Gateway reflex: neural activation-mediated immune cell gateways in the central nervous system

Gateway reflex: neural activation-mediated immune cell gateways in the central nervous system Abstract The neural regulation of organs can be categorized as systemic or local. Whereas systemic regulation by the hypothalamus–pituitary–adrenal gland-mediated release of steroid hormones has been well studied, the mechanisms for local regulation have only recently emerged. Two types of local neural regulation are known, the gateway reflex and the inflammatory reflex. The gateway reflex describes a mechanism that converts regional neural stimulations into inflammatory outputs by changing the state of specific blood vessels. Molecularly, the enhancement of NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) activity in endothelial cells by neurotransmitters, such as noradrenaline and ATP, induces an enhanced production of pro-inflammatory mediators, including chemokines, which form immune cell gateways at specific vessels. Several types of gateway reflex have been identified, and each regulates distinct organs by creating gateways for autoreactive T cells that induce local inflammation. On the other hand, the inflammatory reflex elicits an anti-inflammatory response through vagal nerves. Here, we summarize recent works on these two local neuro-immune interactions, giving special focus to the gateway reflex. blood–brain barrier, chemokines, experimental autoimmune encephalomyelitis, neuroimmunology, NF-κB Introduction All living organisms are exposed to environmental stimulations such as temperature, light, sound and gravity. These stimuli are sensed by the nervous system. In addition to these environmental stimuli, physical and biological changes in our body, including mental stresses, pain, aging, weight, disease and infection, can be sensed by specific neural pathways. These pathways distribute to every organ in the body, suggesting that organ homeostasis is regulated by specific neural pathways. There are two major neuro-immune systems. One is well-known systemic responses via the hypothalamus–pituitary–adrenal axis (1), and the other is local neuro-immune interactions mediated by specific neural activations. We and others have revealed these specific neural pathways that control local immune responses. In this review article, we highlight recent works on these regional neuro-immune interactions, including the gateway reflex and inflammatory reflex. These pathways hold promise for the development of novel therapeutic strategies. Because there are many good reviews about the inflammatory reflex (2–7), we devote most of our attention to the gateway reflex. Gravity is a positive regulator of local inflammation Gravity stimulates all land animals. Gravity stimulation maintains physical functions such as muscle strength and bone mass (8, 9). Some astronauts after flight have experienced ophthalmic changes including optic disc edema, which is indicative of endothelial dysfunction and is related to inflammatory symptoms. Possible mechanisms for these effects are a shift of cerebrospinal fluid or increased cranial pressure under micro-gravity (10, 11). However, the direct contribution of gravity stimulation to local inflammation is not fully understood. We found a new role of gravity in the course of our study about chronic inflammation. Using animal disease models for inflammatory diseases such as multiple sclerosis (MS) and rheumatoid arthritis (RA), we found that non-immune cells including endothelial cells and fibroblasts have an essential role in the induction of inflammatory responses (12, 13). These non-immune cells produce large amounts of pro-inflammatory mediators, such as chemokines, cytokines and growth factors, upon the simultaneous activation of two transcription factors, NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) and STAT3 (signal transducer and activator of transcription 3). The co-activation of NF-κB and STAT3 synergistically activates NF-κB to enhance the production of pro-inflammatory factors, stimulating the pathogenesis of multiple chronic inflammation animal models for RA (12–16), experimental autoimmune encephalomyelitis (EAE), which is a model of MS (12, 17, 18), and allogeneic transplantation (19). We named the synergistic effect of NF-κB and STAT3 the inflammation amplifier (20, 21). During the course of these studies, we considered where and how autoreactive CD4+ T cells in the blood initially invade the central nervous system (CNS) during EAE, because CNS inflammation occurs despite the blood–brain barrier (BBB), which should prevent the entry of large molecules and cells in blood such as immune cells to the CNS. To investigate, we selected an adoptive-transfer EAE mouse model, in which CNS inflammation can be induced by a single intravenous injection of autoreactive, pathogenic CD4+ T cells against myelin oligodendrocyte glycoprotein (22). This model can be induced without Freund’s complete adjuvant and toxin, which could otherwise affect the migration of immune cells by eliciting systemic inflammation and chronic pain. At a preclinical stage of EAE, whole-mount sectioning of recipient mice and subsequent flow cytometry analysis revealed that pathogenic CD4+ T cells exclusively located at the dorsal vessels of the fifth lumbar (L5) spinal cord (17). Consistent with this finding, many chemokines including CCL20, a chemokine that attracts IL-17-secreting CD4+ T (Th17) cells, which are essential for the pathogenesis of EAE (23, 24), were up-regulated at the L5 dorsal vessels compared with other dorsal vessels of the spinal cord (17). This preferential accumulation of pathogenic CD4+ T cells at L5 was not observed with the conditional deletion of STAT3 in non-immune cells such as endothelial cells, indicating that the up-regulation of chemokines at the L5 vessels was dependent on the inflammation amplifier (20, 21). Therefore, the L5 dorsal vessels are the first breach point of the BBB for pathogenic CD4+ T cells in the transfer EAE model (Fig. 1). We also found that the L5 region is severely damaged in an active immunization EAE model in mice (unpublished data by Masaaki Murakami), although we do not exclude a possibility that other areas such as the choroid plexus in the brain are also concomitantly affected in this model, as reported previously (25). On the other hand, preferential demyelination in the L5 region at very early stage of MS in humans remains to be elucidated. Unlike our EAE model, which typically shows a loss of tonicity of the tail tip as a first sign, MS patients have heterogeneous clinical presentations including visual disturbances, motor impairments, pain and/or cognitive deficits (26), suggesting neuroinflammation in various sites of the CNS. As discussed in detail below, because specific local immune interactions define the first breach point of the BBB by immune cells in our EAE model, we hypothesize that compared to mice, humans have abilities for more social interactions and sophisticated movements, which may induce patients’ specific neural activations to form respective BBB breaching points. Fig. 1. View largeDownload slide Gravity gateway reflex. Gravity stimulates the soleus muscles to activate sensory nerves whose cell bodies are located at the DRG of the fifth lumbar (L5) spinal cord. Neural signals to the L5 DRG neurons transmit to the L5 sympathetic ganglion, followed by NE secretion at the L5 dorsal vessels. Although anatomical or electrophysiological evidence for neural connections between neurons in the L5 DRG and L5 sympathetic ganglions remains to be established, these experimental data suggest their functional connection. NE enhances the inflammation amplifier to up-regulate chemokine expression in the L5 dorsal vessels, causing the recruitment of pathogenic CD4+ T cells. Fig. 1. View largeDownload slide Gravity gateway reflex. Gravity stimulates the soleus muscles to activate sensory nerves whose cell bodies are located at the DRG of the fifth lumbar (L5) spinal cord. Neural signals to the L5 DRG neurons transmit to the L5 sympathetic ganglion, followed by NE secretion at the L5 dorsal vessels. Although anatomical or electrophysiological evidence for neural connections between neurons in the L5 DRG and L5 sympathetic ganglions remains to be established, these experimental data suggest their functional connection. NE enhances the inflammation amplifier to up-regulate chemokine expression in the L5 dorsal vessels, causing the recruitment of pathogenic CD4+ T cells. We investigated how these cells accumulate at the L5 cord and found that a neuro-immune interaction explains the selective accumulation. The L5 dorsal root ganglion (DRG) is the largest of DRGs in both human and mice, partly because the L5 DRG contains neurons that connect to the soleus muscles, the main anti-gravity muscles, and are constantly activated by gravity (27, 28). The tail-suspension assay, a ground experiment employed by the National Aeronautics and Space Administration (NASA) that releases the hindlegs from gravity burden by tail suspension (29), abrogated the selective accumulation of immune cells at L5 in EAE mice (17). Consistently, the tail suspension significantly decreased the expression of chemokine levels in the L5 vessels, and instead up-regulated them in the cervical vessels because of an increased gravity burden on the forelegs. When the soleus muscles were electrically stimulated during the tail suspension, the expression of chemokines was restored in the L5 vessels. Furthermore, electric stimulation to other muscles, including quadriceps and triceps, induced an up-regulation of chemokines at the specific dorsal vessels of the spinal levels corresponding to the site of the stimulation (17). These results indicated that activation of a specific sensory pathway enhances chemokine expression at specific vessels. Furthermore, we found that nerves in the sympathetic ganglions were more activated at the L5 level than the L1 level and that chemical blockade of adrenergic receptors inhibited chemokine expression at L5 and lowered EAE clinical stores. Although anatomical or electrophysiological evidence for neural connections between neurons in the L5 DRG and L5 sympathetic ganglions remains to be established, these experimental data suggest their functional connection. On the basis of these observations, we concluded that a specific regional sensory–sympathetic nerve interaction from the soleus muscles creates the initial gateway for immune cells to enter the CNS at the L5 level (Fig. 1) (17). These results revealed a novel neuro-immune interaction, where regional neural activation regulates the blood vessels to secrete chemokines, creating the gateway for immune cells to enter and locally accumulate. This phenomenon is now called the gateway reflex (3, 5–7, 30–34). The gateway reflex can be controlled by electric stimulation. Weak electric pulses to the quadriceps, whose sensory neurons connect to L3 DRG, induced chemokine expression at the L3 dorsal vessels, and those to the triceps, whose sensory pathway connects to the DRGs of the lower cervical to upper thoracic levels, induced chemokine expression at the dorsal vessels of the respective spinal cord levels (Fig. 2) (17). This electric gateway reflex may be utilized as a therapeutic option or drug delivery to enhance an immune reaction against tumor cells, for example. Fig. 2. View largeDownload slide Electric gateway reflex. Weak electric stimulation to muscles can induce the gateway reflex via activation of specific sensory and sympathetic nerves. Electric pulses to the triceps up-regulate chemokine expression at the dorsal vessels of the lower cervical to upper thoracic [fifth cervical (C5) to fifth thoracic (T5)] spinal cord. Similarly, stimulation of the quadriceps and soleus muscles induces a gateway at the L3 and L5 dorsal vessels, respectively. Fig. 2. View largeDownload slide Electric gateway reflex. Weak electric stimulation to muscles can induce the gateway reflex via activation of specific sensory and sympathetic nerves. Electric pulses to the triceps up-regulate chemokine expression at the dorsal vessels of the lower cervical to upper thoracic [fifth cervical (C5) to fifth thoracic (T5)] spinal cord. Similarly, stimulation of the quadriceps and soleus muscles induces a gateway at the L3 and L5 dorsal vessels, respectively. Pain sensation can trigger pathology Because gravity and electric stimulations create specific blood vessel gateways for immune cells at different locations of the CNS, we tested other neural stimulations to extend the gateway reflex. We chose pain sensation as another neural input, because pain sensation causes a tonic sensory stimulation (35, 36) and is a common unwanted symptom that significantly compromises the quality of life of patients with various diseases and injuries (37). A positive correlation between disease symptoms and pain sensation is reported in MS patients (38–40), and a change in pain sensitivity is also reported during EAE (41). The trigeminal nerves are composed of three main branches, with the middle branch known to contain sensory nerves exclusively. We performed a partial ligation of the middle branch of the trigeminal nerves to induce pain sensation at the same day of pathogenic CD4+ T cell transfer and found that the pain sensation prolonged the EAE disease symptom (42). The transfer EAE model we used shows only transient paralysis that never relapses under normal rearing (22, 42). However, the induction of pain during a remission phase of the transfer EAE clearly caused relapse of the disease. Because pain-mediated relapse was also transient, and the mice under the remission phase even with the nerve ligation tolerated the pain, we hypothesized that the sensitivity to pain might have different phenotypes in MS patients and the EAE model. Indeed, the injection of capsaicin, which activates nociceptors including TRPV1 (transient receptor potential vanilloid 1) to cause pain sensation (43), at the cheek or forepaw also induced EAE relapse (42). These results demonstrated that pain sensation is not simply an alert for the disease status or injury, but also a positive regulator of local inflammation at a remote site via specific neural pathways. To identify the blood vessel gateway for immune cells during the pain-induced relapse, we examined the L5 cord of EAE mice under the remission phase (hereinafter referred to as EAE-recovered mice). Although EAE-recovered mice showed complete remission without any clinical signs in appearance, a high number of activated monocytes with high levels of major histocompatibility complex (MHC) class II from the periphery (MHC class II high monocytes) and not derived from microglia were present around the meninges of the L5 spinal cord. Interestingly, after pain induction, these cells accumulated at the ventral vessels of the L5 cord, followed by infiltration of pathogenic CD4+ T cells from the ventral vessels. Interestingly, removal of MHC class II high monocytes by clodronate liposome or neutralization of chemokine CX3CL1 before pain induction prevented not only the accumulation of the activated monocytes at the vessels, but also the infiltration of pathogenic CD4+ T cells and relapse of EAE, indicating a key role of MHC class II high monocytes (42). In vitro data demonstrated that these monocytes produce CX3CL1 upon stimulation with norepinephrine (NE), express the CX3CL1 receptor CX3CR1 and have antigen-presenting ability, suggesting the following scheme: (i) sensory–sympathetic nerve interactions by pain induce NE secretion around the ventral vessels of the L5 cord, (ii) auto/paracrine actions of the CX3CL1-CX3CR1 axis in MHC class II high monocytes accumulate at the ventral vessels and (iii) activation of pathogenic CD4+ T cells by the MHC class II high monocytes, followed by EAE relapse (Fig. 3). Because these monocytes persisted in the spinal cord for a long time (even 1 year), we hypothesized that survival factors for these cells could be novel targets for the treatment of MS relapse. Collectively, these findings indicate that the gateway for immune cells during pain sensation is the ventral blood vessels of the spinal cord (Fig. 3). The gateway reflex by gravity or electric stimulation targeted the dorsal vessels of the spinal cord. Therefore, the gateway reflex by pain stimulation reveals distinct neural activation at different sites. Further experiments using gene knockout mice and chemical inhibitors defined a specific neural pathway that involves TRPV1/Nav1.8-positive sensory neurons, the anterior cingulate cortex, which is a pain-processing area in the brain, and sympathetic neurons that distribute to the ventral vessels of the spinal cord (Fig. 3) (42). Blood corticosterone and catecholamine levels were elevated in mice with pain sensation, suggesting a systemic activation of sympathetic neurons. Indeed, pain induction in mice stimulates neural activations in L1 as well as L5 sympathetic ganglion, suggesting broader spinal cord levels are affected. Because L5 cord is the first inflammation site and contains higher numbers of MHC class II high monocytes even during the remission phase of EAE, the relapse response takes place mostly at L5 cord dependently on pathogenic CD4+ T cells in the blood (42). Following the gravity and electric gateway reflexes, this pain gateway reflex is the third example of the gateway reflex. Fig. 3. View largeDownload slide Pain gateway reflex. Sensory nerve stimulation by pain activates the anterior cingulate cortex (ACC), a pain-processing area in the brain. Subsequent activation of specific sympathetic nerves causes NE-dependent up-regulation of chemokine CX3CL1 expression around the ventral vessels of the spinal cord. Because MHC class II high monocytes reside within the L5 spinal cord, which is the initial inflammatory lesion of transfer EAE, this region is affected most after pain induction. NE induces the production of chemokine CX3CL1 from MHC class II high monocytes. CX3CL1 further recruits these cells around the ventral vessels. MHC class II high monocytes have a capacity to present myelin oligodendrocyte glycoprotein (MOG) antigens to pathogenic CD4+ T cells, causing disease relapse. Fig. 3. View largeDownload slide Pain gateway reflex. Sensory nerve stimulation by pain activates the anterior cingulate cortex (ACC), a pain-processing area in the brain. Subsequent activation of specific sympathetic nerves causes NE-dependent up-regulation of chemokine CX3CL1 expression around the ventral vessels of the spinal cord. Because MHC class II high monocytes reside within the L5 spinal cord, which is the initial inflammatory lesion of transfer EAE, this region is affected most after pain induction. NE induces the production of chemokine CX3CL1 from MHC class II high monocytes. CX3CL1 further recruits these cells around the ventral vessels. MHC class II high monocytes have a capacity to present myelin oligodendrocyte glycoprotein (MOG) antigens to pathogenic CD4+ T cells, causing disease relapse. A molecular basis for ‘illness starts in the mind’ It is known that mental stresses deteriorate a pre-existing disease status and make one predisposed to gastrointestinal (44) and cardiovascular dysfunctions (45). These phenomena are the roots of proverbs like ‘illness starts in the mind’ and ‘care killed the cat’, but the underlying molecular mechanisms remain undefined. Because mental stresses are perceived as neural stimulations in specific brain areas, including the paraventricular nucleus (PVN), dorsomedial nucleus of hypothalamus (DMH), dorsal motor nucleus of the vagal nerve (DMX) and the vagal nerve pathway (46), and because MS has been associated with mental illnesses and gastrointestinal failures (47–53), we pursued a proof-of-concept for the stress gateway reflex using the transfer EAE model. To examine the effect of stress conditions during transfer EAE, a sleep disorder model was employed, in which chronic stress can be imposed on mice reared on a free rotation wheel by the perpetual avoidance of water (54, 55). To our surprise, the combination of sleep disorder and transfer EAE resulted in a high rate of sudden death of the mice (56). Neither sleep disorder nor transfer EAE alone, however, induced death. Another chronic stress model, such as water in the bedding, which gives the mice discomfort, in combination with transfer EAE also increased the rate of sudden death, suggesting that chronic stress, not just sleep disturbance, is a determining factor. As mentioned earlier, transfer EAE develops from the L5 spinal cord because of the gravity gateway reflex (17). However, stress plus pathogenic T-cell transfer in mice caused almost no immune cell accumulation at the L5 cord. Instead, donor pathogenic CD4+ T cells and periphery-derived MHC class II high monocytes during the early phase were found around blood vessels surrounded by the dentate gyrus (DG, a part of the hippocampus), thalamus and third ventricle (3V) in the brain. Because an essential element of the blood vessel gateway formation is chemokine induction, we neutralized various chemokines and found that anti-CCL5 antibody treatment suppressed the sudden death by transfer EAE under stress condition. Indeed, CCL5 expression in specific vessels of the boundary area of the DG, thalamus and 3V was induced by chronic stress alone but not by pathogenic T cells transfer (56). Neural tracing experiments revealed a neural connection with tyrosine hydroxylase-positive noradrenergic nerves between the specific vessels and PVN. These results demonstrate that chronic stress can change the immune cell gateway from the L5 dorsal vessels to specific brain vessels by the induction of CCL5 via a specific neural pathway from the PVN to the vessels in the presence of stress, thus defining the fourth example of the gateway reflex, the stress gateway reflex (Fig. 4) (56). Fig. 4. View largeDownload slide Stress gateway reflex. Under a chronic stress condition, EAE mice develop severe gastrointestinal (GI) failure with high mortality. Chronic stress induces activation of the PVN, which in turn stimulates tyrosine hydroxylase (TH)+ neurons connecting to specific vessels surrounded by the third ventricle, thalamus and DG to cause micro-inflammation in the brain. This micro-inflammation activates an otherwise resting neural pathway to the DMH through the production of ATP. DMH signals transmit to the DMX to activate efferent vagal nerves and severe upper GI tract failure. Potassium ion levels in the blood increase with severe GI bleeding, which eventually induces heart failure associated with cardiac myocyte death. Experimental evidence showing that afferent vagal nerves prevented the fatal symptom also suggests an involvement of the afferent vagal nerve pathway for the stress gateway reflex (not shown in the figure). Fig. 4. View largeDownload slide Stress gateway reflex. Under a chronic stress condition, EAE mice develop severe gastrointestinal (GI) failure with high mortality. Chronic stress induces activation of the PVN, which in turn stimulates tyrosine hydroxylase (TH)+ neurons connecting to specific vessels surrounded by the third ventricle, thalamus and DG to cause micro-inflammation in the brain. This micro-inflammation activates an otherwise resting neural pathway to the DMH through the production of ATP. DMH signals transmit to the DMX to activate efferent vagal nerves and severe upper GI tract failure. Potassium ion levels in the blood increase with severe GI bleeding, which eventually induces heart failure associated with cardiac myocyte death. Experimental evidence showing that afferent vagal nerves prevented the fatal symptom also suggests an involvement of the afferent vagal nerve pathway for the stress gateway reflex (not shown in the figure). Immune cell infiltration at specific brain vessels could not directly explain the sudden death of EAE mice with stress. We further looked for neural connections from specific vessels adjacent to the DG, thalamus and 3V and found that there is a neural circuit from the vessels to the DMH, a hypothalamus region that is hyper-activated in mice with stress plus pathogenic T-cell transfer and communicates with the DMX (56). In other words, a new neural pathway, specific vessels–DMH–DMX, is hyper-activated in mice with stress plus pathogenic T cells. To activate this pathway, a neurotransmitter is required. We identified this neurotransmitter as ATP (57), which can also act as a pro-inflammatory factor (58). Intracranial injection of ATP or an antagonist for P2RX7, an ATP receptor, to blood vessels at the boundary area of the DG, thalamus and 3V induced mortality in stressed mice without transfer EAE or suppressed it with transfer EAE. Neural activation in the DMH was also significantly suppressed by P2RX7 antagonism at the specific vessels in mice with stress and pathogenic T cells. Cytokine injection around these vessels also caused sudden death in stressed mice (56). Thus, micro-inflammation in the specific vessels adjacent to the DG, thalamus and 3V is sufficient to mimic the effect of transfer EAE under stress condition. Finally, we surgically blocked the vagal nerve pathway, whose hyper-activation is known to produce gastric acid and epithelial damage in the stomach (59, 60). As expected, the vagotomy prevented mortality in EAE mice under stress. Therefore, the stress gateway reflex strongly enhances a stress response via the formation of a blood vessel gateway at specific blood vessels at the boundary of the DG, thalamus and 3V to activate a new neural circuit involving the DMH, DMX and vagal nerves. Enhanced activation of the vagal nerves in stressed EAE mice led to epithelial damage in the upper gastrointestinal tract and a subsequent increase in plasma potassium levels caused by gastrointestinal bleeding, which eventually led to heart failure (Fig. 4) (56). Blood aldosterone and cortisol levels were also elevated in mice with chronic stress, but these levels were not significantly changed between groups with or without pathogenic CD4+ T-cell transfer. Moreover, corticosteroid receptor antagonists, mifepristone and guggulsterone, had no effect on the phenotypes of the stress gateway reflex (56). These results suggest that the stress gateway reflex enhances the specific stress responses systemically elicited by activating specific neural circuits via micro-inflammation at specific vessels. We also observed high levels of circulating troponin I and creatine kinase-MB, markers for the necrosis of cardiac myocytes, and the involvement of afferent vagal nerves in the sudden death, since desensitization of the afferent vagal nerves by capsaicin prevented the fatal symptom (unpublished data by Masaaki Murakami). Micro-inflammation in the brain can modulate the function of organs in the periphery and maybe brain function itself by activating otherwise quiescent neurons. Neural activations generally require energy supply. It is reported that neural activity and cerebral blood flow are functionally coupled: increases of neural activity lead to increases of cerebral blood flow in highly restricted areas of neural activation by releasing substances such as NO, PGE2 and ATP, a phenomenon called neurovascular coupling (61). This coupling is thought to reflect the lack of energy resources in the activated areas of the brain (61). Since ATP mediates the stress gateway reflex (56) and neurovascular coupling in some cases (61, 62), we propose that a mechanism similar to neurovascular coupling satisfies the additional energy demands upon the activation of new neural pathways by the stress gateway reflex. Because stress can cause brain micro-inflammation if CNS-reactive CD4+ T cells are present, this mechanism can be regarded as one molecular basis for ‘illness starts in the mind’. Indeed, brain micro-inflammation is observed in pathological conditions such as Alzheimer’s disease, non-Alzheimer-type dementia, Parkinson’s disease, psychological disorder and epilepsy (63–66). Therefore, it could potentially activate new neural pathways to disturb brain function by establishing new neural networks and dysregulating existing ones and/or certain organ functions in the periphery, which is consistent with the stress gateway reflex. Neuro-immune interactions with anti-inflammatory effects The gateway reflex explains how specific neural activations change the status of specific blood vessels and have pro-inflammatory effects. On the other hand, neuro-immune interactions, which inhibit inflammatory responses, have also been reported. Kevin J. Tracey and his group demonstrated that vagal nerve stimulation inhibits endotoxin shock in mice, what they called the inflammatory reflex (3–7). Pro-inflammatory cytokines and/or pathogen-associated molecular patterns stimulate afferent vagal nerves to activate efferent vagal nerves via brain stem nuclei. The efferent neural signals are transmitted to splenic nerves, releasing noradrenaline in the spleen. CD4+ T cells expressing β1/2 adrenaline receptor were shown to produce acetylcholine in response to noradrenaline, which suppressed the production of pro-inflammatory cytokines from macrophages in a manner dependent on α7 nicotinic acetylcholine receptor (Fig. 5) (3–7, 67). It was recently reported that C1 neurons, which are glutamatergic, catecholaminergic and peptidergic neurons in the medulla oblongata, can mediate the cholinergic anti-inflammatory effect in mice during ischemia-reperfusion injury (68, 69). On the other hand, the responsible neural pathway(s) between efferent vagal and splenic nerves is still a matter of debate (70, 71). The cholinergic anti-inflammatory effects can be manipulated artificially. Electro-acupuncture of mice at the ST36 Zusanli acupoint inhibited a septic shock response through vagal activation (72). Furthermore, local ultrasound stimulated the anti-inflammatory splenic neuro-immune interaction in mice and improved kidney ischemia-reperfusion injury (73). Interestingly, electric vagal stimulation in brain-dead animals also exerted anti-inflammatory effects, and a kidney graft from vagal-stimulated donor rats maintained significantly better long-term renal function in recipient rats (74). The Tracey group developed a strategy to stimulate vagal nerves known as bioelectric medicine (75, 76), which was found effective in patients with RA (77). An implantable vagal nerve stimulator is also effective in Crohn’s disease (78). These translational researches hold promise for treating various inflammatory diseases by modulating specific neural activation to regulate immune responses. Fig. 5. View largeDownload slide Inflammatory reflex. After afferent and efferent vagal nerves are activated during septic shock or ischemic reperfusion injury, the production of NE from the splenic nerves is induced. NE stimulates the release of acetylcholine (ACh) from a subset of CD4+ T cells expressing β1/2 adrenaline receptor. ACh binds to α7 nicotinic acetylcholine receptor (α7nAChR)-positive macrophages to down-regulate the expression of pro-inflammatory cytokines including tumor necrosis factor α (TNFα). It is reported that C1 neurons in the brain mediate this cholinergic anti-inflammatory effect. Fig. 5. View largeDownload slide Inflammatory reflex. After afferent and efferent vagal nerves are activated during septic shock or ischemic reperfusion injury, the production of NE from the splenic nerves is induced. NE stimulates the release of acetylcholine (ACh) from a subset of CD4+ T cells expressing β1/2 adrenaline receptor. ACh binds to α7 nicotinic acetylcholine receptor (α7nAChR)-positive macrophages to down-regulate the expression of pro-inflammatory cytokines including tumor necrosis factor α (TNFα). It is reported that C1 neurons in the brain mediate this cholinergic anti-inflammatory effect. Conclusion To date, four types of gateway reflex have been reported. We are investigating new types of gateway reflex, including light-mediated changes in eye pathology (light gateway reflex) and a local neural network to explain symmetrical inflammation (symmetrical gateway reflex). Recent advances in analytic technologies including tissue clearing reagents such as CUBIC, PACT, Scale and CLARITY (79–84), transgenic mice for the study of neural activations including Arc-dVenus, GCaMP and cFos-GFP mice (85–87), opto/chemogenetics (88, 89), various viral and chemical tracers (90), and neuron mapping with an RNA barcoding system (91) are expected to facilitate the identification of more neural pathways in the gateway reflex. Because neural circuits are distributed throughout the body, identifying tissue-specific, local neuro-immune interactions should contribute to novel therapies, similar to the inflammatory reflex (77, 78). In this article, we mainly summarized recent findings about the gateway reflex. The discovery of the stress gateway reflex (56) extended this research field to one that not only investigates neural regulation of the immune system, but also enables unexpected organ function as a consequence of interplay between the nervous and immune systems. Funding Our work described here was supported by KAKENHI (D.K., Y.A. and M.M.), Takeda Science Foundation (M.M.), Institute for Fermentation Osaka (M.M.), Mitsubishi Foundation (M.M.), Mochida Memorial Foundation for Medical and Pharmaceutical Research (D.K.), Suzuken Memorial Foundation (Y.A. and D.K.), Japan Prize Foundation (Y.A.), Ono Medical Research Foundation (Y.A.), Kanzawa Medical Research Foundation (Y.A.), Kishimoto Foundation (Y.A.), Nagao Takeshi Research Foundation (Y.A.), Japan Multiple Sclerosis Society (Y.A.), Kanae Fundation (Y.A.) and Tokyo Medical Research Foundation (M.M. and Y.A.). Acknowledgements We appreciate the excellent technical assistances provided by Ms Mitsue Ezawa and Ms Chiemi Nakayama for our work described here, and we thank Ms Satomi Fukumoto for her excellent assistance. We thank Dr P. Karagiannis (CiRA, Kyoto University, Kyoto, Japan) for carefully reading the manuscript. Conflicts of interest statement: The authors declared no conflicts of interest. References 1 Bellavance , M. A. and Rivest , S . 2014 . The HPA – immune axis and the immunomodulatory actions of glucocorticoids in the brain . Front. Immunol . 5 : 136 . Google Scholar CrossRef Search ADS PubMed 2 Tracey , K. J . 2007 . Physiology and immunology of the cholinergic antiinflammatory pathway . J. Clin. Invest . 117 : 289 . Google Scholar CrossRef Search ADS PubMed 3 Andersson , U. and Tracey , K. J . 2012 . Neural reflexes in inflammation and immunity . J. Exp. Med . 209 : 1057 . Google Scholar CrossRef Search ADS PubMed 4 Andersson , U. and Tracey , K. J . 2012 . Reflex principles of immunological homeostasis . Annu. Rev. Immunol . 30 : 313 . Google Scholar CrossRef Search ADS PubMed 5 Tracey , K. J . 2016 . Reflexes in immunity . Cell 164 : 343 . Google Scholar CrossRef Search ADS PubMed 6 Pavlov , V. A. and Tracey , K. J . 2017 . Neural regulation of immunity: molecular mechanisms and clinical translation . Nat. Neurosci . 20 : 156 . Google Scholar CrossRef Search ADS PubMed 7 Chavan , S. S. , Pavlov , V. A. and Tracey , K. J . 2017 . Mechanisms and therapeutic relevance of neuro-immune communication . Immunity 46 : 927 . Google Scholar CrossRef Search ADS PubMed 8 Sibonga , J. D . 2013 . Spaceflight-induced bone loss: is there an osteoporosis risk ? Curr. Osteoporos. Rep . 11 : 92 . Google Scholar CrossRef Search ADS PubMed 9 Petersen , N. , Lambrecht , G. , Scott , J. , Hirsch , N. , Stokes , M. and Mester , J . 2017 . Postflight reconditioning for European Astronauts—a case report of recovery after six months in space . Musculoskelet. Sci. Pract . 27 ( Suppl. 1 ): S23 . Google Scholar CrossRef Search ADS PubMed 10 Mader , T. H. , Gibson , C. R. , Pass , A. F. , et al. 2011 . Optic disc edema, globe flattening, choroidal folds, and hyperopic shifts observed in astronauts after long-duration space flight . Ophthalmology 118 : 2058 . Google Scholar CrossRef Search ADS PubMed 11 Zwart , S. R. , Gibson , C. R. , Gregory , J. F. , et al. 2017 . Astronaut ophthalmic syndrome . FASEB J . 31 : 3746 . Google Scholar CrossRef Search ADS PubMed 12 Ogura , H. , Murakami , M. , Okuyama , Y. , et al. 2008 . Interleukin-17 promotes autoimmunity by triggering a positive-feedback loop via interleukin-6 induction . Immunity 29 : 628 . Google Scholar CrossRef Search ADS PubMed 13 Sawa , S. , Kamimura , D. , Jin , G. H. , et al. 2006 . Autoimmune arthritis associated with mutated interleukin (IL)-6 receptor gp130 is driven by STAT3/IL-7-dependent homeostatic proliferation of CD4+ T cells . J. Exp. Med . 203 : 1459 . Google Scholar CrossRef Search ADS PubMed 14 Murakami , M. , Okuyama , Y. , Ogura , H. , et al. 2011 . Local microbleeding facilitates IL-6- and IL-17-dependent arthritis in the absence of tissue antigen recognition by activated T cells . J. Exp. Med . 208 : 103 . Google Scholar CrossRef Search ADS PubMed 15 Meng , J. , Jiang , J. J. , Atsumi , T. , et al. 2016 . Breakpoint cluster region-mediated inflammation is dependent on casein kinase II . J. Immunol . 197 : 3111 . Google Scholar CrossRef Search ADS PubMed 16 Atsumi , T. , Suzuki , H. , Jiang , J. J. , et al. 2017 . Rbm10 regulates inflammation development via alternative splicing of Dnmt3b . Int. Immunol . 29 : 581 . Google Scholar CrossRef Search ADS PubMed 17 Arima , Y. , Harada , M. , Kamimura , D. , et al. 2012 . Regional neural activation defines a gateway for autoreactive T cells to cross the blood–brain barrier . Cell 148 : 447 . Google Scholar CrossRef Search ADS PubMed 18 Murakami , M. , Harada , M. , Kamimura , D. , et al. 2013 . Disease-association analysis of an inflammation-related feedback loop . Cell Rep . 3 : 946 . Google Scholar CrossRef Search ADS PubMed 19 Lee , J. , Nakagiri , T. , Oto , T. , et al. 2012 . IL-6 amplifier, NF-κB-triggered positive feedback for IL-6 signaling, in grafts is involved in allogeneic rejection responses . J. Immunol . 189 : 1928 . Google Scholar CrossRef Search ADS PubMed 20 Atsumi , T. , Singh , R. , Sabharwal , L. , et al. 2014 . Inflammation amplifier, a new paradigm in cancer biology . Cancer Res . 74 : 8 . Google Scholar CrossRef Search ADS PubMed 21 Nakagawa , I. , Kamimura , D. , Atsumi , T. , Arima , Y. and Murakami , M . 2015 . Role of inflammation amplifier-induced growth factor expression in the development of inflammatory diseases . Crit. Rev. Immunol . 35 : 365 . Google Scholar CrossRef Search ADS PubMed 22 Tanaka , Y. , Arima , Y. , Higuchi , K. , et al. 2017 . EAE induction by passive transfer of MOG-specific CD4+ T cells . Bio-protocol 7 :e2370. 23 Komiyama , Y. , Nakae , S. , Matsuki , T. , et al. 2006 . IL-17 plays an important role in the development of experimental autoimmune encephalomyelitis . J. Immunol . 177 : 566 . Google Scholar CrossRef Search ADS PubMed 24 Korn , T. , Bettelli , E. , Oukka , M. and Kuchroo , V. K . 2009 . IL-17 and Th17 Cells . Annu. Rev. Immunol . 27 : 485 . Google Scholar CrossRef Search ADS PubMed 25 Reboldi , A. , Coisne , C. , Baumjohann , D. , et al. 2009 . C-C chemokine receptor 6-regulated entry of TH-17 cells into the CNS through the choroid plexus is required for the initiation of EAE . Nat. Immunol . 10 : 514 . Google Scholar CrossRef Search ADS PubMed 26 Dendrou , C. A. , Fugger , L. and Friese , M. A . 2015 . Immunopathology of multiple sclerosis . Nat. Rev. Immunol . 15 : 545 . Google Scholar CrossRef Search ADS PubMed 27 Ohira , Y. , Kawano , F. , Stevens , J. L. , Wang , X. D. and Ishihara , A . 2004 . Load-dependent regulation of neuromuscular system . J. Gravit. Physiol . 11 : P127 . Google Scholar PubMed 28 Shen , J. , Wang , H. Y. , Chen , J. Y. and Liang , B. L . 2006 . Morphologic analysis of normal human lumbar dorsal root ganglion by 3D MR imaging . AJNR Am. J. Neuroradiol . 27 : 2098 . Google Scholar PubMed 29 Morey-Holton , E. R. and Globus , R. K . 2002 . Hindlimb unloading rodent model: technical aspects . J. Appl. Physiol. (1985) . 92 : 1367 . Google Scholar CrossRef Search ADS PubMed 30 Tracey , K. J . 2012 . Immune cells exploit a neural circuit to enter the CNS . Cell 148 : 392 . Google Scholar CrossRef Search ADS PubMed 31 Deutschman , C. S. and Tracey , K. J . 2014 . Sepsis: current dogma and new perspectives . Immunity 40 : 463 . Google Scholar CrossRef Search ADS PubMed 32 Sabharwal , L. , Kamimura , D. , Meng , J. , et al. 2014 . The Gateway Reflex, which is mediated by the inflammation amplifier, directs pathogenic immune cells into the CNS . J. Biochem . 156 : 299 . Google Scholar CrossRef Search ADS PubMed 33 Ohki , T. , Kamimura , D. , Arima , Y. and Murakami , M . 2017 . Gateway reflex, a new paradigm of neuro-immune interaction . Clin. Exp. Neuroimmunol . 8 : 23 . Google Scholar CrossRef Search ADS 34 Tanaka , Y. , Arima , Y. , Kamimura , D. and Murakami , M . 2017 . The gateway reflex, a novel neuro-immune interaction for the regulation of regional vessels . Front. Immunol . 8 : 1321 . Google Scholar CrossRef Search ADS PubMed 35 Morales-Lázaro , S. L. , Simon , S. A. and Rosenbaum , T . 2013 . The role of endogenous molecules in modulating pain through transient receptor potential vanilloid 1 (TRPV1) . J. Physiol . 591 : 3109 . Google Scholar CrossRef Search ADS PubMed 36 Bennett , D. L. and Woods , C. G . 2014 . Painful and painless channelopathies . Lancet Neurol . 13 : 587 . Google Scholar CrossRef Search ADS PubMed 37 Feinstein , A. , Magalhaes , S. , Richard , J. F. , Audet , B. and Moore , C . 2014 . The link between multiple sclerosis and depression . Nat. Rev. Neurol . 10 : 507 . Google Scholar CrossRef Search ADS PubMed 38 Ehde , D. M. , Gibbons , L. E. , Chwastiak , L. , Bombardier , C. H. , Sullivan , M. D. and Kraft , G. H . 2003 . Chronic pain in a large community sample of persons with multiple sclerosis . Mult. Scler . 9 : 605 . Google Scholar CrossRef Search ADS PubMed 39 Ehde , D. M. , Osborne , T. L. , Hanley , M. A. , Jensen , M. P. and Kraft , G. H . 2006 . The scope and nature of pain in persons with multiple sclerosis . Mult. Scler . 12 : 629 . Google Scholar CrossRef Search ADS PubMed 40 O’Connor , A. B. , Schwid , S. R. , Herrmann , D. N. , Markman , J. D. and Dworkin , R. H . 2008 . Pain associated with multiple sclerosis: systematic review and proposed classification . Pain 137 : 96 . Google Scholar CrossRef Search ADS PubMed 41 Khan , N. and Smith , M. T . 2014 . Multiple sclerosis-induced neuropathic pain: pharmacological management and pathophysiological insights from rodent EAE models . Inflammopharmacology 22 : 1 . Google Scholar CrossRef Search ADS PubMed 42 Arima , Y. , Kamimura , D. , Atsumi , T. , et al. 2015 . A pain-mediated neural signal induces relapse in murine autoimmune encephalomyelitis, a multiple sclerosis model . eLife 4 : e08733 . Google Scholar CrossRef Search ADS 43 Davis , J. B. , Gray , J. , Gunthorpe , M. J. , et al. 2000 . Vanilloid receptor-1 is essential for inflammatory thermal hyperalgesia . Nature 405 : 183 . Google Scholar CrossRef Search ADS PubMed 44 Konturek , P. C. , Brzozowski , T. and Konturek , S. J . 2011 . Stress and the gut: pathophysiology, clinical consequences, diagnostic approach and treatment options . J. Physiol. Pharmacol . 62 : 591 . Google Scholar PubMed 45 Esler , M . 2017 . Mental stress and human cardiovascular disease . Neurosci. Biobehav. Rev . 74 : 269 . Google Scholar CrossRef Search ADS PubMed 46 Ulrich-Lai , Y. M. and Herman , J. P . 2009 . Neural regulation of endocrine and autonomic stress responses . Nat. Rev. Neurosci . 10 : 397 . Google Scholar CrossRef Search ADS PubMed 47 Goldman Consensus , Group . 2005 . The Goldman Consensus statement on depression in multiple sclerosis . Mult. Scler . 11 : 328 . Google Scholar CrossRef Search ADS PubMed 48 Marrie , R. A. , Reingold , S. , Cohen , J. , et al. 2015 . The incidence and prevalence of psychiatric disorders in multiple sclerosis: a systematic review . Mult. Scler . 21 : 305 . Google Scholar CrossRef Search ADS PubMed 49 Rang , E. H. , Brooke , B. N. and Hermon-Taylor , J . 1982 . Association of ulcerative colitis with multiple sclerosis . Lancet 2 : 555 . Google Scholar CrossRef Search ADS PubMed 50 Sadovnick , A. D. , Paty , D. W. and Yannakoulias , G . 1989 . Concurrence of multiple sclerosis and inflammatory bowel disease . N. Engl. J. Med . 321 : 762 . Google Scholar PubMed 51 Gupta , G. , Gelfand , J. M. and Lewis , J. D . 2005 . Increased risk for demyelinating diseases in patients with inflammatory bowel disease . Gastroenterology 129 : 819 . Google Scholar CrossRef Search ADS PubMed 52 Kimura , K. , Hunter , S. F. , Thollander , M. S. , et al. 2000 . Concurrence of inflammatory bowel disease and multiple sclerosis . Mayo Clin. Proc . 75 : 802 . Google Scholar CrossRef Search ADS PubMed 53 Pokorny , C. S. , Beran , R. G. and Pokorny , M. J . 2007 . Association between ulcerative colitis and multiple sclerosis . Intern. Med. J . 37 : 721 . Google Scholar CrossRef Search ADS PubMed 54 Miyazaki , K. , Itoh , N. , Ohyama , S. , Kadota , K. and Oishi , K . 2013 . Continuous exposure to a novel stressor based on water aversion induces abnormal circadian locomotor rhythms and sleep-wake cycles in mice . PLoS One 8 : e55452 . Google Scholar CrossRef Search ADS PubMed 55 Oishi , K. , Yamamoto , S. , Itoh , N. , et al. 2014 . Disruption of behavioral circadian rhythms induced by psychophysiological stress affects plasma free amino acid profiles without affecting peripheral clock gene expression in mice . Biochem. Biophys. Res. Commun . 450 : 880 . Google Scholar CrossRef Search ADS PubMed 56 Arima , Y. , Ohki , T. , Nishikawa , N. , et al. 2017 . Brain micro-inflammation at specific vessels dysregulates organ-homeostasis via the activation of a new neural circuit . eLife 6 : e25517 . Google Scholar CrossRef Search ADS PubMed 57 Burnstock , G . 2006 . Historical review: ATP as a neurotransmitter . Trends Pharmacol. Sci . 27 : 166 . Google Scholar CrossRef Search ADS PubMed 58 Di Virgilio , F. , Dal Ben , D. , Sarti , A. C. , Giuliani , A. L. and Falzoni , S . 2017 . The P2X7 receptor in infection and inflammation . Immunity 47 : 15 . Google Scholar CrossRef Search ADS PubMed 59 Okumura , T. , Uehara , A. , Okamura , K. and Namiki , M . 1990 . Site-specific formation of gastric ulcers by the electric stimulation of the left or right gastric branch of the vagus nerve in the rat . Scand. J. Gastroenterol . 25 : 834 . Google Scholar CrossRef Search ADS PubMed 60 Schubert , M. L . 2003 . Gastric secretion . Curr. Opin. Gastroenterol . 19 : 519 . Google Scholar CrossRef Search ADS PubMed 61 Iadecola , C . 2017 . The neurovascular unit coming of age: a journey through neurovascular coupling in health and disease . Neuron 96 : 17 . Google Scholar CrossRef Search ADS PubMed 62 Mishra , A. , Reynolds , J. P. , Chen , Y. , Gourine , A. V. , Rusakov , D. A. and Attwell , D . 2016 . Astrocytes mediate neurovascular signaling to capillary pericytes but not to arterioles . Nat. Neurosci . 19 : 1619 . Google Scholar CrossRef Search ADS PubMed 63 Togo , T. , Akiyama , H. , Iseki , E. , et al. 2002 . Occurrence of T cells in the brain of Alzheimer’s disease and other neurological diseases . J. Neuroimmunol . 124 : 83 . Google Scholar CrossRef Search ADS PubMed 64 Appel , S. H. , Beers , D. R. and Henkel , J. S . 2010 . T cell-microglial dialogue in Parkinson’s disease and amyotrophic lateral sclerosis: are we listening ? Trends Immunol . 31 : 7 . Google Scholar CrossRef Search ADS PubMed 65 Vezzani , A. , French , J. , Bartfai , T. and Baram , T. Z . 2011 . The role of inflammation in epilepsy . Nat. Rev. Neurol . 7 : 31 . Google Scholar CrossRef Search ADS PubMed 66 Najjar , S. , Pearlman , D. M. , Alper , K. , Najjar , A. and Devinsky , O . 2013 . Neuroinflammation and psychiatric illness . J. Neuroinflammation 10 : 43 . Google Scholar PubMed 67 Okusa , M. D. , Rosin , D. L. and Tracey , K. J . 2017 . Targeting neural reflex circuits in immunity to treat kidney disease . Nat. Rev. Nephrol . 13 : 669 . Google Scholar CrossRef Search ADS PubMed 68 Inoue , T. , Abe , C. , Sung , S. S. , et al. 2016 . Vagus nerve stimulation mediates protection from kidney ischemia-reperfusion injury through α7nAChR+ splenocytes . J. Clin. Invest . 126 : 1939 . Google Scholar CrossRef Search ADS PubMed 69 Abe , C. , Inoue , T. , Inglis , M. A. , et al. 2017 . C1 neurons mediate a stress-induced anti-inflammatory reflex in mice . Nat. Neurosci . 20 : 700 . Google Scholar CrossRef Search ADS PubMed 70 Bratton , B. O. , Martelli , D. , McKinley , M. J. , Trevaks , D. , Anderson , C. R. and McAllen , R. M . 2012 . Neural regulation of inflammation: no neural connection from the vagus to splenic sympathetic neurons . Exp. Physiol . 97 : 1180 . Google Scholar CrossRef Search ADS PubMed 71 Bonaz , B. , Sinniger , V. and Pellissier , S . 2016 . Anti-inflammatory properties of the vagus nerve: potential therapeutic implications of vagus nerve stimulation . J. Physiol . 594 : 5781 . Google Scholar CrossRef Search ADS PubMed 72 Torres-Rosas , R. , Yehia , G. , Peña , G. , et al. 2014 . Dopamine mediates vagal modulation of the immune system by electroacupuncture . Nat. Med . 20 : 291 . Google Scholar CrossRef Search ADS PubMed 73 Gigliotti , J. C. , Huang , L. , Bajwa , A. , et al. 2015 . Ultrasound modulates the splenic neuroimmune axis in attenuating AKI . J. Am. Soc. Nephrol . 26 : 2470 . Google Scholar CrossRef Search ADS PubMed 74 Hoeger , S. , Fontana , J. , Jarczyk , J. , et al. 2014 . Vagal stimulation in brain dead donor rats decreases chronic allograft nephropathy in recipients . Nephrol. Dial. Transplant . 29 : 544 . Google Scholar CrossRef Search ADS PubMed 75 Tracey , K. J . 2015 . Electronic medicine fights disease . Sci. Am . 312 : 28 . Google Scholar CrossRef Search ADS 76 Olofsson , P. S. and Tracey , K. J . 2017 . Bioelectronic medicine: technology targeting molecular mechanisms for therapy . J. Intern. Med . 282 : 3 . Google Scholar CrossRef Search ADS PubMed 77 Koopman , F. A. , Chavan , S. S. , Miljko , S. , et al. 2016 . Vagus nerve stimulation inhibits cytokine production and attenuates disease severity in rheumatoid arthritis . Proc. Natl Acad. Sci. USA 113 : 8284 . Google Scholar CrossRef Search ADS 78 Bonaz , B. , Sinniger , V. , Hoffmann , D. , et al. 2016 . Chronic vagus nerve stimulation in Crohn’s disease: a 6-month follow-up pilot study . Neurogastroenterol. Motil . 28 : 948 . Google Scholar CrossRef Search ADS PubMed 79 Hama , H. , Kurokawa , H. , Kawano , H. , et al. 2011 . Scale: a chemical approach for fluorescence imaging and reconstruction of transparent mouse brain . Nat. Neurosci . 14 : 1481 . Google Scholar CrossRef Search ADS PubMed 80 Chung , K. , Wallace , J. , Kim , S. Y. , et al. 2013 . Structural and molecular interrogation of intact biological systems . Nature 497 : 332 . Google Scholar CrossRef Search ADS PubMed 81 Susaki , E. A. , Tainaka , K. , Perrin , D. , et al. 2014 . Whole-brain imaging with single-cell resolution using chemical cocktails and computational analysis . Cell 157 : 726 . Google Scholar CrossRef Search ADS PubMed 82 Tainaka , K. , Kubota , S. I. , Suyama , T. Q. , et al. 2014 . Whole-body imaging with single-cell resolution by tissue decolorization . Cell 159 : 911 . Google Scholar CrossRef Search ADS PubMed 83 Yang , B. , Treweek , J. B. , Kulkarni , R. P. , et al. 2014 . Single-cell phenotyping within transparent intact tissue through whole-body clearing . Cell 158 : 945 . Google Scholar CrossRef Search ADS PubMed 84 Tainaka , K. , Kuno , A. , Kubota , S. I. , Murakami , T. and Ueda , H. R . 2016 . Chemical principles in tissue clearing and staining protocols for whole-body cell profiling . Annu. Rev. Cell Dev. Biol . 32 : 713 . Google Scholar CrossRef Search ADS PubMed 85 Barth , A. L. , Gerkin , R. C. and Dean , K. L . 2004 . Alteration of neuronal firing properties after in vivo experience in a FosGFP transgenic mouse . J. Neurosci . 24 : 6466 . Google Scholar CrossRef Search ADS PubMed 86 Eguchi , M. and Yamaguchi , S . 2009 . In vivo and in vitro visualization of gene expression dynamics over extensive areas of the brain . Neuroimage 44 : 1274 . Google Scholar CrossRef Search ADS PubMed 87 Chen , Q. , Cichon , J. , Wang , W. , et al. 2012 . Imaging neural activity using Thy1-GCaMP transgenic mice . Neuron 76 : 297 . Google Scholar CrossRef Search ADS PubMed 88 Kim , C. K. , Adhikari , A. and Deisseroth , K . 2017 . Integration of optogenetics with complementary methodologies in systems neuroscience . Nat. Rev. Neurosci . 18 : 222 . Google Scholar CrossRef Search ADS PubMed 89 Campbell , E. J. and Marchant , N. J . 2018 . The use of chemogenetics in behavioural neuroscience: receptor variants, targeting approaches and caveats . Br. J. Pharmacol . 175 : 994 . Google Scholar CrossRef Search ADS PubMed 90 Zeng , H . 2018 . Mesoscale connectomics . Curr. Opin. Neurobiol . 50 : 154 . Google Scholar CrossRef Search ADS PubMed 91 Kebschull , J. M. , Garcia da Silva , P. , Reid , A. P. , Peikon , I. D. , Albeanu , D. F. and Zador , A. M . 2016 . High-throughput mapping of single-neuron projections by sequencing of barcoded RNA . Neuron 91 : 975 . Google Scholar CrossRef Search ADS PubMed © The Japanese Society for Immunology. 2018. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/about_us/legal/notices) http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png International Immunology Oxford University Press

Gateway reflex: neural activation-mediated immune cell gateways in the central nervous system

International Immunology , Volume Advance Article (7) – May 15, 2018

Loading next page...
 
/lp/ou_press/gateway-reflex-neural-activation-mediated-immune-cell-gateways-in-the-0ekz68dR7r
Publisher
Oxford University Press
Copyright
© The Japanese Society for Immunology. 2018. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com
ISSN
0953-8178
eISSN
1460-2377
D.O.I.
10.1093/intimm/dxy034
Publisher site
See Article on Publisher Site

Abstract

Abstract The neural regulation of organs can be categorized as systemic or local. Whereas systemic regulation by the hypothalamus–pituitary–adrenal gland-mediated release of steroid hormones has been well studied, the mechanisms for local regulation have only recently emerged. Two types of local neural regulation are known, the gateway reflex and the inflammatory reflex. The gateway reflex describes a mechanism that converts regional neural stimulations into inflammatory outputs by changing the state of specific blood vessels. Molecularly, the enhancement of NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) activity in endothelial cells by neurotransmitters, such as noradrenaline and ATP, induces an enhanced production of pro-inflammatory mediators, including chemokines, which form immune cell gateways at specific vessels. Several types of gateway reflex have been identified, and each regulates distinct organs by creating gateways for autoreactive T cells that induce local inflammation. On the other hand, the inflammatory reflex elicits an anti-inflammatory response through vagal nerves. Here, we summarize recent works on these two local neuro-immune interactions, giving special focus to the gateway reflex. blood–brain barrier, chemokines, experimental autoimmune encephalomyelitis, neuroimmunology, NF-κB Introduction All living organisms are exposed to environmental stimulations such as temperature, light, sound and gravity. These stimuli are sensed by the nervous system. In addition to these environmental stimuli, physical and biological changes in our body, including mental stresses, pain, aging, weight, disease and infection, can be sensed by specific neural pathways. These pathways distribute to every organ in the body, suggesting that organ homeostasis is regulated by specific neural pathways. There are two major neuro-immune systems. One is well-known systemic responses via the hypothalamus–pituitary–adrenal axis (1), and the other is local neuro-immune interactions mediated by specific neural activations. We and others have revealed these specific neural pathways that control local immune responses. In this review article, we highlight recent works on these regional neuro-immune interactions, including the gateway reflex and inflammatory reflex. These pathways hold promise for the development of novel therapeutic strategies. Because there are many good reviews about the inflammatory reflex (2–7), we devote most of our attention to the gateway reflex. Gravity is a positive regulator of local inflammation Gravity stimulates all land animals. Gravity stimulation maintains physical functions such as muscle strength and bone mass (8, 9). Some astronauts after flight have experienced ophthalmic changes including optic disc edema, which is indicative of endothelial dysfunction and is related to inflammatory symptoms. Possible mechanisms for these effects are a shift of cerebrospinal fluid or increased cranial pressure under micro-gravity (10, 11). However, the direct contribution of gravity stimulation to local inflammation is not fully understood. We found a new role of gravity in the course of our study about chronic inflammation. Using animal disease models for inflammatory diseases such as multiple sclerosis (MS) and rheumatoid arthritis (RA), we found that non-immune cells including endothelial cells and fibroblasts have an essential role in the induction of inflammatory responses (12, 13). These non-immune cells produce large amounts of pro-inflammatory mediators, such as chemokines, cytokines and growth factors, upon the simultaneous activation of two transcription factors, NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) and STAT3 (signal transducer and activator of transcription 3). The co-activation of NF-κB and STAT3 synergistically activates NF-κB to enhance the production of pro-inflammatory factors, stimulating the pathogenesis of multiple chronic inflammation animal models for RA (12–16), experimental autoimmune encephalomyelitis (EAE), which is a model of MS (12, 17, 18), and allogeneic transplantation (19). We named the synergistic effect of NF-κB and STAT3 the inflammation amplifier (20, 21). During the course of these studies, we considered where and how autoreactive CD4+ T cells in the blood initially invade the central nervous system (CNS) during EAE, because CNS inflammation occurs despite the blood–brain barrier (BBB), which should prevent the entry of large molecules and cells in blood such as immune cells to the CNS. To investigate, we selected an adoptive-transfer EAE mouse model, in which CNS inflammation can be induced by a single intravenous injection of autoreactive, pathogenic CD4+ T cells against myelin oligodendrocyte glycoprotein (22). This model can be induced without Freund’s complete adjuvant and toxin, which could otherwise affect the migration of immune cells by eliciting systemic inflammation and chronic pain. At a preclinical stage of EAE, whole-mount sectioning of recipient mice and subsequent flow cytometry analysis revealed that pathogenic CD4+ T cells exclusively located at the dorsal vessels of the fifth lumbar (L5) spinal cord (17). Consistent with this finding, many chemokines including CCL20, a chemokine that attracts IL-17-secreting CD4+ T (Th17) cells, which are essential for the pathogenesis of EAE (23, 24), were up-regulated at the L5 dorsal vessels compared with other dorsal vessels of the spinal cord (17). This preferential accumulation of pathogenic CD4+ T cells at L5 was not observed with the conditional deletion of STAT3 in non-immune cells such as endothelial cells, indicating that the up-regulation of chemokines at the L5 vessels was dependent on the inflammation amplifier (20, 21). Therefore, the L5 dorsal vessels are the first breach point of the BBB for pathogenic CD4+ T cells in the transfer EAE model (Fig. 1). We also found that the L5 region is severely damaged in an active immunization EAE model in mice (unpublished data by Masaaki Murakami), although we do not exclude a possibility that other areas such as the choroid plexus in the brain are also concomitantly affected in this model, as reported previously (25). On the other hand, preferential demyelination in the L5 region at very early stage of MS in humans remains to be elucidated. Unlike our EAE model, which typically shows a loss of tonicity of the tail tip as a first sign, MS patients have heterogeneous clinical presentations including visual disturbances, motor impairments, pain and/or cognitive deficits (26), suggesting neuroinflammation in various sites of the CNS. As discussed in detail below, because specific local immune interactions define the first breach point of the BBB by immune cells in our EAE model, we hypothesize that compared to mice, humans have abilities for more social interactions and sophisticated movements, which may induce patients’ specific neural activations to form respective BBB breaching points. Fig. 1. View largeDownload slide Gravity gateway reflex. Gravity stimulates the soleus muscles to activate sensory nerves whose cell bodies are located at the DRG of the fifth lumbar (L5) spinal cord. Neural signals to the L5 DRG neurons transmit to the L5 sympathetic ganglion, followed by NE secretion at the L5 dorsal vessels. Although anatomical or electrophysiological evidence for neural connections between neurons in the L5 DRG and L5 sympathetic ganglions remains to be established, these experimental data suggest their functional connection. NE enhances the inflammation amplifier to up-regulate chemokine expression in the L5 dorsal vessels, causing the recruitment of pathogenic CD4+ T cells. Fig. 1. View largeDownload slide Gravity gateway reflex. Gravity stimulates the soleus muscles to activate sensory nerves whose cell bodies are located at the DRG of the fifth lumbar (L5) spinal cord. Neural signals to the L5 DRG neurons transmit to the L5 sympathetic ganglion, followed by NE secretion at the L5 dorsal vessels. Although anatomical or electrophysiological evidence for neural connections between neurons in the L5 DRG and L5 sympathetic ganglions remains to be established, these experimental data suggest their functional connection. NE enhances the inflammation amplifier to up-regulate chemokine expression in the L5 dorsal vessels, causing the recruitment of pathogenic CD4+ T cells. We investigated how these cells accumulate at the L5 cord and found that a neuro-immune interaction explains the selective accumulation. The L5 dorsal root ganglion (DRG) is the largest of DRGs in both human and mice, partly because the L5 DRG contains neurons that connect to the soleus muscles, the main anti-gravity muscles, and are constantly activated by gravity (27, 28). The tail-suspension assay, a ground experiment employed by the National Aeronautics and Space Administration (NASA) that releases the hindlegs from gravity burden by tail suspension (29), abrogated the selective accumulation of immune cells at L5 in EAE mice (17). Consistently, the tail suspension significantly decreased the expression of chemokine levels in the L5 vessels, and instead up-regulated them in the cervical vessels because of an increased gravity burden on the forelegs. When the soleus muscles were electrically stimulated during the tail suspension, the expression of chemokines was restored in the L5 vessels. Furthermore, electric stimulation to other muscles, including quadriceps and triceps, induced an up-regulation of chemokines at the specific dorsal vessels of the spinal levels corresponding to the site of the stimulation (17). These results indicated that activation of a specific sensory pathway enhances chemokine expression at specific vessels. Furthermore, we found that nerves in the sympathetic ganglions were more activated at the L5 level than the L1 level and that chemical blockade of adrenergic receptors inhibited chemokine expression at L5 and lowered EAE clinical stores. Although anatomical or electrophysiological evidence for neural connections between neurons in the L5 DRG and L5 sympathetic ganglions remains to be established, these experimental data suggest their functional connection. On the basis of these observations, we concluded that a specific regional sensory–sympathetic nerve interaction from the soleus muscles creates the initial gateway for immune cells to enter the CNS at the L5 level (Fig. 1) (17). These results revealed a novel neuro-immune interaction, where regional neural activation regulates the blood vessels to secrete chemokines, creating the gateway for immune cells to enter and locally accumulate. This phenomenon is now called the gateway reflex (3, 5–7, 30–34). The gateway reflex can be controlled by electric stimulation. Weak electric pulses to the quadriceps, whose sensory neurons connect to L3 DRG, induced chemokine expression at the L3 dorsal vessels, and those to the triceps, whose sensory pathway connects to the DRGs of the lower cervical to upper thoracic levels, induced chemokine expression at the dorsal vessels of the respective spinal cord levels (Fig. 2) (17). This electric gateway reflex may be utilized as a therapeutic option or drug delivery to enhance an immune reaction against tumor cells, for example. Fig. 2. View largeDownload slide Electric gateway reflex. Weak electric stimulation to muscles can induce the gateway reflex via activation of specific sensory and sympathetic nerves. Electric pulses to the triceps up-regulate chemokine expression at the dorsal vessels of the lower cervical to upper thoracic [fifth cervical (C5) to fifth thoracic (T5)] spinal cord. Similarly, stimulation of the quadriceps and soleus muscles induces a gateway at the L3 and L5 dorsal vessels, respectively. Fig. 2. View largeDownload slide Electric gateway reflex. Weak electric stimulation to muscles can induce the gateway reflex via activation of specific sensory and sympathetic nerves. Electric pulses to the triceps up-regulate chemokine expression at the dorsal vessels of the lower cervical to upper thoracic [fifth cervical (C5) to fifth thoracic (T5)] spinal cord. Similarly, stimulation of the quadriceps and soleus muscles induces a gateway at the L3 and L5 dorsal vessels, respectively. Pain sensation can trigger pathology Because gravity and electric stimulations create specific blood vessel gateways for immune cells at different locations of the CNS, we tested other neural stimulations to extend the gateway reflex. We chose pain sensation as another neural input, because pain sensation causes a tonic sensory stimulation (35, 36) and is a common unwanted symptom that significantly compromises the quality of life of patients with various diseases and injuries (37). A positive correlation between disease symptoms and pain sensation is reported in MS patients (38–40), and a change in pain sensitivity is also reported during EAE (41). The trigeminal nerves are composed of three main branches, with the middle branch known to contain sensory nerves exclusively. We performed a partial ligation of the middle branch of the trigeminal nerves to induce pain sensation at the same day of pathogenic CD4+ T cell transfer and found that the pain sensation prolonged the EAE disease symptom (42). The transfer EAE model we used shows only transient paralysis that never relapses under normal rearing (22, 42). However, the induction of pain during a remission phase of the transfer EAE clearly caused relapse of the disease. Because pain-mediated relapse was also transient, and the mice under the remission phase even with the nerve ligation tolerated the pain, we hypothesized that the sensitivity to pain might have different phenotypes in MS patients and the EAE model. Indeed, the injection of capsaicin, which activates nociceptors including TRPV1 (transient receptor potential vanilloid 1) to cause pain sensation (43), at the cheek or forepaw also induced EAE relapse (42). These results demonstrated that pain sensation is not simply an alert for the disease status or injury, but also a positive regulator of local inflammation at a remote site via specific neural pathways. To identify the blood vessel gateway for immune cells during the pain-induced relapse, we examined the L5 cord of EAE mice under the remission phase (hereinafter referred to as EAE-recovered mice). Although EAE-recovered mice showed complete remission without any clinical signs in appearance, a high number of activated monocytes with high levels of major histocompatibility complex (MHC) class II from the periphery (MHC class II high monocytes) and not derived from microglia were present around the meninges of the L5 spinal cord. Interestingly, after pain induction, these cells accumulated at the ventral vessels of the L5 cord, followed by infiltration of pathogenic CD4+ T cells from the ventral vessels. Interestingly, removal of MHC class II high monocytes by clodronate liposome or neutralization of chemokine CX3CL1 before pain induction prevented not only the accumulation of the activated monocytes at the vessels, but also the infiltration of pathogenic CD4+ T cells and relapse of EAE, indicating a key role of MHC class II high monocytes (42). In vitro data demonstrated that these monocytes produce CX3CL1 upon stimulation with norepinephrine (NE), express the CX3CL1 receptor CX3CR1 and have antigen-presenting ability, suggesting the following scheme: (i) sensory–sympathetic nerve interactions by pain induce NE secretion around the ventral vessels of the L5 cord, (ii) auto/paracrine actions of the CX3CL1-CX3CR1 axis in MHC class II high monocytes accumulate at the ventral vessels and (iii) activation of pathogenic CD4+ T cells by the MHC class II high monocytes, followed by EAE relapse (Fig. 3). Because these monocytes persisted in the spinal cord for a long time (even 1 year), we hypothesized that survival factors for these cells could be novel targets for the treatment of MS relapse. Collectively, these findings indicate that the gateway for immune cells during pain sensation is the ventral blood vessels of the spinal cord (Fig. 3). The gateway reflex by gravity or electric stimulation targeted the dorsal vessels of the spinal cord. Therefore, the gateway reflex by pain stimulation reveals distinct neural activation at different sites. Further experiments using gene knockout mice and chemical inhibitors defined a specific neural pathway that involves TRPV1/Nav1.8-positive sensory neurons, the anterior cingulate cortex, which is a pain-processing area in the brain, and sympathetic neurons that distribute to the ventral vessels of the spinal cord (Fig. 3) (42). Blood corticosterone and catecholamine levels were elevated in mice with pain sensation, suggesting a systemic activation of sympathetic neurons. Indeed, pain induction in mice stimulates neural activations in L1 as well as L5 sympathetic ganglion, suggesting broader spinal cord levels are affected. Because L5 cord is the first inflammation site and contains higher numbers of MHC class II high monocytes even during the remission phase of EAE, the relapse response takes place mostly at L5 cord dependently on pathogenic CD4+ T cells in the blood (42). Following the gravity and electric gateway reflexes, this pain gateway reflex is the third example of the gateway reflex. Fig. 3. View largeDownload slide Pain gateway reflex. Sensory nerve stimulation by pain activates the anterior cingulate cortex (ACC), a pain-processing area in the brain. Subsequent activation of specific sympathetic nerves causes NE-dependent up-regulation of chemokine CX3CL1 expression around the ventral vessels of the spinal cord. Because MHC class II high monocytes reside within the L5 spinal cord, which is the initial inflammatory lesion of transfer EAE, this region is affected most after pain induction. NE induces the production of chemokine CX3CL1 from MHC class II high monocytes. CX3CL1 further recruits these cells around the ventral vessels. MHC class II high monocytes have a capacity to present myelin oligodendrocyte glycoprotein (MOG) antigens to pathogenic CD4+ T cells, causing disease relapse. Fig. 3. View largeDownload slide Pain gateway reflex. Sensory nerve stimulation by pain activates the anterior cingulate cortex (ACC), a pain-processing area in the brain. Subsequent activation of specific sympathetic nerves causes NE-dependent up-regulation of chemokine CX3CL1 expression around the ventral vessels of the spinal cord. Because MHC class II high monocytes reside within the L5 spinal cord, which is the initial inflammatory lesion of transfer EAE, this region is affected most after pain induction. NE induces the production of chemokine CX3CL1 from MHC class II high monocytes. CX3CL1 further recruits these cells around the ventral vessels. MHC class II high monocytes have a capacity to present myelin oligodendrocyte glycoprotein (MOG) antigens to pathogenic CD4+ T cells, causing disease relapse. A molecular basis for ‘illness starts in the mind’ It is known that mental stresses deteriorate a pre-existing disease status and make one predisposed to gastrointestinal (44) and cardiovascular dysfunctions (45). These phenomena are the roots of proverbs like ‘illness starts in the mind’ and ‘care killed the cat’, but the underlying molecular mechanisms remain undefined. Because mental stresses are perceived as neural stimulations in specific brain areas, including the paraventricular nucleus (PVN), dorsomedial nucleus of hypothalamus (DMH), dorsal motor nucleus of the vagal nerve (DMX) and the vagal nerve pathway (46), and because MS has been associated with mental illnesses and gastrointestinal failures (47–53), we pursued a proof-of-concept for the stress gateway reflex using the transfer EAE model. To examine the effect of stress conditions during transfer EAE, a sleep disorder model was employed, in which chronic stress can be imposed on mice reared on a free rotation wheel by the perpetual avoidance of water (54, 55). To our surprise, the combination of sleep disorder and transfer EAE resulted in a high rate of sudden death of the mice (56). Neither sleep disorder nor transfer EAE alone, however, induced death. Another chronic stress model, such as water in the bedding, which gives the mice discomfort, in combination with transfer EAE also increased the rate of sudden death, suggesting that chronic stress, not just sleep disturbance, is a determining factor. As mentioned earlier, transfer EAE develops from the L5 spinal cord because of the gravity gateway reflex (17). However, stress plus pathogenic T-cell transfer in mice caused almost no immune cell accumulation at the L5 cord. Instead, donor pathogenic CD4+ T cells and periphery-derived MHC class II high monocytes during the early phase were found around blood vessels surrounded by the dentate gyrus (DG, a part of the hippocampus), thalamus and third ventricle (3V) in the brain. Because an essential element of the blood vessel gateway formation is chemokine induction, we neutralized various chemokines and found that anti-CCL5 antibody treatment suppressed the sudden death by transfer EAE under stress condition. Indeed, CCL5 expression in specific vessels of the boundary area of the DG, thalamus and 3V was induced by chronic stress alone but not by pathogenic T cells transfer (56). Neural tracing experiments revealed a neural connection with tyrosine hydroxylase-positive noradrenergic nerves between the specific vessels and PVN. These results demonstrate that chronic stress can change the immune cell gateway from the L5 dorsal vessels to specific brain vessels by the induction of CCL5 via a specific neural pathway from the PVN to the vessels in the presence of stress, thus defining the fourth example of the gateway reflex, the stress gateway reflex (Fig. 4) (56). Fig. 4. View largeDownload slide Stress gateway reflex. Under a chronic stress condition, EAE mice develop severe gastrointestinal (GI) failure with high mortality. Chronic stress induces activation of the PVN, which in turn stimulates tyrosine hydroxylase (TH)+ neurons connecting to specific vessels surrounded by the third ventricle, thalamus and DG to cause micro-inflammation in the brain. This micro-inflammation activates an otherwise resting neural pathway to the DMH through the production of ATP. DMH signals transmit to the DMX to activate efferent vagal nerves and severe upper GI tract failure. Potassium ion levels in the blood increase with severe GI bleeding, which eventually induces heart failure associated with cardiac myocyte death. Experimental evidence showing that afferent vagal nerves prevented the fatal symptom also suggests an involvement of the afferent vagal nerve pathway for the stress gateway reflex (not shown in the figure). Fig. 4. View largeDownload slide Stress gateway reflex. Under a chronic stress condition, EAE mice develop severe gastrointestinal (GI) failure with high mortality. Chronic stress induces activation of the PVN, which in turn stimulates tyrosine hydroxylase (TH)+ neurons connecting to specific vessels surrounded by the third ventricle, thalamus and DG to cause micro-inflammation in the brain. This micro-inflammation activates an otherwise resting neural pathway to the DMH through the production of ATP. DMH signals transmit to the DMX to activate efferent vagal nerves and severe upper GI tract failure. Potassium ion levels in the blood increase with severe GI bleeding, which eventually induces heart failure associated with cardiac myocyte death. Experimental evidence showing that afferent vagal nerves prevented the fatal symptom also suggests an involvement of the afferent vagal nerve pathway for the stress gateway reflex (not shown in the figure). Immune cell infiltration at specific brain vessels could not directly explain the sudden death of EAE mice with stress. We further looked for neural connections from specific vessels adjacent to the DG, thalamus and 3V and found that there is a neural circuit from the vessels to the DMH, a hypothalamus region that is hyper-activated in mice with stress plus pathogenic T-cell transfer and communicates with the DMX (56). In other words, a new neural pathway, specific vessels–DMH–DMX, is hyper-activated in mice with stress plus pathogenic T cells. To activate this pathway, a neurotransmitter is required. We identified this neurotransmitter as ATP (57), which can also act as a pro-inflammatory factor (58). Intracranial injection of ATP or an antagonist for P2RX7, an ATP receptor, to blood vessels at the boundary area of the DG, thalamus and 3V induced mortality in stressed mice without transfer EAE or suppressed it with transfer EAE. Neural activation in the DMH was also significantly suppressed by P2RX7 antagonism at the specific vessels in mice with stress and pathogenic T cells. Cytokine injection around these vessels also caused sudden death in stressed mice (56). Thus, micro-inflammation in the specific vessels adjacent to the DG, thalamus and 3V is sufficient to mimic the effect of transfer EAE under stress condition. Finally, we surgically blocked the vagal nerve pathway, whose hyper-activation is known to produce gastric acid and epithelial damage in the stomach (59, 60). As expected, the vagotomy prevented mortality in EAE mice under stress. Therefore, the stress gateway reflex strongly enhances a stress response via the formation of a blood vessel gateway at specific blood vessels at the boundary of the DG, thalamus and 3V to activate a new neural circuit involving the DMH, DMX and vagal nerves. Enhanced activation of the vagal nerves in stressed EAE mice led to epithelial damage in the upper gastrointestinal tract and a subsequent increase in plasma potassium levels caused by gastrointestinal bleeding, which eventually led to heart failure (Fig. 4) (56). Blood aldosterone and cortisol levels were also elevated in mice with chronic stress, but these levels were not significantly changed between groups with or without pathogenic CD4+ T-cell transfer. Moreover, corticosteroid receptor antagonists, mifepristone and guggulsterone, had no effect on the phenotypes of the stress gateway reflex (56). These results suggest that the stress gateway reflex enhances the specific stress responses systemically elicited by activating specific neural circuits via micro-inflammation at specific vessels. We also observed high levels of circulating troponin I and creatine kinase-MB, markers for the necrosis of cardiac myocytes, and the involvement of afferent vagal nerves in the sudden death, since desensitization of the afferent vagal nerves by capsaicin prevented the fatal symptom (unpublished data by Masaaki Murakami). Micro-inflammation in the brain can modulate the function of organs in the periphery and maybe brain function itself by activating otherwise quiescent neurons. Neural activations generally require energy supply. It is reported that neural activity and cerebral blood flow are functionally coupled: increases of neural activity lead to increases of cerebral blood flow in highly restricted areas of neural activation by releasing substances such as NO, PGE2 and ATP, a phenomenon called neurovascular coupling (61). This coupling is thought to reflect the lack of energy resources in the activated areas of the brain (61). Since ATP mediates the stress gateway reflex (56) and neurovascular coupling in some cases (61, 62), we propose that a mechanism similar to neurovascular coupling satisfies the additional energy demands upon the activation of new neural pathways by the stress gateway reflex. Because stress can cause brain micro-inflammation if CNS-reactive CD4+ T cells are present, this mechanism can be regarded as one molecular basis for ‘illness starts in the mind’. Indeed, brain micro-inflammation is observed in pathological conditions such as Alzheimer’s disease, non-Alzheimer-type dementia, Parkinson’s disease, psychological disorder and epilepsy (63–66). Therefore, it could potentially activate new neural pathways to disturb brain function by establishing new neural networks and dysregulating existing ones and/or certain organ functions in the periphery, which is consistent with the stress gateway reflex. Neuro-immune interactions with anti-inflammatory effects The gateway reflex explains how specific neural activations change the status of specific blood vessels and have pro-inflammatory effects. On the other hand, neuro-immune interactions, which inhibit inflammatory responses, have also been reported. Kevin J. Tracey and his group demonstrated that vagal nerve stimulation inhibits endotoxin shock in mice, what they called the inflammatory reflex (3–7). Pro-inflammatory cytokines and/or pathogen-associated molecular patterns stimulate afferent vagal nerves to activate efferent vagal nerves via brain stem nuclei. The efferent neural signals are transmitted to splenic nerves, releasing noradrenaline in the spleen. CD4+ T cells expressing β1/2 adrenaline receptor were shown to produce acetylcholine in response to noradrenaline, which suppressed the production of pro-inflammatory cytokines from macrophages in a manner dependent on α7 nicotinic acetylcholine receptor (Fig. 5) (3–7, 67). It was recently reported that C1 neurons, which are glutamatergic, catecholaminergic and peptidergic neurons in the medulla oblongata, can mediate the cholinergic anti-inflammatory effect in mice during ischemia-reperfusion injury (68, 69). On the other hand, the responsible neural pathway(s) between efferent vagal and splenic nerves is still a matter of debate (70, 71). The cholinergic anti-inflammatory effects can be manipulated artificially. Electro-acupuncture of mice at the ST36 Zusanli acupoint inhibited a septic shock response through vagal activation (72). Furthermore, local ultrasound stimulated the anti-inflammatory splenic neuro-immune interaction in mice and improved kidney ischemia-reperfusion injury (73). Interestingly, electric vagal stimulation in brain-dead animals also exerted anti-inflammatory effects, and a kidney graft from vagal-stimulated donor rats maintained significantly better long-term renal function in recipient rats (74). The Tracey group developed a strategy to stimulate vagal nerves known as bioelectric medicine (75, 76), which was found effective in patients with RA (77). An implantable vagal nerve stimulator is also effective in Crohn’s disease (78). These translational researches hold promise for treating various inflammatory diseases by modulating specific neural activation to regulate immune responses. Fig. 5. View largeDownload slide Inflammatory reflex. After afferent and efferent vagal nerves are activated during septic shock or ischemic reperfusion injury, the production of NE from the splenic nerves is induced. NE stimulates the release of acetylcholine (ACh) from a subset of CD4+ T cells expressing β1/2 adrenaline receptor. ACh binds to α7 nicotinic acetylcholine receptor (α7nAChR)-positive macrophages to down-regulate the expression of pro-inflammatory cytokines including tumor necrosis factor α (TNFα). It is reported that C1 neurons in the brain mediate this cholinergic anti-inflammatory effect. Fig. 5. View largeDownload slide Inflammatory reflex. After afferent and efferent vagal nerves are activated during septic shock or ischemic reperfusion injury, the production of NE from the splenic nerves is induced. NE stimulates the release of acetylcholine (ACh) from a subset of CD4+ T cells expressing β1/2 adrenaline receptor. ACh binds to α7 nicotinic acetylcholine receptor (α7nAChR)-positive macrophages to down-regulate the expression of pro-inflammatory cytokines including tumor necrosis factor α (TNFα). It is reported that C1 neurons in the brain mediate this cholinergic anti-inflammatory effect. Conclusion To date, four types of gateway reflex have been reported. We are investigating new types of gateway reflex, including light-mediated changes in eye pathology (light gateway reflex) and a local neural network to explain symmetrical inflammation (symmetrical gateway reflex). Recent advances in analytic technologies including tissue clearing reagents such as CUBIC, PACT, Scale and CLARITY (79–84), transgenic mice for the study of neural activations including Arc-dVenus, GCaMP and cFos-GFP mice (85–87), opto/chemogenetics (88, 89), various viral and chemical tracers (90), and neuron mapping with an RNA barcoding system (91) are expected to facilitate the identification of more neural pathways in the gateway reflex. Because neural circuits are distributed throughout the body, identifying tissue-specific, local neuro-immune interactions should contribute to novel therapies, similar to the inflammatory reflex (77, 78). In this article, we mainly summarized recent findings about the gateway reflex. The discovery of the stress gateway reflex (56) extended this research field to one that not only investigates neural regulation of the immune system, but also enables unexpected organ function as a consequence of interplay between the nervous and immune systems. Funding Our work described here was supported by KAKENHI (D.K., Y.A. and M.M.), Takeda Science Foundation (M.M.), Institute for Fermentation Osaka (M.M.), Mitsubishi Foundation (M.M.), Mochida Memorial Foundation for Medical and Pharmaceutical Research (D.K.), Suzuken Memorial Foundation (Y.A. and D.K.), Japan Prize Foundation (Y.A.), Ono Medical Research Foundation (Y.A.), Kanzawa Medical Research Foundation (Y.A.), Kishimoto Foundation (Y.A.), Nagao Takeshi Research Foundation (Y.A.), Japan Multiple Sclerosis Society (Y.A.), Kanae Fundation (Y.A.) and Tokyo Medical Research Foundation (M.M. and Y.A.). Acknowledgements We appreciate the excellent technical assistances provided by Ms Mitsue Ezawa and Ms Chiemi Nakayama for our work described here, and we thank Ms Satomi Fukumoto for her excellent assistance. We thank Dr P. Karagiannis (CiRA, Kyoto University, Kyoto, Japan) for carefully reading the manuscript. Conflicts of interest statement: The authors declared no conflicts of interest. References 1 Bellavance , M. A. and Rivest , S . 2014 . The HPA – immune axis and the immunomodulatory actions of glucocorticoids in the brain . Front. Immunol . 5 : 136 . Google Scholar CrossRef Search ADS PubMed 2 Tracey , K. J . 2007 . Physiology and immunology of the cholinergic antiinflammatory pathway . J. Clin. Invest . 117 : 289 . Google Scholar CrossRef Search ADS PubMed 3 Andersson , U. and Tracey , K. J . 2012 . Neural reflexes in inflammation and immunity . J. Exp. Med . 209 : 1057 . Google Scholar CrossRef Search ADS PubMed 4 Andersson , U. and Tracey , K. J . 2012 . Reflex principles of immunological homeostasis . Annu. Rev. Immunol . 30 : 313 . Google Scholar CrossRef Search ADS PubMed 5 Tracey , K. J . 2016 . Reflexes in immunity . Cell 164 : 343 . Google Scholar CrossRef Search ADS PubMed 6 Pavlov , V. A. and Tracey , K. J . 2017 . Neural regulation of immunity: molecular mechanisms and clinical translation . Nat. Neurosci . 20 : 156 . Google Scholar CrossRef Search ADS PubMed 7 Chavan , S. S. , Pavlov , V. A. and Tracey , K. J . 2017 . Mechanisms and therapeutic relevance of neuro-immune communication . Immunity 46 : 927 . Google Scholar CrossRef Search ADS PubMed 8 Sibonga , J. D . 2013 . Spaceflight-induced bone loss: is there an osteoporosis risk ? Curr. Osteoporos. Rep . 11 : 92 . Google Scholar CrossRef Search ADS PubMed 9 Petersen , N. , Lambrecht , G. , Scott , J. , Hirsch , N. , Stokes , M. and Mester , J . 2017 . Postflight reconditioning for European Astronauts—a case report of recovery after six months in space . Musculoskelet. Sci. Pract . 27 ( Suppl. 1 ): S23 . Google Scholar CrossRef Search ADS PubMed 10 Mader , T. H. , Gibson , C. R. , Pass , A. F. , et al. 2011 . Optic disc edema, globe flattening, choroidal folds, and hyperopic shifts observed in astronauts after long-duration space flight . Ophthalmology 118 : 2058 . Google Scholar CrossRef Search ADS PubMed 11 Zwart , S. R. , Gibson , C. R. , Gregory , J. F. , et al. 2017 . Astronaut ophthalmic syndrome . FASEB J . 31 : 3746 . Google Scholar CrossRef Search ADS PubMed 12 Ogura , H. , Murakami , M. , Okuyama , Y. , et al. 2008 . Interleukin-17 promotes autoimmunity by triggering a positive-feedback loop via interleukin-6 induction . Immunity 29 : 628 . Google Scholar CrossRef Search ADS PubMed 13 Sawa , S. , Kamimura , D. , Jin , G. H. , et al. 2006 . Autoimmune arthritis associated with mutated interleukin (IL)-6 receptor gp130 is driven by STAT3/IL-7-dependent homeostatic proliferation of CD4+ T cells . J. Exp. Med . 203 : 1459 . Google Scholar CrossRef Search ADS PubMed 14 Murakami , M. , Okuyama , Y. , Ogura , H. , et al. 2011 . Local microbleeding facilitates IL-6- and IL-17-dependent arthritis in the absence of tissue antigen recognition by activated T cells . J. Exp. Med . 208 : 103 . Google Scholar CrossRef Search ADS PubMed 15 Meng , J. , Jiang , J. J. , Atsumi , T. , et al. 2016 . Breakpoint cluster region-mediated inflammation is dependent on casein kinase II . J. Immunol . 197 : 3111 . Google Scholar CrossRef Search ADS PubMed 16 Atsumi , T. , Suzuki , H. , Jiang , J. J. , et al. 2017 . Rbm10 regulates inflammation development via alternative splicing of Dnmt3b . Int. Immunol . 29 : 581 . Google Scholar CrossRef Search ADS PubMed 17 Arima , Y. , Harada , M. , Kamimura , D. , et al. 2012 . Regional neural activation defines a gateway for autoreactive T cells to cross the blood–brain barrier . Cell 148 : 447 . Google Scholar CrossRef Search ADS PubMed 18 Murakami , M. , Harada , M. , Kamimura , D. , et al. 2013 . Disease-association analysis of an inflammation-related feedback loop . Cell Rep . 3 : 946 . Google Scholar CrossRef Search ADS PubMed 19 Lee , J. , Nakagiri , T. , Oto , T. , et al. 2012 . IL-6 amplifier, NF-κB-triggered positive feedback for IL-6 signaling, in grafts is involved in allogeneic rejection responses . J. Immunol . 189 : 1928 . Google Scholar CrossRef Search ADS PubMed 20 Atsumi , T. , Singh , R. , Sabharwal , L. , et al. 2014 . Inflammation amplifier, a new paradigm in cancer biology . Cancer Res . 74 : 8 . Google Scholar CrossRef Search ADS PubMed 21 Nakagawa , I. , Kamimura , D. , Atsumi , T. , Arima , Y. and Murakami , M . 2015 . Role of inflammation amplifier-induced growth factor expression in the development of inflammatory diseases . Crit. Rev. Immunol . 35 : 365 . Google Scholar CrossRef Search ADS PubMed 22 Tanaka , Y. , Arima , Y. , Higuchi , K. , et al. 2017 . EAE induction by passive transfer of MOG-specific CD4+ T cells . Bio-protocol 7 :e2370. 23 Komiyama , Y. , Nakae , S. , Matsuki , T. , et al. 2006 . IL-17 plays an important role in the development of experimental autoimmune encephalomyelitis . J. Immunol . 177 : 566 . Google Scholar CrossRef Search ADS PubMed 24 Korn , T. , Bettelli , E. , Oukka , M. and Kuchroo , V. K . 2009 . IL-17 and Th17 Cells . Annu. Rev. Immunol . 27 : 485 . Google Scholar CrossRef Search ADS PubMed 25 Reboldi , A. , Coisne , C. , Baumjohann , D. , et al. 2009 . C-C chemokine receptor 6-regulated entry of TH-17 cells into the CNS through the choroid plexus is required for the initiation of EAE . Nat. Immunol . 10 : 514 . Google Scholar CrossRef Search ADS PubMed 26 Dendrou , C. A. , Fugger , L. and Friese , M. A . 2015 . Immunopathology of multiple sclerosis . Nat. Rev. Immunol . 15 : 545 . Google Scholar CrossRef Search ADS PubMed 27 Ohira , Y. , Kawano , F. , Stevens , J. L. , Wang , X. D. and Ishihara , A . 2004 . Load-dependent regulation of neuromuscular system . J. Gravit. Physiol . 11 : P127 . Google Scholar PubMed 28 Shen , J. , Wang , H. Y. , Chen , J. Y. and Liang , B. L . 2006 . Morphologic analysis of normal human lumbar dorsal root ganglion by 3D MR imaging . AJNR Am. J. Neuroradiol . 27 : 2098 . Google Scholar PubMed 29 Morey-Holton , E. R. and Globus , R. K . 2002 . Hindlimb unloading rodent model: technical aspects . J. Appl. Physiol. (1985) . 92 : 1367 . Google Scholar CrossRef Search ADS PubMed 30 Tracey , K. J . 2012 . Immune cells exploit a neural circuit to enter the CNS . Cell 148 : 392 . Google Scholar CrossRef Search ADS PubMed 31 Deutschman , C. S. and Tracey , K. J . 2014 . Sepsis: current dogma and new perspectives . Immunity 40 : 463 . Google Scholar CrossRef Search ADS PubMed 32 Sabharwal , L. , Kamimura , D. , Meng , J. , et al. 2014 . The Gateway Reflex, which is mediated by the inflammation amplifier, directs pathogenic immune cells into the CNS . J. Biochem . 156 : 299 . Google Scholar CrossRef Search ADS PubMed 33 Ohki , T. , Kamimura , D. , Arima , Y. and Murakami , M . 2017 . Gateway reflex, a new paradigm of neuro-immune interaction . Clin. Exp. Neuroimmunol . 8 : 23 . Google Scholar CrossRef Search ADS 34 Tanaka , Y. , Arima , Y. , Kamimura , D. and Murakami , M . 2017 . The gateway reflex, a novel neuro-immune interaction for the regulation of regional vessels . Front. Immunol . 8 : 1321 . Google Scholar CrossRef Search ADS PubMed 35 Morales-Lázaro , S. L. , Simon , S. A. and Rosenbaum , T . 2013 . The role of endogenous molecules in modulating pain through transient receptor potential vanilloid 1 (TRPV1) . J. Physiol . 591 : 3109 . Google Scholar CrossRef Search ADS PubMed 36 Bennett , D. L. and Woods , C. G . 2014 . Painful and painless channelopathies . Lancet Neurol . 13 : 587 . Google Scholar CrossRef Search ADS PubMed 37 Feinstein , A. , Magalhaes , S. , Richard , J. F. , Audet , B. and Moore , C . 2014 . The link between multiple sclerosis and depression . Nat. Rev. Neurol . 10 : 507 . Google Scholar CrossRef Search ADS PubMed 38 Ehde , D. M. , Gibbons , L. E. , Chwastiak , L. , Bombardier , C. H. , Sullivan , M. D. and Kraft , G. H . 2003 . Chronic pain in a large community sample of persons with multiple sclerosis . Mult. Scler . 9 : 605 . Google Scholar CrossRef Search ADS PubMed 39 Ehde , D. M. , Osborne , T. L. , Hanley , M. A. , Jensen , M. P. and Kraft , G. H . 2006 . The scope and nature of pain in persons with multiple sclerosis . Mult. Scler . 12 : 629 . Google Scholar CrossRef Search ADS PubMed 40 O’Connor , A. B. , Schwid , S. R. , Herrmann , D. N. , Markman , J. D. and Dworkin , R. H . 2008 . Pain associated with multiple sclerosis: systematic review and proposed classification . Pain 137 : 96 . Google Scholar CrossRef Search ADS PubMed 41 Khan , N. and Smith , M. T . 2014 . Multiple sclerosis-induced neuropathic pain: pharmacological management and pathophysiological insights from rodent EAE models . Inflammopharmacology 22 : 1 . Google Scholar CrossRef Search ADS PubMed 42 Arima , Y. , Kamimura , D. , Atsumi , T. , et al. 2015 . A pain-mediated neural signal induces relapse in murine autoimmune encephalomyelitis, a multiple sclerosis model . eLife 4 : e08733 . Google Scholar CrossRef Search ADS 43 Davis , J. B. , Gray , J. , Gunthorpe , M. J. , et al. 2000 . Vanilloid receptor-1 is essential for inflammatory thermal hyperalgesia . Nature 405 : 183 . Google Scholar CrossRef Search ADS PubMed 44 Konturek , P. C. , Brzozowski , T. and Konturek , S. J . 2011 . Stress and the gut: pathophysiology, clinical consequences, diagnostic approach and treatment options . J. Physiol. Pharmacol . 62 : 591 . Google Scholar PubMed 45 Esler , M . 2017 . Mental stress and human cardiovascular disease . Neurosci. Biobehav. Rev . 74 : 269 . Google Scholar CrossRef Search ADS PubMed 46 Ulrich-Lai , Y. M. and Herman , J. P . 2009 . Neural regulation of endocrine and autonomic stress responses . Nat. Rev. Neurosci . 10 : 397 . Google Scholar CrossRef Search ADS PubMed 47 Goldman Consensus , Group . 2005 . The Goldman Consensus statement on depression in multiple sclerosis . Mult. Scler . 11 : 328 . Google Scholar CrossRef Search ADS PubMed 48 Marrie , R. A. , Reingold , S. , Cohen , J. , et al. 2015 . The incidence and prevalence of psychiatric disorders in multiple sclerosis: a systematic review . Mult. Scler . 21 : 305 . Google Scholar CrossRef Search ADS PubMed 49 Rang , E. H. , Brooke , B. N. and Hermon-Taylor , J . 1982 . Association of ulcerative colitis with multiple sclerosis . Lancet 2 : 555 . Google Scholar CrossRef Search ADS PubMed 50 Sadovnick , A. D. , Paty , D. W. and Yannakoulias , G . 1989 . Concurrence of multiple sclerosis and inflammatory bowel disease . N. Engl. J. Med . 321 : 762 . Google Scholar PubMed 51 Gupta , G. , Gelfand , J. M. and Lewis , J. D . 2005 . Increased risk for demyelinating diseases in patients with inflammatory bowel disease . Gastroenterology 129 : 819 . Google Scholar CrossRef Search ADS PubMed 52 Kimura , K. , Hunter , S. F. , Thollander , M. S. , et al. 2000 . Concurrence of inflammatory bowel disease and multiple sclerosis . Mayo Clin. Proc . 75 : 802 . Google Scholar CrossRef Search ADS PubMed 53 Pokorny , C. S. , Beran , R. G. and Pokorny , M. J . 2007 . Association between ulcerative colitis and multiple sclerosis . Intern. Med. J . 37 : 721 . Google Scholar CrossRef Search ADS PubMed 54 Miyazaki , K. , Itoh , N. , Ohyama , S. , Kadota , K. and Oishi , K . 2013 . Continuous exposure to a novel stressor based on water aversion induces abnormal circadian locomotor rhythms and sleep-wake cycles in mice . PLoS One 8 : e55452 . Google Scholar CrossRef Search ADS PubMed 55 Oishi , K. , Yamamoto , S. , Itoh , N. , et al. 2014 . Disruption of behavioral circadian rhythms induced by psychophysiological stress affects plasma free amino acid profiles without affecting peripheral clock gene expression in mice . Biochem. Biophys. Res. Commun . 450 : 880 . Google Scholar CrossRef Search ADS PubMed 56 Arima , Y. , Ohki , T. , Nishikawa , N. , et al. 2017 . Brain micro-inflammation at specific vessels dysregulates organ-homeostasis via the activation of a new neural circuit . eLife 6 : e25517 . Google Scholar CrossRef Search ADS PubMed 57 Burnstock , G . 2006 . Historical review: ATP as a neurotransmitter . Trends Pharmacol. Sci . 27 : 166 . Google Scholar CrossRef Search ADS PubMed 58 Di Virgilio , F. , Dal Ben , D. , Sarti , A. C. , Giuliani , A. L. and Falzoni , S . 2017 . The P2X7 receptor in infection and inflammation . Immunity 47 : 15 . Google Scholar CrossRef Search ADS PubMed 59 Okumura , T. , Uehara , A. , Okamura , K. and Namiki , M . 1990 . Site-specific formation of gastric ulcers by the electric stimulation of the left or right gastric branch of the vagus nerve in the rat . Scand. J. Gastroenterol . 25 : 834 . Google Scholar CrossRef Search ADS PubMed 60 Schubert , M. L . 2003 . Gastric secretion . Curr. Opin. Gastroenterol . 19 : 519 . Google Scholar CrossRef Search ADS PubMed 61 Iadecola , C . 2017 . The neurovascular unit coming of age: a journey through neurovascular coupling in health and disease . Neuron 96 : 17 . Google Scholar CrossRef Search ADS PubMed 62 Mishra , A. , Reynolds , J. P. , Chen , Y. , Gourine , A. V. , Rusakov , D. A. and Attwell , D . 2016 . Astrocytes mediate neurovascular signaling to capillary pericytes but not to arterioles . Nat. Neurosci . 19 : 1619 . Google Scholar CrossRef Search ADS PubMed 63 Togo , T. , Akiyama , H. , Iseki , E. , et al. 2002 . Occurrence of T cells in the brain of Alzheimer’s disease and other neurological diseases . J. Neuroimmunol . 124 : 83 . Google Scholar CrossRef Search ADS PubMed 64 Appel , S. H. , Beers , D. R. and Henkel , J. S . 2010 . T cell-microglial dialogue in Parkinson’s disease and amyotrophic lateral sclerosis: are we listening ? Trends Immunol . 31 : 7 . Google Scholar CrossRef Search ADS PubMed 65 Vezzani , A. , French , J. , Bartfai , T. and Baram , T. Z . 2011 . The role of inflammation in epilepsy . Nat. Rev. Neurol . 7 : 31 . Google Scholar CrossRef Search ADS PubMed 66 Najjar , S. , Pearlman , D. M. , Alper , K. , Najjar , A. and Devinsky , O . 2013 . Neuroinflammation and psychiatric illness . J. Neuroinflammation 10 : 43 . Google Scholar PubMed 67 Okusa , M. D. , Rosin , D. L. and Tracey , K. J . 2017 . Targeting neural reflex circuits in immunity to treat kidney disease . Nat. Rev. Nephrol . 13 : 669 . Google Scholar CrossRef Search ADS PubMed 68 Inoue , T. , Abe , C. , Sung , S. S. , et al. 2016 . Vagus nerve stimulation mediates protection from kidney ischemia-reperfusion injury through α7nAChR+ splenocytes . J. Clin. Invest . 126 : 1939 . Google Scholar CrossRef Search ADS PubMed 69 Abe , C. , Inoue , T. , Inglis , M. A. , et al. 2017 . C1 neurons mediate a stress-induced anti-inflammatory reflex in mice . Nat. Neurosci . 20 : 700 . Google Scholar CrossRef Search ADS PubMed 70 Bratton , B. O. , Martelli , D. , McKinley , M. J. , Trevaks , D. , Anderson , C. R. and McAllen , R. M . 2012 . Neural regulation of inflammation: no neural connection from the vagus to splenic sympathetic neurons . Exp. Physiol . 97 : 1180 . Google Scholar CrossRef Search ADS PubMed 71 Bonaz , B. , Sinniger , V. and Pellissier , S . 2016 . Anti-inflammatory properties of the vagus nerve: potential therapeutic implications of vagus nerve stimulation . J. Physiol . 594 : 5781 . Google Scholar CrossRef Search ADS PubMed 72 Torres-Rosas , R. , Yehia , G. , Peña , G. , et al. 2014 . Dopamine mediates vagal modulation of the immune system by electroacupuncture . Nat. Med . 20 : 291 . Google Scholar CrossRef Search ADS PubMed 73 Gigliotti , J. C. , Huang , L. , Bajwa , A. , et al. 2015 . Ultrasound modulates the splenic neuroimmune axis in attenuating AKI . J. Am. Soc. Nephrol . 26 : 2470 . Google Scholar CrossRef Search ADS PubMed 74 Hoeger , S. , Fontana , J. , Jarczyk , J. , et al. 2014 . Vagal stimulation in brain dead donor rats decreases chronic allograft nephropathy in recipients . Nephrol. Dial. Transplant . 29 : 544 . Google Scholar CrossRef Search ADS PubMed 75 Tracey , K. J . 2015 . Electronic medicine fights disease . Sci. Am . 312 : 28 . Google Scholar CrossRef Search ADS 76 Olofsson , P. S. and Tracey , K. J . 2017 . Bioelectronic medicine: technology targeting molecular mechanisms for therapy . J. Intern. Med . 282 : 3 . Google Scholar CrossRef Search ADS PubMed 77 Koopman , F. A. , Chavan , S. S. , Miljko , S. , et al. 2016 . Vagus nerve stimulation inhibits cytokine production and attenuates disease severity in rheumatoid arthritis . Proc. Natl Acad. Sci. USA 113 : 8284 . Google Scholar CrossRef Search ADS 78 Bonaz , B. , Sinniger , V. , Hoffmann , D. , et al. 2016 . Chronic vagus nerve stimulation in Crohn’s disease: a 6-month follow-up pilot study . Neurogastroenterol. Motil . 28 : 948 . Google Scholar CrossRef Search ADS PubMed 79 Hama , H. , Kurokawa , H. , Kawano , H. , et al. 2011 . Scale: a chemical approach for fluorescence imaging and reconstruction of transparent mouse brain . Nat. Neurosci . 14 : 1481 . Google Scholar CrossRef Search ADS PubMed 80 Chung , K. , Wallace , J. , Kim , S. Y. , et al. 2013 . Structural and molecular interrogation of intact biological systems . Nature 497 : 332 . Google Scholar CrossRef Search ADS PubMed 81 Susaki , E. A. , Tainaka , K. , Perrin , D. , et al. 2014 . Whole-brain imaging with single-cell resolution using chemical cocktails and computational analysis . Cell 157 : 726 . Google Scholar CrossRef Search ADS PubMed 82 Tainaka , K. , Kubota , S. I. , Suyama , T. Q. , et al. 2014 . Whole-body imaging with single-cell resolution by tissue decolorization . Cell 159 : 911 . Google Scholar CrossRef Search ADS PubMed 83 Yang , B. , Treweek , J. B. , Kulkarni , R. P. , et al. 2014 . Single-cell phenotyping within transparent intact tissue through whole-body clearing . Cell 158 : 945 . Google Scholar CrossRef Search ADS PubMed 84 Tainaka , K. , Kuno , A. , Kubota , S. I. , Murakami , T. and Ueda , H. R . 2016 . Chemical principles in tissue clearing and staining protocols for whole-body cell profiling . Annu. Rev. Cell Dev. Biol . 32 : 713 . Google Scholar CrossRef Search ADS PubMed 85 Barth , A. L. , Gerkin , R. C. and Dean , K. L . 2004 . Alteration of neuronal firing properties after in vivo experience in a FosGFP transgenic mouse . J. Neurosci . 24 : 6466 . Google Scholar CrossRef Search ADS PubMed 86 Eguchi , M. and Yamaguchi , S . 2009 . In vivo and in vitro visualization of gene expression dynamics over extensive areas of the brain . Neuroimage 44 : 1274 . Google Scholar CrossRef Search ADS PubMed 87 Chen , Q. , Cichon , J. , Wang , W. , et al. 2012 . Imaging neural activity using Thy1-GCaMP transgenic mice . Neuron 76 : 297 . Google Scholar CrossRef Search ADS PubMed 88 Kim , C. K. , Adhikari , A. and Deisseroth , K . 2017 . Integration of optogenetics with complementary methodologies in systems neuroscience . Nat. Rev. Neurosci . 18 : 222 . Google Scholar CrossRef Search ADS PubMed 89 Campbell , E. J. and Marchant , N. J . 2018 . The use of chemogenetics in behavioural neuroscience: receptor variants, targeting approaches and caveats . Br. J. Pharmacol . 175 : 994 . Google Scholar CrossRef Search ADS PubMed 90 Zeng , H . 2018 . Mesoscale connectomics . Curr. Opin. Neurobiol . 50 : 154 . Google Scholar CrossRef Search ADS PubMed 91 Kebschull , J. M. , Garcia da Silva , P. , Reid , A. P. , Peikon , I. D. , Albeanu , D. F. and Zador , A. M . 2016 . High-throughput mapping of single-neuron projections by sequencing of barcoded RNA . Neuron 91 : 975 . Google Scholar CrossRef Search ADS PubMed © The Japanese Society for Immunology. 2018. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/about_us/legal/notices)

Journal

International ImmunologyOxford University Press

Published: May 15, 2018

There are no references for this article.

You’re reading a free preview. Subscribe to read the entire article.


DeepDyve is your
personal research library

It’s your single place to instantly
discover and read the research
that matters to you.

Enjoy affordable access to
over 18 million articles from more than
15,000 peer-reviewed journals.

All for just $49/month

Explore the DeepDyve Library

Search

Query the DeepDyve database, plus search all of PubMed and Google Scholar seamlessly

Organize

Save any article or search result from DeepDyve, PubMed, and Google Scholar... all in one place.

Access

Get unlimited, online access to over 18 million full-text articles from more than 15,000 scientific journals.

Your journals are on DeepDyve

Read from thousands of the leading scholarly journals from SpringerNature, Elsevier, Wiley-Blackwell, Oxford University Press and more.

All the latest content is available, no embargo periods.

See the journals in your area

DeepDyve

Freelancer

DeepDyve

Pro

Price

FREE

$49/month
$360/year

Save searches from
Google Scholar,
PubMed

Create lists to
organize your research

Export lists, citations

Read DeepDyve articles

Abstract access only

Unlimited access to over
18 million full-text articles

Print

20 pages / month

PDF Discount

20% off