm). There are doors in the middle of the bottoms of the two middle partitions, allowing mice to freely enter between the three boxes. Two empty round wire metal cages (height 10 cm, bottom diameter 10 cm, wire bar spacing 2 cm) are placed in the two side chambers. The shape and structure of the box are shown in Figure 1 . This experiment has a total of five stages, namely drug administration, adaptation stage, social ability, 2h social memory, and 24h social memory.

[0030] (1) Administer ANP and BNP (Gil Biochemical (Shanghai) Co., Ltd.), and conduct the three-chamber experiment 30 minutes later.

[0031] (2) Adaptation stage: Place the mouse in the middle box and let it freely explore among the three boxes for 5 minutes (at this time, the cages in the left and right boxes are empty).

[0032] (3) Social ability: After the adaptation stage, close the small doors on both sides and place the mouse back in the middle box. Then place a strange mouse M1 from a different cage from the experimental mouse in one of the metal cages, open the small door, and let the experimental mouse freely explore among the three boxes for 5 minutes. After that, return the experimental mouse and the strange mouse to their original cages (normal mice will spend more time staying in the box on the side with the mouse).

[0033] (4) 2h social memory: Two hours later, place the experimental mouse back in the middle box, and the small doors on both sides are still closed. Place the mouse M1 familiar to the experimental mouse in one of the two metal cages, and place a new strange mouse M2 in the other metal cage. Open the small door and let the experimental mouse freely explore among the three boxes for 5 minutes. After that, return the experimental mouse and the strange mouse to their original cages (normal mice still remember that M1 is a familiar mouse at 2h and will spend more time staying in the box on the side with the strange mouse M2). Note: If it is determined that it has the effect of enhancing memory, this can be skipped. If it is uncertain, it still needs to be done.

[0034] (5) 24h social memory: Twenty-four hours later, place the mouse M1 familiar to the experimental mouse in one of the two metal cages, and place a new strange mouse M3 in the other metal cage. Open the small door and let the experimental mouse freely explore among the three boxes for 5 minutes. This tests the long-term social memory of the mouse (normal mice don't remember that M1 is a familiar mouse at 24h, so there is no difference in the time spent in the boxes on both sides of M1 and M3 mice. If there is an effect of enhancing memory, then it will stay longer on the side of the M3 mouse).

[0035] At the end of each experiment, each experimental mouse's cage and the two cages were thoroughly wiped with 75% alcohol to prevent the smell from affecting the behavior of the subsequent mice. The light intensity during the test was maintained at about 600 lux.

[0036] 1.3 Intracerebral cannula drug administration technique (ventral hippocampal CA1 region of the brain)

[0037] 6 - 8-week-old mice were anesthetized intraperitoneally with Zoletil. After the mice were anesthetized, the hair on their heads was shaved off, and they were fixed on a stereotaxic apparatus. The scalp was incised with sterilized surgical instruments, and the surface of the skull was wiped with sterilized saline to level the cranial surface. According to the Allen brain atlas, with the Bregma point of the anterior fontanelle as the zero point, brain region localization (ventral hippocampal CA1 region of the brain) was performed according to the corresponding coordinates of anterior-posterior (AP), medial-lateral (ML), and dorsal-ventral (DV). A cranial drill was used to drill a hole at the corresponding coordinate position. Then, the drug administration cannula (RWD) was placed in the target area and fixed with dental cement. After the operation, the mice were placed on a heating blanket to recover and keep warm until they woke up.

[0038] 1.4 Drug administration

[0039] 30 minutes before the behavior, BNP was administered through the cannula (the dosage was 0.8 μl bilaterally, the concentration was 2.5 mM, and the administration rate was 0.2 μl / min), and then the behavioral experiment of mouse social memory test was carried out.

[0040] 1.5 Data analysis

[0041] All results were expressed as mean ± standard error. Statistical comparisons were made using unpaired or paired Student's t-tests, one-way ANOVA, or two-way repeated measures ANOVA. Among them, P < 0.05, P < 0.01, and P < 0.001 were considered to be significantly different.

[0042] 2. Results

[0043] As Figure 1 shown, administering brain natriuretic peptide (BNP) through the cannula 30 minutes before the behavior did not affect the social ability of the mice, but significantly inhibited the 2-hour social memory of the mice.

[0044] Example 2 Knocking out the NPR3 receptor inhibits the social memory of mice

[0045] Transgenic mice NPR3-flox were used to conduct the behavioral experiment of mouse social memory test to see whether knocking out the natriuretic peptide receptor NPR3 would affect the social memory of the mice.

[0046] Results: As Figure 2 shown, by using transgenic mice NPR3-flox, after knocking out the NPR3 receptor, the social memory of mice will also be inhibited, indicating that after knocking out the NPR3 receptor, when the natriuretic peptide receptor NPR1 receives an excessive amount of its own BNP, the 2-hour social memory of mice will also be inhibited.

[0047] Example 3: After administering brain natriuretic peptide (BNP), a social memory test was performed, and 24 hours later, the relevant memory cells were optogenetically activated, but the relevant social memory could not be re-evoked.

[0048] 1. Method

[0049] 1.1 Optogenetic manipulation

[0050] Two to three days before the formal experiment, fiber optic jumpers were worn on the mice (5 minutes per day) to reduce their stress response to the weight of the fiber optic jumpers themselves. One end of the jumper was connected to the ceramic ferrule implanted in the mouse's head, and the other end was connected to an optogenetic stimulator (Hangzhou Newton Technology Co., Ltd.). Blue light (473 nm, terminal power of 10 mW, 20 Hz, 20 ms pulses) was given during the social stage to activate specific neurons (the illumination duration lasted for 10 minutes to ensure the effectiveness of optogenetic intervention). Red light (638 nm, terminal power of 10 mW, 10 ms pulses) was given at 24 hours to activate specific neurons (the illumination duration was 2 minutes on and 1 minute off to ensure the effectiveness of optogenetic intervention).

[0051] 1.2 Brain natriuretic peptide (BNP) was administered through a cannula in the brain region (ventral hippocampal CA1 region of the brain), and then the behavioral experiment of the mice was carried out according to the social memory test described in Example 1. At the same time, blue light (473 nm, terminal power of 10 mW, 20 Hz, 20 ms pulses) was given during the social stage to activate specific neurons, that is, the social stage was marked first.

[0052] 1.3 At 24 hours later, red light (638 nm, terminal power of 10 mW, 10 ms pulses) was given to activate specific neurons of the relevant memory cells, and the illumination duration was 2 minutes on and 1 minute off.

[0053] 2. Results

[0054] As Figure 3 shown, through optogenetic technology, after BNP administration, the social stage was marked first, and 24 hours later, the relevant memory cells were optogenetically activated, but the relevant social memory could not be re-evoked. This indicates that the social memory inhibited by BNP cannot be re-evoked by optogenetic activation.

[0055] The above-described embodiments merely represent the implementation manners of the present invention. The description thereof is relatively specific and detailed, but it should not be construed as a limitation on the scope of the patent for the present invention. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can still be made, and these all fall within the protection scope of the present invention. Therefore, the protection scope of the patent for the present invention shall be subject to the appended claims.

Claims

1. A preparation for a natriuretic peptide signaling pathway, which maintains the content of natriuretic peptide metabolites in the body at a normal level.

2. The preparation according to claim 1, characterized in that The preparation includes small molecule compounds, polypeptides, proteins, or nucleic acid drugs.

3. The preparation according to claim 1, characterized in that The agent acts on nerve cells in the CA1 region of the ventral hippocampus of the brain.

4. The preparation according to claim 1, characterized in that The natriuretic peptide is brain natriuretic peptide.

5. Use of drugs targeting brain natriuretic peptide or its related receptors in the preparation of drugs for the treatment of Alzheimer's disease or social disorder-related diseases.

6. Use of drugs that regulate natriuretic peptide signaling pathways in the preparation of drugs for the treatment of Alzheimer's disease or social disorder-related diseases.

7. The use according to claim 5 or 6, characterized in that: The drug is a brain-targeted drug.

8. The use according to claim 7, characterized in that: The drug targets nerve cells in the CA1 region of the ventral hippocampus in the brain.

9. Use of the preparation according to any one of claims 1 to 4 in the preparation of a medicament for treating Alzheimer's disease or diseases related to social disorders.

10. A neuromodulatory substance for treating Alzheimer's disease or social disorder-related diseases, wherein the neuromodulatory substance is a substance that regulates the natriuretic peptide signaling pathway in the CA1 region of the ventral hippocampus of the brain.

Images