Neuromodulation techniques for perturbation of physiological systems

Abstract

To provide techniques for assessing physiological systems via intentional perturbations.SOLUTION: As a perturbation, a pulse from an energy application device (ultrasound transducer) 12 is directed toward a target region, thereby applying a physiological perturbation. This induces local changes in tissue or function, such as local changes in the concentration of specific molecules in the target region, tissue displacement, or increased fluid movement. One of the specific molecules is glucose. Based on changes in the concentration of the target molecules, the presence or absence of exposure to a pathogen is determined.SELECTED DRAWING: Figure 1

Description

Statement Regarding Research and Development Funded by the Federal Government

[0001] This invention was made with government support under Contract No. HR0011-18-C-0040 awarded by the Defense Advanced Research Projects Agency (DARPA). The government has certain rights in this invention.

Technical Field

[0002] The subject matter disclosed herein relates to techniques for evaluating physiological systems through intentional perturbation of such systems, as well as the evaluation of recovery from perturbation and / or characteristics of the perturbation. In particular, the perturbation can be the result of targeted neuromodulation.

Background Art

[0003] Healthcare providers use various methods to examine and diagnose clinical conditions. In certain cases, a subject may be subjected to a state that induces some kind of stress or perturbation, and the evaluation of the subject's response to the stress or perturbation can indicate the clinical condition or overall health. Such perturbations can be induced by exercise, drugs, fasting, etc. For example, a glucose tolerance test may involve administering a glucose load to a subject and evaluating a blood sample taken from the subject after administration to determine how the subject responds. However, such tests are complex and may require dietary restrictions (e.g., prior fasting), and there may be subjects who do not comply, which can affect the results. Furthermore, the administration of drugs can be accompanied by undesirable side effects. Therefore, improved techniques for diagnosing specific clinical conditions are beneficial.

Summary of the Invention

[0004] The disclosed embodiments are not intended to limit the scope of the claimed subject matter; rather, these embodiments are only intended to provide a brief overview of possible embodiments. In fact, the present disclosure may encompass various forms that may be the same as or different from the embodiments described below.

[0005] In one embodiment, a regulation system is provided that includes an energy application device configured to apply energy to a target region to cause a physiological perturbation in a subject. The system is also configured to control the application of energy to the target region via the energy application device to induce a physiological perturbation to cause a change in the concentration of one or more target molecules relative to a baseline concentration. The controller is also configured to receive information indicative of the concentration of one or more target molecules and, based on the change in the concentration of one or more target molecules relative to the baseline concentration over a period of time, determine that the subject is in a category selected from two or more categories. selected category.

[0006] In another embodiment, a method for inducing a physiological perturbation in a subject is provided. The method includes directing an energy application device toward a target region of the subject, applying energy to the target region such that, as a result of the application, the activity of at least one axon terminal within the target region is modulated, evaluating one or more characteristics of the perturbation at one or more time points after the perturbation, and providing an indication of the clinical state of the subject based on the evaluation.

[0007] In another embodiment, a regulation system is provided that includes an energy application device configured to apply energy to a first target region of a first organ and a second target region of a second organ. The system also includes a controller configured to control a first application of energy to the first target region via the energy application device to cause a first perturbation as a result of the first application of energy, receive information indicative of a first perturbation characteristic, control a second application of energy to the second target region via the energy application device to cause a second perturbation as a result of the second application of energy, receive information indicative of a second perturbation characteristic, and determine the clinical state of the subject and provide an indication of the clinical state based on the first perturbation characteristic and the second perturbation characteristic.

[0008] In another embodiment, a method for evaluating a physiological perturbation of a subject is provided. The method includes applying ultrasonic energy to a target region within the subject to cause an approximated fasting state within the subject via neuromodulation, receiving glucose and insulin concentration data from the subject in the approximated fasting state, applying the data to a model, wherein the model is based on the relationship between the concentrations of glucose and insulin in an approximated fasting state for a plurality of normal subjects, receiving a display of the insulin resistance of the subject using the model, and providing a treatment recommendation based on the display.

Brief Description of the Drawings

[0009] These and other features, aspects, and advantages of the present invention will be better understood when the following detailed description is read with reference to the accompanying drawings in which like characters represent like parts throughout the drawings.

[0010]

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DETAILED DESCRIPTION OF THE INVENTION

[0011] One or more specific embodiments are described below. To provide a concise description of these embodiments, not all features of an actual implementation are described herein. In the development of an actual implementation, such as an engineering or design project, numerous decisions specific to the implementation must be made in order to achieve the developer's particular goals, such as accommodating system - related and business - related constraints, which may vary from one implementation to another. It should be understood that, although such development efforts may be complex and time - consuming, they are routine tasks in design, fabrication, and manufacture for those skilled in the art who benefit from the present disclosure.

[0012] Any example or illustration given herein should never be construed as representing a limitation, restriction, or definition for any one or more of the terms for which they are used. Instead, these examples or figures should be regarded as being described with respect to various specific embodiments and as being merely illustrative. One of ordinary skill in the art will understand that any one or more of the terms utilized in these examples or figures encompasses other embodiments, whether or not given with them or elsewhere in this specification, and that all such embodiments are intended to be included within the scope of one or more of those terms. Languages specifying such non - limiting examples and figures include, but are not limited to, "for example", "for instance", "such as", "e.g.", "including", "in certain embodiments", "in some embodiments", and "in one (an) embodiment".

[0013] This specification provides techniques for physiological perturbation via neuromodulation of a targeted region of interest and for causing a targeted physiological perturbation that is the result of the modulation. Such physiological perturbations can be useful for evaluating a subject's response to perturbations in a controlled environment. For example, the physiological perturbation can adjust a metabolic pathway to achieve homeostasis in response to the effect of the perturbation. The quality or effectiveness of the response can be a measure of the overall state of the subject. In one embodiment, neuromodulation is used as an alternative or in addition to other techniques (e.g., exercise stress test, drug administration) for inducing physiological changes. Drugs can disrupt physiological systems, but their effects can vary from subject to subject due to variability in a patient's comorbidities or variability in physiological processes that respond to the drug and are not connected to the system being tested. For example, a particular subject can metabolize a drug at a different rate due to pharmacogenetic differences in the drug metabolism pathway. Differences in metabolism rates can result in differences in drug effects. Thus, such tests using drugs may not be able to identify patients with a particular clinical condition. That is, a particular patient can have a response that is difficult to fit into an existing model of indicators or response types.

[0014] The neuromodulation applied to cause the physiological perturbation can be targeted, enabling a healthcare provider to induce a desired change and evaluate one or more characteristics of the perturbation. Such characteristics can include changes in the concentration of one or more target molecules in response to the perturbation and the time to recovery (e.g., evaluation of the persistence of the physiological perturbation). Thus, the evaluated characteristics of the perturbation can be changes in the concentration of one or more target molecules over a predetermined period and / or total time to achieve a predetermined baseline concentration of the one or more target molecules. In certain embodiments, based on the evaluation, a diagnosis of a clinical condition can be made and / or treatment or treatment recommendations can be provided to the subject.

[0015] Neural modulation of a target region can cause physiological perturbations in healthy or diseased subjects. In one embodiment, neural modulation that does not cause characteristic changes in a subject may be associated with a clinical condition or disease diagnosis. In another embodiment, different clinical conditions or categories may be associated with specific characteristic responses to neural modulation. That is, the lack of a predicted physiological response to the application of energy and perturbation to a target region may indicate an affected organ that includes metabolic or inflammatory / anti-inflammatory pathways that cannot respond to neural modulation, such as a healthy patient. In one example, neural modulation energy applied to the pancreas without an accompanying increase in circulating insulin levels indicates a non-responsive subject that can no longer produce insulin. Thus, a neural modulation therapy may not function, and a treatment recommendation may be provided that insulin should be provided.

[0016] Perturbations can be induced by challenges such as exercise, administration of drugs, and / or changes in diet (fasting), but such perturbations are typically broad and not targeted. In contrast, as provided herein, neural modulation can target one or more specific structures in the body to cause targeted or predictable physiological perturbations. Further, as long as multiple organs can be involved in metabolic processes, neural modulation of one or more target regions in a specific organ can be useful in identifying portions of pathways that may be dysfunctional.

[0017] As provided herein, neuromodulation can induce a physiological perturbation through modulation of activity at axonal terminals within a target region. The targeted or target region can be any tissue or structure in the body having axonal terminals that form synapses with non-neuronal cells or fluids. In one example, the target region may be within an organ or structure such as the spleen, liver, pancreas, or gastrointestinal tissue. In another example, the target region can be within lymphoid tissue. Neuromodulation of the target region enables local, limited, and non-excisive application of energy to only the target region without applying energy outside the target region. Energy application can induce a centrifugal perturbation outside the targeted target region, such as in the same organ, tissue, or structure containing the target region, or in other organs and structures not containing the targeted target region. In some embodiments, a centripetal effect can be induced, for example, in a region of the hypothalamus. Energy application can also induce a centripetal effect along the target nerve upstream of the energy application site. In some embodiments, effects outside the targeted target region(s) can be achieved without directly applying energy to regions outside the target region(s) in which the effect is induced. Thus, local energy application can be used to achieve or effect a systemic perturbation that can include local, downstream, and / or upstream effects.

[0018] The disclosed techniques can be used to exert a perturbation of a physiological process of the body using an external or extracorporeal source to cause a targeted physiological perturbation in a subject. Through neuromodulation of a targeted target region, a physiological process can be altered, slowed, stopped, or reversed. Also provided herein are techniques that can be applied to a subject to disrupt or temporarily challenge the homeostasis or constancy of a physiological process, such as glucose regulation, or to stimulate a physiological system to return to a steady state. Neuromodulation of a targeted target region can exert a change in a physiological process to interrupt, reduce, or enhance one or more physiological pathways in a subject to effect a desired physiological perturbation.

[0019] The neuromodulation techniques discussed herein can be used to cause physiological perturbations in a neuromodulated subject. Physiological perturbations can include local changes in a target region or tissue to which energy is applied, as well as systemic changes that are a result of neuromodulation. In certain embodiments, one or more characteristics of the perturbation can be evaluated as part of the present technology. These changes can include changes in one or more target molecules, changes in the physiological parameters of the subject, displacement, enlargement, or other morphological changes in one or more tissue structures of the subject, changes in the cell populations of the subject, changes in the flow parameters of blood or other fluids, and the like. In one embodiment, the physiological perturbation causes a change in the concentration (e.g., increase, decrease) of a target molecule and / or a change in the characteristics of the target molecule. That is, the perturbation can include selective regulation of tissue production or release of one or more target molecules (e.g., a first target molecule, a second target molecule, etc.), and can refer to the regulation or influence of the concentration (circulating, tissue) or characteristics (covalent modification) of a molecule as a result of applying energy to one or more target regions (e.g., a first target region, a second target region, etc.) in one or more tissues (e.g., a first tissue, a second tissue, etc.). The regulation of a target molecule can include changes in the characteristics of the molecule such as expression, secretion, protein translocation, and direct activity changes. The regulation can be driven either to drive the nerve activity and function itself or based on the effect of the applied energy on ion channels for the regulation of adjacent non-neuronal cells as a result of molecules derived from direct activation within the nerve activity or non-neuronal cells. The regulation of a target molecule can also refer to maintaining the desired concentration of the molecule so that changes or fluctuations in concentration that would be expected as a result of neuromodulation do not occur. The regulation of a target molecule can refer to causing changes in the characteristics of the molecule such as enzyme-mediated covalent modifications (changes such as phosphorylation, acetylation, ribosylation, etc.). That is, it should be understood that the selective regulation of a target molecule can refer to the molecular concentration and / or the molecular characteristics. The target molecule can be a biological molecule such as one or more of carbohydrates (monosaccharides, polysaccharides), lipids, nucleic acids (DNA, RNA), or proteins.In certain embodiments, the target molecule can be a signaling molecule such as a hormone (amine hormone, peptide hormone, or steroid hormone).

[0020] Certain embodiments described herein provide neuromodulation techniques for the diagnosis of metabolic dysfunction. In one embodiment, the diagnosis can be the presence or absence of glucose metabolic dysfunction and related disorders. Glucose regulation is complex and involves different local and systemic metabolic pathways. Applying energy to a targeted region(s) of interest results in characteristic changes in these metabolic pathways that affect glucose regulation. In some embodiments, modulation at one or more regions of interest is used to identify disorders including, but not limited to, diabetes (i.e., type 1 or type 2 diabetes), hyperglycemia, hyperlipidemia, sepsis, trauma, infections, physiological stress, diabetes-related dementia, obesity, or other eating disorders or metabolic disorders. In one example, physiological stress can be medically defined to include various acute medical conditions (infections, severe injury / trauma, heart attack, bypass) as well as surgical cases presenting hyperglycemia. Targeted perturbation via neuromodulation can induce changes in glycemic hormones in the blood or tissue, causing deviation from normal glucose concentrations. Targeted perturbation via neuromodulation can also induce changes in the activity of sensory or effector neurons within the metabolic physiological control system, and the response of this system can be analyzed by imaging (e.g., MRI) or electrical recording (e.g., ECG) or other physiological monitoring. These changes to these systems as a result of neuromodulation can be different in healthy patients and diseased patients. In healthy subjects, the perturbation can be cleared over time such that glucose concentrations return to normal. Additionally, physiological perturbation can be used to identify subjects who have not received a disease diagnosis but are prediabetic. In one embodiment, physiological perturbation is used to identify subjects with insulin resistance, who may or may not be identified as diabetic.

[0021] The specific embodiments described herein provide neuromodulation techniques for the diagnosis of the control of inflammation and immune function and related disorders. The regulation of the inflammatory and activation states of immune cells involves different local and systemic neural, humoral, and cellular pathways. The application of energy to a targeted area of interest causes characteristic changes in these inflammatory and anti-inflammatory pathways that affect the concentrations of circulating cytokines and neurotransmitters, and thus the activity and state of immune cells. In some embodiments, modulation at one or more areas of interest is used to identify disorders including, but not limited to, rheumatoid arthritis, inflammatory bowel disease, psoriasis, or other chronic inflammatory or related disorders. Targeted perturbation via neuromodulation can induce changes in immune markers such as inflammatory cytokines, hormones, or neurotransmitters in the blood or tissue, causing changes in the activity of resident or circulating immune cells. Targeted perturbation via neuromodulation may also induce changes in the activity of sensory or effector neurons within the inflammation control system, and the response of this system can be analyzed by imaging or electrical recording. These changes to these systems as a result of neuromodulation can differ between healthy and diseased patients. In healthy patients, the perturbation may cause transient or smaller changes to physiology, while in diseased patients the perturbation may last longer. Additionally, physiological perturbations can be used to identify patients who are not currently receiving a disease diagnosis, or pre-symptomatic or symptomatic patients with chronic inflammatory diseases.

[0022] For that purpose, the present technique relates to the targeted regulation of synapses at axonal terminals within tissue via the direct application of energy by an energy source that causes changes that result in a measurable physiological perturbation (e.g., a change in circulating molecular concentration or a physiological profile characterized by a series of concentration changes). The targeted synapses can include extra-axonal cell synapses formed between presynaptic axonal terminals and postsynaptic non-neuronal cells. Further, while certain disclosed embodiments are discussed in the context of extra-axonal cell synapses, axonal terminals can form axonal secretions, axonal synapses, between axonal cell bodies or extra-axonal cell synapses, and additionally or alternatively, it should be understood that these synapse types are thought to be selectively regulated as provided herein. Further, certain axonal terminals can terminate in interstitial or body fluids that can also experience neurotransmitter release as a result of regulation. The disclosed synapses can be regulated to change activity at the synapse, e.g., the release of neurotransmitters from presynaptic axonal terminals, as a result of energy application. Thus, the change in activity can result in local and / or non-local (e.g., systemic) effects that cause the overall profile of physiological changes associated with a desired or targeted physiological perturbation. The present technique enables targeting and focusing energy on the volume of tissue containing a particular axonal terminal to preferentially and directly activate the targeted axonal terminal to achieve a desired physiological perturbation. In this way, in certain embodiments, axonal terminals that are within the same organ or tissue structure but outside the target region are not activated, while the targeted axonal terminals within the target region are activated. Since organs and tissue structures can include different types of axonal terminals that form synapses with different types of postsynaptic non-neuronal cells, a target region can be selected that includes a particular type of axonal terminal that, when activated, results in a desired targeted physiological perturbation. Thus, the regulation can target a particular type of axonal terminal based on the presynaptic neuron type, the postsynaptic cell type, or both.

[0023] For example, in one embodiment, the type of axon terminal can be an axon terminal that forms an axonal extracellular cell synapse with endogenous (i.e., tissue endogenous or non-circulating) liver, pancreas, or gastrointestinal tissue cells. That is, the axonal extracellular cell synapse is formed at the junction between the axon terminal and a non-neuronal cell or interstitial fluid or body fluid. Thus, the application of energy results in the regulation of function in the target region. However, it should be understood that different targeted physiological effects can be achieved based on the characteristics of the axon terminal type population and the presynaptic neuron type and postsynaptic cells (e.g., immune cells, lymphocytes, mucosal cells, muscle cells, etc.) of the axonal extracellular cell synapse. Further, as described above, the axon terminal can terminate in interstitial fluid or body fluid that can also experience neurotransmitter release as a result of regulation. Thus, when energy is applied to a target region within the tissue of a subject, the axon terminals (and, where applicable, their associated axonal extracellular cell synapses) within the target region can be activated, but non-target axon terminals (and associated synapses) outside the target region may not be affected. However, since the regulation can result in a systemic effect, certain systemic changes can occur in non-target axon terminals outside the target region as a result of the activation of the axon terminals within the target region. As provided herein, preferential activation or direct activation can refer to a cell or structure (e.g., a synapse) within the target region that receives direct application of energy (e.g., the energy is applied directly to the cell or structure). For example, an axon terminal, an axonal extracellular cell synapse, and / or a postsynaptic non-neuronal cell within the target region, or interstitial fluid or body fluid that can directly receive the applied energy provided herein. Even in a region that undergoes physiological changes as a result of energy application, preferential or direct activation can be considered in contrast to regions outside the target region that do not receive direct energy application.

[0024] Neuromodulation is a technique that applies energy from an external energy source to specific regions of the nervous system to activate or enhance nerves or nerve functions and / or to block or reduce nerves or nerve functions. In certain neuromodulation techniques, one or more electrodes are applied to or near the target nerve, and the application of energy is carried through the nerve (e.g., as an action potential) to cause a physiological response in regions downstream of the energy application site. However, because the nervous system is complex, it is difficult to predict the range and ultimate endpoints of the physiological response to a given energy application site. In one example, stimulation of axonal terminals releases neurotransmitters / neuropeptides or induces altered neurotransmitter release in the vicinity of adjacent non-neuronal cells such as secretory cells or other cells, regulating the cell activity of adjacent or nearby non-neuronal cells including postsynaptic cells.

[0025] Advantages of the technology include local regulation in a target region of tissue to achieve a physiological perturbation that can be used to evaluate the state of a subject. Further, the local regulation can include direct activation of a relatively small region of tissue (e.g., less than 25% of the total tissue volume) to achieve these effects. In this way, the total applied energy to achieve the desired physiological perturbation is relatively small. In certain embodiments, the applied energy can be from a non-invasive extracorporeal energy source (e.g., an ultrasonic energy source, a mechanical vibrator). For example, a focused energy probe can apply energy through the skin of a subject and be focused on a target region of internal tissue. Such embodiments achieve the desired physiological perturbation without invasive procedures or side effects associated with other types of procedures or treatments.

[0026] In certain embodiments, techniques for neuromodulation are provided in which energy from an energy source (e.g., an external or extracorporeal energy source) is applied to an axon terminal such that the site of energy application focus, e.g., an induced physiological perturbation at the axon terminal such as neurotransmitter release, is caused in response to the energy application rather than in response to an action potential. That is, direct application of energy to the axon terminal acts instead of an action potential to facilitate the release of neurotransmitters to a neural junction with a non-neural cell (i.e., a synapse). Direct application of energy to the axon terminal further induces a changed release of neurotransmitters from the axon terminal within the synapse (e.g., an extra-axonal cell synapse) to the vicinity of an adjacent non-neural cell. In one embodiment, the energy source is an extracorporeal energy source such as an ultrasonic energy source or a mechanical oscillator. In this way, non-invasive and targeted neuromodulation can be achieved directly at the energy concentration site rather than through stimulation at an upstream site that induces an action potential and propagates it to a downstream site target to activate the downstream target.

[0027] In certain embodiments, the target tissue is an internal tissue or organ that is difficult to access using electrical stimulation techniques using electrodes. Envisioned tissue targets include gastrointestinal (GI) tissue (stomach, intestine), muscle tissue (heart, smooth and skeletal), epithelial tissue (epidermis, organ / GI inner layer), connective tissue, glandular tissue (exocrine / endocrine), organ tissue, etc. In one example, focused application of energy at the neuromuscular junction facilitates the release of neurotransmitters at the neuromuscular junction without an upstream action potential. In one embodiment, the intended target or region for modulation may include a part of the pancreas responsible for insulin release or a part of the liver responsible for glucose regulation. In another embodiment, the intended target region may be located in the liver. In another embodiment, the intended target region may be located in the spleen. However, it should be understood that these embodiments are examples.

[0028] For that purpose, the disclosed neuromodulation techniques can be used with a neuromodulation system. FIG. 1 is a schematic diagram of a system 10 for neuromodulation to achieve the release of neurotransmitters in response to the application of energy and / or to activate components of a synapse (e.g., presynaptic cell, postsynaptic cell). The illustrated system includes a pulse generator 14 coupled to an energy application device 12 (e.g., an ultrasonic transducer). The energy application device 12 is configured to receive energy pulses, for example, via a lead wire or a wireless connection, and is directed at a target region of the internal tissue or organ of a subject during use, resulting in a targeted physiological perturbation. In certain embodiments, the pulse generator 14 and / or the energy application device 12 may be implanted at a biocompatible site (e.g., the abdomen), and one or more lead wires internally couple the energy application device 12 and the pulse generator 14. For example, the energy application device 12 may be a MEMS transducer such as a capacitive micromachine ultrasonic transducer.

[0029] In certain embodiments, the energy application device 12 and / or the pulse generator 14 can communicate wirelessly with a controller 16 that can, for example, provide commands to the pulse generator 14 in sequence. In other embodiments, the pulse generator 14 may be an extracorporeal device and may operate, for example, to apply energy transcutaneously or non-invasively from a location outside the subject's body, and in certain embodiments, may be incorporated within the controller 16. In embodiments where the pulse generator 14 is extracorporeal, the energy application device 12 may be operated by a caregiver and may be placed at a point on or above the subject's skin such that the energy pulses are delivered transcutaneously to the desired internal tissue. When arranged to apply energy pulses to the desired site, the system 10 can initiate neuromodulation to achieve a targeted physiological perturbation or clinical effect.

[0030] In certain embodiments, system 10 may include an evaluation device 20, which is coupled to controller 16 and evaluates a feature indicating whether a targeted physiological perturbation of regulation has been achieved. In one embodiment, the targeted physiological perturbation may be local. For example, the regulation may result in local changes in tissue structure, local changes in the concentration of a particular molecule, tissue displacement, increased fluid movement, or other local tissue or function changes.

[0031] The regulation may result in systemic or non-local changes, and the targeted physiological perturbation may be related to changes in the concentration of circulating molecules or changes in the characteristics of tissues not including the region where energy is directly applied. In one example, tissue displacement may be a surrogate measure of the desired regulation, and a displacement measurement value less than the expected displacement value can result in modification of the regulation parameters until the expected displacement value is induced. Thus, in some embodiments, evaluation device 20 may be configured to evaluate changes in molecular concentration or activity. For example, evaluation device 20 can include a chemical sensor for the target molecule. In some embodiments, evaluation device 20 may be an imaging device configured to evaluate changes in the size and / or position of an organ, changes in body fluid flow, changes in RNA or protein expression, or other indicators of physiological perturbation. In another embodiment, evaluation device 20 may be a circulating glucose monitor and / or a continuous glucose monitor that measures interstitial fluid glucose. Evaluation device 20 may be an MRI device or other device capable of acquiring image data of a subject to identify systemic changes. Evaluation device 20 may be an ECG or other physiological monitor. It should be understood that although the illustrated elements of system 10 are shown separately, some or all of the elements may be combined with each other. Further, some or all of the elements can communicate with each other in a wired or wireless manner.

[0032] Based on the evaluation, the adjustment parameters of the controller 16 can be changed. For example, if the desired adjustment is related to the change in concentration (circulating concentration or tissue concentration of one or more molecules) within a defined time window (e.g., 5 minutes after starting the energy application treatment, 30 minutes after), or relative to the baseline at the start of the treatment, a change in adjustment parameters such as pulse frequency or other parameters may be desired, which can be provided to the controller 16 in turn by the operator or via an automatic feedback loop to define or adjust the energy application parameters or adjustment parameters of the pulse generator 14.

[0033] The system 10 provided herein can provide energy pulses according to various adjustment parameters. For example, the adjustment parameters can include various stimulation time patterns ranging from continuous to intermittent. In intermittent stimulation, energy is delivered at a constant frequency for a certain period during the signal on-time. After the signal on-time, a period without energy supply, called the signal off-time, follows. The adjustment parameters can also include the frequency and duration of the stimulation application. The application frequency can be continuous or delivered at various intervals, for example, within one day or within one week. In one example, the adjustment can be specified with respect to time (morning, evening, night) or diet (fasting, non-fasting). The treatment period for causing a physiological perturbation can continue for various periods including, but not limited to, several minutes to several hours. In certain embodiments, the duration of the treatment with a specified stimulation pattern can be 1 hour and repeated, for example, at 72-hour intervals. In certain embodiments, the energy can be delivered at a higher frequency, for example, every 3 hours, with a shorter duration, for example, for 30 minutes. According to adjustment parameters such as the duration and frequency of the treatment, the application of energy can be controllably adjusted to achieve the desired result.

[0034] FIG. 2 is a block diagram of certain components of system 10. As provided herein, system 10 for neuromodulation can include a pulse generator 14 adapted to generate a plurality of energy pulses for application to a subject's tissue. The pulse generator 14 may be separate or may be integrated into an external device such as a controller 16. The controller 16 includes a processor 30 for controlling the device. Software code or instructions are stored in the memory 32 of the controller 16 and executed by the processor 30 to control the various components of the device. The controller 16 and / or the pulse generator 14 can be connected to the energy application device 12 via one or more leads 33 or wirelessly.

[0035] The controller 16 also includes a user interface having an input / output circuit 34 and a display 36 adapted to enable a clinician to provide selection inputs or adjustment parameters to the adjustment program. Each adjustment program can include one or more sets of adjustment parameters including, but not limited to, pulse amplitude, pulse width, pulse frequency, and the like. The pulse generator 14 changes its internal parameters in response to a control signal from the controller device 16 to vary the stimulation characteristics of the energy pulses transmitted via the lead wire 33 to the subject to which the energy application device 12 is applied. Any suitable type of pulse generation circuit can be used, including, but not limited to, constant current, constant voltage, multiple independent current or voltage sources. The energy applied is a function of the current amplitude and the pulse width duration. The controller 16 enables the energy to be adjustably controlled by changing the adjustment parameters and / or starting the energy application at a certain time or canceling / suppressing the energy application at a certain time. In one embodiment, the adjustable control of the energy application device is based on information regarding the concentration of one or more molecules in the subject (e.g., circulating molecules). If the information is from the evaluation device 20, a feedback loop can drive the adjustable control. For example, a diagnosis can be made based on the circulating glucose concentration measured by the evaluation device 20 in response to neuromodulation. When the concentration exceeds a predetermined threshold or range, the controller 16 can initiate a treatment protocol for energy application to a target area (e.g., the liver) and may use adjustment parameters related to the decrease in circulating glucose. The treatment protocol can use different adjustment parameters (e.g., higher energy levels, more frequent applications) than those used in the diagnostic protocol. The controller can initiate a treatment protocol for energy application to a target area (e.g., the spleen or immune tissue) and may use adjustment parameters related to an increase or decrease in circulating cytokines or immune cell number, activity, or phenotype.

[0036] In one embodiment, the memory 32 stores different operation modes selectable by an operator. For example, the stored operation modes can include instructions for executing a set of adjustment parameters related to a specific treatment site such as a target region in the liver, pancreas, digestive tract, spleen, etc. Different sites may have different associated adjustment parameters. Instead of having the operator manually enter the mode, the controller 16 may be configured to execute appropriate instructions based on the selection. In another embodiment, the memory 32 stores operation modes for different types of treatments. For example, activation may be related to different stimulation pressures or frequency ranges as compared to those related to suppression or blocking of tissue function. In a specific example, when the energy application device is an ultrasonic transducer, the time-average power (time-average intensity) and peak positive pressure are within the ranges of 1 mW / cm2 to 30,000 mW / cm2 (time-average intensity) and 0.1 MPa to 7 MPa (peak pressure). In one example, the time-average intensity is less than 35 W / cm2 in the target region to avoid levels related to thermal damage and ablation or cavitation. In another specific example, when the energy application device is a mechanical actuator, the amplitude of vibration is within the range of 0.1 to 10 mm. The selected frequency may depend on the mode of energy application, such as ultrasonic or mechanical actuator. The controller 16 can operate in a diagnostic protocol mode, and the results of the diagnosis can induce a change to the treatment operation mode. For example, the diagnostic mode can apply the neuromodulation energy once or repeatedly within a relatively short time window (1 hour), and the treatment operation mode can include repeated neuromodulation events (e.g., daily, hourly) during the treatment protocol.

[0037] The system can also include an imaging device that facilitates focusing of the energy application device 12. In one embodiment, the imaging device is such that different ultrasonic parameters (frequency, aperture, or energy) are applied to select (e.g., spatially select) a target region and to focus the energy on the selected target region for targeting and subsequent neuromodulation, and may be integrated with the energy application device 12 or be the same device as the energy application device 12. In another embodiment, the memory 32 stores one or more targeting modes or focusing modes used to spatially select a target region within an organ or tissue structure. The spatial selection can include selecting a sub-region of the organ to identify the volume of the organ corresponding to the target region. The spatial selection can depend on the image data provided herein. Based on the spatial selection, the energy application device 12 can be focused on the selected volume corresponding to the target region. For example, the energy application device 12 may first be configured to operate in a targeting mode to apply targeting mode energy for capturing the image data used to identify the target region. The targeting mode energy is not at a level suitable for preferential activation and / or is not applied with regulatory parameters. However, once the target region is identified, the controller 16 can operate in a treatment mode according to the regulatory parameters associated with preferential activation.

[0038] The controller 16 may also be configured to receive an input related to a targeted physiological perturbation as an input for the selection of adjustment parameters. For example, if an imaging modality is used to evaluate tissue characteristics, the controller 16 may be configured to receive the calculated metrics or parameters of the characteristics. Based on whether the metrics or parameters exceed or fall below a predetermined threshold, a diagnosis can be made, and a display of the diagnosis can be provided (e.g., via a display). In one embodiment, the parameter may be a measure of tissue displacement of diseased tissue or a measure of the depth of the diseased tissue. Other parameters may include evaluating the concentration of one or more target molecules (e.g., evaluating one or more of the change in concentration relative to a threshold or baseline / control, the rate of change, determining whether the concentration is within a desired range). Further, the energy application device 12 (e.g., an ultrasonic transducer) operates under the control of the controller 16 to a) acquire image data of tissue that can be used to spatially select a target region within the target tissue, b) apply adjustment energy to the target region, and c) acquire images to determine (e.g., via displacement measurements) that a targeted physiological perturbation has occurred. In such an embodiment, the imaging device, the evaluation device 20, and the energy application device 12 may be the same device.

[0039] Figure 3 is a specific example in which the energy application device 12 includes an ultrasonic transducer 42 that can apply energy to a target tissue 43, such as the liver, spleen, or pancreas. The energy application device 12 can include a control circuit for controlling the ultrasonic transducer 42. The control circuit of the processor 30 (Figure 2) may be integrated with the energy application device 12 (e.g., via the integrated controller 16) or may be a separate component. The ultrasonic transducer 42 may also be configured to acquire image data to help spatially select a desired or targeted target region and focus the applied energy on the target region of the target tissue or structure.

[0040] The desired target tissue 43 can be an internal tissue or organ that includes axonal endings of synapses and non-neuronal cells. The synapse is stimulated by directly applying energy to the axonal endings within the focal region of the ultrasonic transducer 42 focused on the target region 44 of the target tissue 43, causing the release of molecules, such as neurotransmitters, into the synaptic space, and / or changes in ion channel activity can sequentially cause downstream effects. Sensory synapses (or relay cells) can also be stimulated by directly applying energy to the peripheral endings and neurons within the focal region of the ultrasonic transducer focused on the target region of the target tissue, causing direct neural signal transmission back to the CNS (i.e., driving or mimicking sensory input to the CNS control centers, ganglia, and nuclei). The target region can be selected to include specific types of axonal endings, such as those that form synapses with axonal endings of specific neuron types and / or certain types of non-neuronal cells. Thus, the target region 44 can be selected to correspond to a portion of the target tissue 43 having the desired axonal endings (and associated non-neuronal cells). The energy application can be selected to preferentially induce the release of one or more molecules, such as neurotransmitters from neurons within the synapse, or to directly activate the non-neuronal cells themselves through direct energy conversion (i.e., mechanotransduction or voltage-activated proteins within non-neuronal cells), or to cause activation in both neuronal and non-neuronal cells that induces the desired physiological effect. The target region can be selected as a nerve entry site into the organ. In one embodiment, liver stimulation or regulation can refer to the regulation of the porta hepatis or the target region 44 adjacent to the porta hepatis.

[0041] Energy can be focused or substantially concentrated on only the target region 44 and a portion of the internal tissue 43, for example, less than about 50%, less than about 25%, less than about 10%, or less than about 5% of the total volume of the tissue 43. In one embodiment, energy can be applied to two or more target regions 44 within the target tissue 43, and the total volume of the two or more target regions 44 can be less than about 90%, less than about 50%, less than about 25%, less than about 10%, or less than about 5% of the total volume of the tissue 43. In one embodiment, energy is applied to only about 1% to 50% of the total volume of the tissue 43, only about 1% to 25% of the total volume of the tissue 43, only about 1% to 10% of the total volume of the tissue 43, or only about 1% to 5% of the total volume of the tissue 43. In certain embodiments, only the axon terminals of the target region 44 of the target tissue 43 receive the applied energy directly and release neurotransmitters, while the unstimulated axon terminals outside the target region 44 receive no substantial energy and are thus not activated / stimulated in the same way. In some embodiments, the axon terminals of the portion of the tissue that directly receives the energy induce altered neurotransmitter release. In this way, tissue sub-regions can be granularly targeted for neuromodulation, for example, one or more sub-regions can be selected. In some embodiments, the energy application parameters can be selected to induce preferential activation of either the neural or non-neural components within the tissue that directly receives the energy, to induce the desired complex physiological effect. In certain embodiments, the energy may be focused or concentrated within a volume of less than about 25 mm3. In certain embodiments, the energy may be focused or concentrated within a volume of about 0.5 mm3 to 50 mm3. The focal volume and focal depth for focusing or concentrating the energy within the target region 44 can be affected by the size / configuration of the energy application device 12. The focal volume of the energy application may be defined by the focal region of the energy application device 12.

[0042] As provided herein, energy can be applied substantially only to target region 44 in a targeted manner to preferentially activate synapses to achieve a targeted physiological perturbation, and not substantially in a non-specific manner across the entirety or whole of tissue 43. Thus, only a subset of multiple different types of axonal terminals within tissue 43 are directly exposed to energy application. The disclosed techniques can be used to evaluate the state of a subject as a result of a perturbation caused by neuromodulation. The disclosed techniques can use a direct evaluation of the state or function of tissue as a targeted physiological perturbation. These techniques can also be used to detect a deviation from or a return to a steady state or set point of a physiological neuroimmune system, neurohormonal system, and / or neuroreflex system. The evaluation can be performed before (i.e., baseline evaluation), during, and / or after neuromodulation. The evaluation techniques can include at least one of functional magnetic resonance imaging, diffusion tensor magnetic resonance imaging, positron emission tomography, acoustic monitoring, thermal monitoring, or chemical sensing (e.g., immunochemical detection). The evaluation techniques can also include protein and / or molecular concentration evaluation. Images from the evaluation techniques can be received by a system for automated or manual evaluation. Based on the image data, adjustment parameters can also be modified. For example, changes in the size or displacement of an organ can be effectively used as a marker for local neurotransmitter concentration, as a surrogate marker for local cell exposure to phenotypic modulating neurotransmitters, and as a marker for the predicted effect on glucose metabolic pathways. Local concentration can refer to the concentration within the focal region of energy application.

[0043] Additionally or alternatively, the system can evaluate the presence or concentration of one or more molecules circulating in the tissue or in the blood. The concentration within the tissue can be referred to as the local concentration or the standing concentration. The tissue can be obtained by fine needle aspiration, and the evaluation of the presence or level of the target molecule (e.g., a metabolite molecule, a marker of a metabolic pathway, a peptide transmitter, a catecholamine) can be performed by any suitable technique known to those skilled in the art.

[0044] In other embodiments, the targeted physiological perturbation can include, but is not limited to, tissue displacement, changes in tissue size, changes in the concentration of one or more molecules (either local, non-local, or circulating concentrations), changes in gene or marker expression, afferent activity, and cell migration. For example, tissue displacement (e.g., displacement of the liver) can occur as a result of the application of energy to the tissue. By evaluating the tissue displacement (e.g., via imaging), other effects can be inferred. For example, a particular displacement can be characteristic of a particular change in molecular concentration. In one example, a 5% displacement of the liver can indicate or be associated with a desired decrease in circulating glucose concentration based on empirical data. In another example, the tissue displacement can be evaluated by determining the displacement parameter by comparing the reference image data (the tissue image before applying energy to the tissue) with the post-treatment image data (the tissue image taken after applying energy to the tissue). The parameter can be the maximum displacement value or the average displacement value of the tissue. If the displacement parameter is greater than the threshold displacement, it can be evaluated that the application of energy is likely to have caused the desired targeted physiological perturbation.

[0045] In one example, the present technique can be used to diagnose a subject having a metabolic disorder. The present technique can also be used to diagnose and / or treat a subject having a glucose regulation disorder. Thus, the present technique can be used to promote the homeostasis of a target molecule or to promote a desired circulating concentration or concentration range of one or more target molecules (e.g., glucose, insulin, glucagon, or combinations thereof). In one embodiment, the present technique is used to evaluate a clinical condition related to circulating (i.e., blood) glucose levels.

[0046] In one embodiment, a threshold of glucose concentration can be used to identify blood glucose levels outside or within the normal range as part of the diagnosis of metabolic dysfunction. Fasting: Less than 50 mg / dL (2.8 mmol / L): Insulin shock 50 - 70 mg / dL (2.8 - 3.9 mmol / L): Hypoglycemia / hypoglycaemia 70 - 110 mg / dL (3.9 - 6.1 mmol / L): Normal 110 - 125 mg / dL (6.1 - 6.9 mmol / L): Elevated / impaired (prediabetes) 125 (7 mmol / L) or higher: Diabetes Non - fasting (about 2 hours after a meal): 70 - 140 mg / dL: Normal 140 - 199 mg / dL (8 - 11 mmol / L): Elevated or “borderline” / diabetes Greater than 200 mg / dL (11 mmol / L): Diabetes

[0047] However, as provided herein, the present technology enables the assessment of perturbations as a result of neuromodulation even in the absence of glucose information during fasting or non-fasting. In this way, patient non-compliance or false reporting can be excluded as confounding factors for the assessment of metabolic dysfunction. That is, a subject with an unknown fasting or non-fasting state can be perturbed into an approximate or pseudo-fasting state via neuromodulation, even if the subject has not fasted, for example, has eaten recently. As provided herein, the approximate fasting state can refer to the state of a patient after neuromodulatory energy has been applied (e.g., within 1 to 6 hours later, within 30 minutes later) to a target region related to metabolism, such as the liver, pancreas, GI. Based on the profile of the subject or the response to neuromodulation, the presence or absence of metabolic dysfunction can be identified.

[0048] For example, the above normal glucose range can be used as a recovery target for tracking the time to recovery after a perturbation-induced change in glucose level caused by neuromodulation. In another example, a subject with elevated fasting glucose levels who does not experience a change in glucose level after neuromodulation is diagnosed with insufficient insulin production (e.g., pancreatic disorder). In contrast, if the same subject significantly (more than the expected amount) decreases their glucose level after neuromodulation, it may indicate a problem in the brain control center (e.g., glucose set point). Thus, in one embodiment, the response to a perturbation enables the subject to be classified based on those specific response profiles (e.g., lack of change in glucose versus change in glucose concentration). In one embodiment, a perturbation is induced and the concentration of one or more target molecules is evaluated. Based on whether the concentration is above or below a threshold, the subject is classified into a first category or a second category. Further, those response profiles can be classified relative to their baseline state. A subject with normal glucose levels who does not experience a change in glucose after a neuromodulation-induced perturbation is classified differently (e.g., placed in a first category) from a subject with high glucose levels who does not experience a change in glucose after a neuromodulation-induced perturbation (e.g., placed in a second category). Thus, the level of change relative to the baseline is used to classify the subject. A change relative to the baseline that is less than a predetermined threshold is associated with the first category, and a change greater than the predetermined threshold is associated with the second category. It should be understood that the disclosed categories may include additional subcategories for the subject.

[0049] In another example, the technology induces a perturbation in both fasting and non-fasting subjects. Thus, a subject may prefer to undergo diagnostic neuromodulation without a change in diet content. Further, in certain embodiments, the subject also does not need to receive an intravenous administration of a drug (e.g., glucose, insulin).

[0050] In another example, as provided herein, neuromodulation of lymphoid tissue can result in perturbation of immune activity or function. In certain embodiments, neuromodulation of lymph nodes results in local enlargement of the lymph nodes as compared to contralateral lymph nodes. Hypertrophy or dysplasia may be associated with molecular changes in key barrier tissues (such as high endothelial venules (HEVs) within lymph nodes), including changes in perilymphatic muscle cell tone, long-term mobilization and rearrangement of lymphatic vessels around lymph nodes, and / or changes in key transport proteins (such as aquaporins (water / fluid transport) or CCL 21 / CXCL 13 secretion (cellular chemokines)). Perturbation can result in dramatic changes in the overall lymph node tissue microenvironment, including cell density, cell number, and enlargement. Furthermore, activation of lymph nodes can result in an activation cascade that amplifies or augments local activation to systemically activate the lymphatic system. That is, local stimulation can result in downstream and upstream activation of the lymphatic system. Thus, local perturbation can be used to activate a systemic immune response. Further perturbations can include changes in lymphatic fluid flow, immune cell transport to / from lymphoid tissue, changes in immune cell phenotype or local immune response, and / or antigen transport to / from lymphoid tissue. Local stimulation can enable tissue- or location-specific increases in the mobilization of lymphatic fluid or immune cells. Thus, in one embodiment, evaluation of perturbations to neuromodulation and immune activity (or lack thereof) of lymphoid tissue can be used to diagnose immunodysfunction or immune disorders.

[0051] FIG. 4 is a flow diagram of a method 50 of causing perturbation via neuromodulation that can be used as part of a diagnostic protocol. In method 50, a target region is spatially selected 52. An energy application device is arranged in step 54 such that an energy pulse is focused on the desired target region, and a pulse generator applies a plurality of energy pulses to the target region of the target tissue in step 56 to preferentially activate a subset of synapses within the target tissue, for example, stimulate axonal terminals to release neurotransmitters, and / or induce a change in neurotransmitter release, and / or induce a change in the activity of non-neuronal cells (within the synapse) to cause a physiological perturbation targeted in step 58 as provided herein. In certain embodiments, the method can include a step of evaluating the effect of the perturbation. For example, one or more direct or indirect evaluations of the function or state of the tissue can be used.

[0052] In one embodiment, an evaluation is performed before and after applying an energy pulse to evaluate the change in glucose concentration as a result of the adjustment. Further, the characteristics or states to be evaluated may be values or metrics, such as flow rate, concentration, cell population, or any combination thereof, and these may be analyzed by appropriate techniques. For example, a relative change exceeding a threshold is used for diagnosing a subject. The diagnosis is evaluated through measured perturbations such as the presence or absence of an increase in tissue structure size (e.g., lymph node size) or a change in the concentration of one or more release molecules (e.g., relative to the baseline concentration before neuromodulation). In one embodiment, the perturbation includes an increase in concentration exceeding a threshold, such as exceeding about 50%, 100%, 200%, 400%, 1000% compared to the baseline. The evaluation includes tracking a decrease in the concentration of the molecule over time, such as a decrease of at least 10%, 20%, 30%, 50%, or 75% of the target molecule. Further, for a particular subject, the diagnosis may be related to a relatively stable concentration of a particular molecule in the context of other clinical events that may tend to increase the concentration of the molecule. That is, a normal subject may respond to glucose administration simultaneously or after neuromodulation in a manner distinguishable from an insulin-resistant subject. The increase or decrease or other induced and measurable effects are measured within a certain time frame from the start of the treatment, such as within about 5 minutes, within about 30 minutes.

[0053] FIG. 5 is a flow diagram of a method 60 for evaluating perturbations of one or more target molecules relative to a baseline to diagnose a subject or part of a diagnostic protocol. For example, the disclosed techniques can be used to classify a subject into categories such as first, second, third, etc. based on the level of perturbation response and / or recovery back to baseline. The baseline concentration (block 62) can be determined based on the molecular concentration at or before perturbation and can be a single baseline concentration or an average over several time points before perturbation. The diagnostic protocol can also include steps to cause a physiological perturbation via neuromodulation of a target region of the target tissue of the subject (e.g., as in method 50 of FIG. 4) (block 64). The target tissue can be selected based on the diagnostic protocol. For example, the diagnosis of glucose metabolism disorders can involve neuromodulation of the liver and / or pancreas, while the diagnosis of immune function disorders can involve neuromodulation of the spleen and / or lymphoid tissue. The target region is selected based on the target tissue disclosed with respect to FIG. 4. Deviations in the concentration of one or more target molecules relative to the baseline are determined at one or more time points (block 66). The clinical state of the subject can be determined based on an evaluation at one or more time points after application of energy to the target region (block 68). The time points of change can be determined based on empirical evidence. For example, a particular subject can exhibit a change in the concentration of the target molecule by one minute, five minutes, ten minutes, or up to sixty minutes after neuromodulation. Additionally, such changes may dissipate towards recovery after a certain period (e.g., three hours later, twelve hours later, twenty-four hours later). Thus, the evaluation of the change is performed during the period in which a physiological perturbation is expected to be observed, at least in a particular population of subjects or for a particular diagnosis. Additionally, it should be understood that the concentration at additional time points during the recovery period can be evaluated. Further, the concentration of the target molecule over a period of time can be evaluated for a predicted increase or decrease (or lack of change) associated with one or more clinical states.

[0054] Method 60 can be used to diagnose glucose metabolism disorders. In one embodiment, the technique can cause a physiological perturbation that results in an overall decrease in circulating glucose, or a change in the concentration or rate of change of glucose relative to circulating hormone concentrations (such as insulin or glucagon), after neuromodulation of the liver (e.g., in a target region near or at the porta hepatis). Such changes can be related to the clinical state of the subject. For example, in a diabetic or prediabetic subject, the baseline circulating glucose level can be a level exceeding 140 mg / dL. After the perturbation caused by liver neuromodulation, the level of perturbed glucose can decrease to a level consistent with a non-diabetic subject, e.g., a level less than 140 mg / dL. Thus, a high baseline circulating glucose and a perturbed deviation from baseline caused by liver neuromodulation can be diagnosed as a diabetic or prediabetic subject. In contrast, a normal baseline circulating glucose and no perturbation or a small perturbation of circulating glucose caused by liver neuromodulation can be diagnosed as a healthy or non-diabetic subject. Further, a high baseline circulating glucose, and no deviation or only slightly perturbed deviation from baseline caused by liver neuromodulation, can be diagnosed as an organ dysfunction, regardless of the presence or absence of diabetes. Thus, such subjects are considered non-responsive to neuromodulation, and alternative therapies are recommended.

[0055] Changes in circulating glucose can be evaluated in the context of additional target molecules. For example, the relationship between glucose and insulin and the resulting perturbations can indicate the clinical state of a subject. In one embodiment, physiological perturbations via targeted liver ultrasound cause observable changes in circulating glucose, insulin, cortisol, and triglycerides in diabetic or prediabetic subjects compared to baseline. Perturbations that cause a decrease in glucose without a concomitant change in insulin can indicate a first type of glucose metabolism disorder, and perturbations that cause a decrease in glucose with a concomitant change in insulin can indicate a second type of glucose metabolism disorder. That is, the magnitude of the delta, which can be changes (or lack thereof) in glucose and / or insulin concentrations observed over time, can be a marker of glucose metabolism disorder. In one example, insulin resistance is characterized by subjects with elevated circulating insulin. Due to the likelihood that mechanical waves can cause the release of pancreatic insulin stores in response to the neuromodulation of the pancreas by the present technique, there is an initial increase in circulating insulin. However, following the release of existing pancreatic insulin stores that causes the initial increase, there is an overall subsequent decrease in circulating insulin compared to both the levels seen in the initial increase and the initial or baseline circulating insulin prior to neuromodulation of the pancreas. Observing the response to pancreatic neuromodulation in subjects with elevated circulating insulin and / or elevated circulating glucose that exceeds a threshold either simultaneously with or separate from liver neuromodulation and tracking the initial increase and subsequent decrease in circulating insulin can be used to classify subjects as insulin resistant without the need to fast the subjects. Further, the absence of such characteristic responses can be used to identify subjects who show elevated circulating insulin and / or elevated circulating glucose that exceeds a threshold but do not have insulin resistance and show such concentrations as a result of other dysfunctional metabolic pathways. By observing one or more types of perturbations caused by neuromodulation, subjects can be systematized based on a specific profile of changes or grouped into different categories.The follow-up diagnostic protocol can track deviations from previous classifications and can then be used to track the prognosis of a subject.

[0056] Lymphoid tissue neuromodulation can be used to perturb populations of immune cells produced by lymphoid structures. In one embodiment, neuromodulation of lymph nodes can result in an increase in the population of lymphocytes circulating in lymph fluid. Thus, the local concentration profiles of type 1 (inflammatory) cytokines (e.g., IL-12, TNF-α, IFN-γ, IL-2, TNF-β) and type 2 (anti-inflammatory) cytokines (e.g., IL-4, IL-10, IL-13, IL-6) can be evaluated. In one embodiment, neuromodulation of lymph nodes can result in an increase or decrease in B cells or T cells circulating over lymph fluid, or an increase or decrease in B cells or T cells or dendritic cells mobilized to lymphoid tissue. Thus, neuromodulation of lymphoid tissue can provide opportunities for cell migration patterns. Such migration patterns can be observed using in vivo bioluminescence imaging. Other features can include changes in lymphatic drainage patterns. In certain embodiments, these changes can be features of the perturbation used to diagnose a subject. Healthy subjects, immunocompromised or immunodeficient subjects, and subjects with a hyperactive immune response can experience different perturbations as a result of neuromodulation of lymphoid tissue. The characteristics of these perturbations can be evaluated to create a profile. A subject with an unknown clinical condition can be diagnosed according to the best match to a characteristic profile. For example, an immunodeficient subject can have a characteristic lack or low level of perturbation in response to targeted splenic neuromodulation. In certain embodiments, the occipital lymph nodes, auricular lymph nodes, cervical lymph nodes, axillary lymph nodes, inguinal lymph nodes, pulmonary lymph nodes, mediastinal lymph nodes, intraperitoneal lymph nodes, or supratrochlear lymph nodes can be targeted.

[0057] FIG. 6 is a method 69 for identifying individual portions of an irregular metabolic pathway based on a perturbation pattern. To induce neuromodulation and the resulting perturbations, energy can be applied to individual target regions (e.g., a first target region, a second target region) within different organs (e.g., a first organ, a second organ) (block 70). The energy application may be continuous (e.g., at different times) to enable assessment of the characteristics of the perturbations in each organ (block 71). The characteristics of the perturbations include measurable physiological changes that are evaluated. Based on the evaluation, a clinical condition can be diagnosed (block 72). For example, to identify the point in time of a dysfunction in glucose metabolism, energy can be applied to the liver and pancreas at different times. A subject with an affected pancreas may respond differently than a subject who is insulin resistant. That is, an insulin-resistant subject may respond to an increase in the level of circulating insulin as a result of a targeted pancreatic stimulation. However, a subject with an affected pancreas may not be able to produce insulin and thus may not exhibit the characteristic of an increase in circulating insulin as a result of neuromodulation of the pancreas. Insulin-resistant patients may also respond to a decrease in blood glucose that does not coincide with an increase in insulin concentration after stimulation of these sites, since stimulation of peripheral sensory sites (such as the liver or GI tract) acts on other CNS-based sensory sites that are deficient in glucose sensors and control. Other techniques may not distinguish these scenarios.

[0058] The energy application device 12 may be configured as an extracorporeal non-invasive device or an internal device, such as a minimally invasive device. As described above, the energy application device 12 may be an extracorporeal non-invasive ultrasonic transducer or a mechanical actuator. For example, FIG. 7 shows an embodiment of the energy application device 12 configured as a hand-held ultrasonic probe including an ultrasonic transducer 74. However, it should be understood that other non-invasive implementation forms are contemplated, including other methods for constructing, attaching, or placing an ultrasonic transducer probe on an anatomical target. Further, in addition to the hand-held configuration, the energy application device 12 can include a steering mechanism that responds to commands from the controller 16. The steering mechanism can orient or direct the energy application device 12 towards the target tissue 43 (or structure), and then the controller 16 can focus the energy application on the target area 44. The ultrasonic transducer 74 can include an imaging transducer 74A that is used for spatial selection of the target area 44 within the target tissue 43. The ultrasonic transducer 74 may include a treatment transducer 74B that applies neuromodulation energy.

[0059] In some embodiments, ultrasonic images can be used to induce ultrasonic stimulation and spatially select a target region for targeted delivery of the ultrasonic stimulation. As provided herein, spatial selection or spatially selecting can include acquiring an image of a tissue or organ (or a part of a tissue or organ) and identifying a target region within the organ based on the image (e.g., an ultrasonic image). In some embodiments, the tissue or organ can have anatomical features that are used to induce the selection of the target region within the organ. Such features can include, in some embodiments, by way of non-limiting example, the entry sites of blood vessels or nerves into the organ, tissue types within the organ, the interior or edges of the organ, or sub-organ structures. In certain embodiments, the anatomical features can include the porta hepatis, lower gastrointestinal organs (stomach, small intestine, large intestine), pancreatic ducts, or white splenic pulp. By identifying the anatomical features within the image, the target region can be selected to overlap with, include, or be adjacent to the anatomical features. In other embodiments, the anatomical features may be excluded from the target region. For example, intestinal tissue rather than gastric tissue may be selected as the target region. The identification of the anatomical features can be performed via morphological features visible within the image (e.g., visible in an ultrasonic image) or by the structural recognition features of the imaging modality used to acquire the image. As disclosed herein, the system 10 can be configured such that the energy application device 12 operates in an imaging mode to acquire an image and then operates in an energy application mode after the image is acquired and the target region is spatially selected based on the image.

[0060] In other embodiments, the target region can be identified by the presence or absence of one or more biological markers. Such markers can be evaluated by staining an organ or tissue, obtaining an image showing the staining, and identifying the region of the organ or tissue containing the biological marker. In some embodiments, biological marker information can be obtained by in vivo staining techniques for obtaining real-time position data of biological markers in tissues or organs specific to a subject. In other embodiments, biological marker information may be obtained by in vitro staining techniques to obtain position data of one or more representative images, which can then be used to predict the position of biological markers within the tissues or organs of a subject. In some embodiments, the target region is selected to correspond to a portion of a tissue or organ that is rich in a particular biological marker or lacks a particular biological marker. For example, one or more biological markers can include markers of neural structures (e.g., myelin sheath markers).

[0061] The target region of an organ or tissue can be spatially selected based on an operator's input. For example, the operator can specify the target region on the acquired image by directly manipulating the image (i.e., drawing or writing the target region on the image) or by providing image coordinate information corresponding to the target region. In another embodiment, the target region may be automatically selected based on the image data to achieve spatial selection. In some embodiments, spatial selection includes storing data related to the target region in memory and accessing the data.

[0062] Once spatially selected, the system 10 is configured to apply energy to the target region as provided herein.

[0063] Example: Targeted Physiological Perturbation Before initiating neuromodulation, a GE Vivid E 9 ultrasound system and an 11L probe were used for ultrasound scanning. The focal region corresponding to the internal region of the subject was marked on the animal's skin. The HIFU transducer was placed on the marked region. Another ultrasound scan was also performed using a smaller imaging probe (3S) placed at the opening of the HIFU transducer. The imaging beam of the 3S probe was aligned with the HIFU beam. Thus, using the image of the targeted organ (visualized by the ultrasound scanner), it was possible to confirm that the HIFU beam was targeted at the target region.

[0064] Animal Protocol Adult male obese Zucker rats (250 - 300 g, Charles River Laboratories) at 8 - 12 weeks of age were maintained under conditions supplied by the provider to promote the development of insulin resistance and hyperglycemia. The rats were fed a high-fat diet. Neuromodulation was performed on the liver using the applied ultrasonic energy. The ultrasonic application was performed for 1 minute. Blood samples were collected 15 minutes after the last ultrasonic treatment to analyze changes in circulating catecholamine concentrations (e.g., norepinephrine and dopamine). Terminal blood samples were collected 60 - 90 minutes after the last ultrasonic treatment to analyze changes in circulating molecular concentrations. The blood samples were stored with an anticoagulant (disodium) EDTA to prevent sample coagulation. The protocol used for ultrasonic neuromodulation can be as follows.

[0065] (A) The animal may be anesthetized with 2 - 4% isoflurane. (B) To prevent a rise in body temperature during treatment, the animal may be laid on a water-circulating heating pad. (C) The area above the targeted region (e.g., the target nerve) for ultrasonic stimulation can be shaved with a disposable razor and animal clipper before stimulation. (D) Diagnostic ultrasound imaging can be used to spatially select the target region. (E) The area may be marked with a permanent marker for later identification. (F)Either an FUS ultrasonic probe or a LogiQ E 9 probe can be placed in a designated target area previously identified by diagnostic imaging ultrasound. (G)Subsequently, an ultrasonic pulse can be executed with a total duration of a single stimulus not exceeding a single 1-minute pulse. The energy of the ultrasonic pulse does not reach levels associated with thermal damage and ablation or cavitation (e.g., 35 W / cm2). (H)A second 1-minute ultrasonic pulse can be applied. (I)Subsequently, for acute studies (e.g., 1 hour) and kinetic studies, the animals can be incubated under anesthesia. Then, the animals are sacrificed and tissue and blood samples are collected.

[0066] The incision may start at the base of the peritoneal cavity and extend to the pleural cavity. Organs are quickly removed and homogenized in PBS solution containing phosphatase (0.2 mM phenylmethylsulfonyl fluoride, 5 μg / mL aprotinin, 1 mM benzamidine, 1 mM sodium orthovanadate, and 2 μM cantharidin) and protease (1 μL - 20 mg of tissue by Roche Diagnostics) inhibitors. A targeted final concentration of 0.2 g of tissue per 1 mL of PBS solution was applied to all samples. Blood samples were stored with an anticoagulant (disodium) EDTA to prevent sample clotting. Next the samples are stored at -80 °C until analysis.

[0067] Target Tissue Stimulation and Physiological Perturbation for the Diagnosis of Glucose Metabolism Disorders This example demonstrates a non-invasive method for achieving a physiological perturbation to evaluate a patient's metabolic dysfunction. The disclosed technique offers advantages compared to techniques for evaluating metabolic dysfunction that involve several hours of patient time, including fasting, infusion, and subsequent response / blood sampling steps. For example, in the evaluation of a patient's insulin resistance by a specific procedure, the patient may undergo a fasting period, which enables the patient's physiological system to reach a set point with respect to the population. After fasting, glucose or a metabolically active compound (e.g., orally or by injection) may be administered to the patient. The patient's response is then often measured over a defined period of several hours by monitoring changes in glucose and / or insulin or changes in other hormones that assist in the change of glucose concentration. If either fasting blood glucose or insulin is determined to be outside the range (determined by measuring a previous subject population), the subject may be diagnosed with diabetes. A glucose tolerance test measures how quickly a bolus of glucose is removed from the blood in a fasting state (compared to a population of subjects) and is used to diagnose diabetes in subjects with higher (but not pathologically high) fasting glucose concentrations. The clamp technique is used to quantify how a subject metabolizes glucose. The clamp can include a hyperglycemic clamp (continuous infusion of glucose, quantifying insulin secretory capacity) or a hyperinsulinemic clamp (continuous infusion of insulin, quantifying insulin resistance), which pose problems regarding the cost of evaluation and the excessive complexity affects use in a clinic.

[0068] The HOMA (Homeostatic Model Assessment) calculator uses a mathematical model to estimate insulin sensitivity and B-cell function from plasma insulin and glucose concentrations. This interaction between glucose and insulin in the basal state provides information about the balance between hepatic glucose output and insulin secretion, which is maintained by a feedback loop sustained by the liver and pancreatic B-cells, enabling HOMA to function as a surrogate measure of steady-state beta-cell function (%B) and insulin sensitivity (%S) as a percentage of a normal reference population. The predictions used in the model arise from experimental data in humans and animals.

[0069] The disclosed technique can replace or be used in conjunction with other techniques for the diagnosis of glucose metabolism disorders to eliminate or shorten fasting and subsequent assessment times. In one embodiment, the modified HOMA calculator can more closely approximate the fasting state of a diabetic subject based on the disclosed neuromodulation-induced perturbation. That is, rather than requiring the subject to fast for a certain period of time, for the purpose of evaluating the glucose response, the subject (e.g., diabetic, normal, or unknown) may be perturbed by neuromodulation to lower circulating glucose to a state similar to the fasting state. Further, neuromodulation induces stabilization of the glucose level independent of the feeding state, as indicated by the absence of fluctuations in the glucose level. In summary, this neuromodulation-induced glucose stabilization reduces potential confounding results derived from the feeding state assessment and enables the evaluation of insulin sensitivity in the absence of feeding-derived effects.

[0070] As provided herein, ultrasonic stimulation was performed according to the timeline shown in FIG. 9, showing an induced target physiological perturbation relative to the control. FIG. 10 shows the results of ultrasonic stimulation of the liver (porta hepatis) of obese Zucker rats compared to control rats that received sham ultrasonic treatment. The ultrasonic-treated rats were protected from the increase in circulating glucose in the control rats, even though both groups of rats were raised in conditions associated with the development of insulin resistance and hyperglycemia. Thus, the results demonstrate that ultrasonic treatment causes perturbations in a hyperglycemic rat population detectable via changes in circulating glucose. FIGS. 11 and 12 show that food consumption and weight were similar between the treatment and control groups. While food intake remained constant (FIG. 11), weight gain in the treated animals decelerated relative to the control group (FIG. 12) over the experimental timeline.

[0071] As provided herein, hepatic neuromodulation, as shown in FIG. 13, causes changes in glucose metabolism to provide a controlled / accurate stimulus, modify inter-organ relationships, and affect or change the current clinical score of homeostasis (i.e., the HOMA score). The disclosed techniques can be used to generate metrics that reflect neuromodulation and the resulting perturbations. Typically, the HOMA score indicates the relationship between fasting blood glucose and fasting insulin measurements. Generally, a high HOMA score reflects a high insulin level required for glucose processing. As provided herein, changes in insulin and glucose concentrations can be evaluated in response to hepatic neuromodulation as part of a diagnostic protocol to derive a homeostasis metric that can be used to diagnose and track insulin resistance, even in subjects with normal-range glucose levels or not on a controlled diet.

[0072] The steady-state HOMA model is based on circulating glucose levels and circulating insulin levels according to a population-based formula. Assume the subject is fasting. Further, typical assessments are based on "stimulated" responses such as glucose clamps, insulin clamps, or glucose tolerance tests. In certain embodiments, neuromodulation replaces typical fasting and stimulation (e.g., glucose clamp, insulin clamp, glucose tolerance). In one embodiment, the present technology can be used to replace glucose and / or insulin infusions with neuromodulation so that testing can be performed without the administration of glucose and / or insulin. However, it should be understood that neuromodulation followed by the administration of glucose and / or insulin is also contemplated. In another embodiment, the present technology can be used to eliminate fasting from the measurement protocol (by using neuromodulation) to provide an immediate deviation from baseline organ dynamics / interactions.

[0073] As provided herein, the present technology can be used to construct a population-based model of the relationship between changes in glucose and insulin concentration responses to neuromodulation. The model may be based on the glucose-insulin relationship of a population of healthy individuals perturbed in a fasting state approximated via neuromodulation. In certain embodiments, the model may further be based on the glucose-insulin relationship for a population of individuals with various types of metabolic functions perturbed in a fasting state approximated via neuromodulation. The model can be applied to glucose concentration and insulin concentration data obtained from neuromodulated subjects perturbed in an approximated fasting state to generate a score, which indicates the level of insulin resistance in the subject.

[0074] In one embodiment, FIG. 14 is a flow diagram of a method 90 for metabolic dysfunction assessment technology. This technology obtains baseline concentrations of glucose and / or insulin at step 92, which can be immediately before (within 1 to 6 hours) or simultaneous with the initiation of neuromodulation in a target region (e.g., liver, GI, pancreas) within the target tissue at step 94. As described herein, a continuous glucose monitor can be used to monitor the subject so that glucose concentrations are continuously available. Based on the observed perturbations that can be reflected in the observation of changes in glucose and / or insulin concentrations over time as a result of neuromodulation at step 96, the clinical state of the patient can be determined at step 98.

[0075] This technology can be used to stratify a population of subjects based on the level of post-stimulation glucose and / or insulin changes or to diagnose levels of diabetes type and / or resistance. For example, a subject can be classified as a "responder" or "strong responder" or "low responder" or "non-responder" according to the level of glucose and / or insulin changes. For example, a non-responder may show a concentration change less than a predetermined threshold relative to the baseline concentration, while a responder experiences a concentration change exceeding the predetermined threshold. In one example, a non-responder is considered to show a change in glucose of less than about 5%, less than about 10%, or less than about 15% relative to the baseline. A responder is considered to show a change in glucose exceeding about 10 - 15% relative to the baseline, while a strong responder is considered to show a change in glucose exceeding about 50% relative to the baseline. Responsiveness can be at a time point related to the expected changes in a healthy or responsive population.

[0076] Figure 15 is an example of a neuromodulation response profile for a highly insulin-resistant animal subject given a diet that drives insulin resistance and high glucose levels. The profile shows glucose concentration 100 over several days prior to the onset of neuromodulation, showing a relatively stable high glucose concentration above a threshold shown as a glucose concentration of 225 mg / dL. Neuromodulation is started at baseline 102 and the glucose response to neuromodulation is tracked over time. The illustrated subject is characterized by a rapid decrease in glucose of long duration and magnitude without fasting or glucose administration. As provided herein, the response profile can be evaluated or characterized by one or more measures of the response. In one embodiment, the magnitude of the change from peak baseline concentration 102 to trough 106 can be evaluated. Alternatively or additionally, the gradient 104 can be evaluated (e.g., a steep gradient may indicate insulin resistance). Additional metrics can include the gradient from trough 106 to recovery peak 110, the difference between the baseline concentration and the first recovery peak 110, the time to the first recovery peak 101, and / or the area under the curve 112 relative to the set point. For example, an insulin-resistant subject may have a relatively short period (e.g., less than 225 mg / dL, less than 200 mg / dL) of lower glucose concentration below a predetermined threshold before recovering to a concentration above the predetermined threshold, which is reflected in the area under the curve 112 below the empirically determined threshold. The subject may be monitored during normal activities such as diet and sleep, and the characteristic profile may reflect changes in response to these activities relative to the baseline. Previous techniques have sometimes included complex glucose administration response tracking over several hours in addition to pre-test fasting, but this technique allows for the observation of changes over a relatively short period, which is less burdensome on the subjects being tested. Thus, in one embodiment, the subject is evaluated using the change in glucose concentration observed within 12 hours, 6 hours, 5 hours, 4 hours, 3 hours, 2 hours, or 1 hour after neuromodulation to achieve an approximate fasting state in the subject.Furthermore, glucose monitoring can be achieved using a glucose monitor that enables the subject to be outside the outpatient and clinical environments, so the technology provides a more convenient and rapid assessment of metabolic dysfunction that does not require fasting or administration of a glucose bolus.

[0077] Figure 16 is an example of a neuromodulation response profile of an insulin-resistant subject having an initial glucose concentration lower than that of the subject of Figure 15. This profile shows the pre-treatment glucose concentration 124 above a threshold (e.g., about 250 mg / dL), and the area under the curve 112 to a higher baseline concentration 122 or set point that decreased to a trough 126 in response to a neuromodulation and recovery peak 128. For example, an insulin-resistant subject may have a relatively short period (e.g., less than 225 mg / dL, less than 200 mg / dL) of a lower glucose concentration below a given threshold before recovering to a concentration above the given threshold. The response profile shows a rapid decrease in glucose concentration, albeit of a smaller magnitude and duration. Thus, in certain embodiments, the response profiles in Figures 15 and 16 can both be classified as insulin-resistant, but are different bands or categories. Figure 17 is a response profile of a subject whose initial glucose concentration 130 is below diabetic levels and has little or no response to neuromodulation. Thus, the subject may be considered a non-responder and can be classified differently from the profiles of Figures 15 and 16.

[0078] In another embodiment, the disclosed technology can be used to detect differences in a subject population with respect to various perturbations and their associated clinical states, and / or optimal treatment plans. Differences in the subject population can result in categorizing the subjects into one of several categories and initiating treatment options based on the categorization. For example, drug response can correlate with responsiveness to neuromodulation (e.g., degree of perturbation). Further, the disclosed technology can provide additional data as part of the categorical information (insulin resistant, non-insulin resistant, responder, non-responder) and reporting to caregivers.

[0079] Target tissue stimulation and physiological perturbation for identification of pathogen or toxin exposure As provided herein, the neuromodulation response can indicate exposure to a pathogen, toxin, or environment prior to the time when the pathogen itself is detectable or disease symptoms appear (i.e., such as fever). That is, the technology provides an improved and more rapid assessment of pathogen exposure. Such an assessment can improve the outcome of hospital-associated pathogens that may be more effectively contained if detected more rapidly. Certain techniques for tracking the immune response or systemic response require continuous monitoring to detect exposure to a pathogen, but the technology eliminates the need for baseline monitoring by comparing changes before and after neuromodulation in the expression of specific markers.

[0080] FIG. 18 is a flowchart of a method 150 showing an immune response assessment technique for identifying pathogen exposure by evaluating the response of a region to neuromodulation when interested in immune tissues or structures including lymphoid organs, lymphatic vessels that extend throughout the body and provide flow and drainage, the spleen, lymph nodes, Peyer's patches, and accessory lymphoid tissues (including tonsils and appendix).

[0081] This technique, at step 152, obtains the baseline concentration of one or more markers such as immune markers, which can be just before (within 1 to 6 hours) or simultaneous with the initiation of neuromodulation of the target area (e.g., spleen or immune tissue) at step 154. Based on the observed perturbation, which can be reflected in the observation of the change of one or more markers relative to the baseline at step 156, pathogen exposure can be identified at step 158. Changes in the concentration of one or more markers in response to neuromodulation associated with pathogen exposure or lack of pathogen exposure can be identified according to the techniques provided herein (system changes identified by blood concentration, tissue concentration, sensors and / or image data).

[0082] Figure 19 shows the ultrasonic dose-dependent changes in tumor necrosis factor mRNA expression for animal subjects exposed to endotoxin or lipopolysaccharide (LPS) and treated with neuromodulatory energy applied to the spleen. High-dose treated animals experienced the greatest decrease in tumor necrosis factor mRNA expression compared to the control. Figure 20 shows mRNA expression array data for several markers for control animals as well as LPS-exposed animals and high-dose and low-dose ultrasonic treated animals. Figure 21 is a diagram showing an exemplary protocol for obtaining baseline and potential post-exposure data to evaluate changes in responses based on neuromodulation after exposure to a substance or pathogen. A baseline or characteristic pre-exposure response profile can be obtained and compared to a potential post-exposure response profile to evaluate changes caused by the exposure.

[0083] The disclosed technology provided herein utilizes physiologic outcomes targeted to be achievable by neuromodulation. In addition to implementations in which the physiologic outcomes targeted for treating a subject are used, these outcomes can be used as part of a diagnostic protocol. For example, applying energy to a target region within a target tissue can result in predictable and targeted physiologic perturbations. One or more characteristics of the perturbation itself or the subject's response to the perturbation can indicate the clinical state of the subject. For example, a rapid return to homeostasis within a predetermined period (e.g., returning to the baseline pre-regulation concentration of a target molecule) can indicate a healthy response to the perturbation. In contrast, a slow response can indicate a metabolic pathway with lower performance compared to the metabolic pathways of a healthy subject. Although particular embodiments of the present disclosure have been discussed in the context of glucose regulation, it is to be understood that the technology can be used to induce perturbations in other systems (including acute or chronic inflammatory conditions and related sensory and neuroimmune effector systems), and thus to evaluate the associated clinical states.

[0084] This specification uses examples to disclose the invention, including the best mode, and to enable any person skilled in the art to make and use the invention, including making and using any device or system and performing any incorporated method. The patentable scope of the invention is defined by the claims and can include other examples that occur to those skilled in the art. Such other examples are intended to be within the scope of the claims if they have structural elements that do not differ from the literal language of the claims, or if they include equivalent structural elements that do not differ substantially from the literal language of the claims.

Explanation of Reference Numerals

[0085] 10 System 12 Energy Application Device 14 Pulse Generator 16 Controller 20 Evaluation Device 30 Processor 32 Memory 33 Lead 34 Input / Output Circuit 36 Display 42 Ultrasonic transducer 43 Target tissue 44 Target area 50 Method 60 Method 69 Method 74 Ultrasonic transducer 74A Imaging transducer 74B Treatment transducer 90 Method 122 Baseline concentration 124 Glucose concentration before treatment 126 Trough 128 Recovery peak 130 Glucose concentration at start 150 Method

Claims

1. 1. A regulation system comprising: an energy application device configured to apply energy to a region of interest to cause a physiological perturbation in a subject; A controller, controlling the application of energy to the region of interest via the energy application device to induce the physiological perturbation to cause a change in concentration of one or more molecules of interest in the subject relative to a baseline concentration; receiving information indicative of the concentration of the one or more molecules of interest at one or more time points after the application of the energy; determining that the subject is in a category selected from two or more categories based on a change in the concentration of the one or more molecules of interest relative to the baseline concentration over a period of time; The controller and The adjustment system comprises:

2. The system of claim 1 , wherein the concentration is a glucose concentration received from an evaluation device that includes a glucose monitor.

3. 2. The system of claim 1, wherein the selected category reflects exposure to a pathogen and another of the two or more categories reflects lack of exposure to a pathogen, and the region of interest is a spleen or immune tissue.

4. The system of claim 1 , wherein the energy applicator is an ultrasound probe.

5. 2. The system of claim 1, wherein the subject is determined to be in the selected category if the change in concentration of an individual molecule of the one or more target molecules relative to a baseline concentration is less than a predetermined threshold, and the subject is determined to be in a second category of the two or more categories if the change in concentration is greater than the predetermined threshold.

6. The system of claim 5 , wherein the selected category is not responsive to neuromodulation and the second category is responsive to neuromodulation.

7. The system of claim 5 , wherein the system is configured to provide an indication of a neuromodulation treatment protocol upon determining that the subject is in the second category.

8. The system of claim 5 , wherein the controller is configured to initiate a neuromodulation treatment protocol via the energy application device upon determining that the subject is in the second category.

9. 1. A method of inducing a physiological perturbation in a subject, comprising: The method comprises: Directing energy from an energy application device to a target area of ​​a subject; applying the energy to the region of interest and, as a result of the application, modulating activity of at least one axon terminal within the region of interest; assessing one or more characteristics of the physiological perturbation at one or more time points after the perturbation; providing an indication of a clinical status of the subject based on said evaluation. A method comprising:

10. Evaluating one or more characteristics of the perturbation may include: determining a concentration of the molecule of interest at a first time point after applying the energy; determining a concentration of the molecule of interest at a second time point after applying the energy, the second time point being after the first time point; and 10. The method of claim 9, comprising:

11. 11. The method of claim 10, wherein evaluating one or more characteristics of the perturbation comprises determining a difference between the concentrations of the molecule of interest at the first time point and the second time point, determining a gradient of the concentration of the molecule of interest between the first time point and the second time point, or both.

12. 10. The method of claim 9, wherein evaluating one or more characteristics of the perturbation comprises determining a time between a trough and a recovery peak of the molecule of interest.

13. 10. The method of claim 9, comprising causing the energy application device to deliver treatment energy to the target area, the treatment energy being applied according to a treatment protocol selected based on the clinical condition.

14. 10. The method of claim 9, wherein the region of interest is in the pancreas, and evaluating the one or more characteristics comprises determining a concentration of circulating insulin at the multiple time points after the application and comparing the concentration at the multiple time points to a baseline concentration of insulin to determine the subject's responsiveness to neuromodulation therapy, and the subject is determined to be a non-responder if the change in concentration relative to the baseline concentration is less than a predetermined threshold change.

15. 10. The method of claim 9, wherein the region of interest is in a liver, and evaluating the one or more features comprises determining glucose or insulin concentrations at multiple time points after the application, and determining that a trend in the concentrations at the multiple time points is indicative of a return to a baseline glucose concentration within a predetermined period of time.

16. 10. The method of claim 9, wherein the subject is not fasting and applying the energy induces an approximate fasting state.

17. 10. The method of claim 9, wherein the region of interest is in a spleen, and evaluating the one or more characteristics comprises determining a change in a population of immune cells or a concentration of a cytokine or immune marker following delivery of the energy.

18. The method of claim 9 , wherein the evaluation of the one or more characteristics comprises receiving data to identify a systemic response caused by the application.

19. The method of claim 18 , wherein the data is from an imaging device and / or a physiological monitoring device.

20. 1. A regulation system comprising: an energy application device configured to apply energy to a first target area of ​​a first organ and a second target area of ​​a second organ; A controller, controlling a first application of the energy to the first region of interest via the energy application device to cause a first perturbation as a result of the first application of the energy; receiving information indicative of a first perturbation characteristic; controlling a second application of the energy to the second region of interest via the energy application device to cause a second perturbation as a result of the second application of the energy; receiving information indicative of a second perturbation characteristic; determining a clinical status of the subject based on the first perturbation feature and the second perturbation feature; providing an indication of said clinical condition The controller and The adjustment system comprises:

21. 21. The system of claim 20, wherein the first perturbation characteristic comprises glucose concentrations at multiple time points after the first application of the energy to the first region of interest.

22. 21. The system of claim 20, wherein the second application of energy to the second region of interest is responsive to a determination that the first perturbation feature is associated with a first category.

23. 21. The system of claim 20, wherein the first application of energy is prior to, simultaneous with, or subsequent to the second application of energy.

24. 21. The system of claim 20, wherein the first perturbation feature and the second perturbation feature are combined to identify a combined response to determine the clinical condition of the subject.

25. the first region of interest is in the liver, the second region of interest is in the pancreas, and the clinical condition is a decrease in glucose above a predetermined glucose decrease threshold; 21. The system of claim 20, wherein insulin resistance is determined based on the second perturbation feature being an initial increase in circulating insulin above a predetermined insulin increase threshold followed by a decrease in circulating insulin relative to the initial increase.

26. 1. A method for assessing a physiological perturbation in a subject, the method comprising: applying ultrasound energy to a target area within a subject to induce an approximated fasting state in the subject via neuromodulation; receiving glucose and insulin concentration data from said subject in said approximated fasting state; applying the data to a model, the model being based on a relationship between glucose and insulin concentrations in the approximated fasting state for a plurality of normal subjects; receiving an indication of insulin resistance in the subject using the model; providing a treatment recommendation based on said indication; A method comprising:

27. 27. The method of claim 26, wherein the treatment recommendation is ultrasound treatment.

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