The field of immunology is principally focused on the molecular mechanisms by which hematopoietic cells initiate and maintain innate and adaptive immunity. That cornerstone of attention has been expanded by recent discoveries that neuronal signals occupy a critical regulatory niche in immunity. The discovery is that neuronal circuits operating reflexively regulate innate and adaptive immunity. One particularly well-characterized circuit regulating innate immunity, the inflammatory reflex, is dependent upon action potentials transmitted to the reticuloendothelial system via the vagus and splenic nerves. This field has grown significantly with the identification of several other reflexes regulating discrete immune functions. As outlined in this review, the delineation of these mechanisms revealed a new understanding of immunity, enabled a first-in-class clinical trial using bioelectronic devices to inhibit cytokines and inflammation in rheumatoid arthritis patients, and provided a mosaic view of immunity as the integration of hematopoietic and neural responses to infection and injury.

The history of immunology is a rich story of insights that originated with clinical observations, advanced through technological innovation, and culminated in identification of cellular and molecular mechanisms. Exploiting advances from other fields of science has accelerated progress in immunology, be it growing pathogens in pure cultures, isolating pathogenic toxins and Ags, visualizing phagocytosis and cell-mediated killing, or mass-producing mAbs and cytokines. Most of this work has been focused on hematopoietic cells and the organs they inhabit, the reticuloendothelial system. The products of immunology research are new therapies that benefit millions, including treating autoimmune diseases with mAbs, and treating cancer with modified T cells and check point inhibitors. Today, by exploiting advances in neuroscience, there is a major new opportunity for immunologists to discover new mechanisms in immunity and accelerate efforts to develop new therapeutic treatments.

What happened in neuroscience to enable this possibility? A basic answer is the discovery that neurons are integral components of the immune system, and that a full understanding of immunity requires understanding the role of reflex neural circuits that regulate hematopoietic cells. Sensory neurons are present on the front line of host defense and like macrophages, other phagocytes, and APCs, these neurons interact directly with the inflammatory products of pathogens and host. This interaction initiates sensory neuronal action potentials that are conveyed into the nervous system within milliseconds of the onset of invasion or inflammation. This action-potential signaling mechanism is several orders of magnitude faster than can be accomplished by immune cell release of cytokines and other mediators. Information returning from the nervous system to the reticuloendothelial system reflexively regulates the activity of innate and adaptive immune responses. Recent advances have revealed the molecular mechanisms for the neural control of cytokine production, chemotaxis, germinal center formation, Ag presentation, and other fundamental aspects of immunity.

Considering that it has been known for decades that neurons reside within a few micrometers of virtually every cell in the reticuloendothelial system and within the epithelial system at the front line of host defense, and that neurotransmitters modulate hematopoietic cell functions, it may be appropriate to wonder why neurons have not previously been considered to be an integral component of the immune system. In this review, we review the advances in neuroscience and immunology that reveal a fundamental importance of reflex neural circuits to understanding immunity. A brief overview of the fundamental physiology of reflex circuits precedes a summary of several specific examples of reflex mechanisms in immunity. These are presented as a gentle argument that future students of immunology may be able to advance further by continuing to blur the lines between neuroscience and immunology.

A neural reflex is defined by three components: first, a sensory receptor capable of responding to a change in the environment; second, a sensory or afferent arc that transmits action potentials into the nervous system; and third, a motor or efferent arc that sends action potentials from the nervous system back to the periphery to modulate the environment. Charles Sherrington, one of the fathers of neuroscience, proposed that this basic unit of reflex action provides a fundamental building block upon which an entire nervous system can be assembled (1, 2). Reflexes function at the speed of action potentials, which is sufficient to control the output of organ systems within milliseconds. The cardiovascular, gastrointestinal, respiratory, hepatobiliary, and renal organ systems are all regulated by reflexes that respond to changes in the internal milieu and send corrective and compensatory signals to maintain homeostasis. Reflex integration of physiological output has evolutionarily ancient origins, dating back to Caenorhabditis elegans, a nematode with 302 neurons and a rudimentary immune system (3). Reflexes in mammals and vertebrates evolved to exquisitely maintain organ homeostasis despite a wide range of environmental variability and threat.

One example that perhaps best illustrates the function of a simple reflex controlling the output of a complex system can be found in the neural control of the heart rate. The principle sensory nerve to the visceral organs, including the heart and reticuloendothelial system, is the vagus nerve. It is a paired structure, arising in the brainstem, which descends through the thorax and abdomen to innervate the visceral organs. Comprised of 80,000–100,000 fibers in larger mammals, 80% of these neurons traveling in the vagus nerve are sensory. During exercise or physical exertion, increases in heart rate are sensed by the afferent fibers traveling in the vagus nerve. Activation of this sensory arc to the brainstem activates motor fibers traveling in the efferent arc that descends to the cardiac sinoatrial node. Arrival of action potentials at the heart prolongs the time to subsequent heartbeats, thereby slowing the heart rate. Simple reflexes, such as the regulation of heart rate, are activated when the output of the organ system deviates from a set point.

Within the immune system, sensory and motor nerves, the fundamental components of a simple reflex, were first identified in the organs of the reticuloendothelial system >50 y ago. The thymus, lymph nodes, liver, and spleen are all richly innervated by neurons capable of transmitting information to and from the nervous system to regulate the activity of hematopoietic cells during Ag exposure, infection, or sterile injury. Early studies of fever, inflammation, anorexia, and sickness behavior focused on products of inflammatory and immune responses modulating neuronal signals in the brainstem. For example, IL-1 was extensively studied as a mediator of fever capable of interacting with specific receptors expressed on neurons in the thermal regulatory centers of the brainstem hypothalamus (47). Activation of IL-1 receptor–dependent signaling in the brain also mediates the development of anorexia and behavioral withdrawal, termed sickness syndrome (8, 9). These and other studies with other cytokine mediators established the dogma that cytokines produced in the peripheral tissues that gain access to the brain or are produced locally in the brain by astrocytes and microglial cells are capable of mediating sickness behaviors and changing the neurologic status of the host. Seminal studies by Linda Watkins (10) soon presented a significant challenge to this simple dogma.

In the early 1990s, when Watkins and her colleagues (10, 11) administered IL-1 to rats via the i.p. route they observed, as expected, the onset of hyperthermia. When they repeated these experiments in animals subjected to vagotomy, in which the connections in the vagus nerve to the brain were cut, they found that administration of IL-1 failed to produce a fever. This gave direct evidence that sensory signals traveling in the vagus nerve participate in the onset of the fever response (10). Independent work by Niijima (1214) demonstrated that the presence of IL-1 in the liver activates ascending signals in the vagus nerve that are transmitted to the brainstem, then relayed as efferent signals descending in the vagus nerve to the spleen, thymus and other organs. Together, these findings suggested that cytokines can activate functional afferent neural signals to the brain stem. An independent study and an unexpected experimental result in our laboratory led to the discovery of the inflammatory reflex.

We had developed a cytokine blocking molecule (CNI-1493) (1518) and were studying its effects in the brains of animals subjected to cortical infarctions when we made the striking observation that production of TNF and other cytokines in the reticuloendothelial system was inhibited by the presence of CNI-1493 in the brain (16). We postulated that signals descending from the brain in the vagus nerve comprise a previously unknown motor arc in an inflammatory reflex, completing a sensory arc activated by the presence of IL-1 and other cytokines (19). We repeated the experiments after cutting the vagus nerve, and observed that CNI-1493 in the brain no longer inhibited cytokine production in the reticuloendothelial system (16). The inescapable conclusion from these experiments was that signals descending from the brainstem to the immune system in the vagus nerve regulated cytokine production.

We then began to directly explore the molecular and neurophysiological mechanisms using nerve stimulators positioned on the cervical vagus nerve in anesthetized animals during endotoxemia. The results defined the inflammatory reflex, a neural reflex circuit of action potentials traveling in the sensory and motor vagus nerve that regulate cytokine production in the spleen (1922). The splenic nerve is adrenergic, producing norepinephrine in the vicinity of lymphocytes in the spleen (23). A discrete subset of lymphocytes regulated by splenic nerve signals express choline acetyltransferase, the rate-limiting enzyme in the biosynthesis of acetylcholine (23, 24). Acetylcholine released by lymphocytes under the control of adrenergic splenic nerve signals inhibits macrophage cytokine release, and shifts them toward an M2 anti-inflammatory, tissue-protective phenotype. Acetylcholine-induced signal transduction in monocytes and macrophages is mediated by α-7 nicotinic acetylcholine receptors (25), which inhibits the nuclear translocation of NF-κB (26), activates the JAK2/STAT3 pathway (27), stabilizes mitochondrial membranes, and downregulates the activity of the inflammasome (28) (Fig. 1). The net effect of vagus nerve–mediated signals within the spleen is the inhibition of cytokine release by red pulp and marginal zone macrophages, which together account for 90% of the TNF and IL-1 produced during acute endotoxemia (23, 29).

FIGURE 1.

Mechanisms by which acetylcholine inhibits cytokine release. Extracellular acetylcholine inhibits cytokine production through nicotinic acetylcholine receptor subunit α7 (α7nAChR), which is expressed by cytokine-producing immune cells. α7nAchR signal transduction results in phosphorylation of CREB, which increases expression of cfos that inhibits NF-κB activity leading to suppression of cytokine production. Interaction of α7nAchR with JAK2 leads to phosphorylation of STAT3. Phosphorylated STAT3 dimers translocate to the nucleus to induce suppression of cytokine production. Activation of immune cells with extracellular ATP leads to rapid influx of acetylcholine into the cytoplasm. Cytoplasmic acetylcholine attenuates mitochondrial DNA release via mitochondrial α7nAchR and subsequently inhibits inflammasome activation.

FIGURE 1.

Mechanisms by which acetylcholine inhibits cytokine release. Extracellular acetylcholine inhibits cytokine production through nicotinic acetylcholine receptor subunit α7 (α7nAChR), which is expressed by cytokine-producing immune cells. α7nAchR signal transduction results in phosphorylation of CREB, which increases expression of cfos that inhibits NF-κB activity leading to suppression of cytokine production. Interaction of α7nAchR with JAK2 leads to phosphorylation of STAT3. Phosphorylated STAT3 dimers translocate to the nucleus to induce suppression of cytokine production. Activation of immune cells with extracellular ATP leads to rapid influx of acetylcholine into the cytoplasm. Cytoplasmic acetylcholine attenuates mitochondrial DNA release via mitochondrial α7nAchR and subsequently inhibits inflammasome activation.

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These mechanistic insights have enabled studies of selective nerve stimulation of the inflammatory reflex to block innate immunity and inflammation in animal models of sepsis, endotoxemia, collagen induced arthritis, hemorrhagic shock, ischemia reperfusion injury, autoimmune myocarditis, ileus, and colitis (3040). A comprehensive and unabridged review of these studies is beyond the scope of this review, but this body of work enabled the development of a clinical trial for humans with rheumatoid arthritis. As reported recently, stimulation of the inflammatory reflex in rheumatoid arthritis patients significantly inhibited cytokine production and provided therapeutic benefit in two cohorts of patients: those resistant to methotrexate therapy, and those resistant to combined therapy with methotrexate and biological inhibitors of cytokines (41). This first-in-man study validated the experiments from the basic science laboratories and applied these concepts to patients.

What can be learned from the principles of the inflammatory reflex, and how can they be applied to other questions in immunology? In essence, these studies indicate that neurons have a highly conserved role in modulating the development of immunity. By approaching fundamental questions in immunology from the perspective of neuroscience, a wealth of hypotheses become testable. By focusing experimental design on the principles of basic reflex circuits, it becomes plausible to address the role of immunological mechanisms in activating sensory neurons, modulating neurologic signaling, and responding to input from motor neurons. Including the neuron as an active participant in the development of innate and adaptive immunity provides new understanding of mechanisms and new opportunities for therapy (Fig. 2, Table I).

FIGURE 2.

Neural reflex circuits in immunity. (A) The inflammatory reflex. The efferent signals generated in the vagus nerve are transmitted via the celiac ganglion to the spleen to acetylcholine-producing (ChAT+) T cells. Acetylcholine attenuates cytokine production by macrophages in nicotinic acetylcholine receptor subunit α7 (α7nAChR)-dependent manner. (B) Splenic neural circuits modulating Ab production. Activation of vagus nerve culminates into stimulation of the adrenergic splenic nerve resulting in release of norepinephrine leading to accumulation of marginal zone B cells and decreased Ab production. (C) Vagus nerve-adrenal reflex. Activation of the sciatic nerve results in the efferent vagus nerve signals culminating in the adrenal medulla leading to increased production of dopamine that targets dopaminergic type 1 receptors and suppresses systemic inflammation. (D) Direct electrical stimulation of the vagus nerve results in acetylcholine release by cholinergic myenteric neurons leading to attenuation of inflammation in an α7nAChR manner. Additionally, enteric nerve mediated norepinephrine signals modulate the switch of muscularis macrophages to tissue protective phenotype M2 by signaling through β2-adrenergic receptors. (E) Sensory axon-axon reflex. Bacterial-derived ligands (pathogen-associated molecular patterns [PAMPs] and danger-associated molecular patterns [DAMPs]) activate sensory neurons resulting in a release of neuropeptides that directly suppress innate immune activation. (F) Nociceptive reflexes in lung. The by-products of allergic inflammatory response activate sensory neurons and immune cells in the lung. IL-5 released by activated immune cells directly activates Nav1.8 positive nociceptors to release vasoactive peptide (VIP). VIP then activates immune cells leading to an inflammatory loop. Activated immune cells produce elevated levels of IL-5, IL-13, and other cytokines that amplify the allergic inflammatory responses in the lung. (G) The Gateway reflex. Soleus muscle contractions activate afferent neural signals that culminate into efferent adrenergic signals at the lumbar 5 level modulating the expression of CCL20 by the endothelial cells and providing an important control mechanism that gates the entry of pathogenic T cells into the CNS. (H) Neural reflexes in cancer. In a model of prostate cancer, adrenergic nerves promote tumor growth via norepinephrine release and cholinergic nerves within the tumor tissue enhance tumor metastasis by releasing acetylcholine.

FIGURE 2.

Neural reflex circuits in immunity. (A) The inflammatory reflex. The efferent signals generated in the vagus nerve are transmitted via the celiac ganglion to the spleen to acetylcholine-producing (ChAT+) T cells. Acetylcholine attenuates cytokine production by macrophages in nicotinic acetylcholine receptor subunit α7 (α7nAChR)-dependent manner. (B) Splenic neural circuits modulating Ab production. Activation of vagus nerve culminates into stimulation of the adrenergic splenic nerve resulting in release of norepinephrine leading to accumulation of marginal zone B cells and decreased Ab production. (C) Vagus nerve-adrenal reflex. Activation of the sciatic nerve results in the efferent vagus nerve signals culminating in the adrenal medulla leading to increased production of dopamine that targets dopaminergic type 1 receptors and suppresses systemic inflammation. (D) Direct electrical stimulation of the vagus nerve results in acetylcholine release by cholinergic myenteric neurons leading to attenuation of inflammation in an α7nAChR manner. Additionally, enteric nerve mediated norepinephrine signals modulate the switch of muscularis macrophages to tissue protective phenotype M2 by signaling through β2-adrenergic receptors. (E) Sensory axon-axon reflex. Bacterial-derived ligands (pathogen-associated molecular patterns [PAMPs] and danger-associated molecular patterns [DAMPs]) activate sensory neurons resulting in a release of neuropeptides that directly suppress innate immune activation. (F) Nociceptive reflexes in lung. The by-products of allergic inflammatory response activate sensory neurons and immune cells in the lung. IL-5 released by activated immune cells directly activates Nav1.8 positive nociceptors to release vasoactive peptide (VIP). VIP then activates immune cells leading to an inflammatory loop. Activated immune cells produce elevated levels of IL-5, IL-13, and other cytokines that amplify the allergic inflammatory responses in the lung. (G) The Gateway reflex. Soleus muscle contractions activate afferent neural signals that culminate into efferent adrenergic signals at the lumbar 5 level modulating the expression of CCL20 by the endothelial cells and providing an important control mechanism that gates the entry of pathogenic T cells into the CNS. (H) Neural reflexes in cancer. In a model of prostate cancer, adrenergic nerves promote tumor growth via norepinephrine release and cholinergic nerves within the tumor tissue enhance tumor metastasis by releasing acetylcholine.

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Table I.
Neural reflexes in immunology
Neural ReflexExperimental ModelActivation ModelResultReference
Inflammatory reflex Endotoxin-induced shock Direct electrical stimulation of cervical vagus nerve Decreased circulating TNF, attenuation of shock (23, 25, 29, 31
Cecal ligation and puncture Transcutaneous stimulation of cervical vagus nerve Decreased circulating HMGB1, improved survival (29, 35
Collagen-induced arthritis Direct electrical stimulation of cervical vagus nerve Improved arthritis incidence (32, 36
Ileus Administration of α7-nAChR agonist Decreased intestinal inflammation (40, 57
Myocardial infarction Administration of α7-nAChR agonist Decreased cardiac myoglobin release during reperfusion injury (38
Colitis Administration of α7-nAChR agonist Decreased colitis severity (39
Hemorrhagic shock Direct electrical stimulation of cervical vagus nerve Decreased hypotension severity and circulating TNF, improved survival (33
Pancreatitis Administration of α7-nAChR agonist Decreased pancreatitis severity (30
Ischemia reperfusion injury Direct electrical stimulation of cervical vagus nerve Decreased cytokine production, attenuation of shock and tissue damage (37
Neural circuits in Ab responses Ab response to bacterial Ag Direct electrical stimulation of cervical vagus nerve Impaired migration of B cells, neutrophils, monocytes and dendritic cells, reduced Ag-specific Ab production (48
Vagus nerve-adrenal reflex Endotoxin-induced shock Electrical stimulation of the sciatic nerve Decreased cytokines release, attenuation of organ dysfunction (50, 52
Cecal ligation and puncture Electrical stimulation of the sciatic nerve Decreased cytokines and improved survival (51
Paw edema Laser stimulation of the sciatic nerve Decreased paw edema and temperature (55
Collagen-induced arthritis Electrical stimulation of the sciatic nerve Improved arthritis incidence, attenuated cytokine levels (54
Enteric neural reflex Intestinal peristalsis Selective depletion of muscularis macrophages, luminal infection Disturbed peristaltic activity, reduced neuronal activation (58, 59
Postoperative ileus Direct electrical stimulation of cervical vagus nerve Improved intestinal inflammation and postoperative ileus (57
Sensory axon-axon reflex Bacterial infection model Bacterial infection and mechanical, heat and cold hypersensitivity Bacteria directly activate nociceptors, and immune response is not necessary for bacterial-induced pain (62
Nociceptive reflex in lung Allergic lung inflammation Administration of nociceptor antagonist Reduced airway inflammation and bronchial hyper responsiveness (75
Gateway reflex Experimental allergic encephalomyelitis Tail suspension model Regulates T cell recruitment into the CNS (76
Neural reflex in cancer Prostate cancer Ablation of adrenergic neurons, surgical denervation Attenuated development of tumors in prostate (77
 Administration of cholinergic agonists Enhanced tumor cell invasion and metastasis (77
 Pharmacological blockade or genetic disruption of the stromal type 1 muscarinic receptor Improved survival (77
Gastric cancer Surgical denervation of the stomach Reduced tumor incidence and progression (80
Lung cancer Administration of adrenergic antagonist Reduced metastasis (81
Ovarian cancer Restraint stress Increased tumor growth (82, 83
Melanoma Administration of adrenergic antagonist Decreased tumor growth, increased tumor apoptosis and reduced angiogenesis (84
Neural ReflexExperimental ModelActivation ModelResultReference
Inflammatory reflex Endotoxin-induced shock Direct electrical stimulation of cervical vagus nerve Decreased circulating TNF, attenuation of shock (23, 25, 29, 31
Cecal ligation and puncture Transcutaneous stimulation of cervical vagus nerve Decreased circulating HMGB1, improved survival (29, 35
Collagen-induced arthritis Direct electrical stimulation of cervical vagus nerve Improved arthritis incidence (32, 36
Ileus Administration of α7-nAChR agonist Decreased intestinal inflammation (40, 57
Myocardial infarction Administration of α7-nAChR agonist Decreased cardiac myoglobin release during reperfusion injury (38
Colitis Administration of α7-nAChR agonist Decreased colitis severity (39
Hemorrhagic shock Direct electrical stimulation of cervical vagus nerve Decreased hypotension severity and circulating TNF, improved survival (33
Pancreatitis Administration of α7-nAChR agonist Decreased pancreatitis severity (30
Ischemia reperfusion injury Direct electrical stimulation of cervical vagus nerve Decreased cytokine production, attenuation of shock and tissue damage (37
Neural circuits in Ab responses Ab response to bacterial Ag Direct electrical stimulation of cervical vagus nerve Impaired migration of B cells, neutrophils, monocytes and dendritic cells, reduced Ag-specific Ab production (48
Vagus nerve-adrenal reflex Endotoxin-induced shock Electrical stimulation of the sciatic nerve Decreased cytokines release, attenuation of organ dysfunction (50, 52
Cecal ligation and puncture Electrical stimulation of the sciatic nerve Decreased cytokines and improved survival (51
Paw edema Laser stimulation of the sciatic nerve Decreased paw edema and temperature (55
Collagen-induced arthritis Electrical stimulation of the sciatic nerve Improved arthritis incidence, attenuated cytokine levels (54
Enteric neural reflex Intestinal peristalsis Selective depletion of muscularis macrophages, luminal infection Disturbed peristaltic activity, reduced neuronal activation (58, 59
Postoperative ileus Direct electrical stimulation of cervical vagus nerve Improved intestinal inflammation and postoperative ileus (57
Sensory axon-axon reflex Bacterial infection model Bacterial infection and mechanical, heat and cold hypersensitivity Bacteria directly activate nociceptors, and immune response is not necessary for bacterial-induced pain (62
Nociceptive reflex in lung Allergic lung inflammation Administration of nociceptor antagonist Reduced airway inflammation and bronchial hyper responsiveness (75
Gateway reflex Experimental allergic encephalomyelitis Tail suspension model Regulates T cell recruitment into the CNS (76
Neural reflex in cancer Prostate cancer Ablation of adrenergic neurons, surgical denervation Attenuated development of tumors in prostate (77
 Administration of cholinergic agonists Enhanced tumor cell invasion and metastasis (77
 Pharmacological blockade or genetic disruption of the stromal type 1 muscarinic receptor Improved survival (77
Gastric cancer Surgical denervation of the stomach Reduced tumor incidence and progression (80
Lung cancer Administration of adrenergic antagonist Reduced metastasis (81
Ovarian cancer Restraint stress Increased tumor growth (82, 83
Melanoma Administration of adrenergic antagonist Decreased tumor growth, increased tumor apoptosis and reduced angiogenesis (84

Lymphocytes and monocytes express α-adrenergic and β-adrenergic receptors (β-AR), and binding of norepinephrine modulates cell homing, proliferation, and immune cell function. Catecholamines have been implicated in diverse responses to B cell activation and Ab production. Exposure of naive B cells to norepinephrine can either increase (42) or decrease (43) Th2 cell-dependent Ab production. Norepinephrine-mediated modulation of B cell Ab production requires signaling via β-ARs, but not α-adrenergic receptors (44). Maturity states of B cells govern the effects of norepinephrine on Ab production: specific activation of β2AR in early stages of B cell activation may increase the number of Ab-secreting cells or the level of Ab produced by Ab-secreting cells, whereas B cell differentiation and function is inhibited by β2AR stimulation in more mature cells (45, 46). Animals lacking β2AR expression have normal lymphoid organ cell number, phenotype, and histology, contact sensitivity responses, and T cell-dependent Ab responses as compared with wild type animals. B cells devoid of β2AR fail to respond to norepinephrine, indicating that other adrenergic receptors do not compensate for the absence of β2AR (47). Together, these data indicate that non-adrenergic mechanisms can maintain immune homeostasis.

Reflex neural networks in lymphoid organs maintain the integrity of Ab responses. During pneumococcal invasion, neutrophils, monocytes, and dendritic cells capture and transport the bacteria to the spleen, where resident Ag-specific marginal zone B cells and B1 cells are activated to differentiate into Ab-secreting cells. Marginal zone B cells migrate along splenic nerves, which secrete norepinephrine as the neurotransmitter, arriving at the red pulp venous sinuses where they become Ab-secreting cells, releasing Abs into the blood (48). Adrenergic splenic neurotransmitter release is enhanced by stimulation of the vagus or splenic nerves, providing a critical regulatory step in splenic B cell activation and migration that is key to the efficient secretion of Abs (48).

As noted earlier, the adrenergic splenic nerve also modulates the activity of choline acetyltransferase–expressing T cells that produce acetylcholine in spleen, and are necessary to suppress cytokine production by splenic macrophages under the control of the inflammatory reflex (24). Activation of the inflammatory reflex by direct electrical stimulation of the vagus nerve, or administration of nicotine, arrests neutrophil, dendritic cell and B-cell migration, and decreases Ab secretion. Chemical ablation of the adrenergic splenic nerves decreases Ab titers significantly, indicating that adrenergic signals modulate B cell activation and Ab responses. Importantly, ablation of splenic nerves suppresses the inhibitory effect of nicotine on Ab production, indicating that both cholinergic and adrenergic signals modulate B cell Ab responses (48). Detailed analysis of the neural regulation of Ab responses indicates that cholinergic stimulation induces accumulation of Ag-specific activated B cells in the marginal zone (48). The implications of these results extend well beyond Ab production to other adaptive immune responses, because neural signals also control egress of immune cells from the bone marrow (49).

Torres-Rosas and colleagues (50) mapped a discreet immunoregulatory neural reflex that utilizes cholinergic and dopaminergic neurons. By subjecting animals to adrenalectomy, cervical, or subdiaphragmatic vagotomy, they mapped a sciatic to vagus neural reflex circuit that regulates inflammatory responses. Electrical stimulation of the sciatic nerve sends sensory signals to the brainstem that in turn activate efferent vagus nerve firing to the adrenal medulla. Vagus nerve–mediated activation of the adrenal medulla increased production of dopamine, which suppresses systemic inflammation by signaling through dopaminergic type 1 receptors to improve survival in animals with lethal sepsis (50).

Similar findings were obtained in other models of inflammatory diseases. Electrical stimulation of the sciatic nerve significantly reduced circulating levels of cytokines in a murine endotoxemia model in a vagus nerve–dependent manner. Blocking of vagus nerve activity with either pharmacological antagonists or surgical ablation suppressed the inhibitory effects of electrical stimulation (5153). In a collagen-induced arthritis model, electrical stimulation of the sciatic nerve significantly reduced arthritis incidence, attenuated inflammatory cytokine levels, and prevented joint destruction (54). More recently, activation of the sciatic nerve using low-power laser therapy significantly reduced localized inflammation in a murine paw edema model (55). The precise route by which the activation of the sciatic nerve transmits the signals to the brainstem is not known, but may involve sensory neurons in the sciatic nerve, the lumbosacral plexus, the vagus nerve, and a terminal dopaminergic neurotransmitter. It is interesting to consider the therapeutic implications of a sciatic-vagus-neural circuit because it may be possible to stimulate afferent neural responses in the sciatic nerve to induce anti-inflammatory efferent vagus nerve signaling to suppress inflammatory responses in infection and autoimmunity.

Macrophages are anatomically stratified in the layers of the intestinal wall, and play a vital role in maintaining tissue homeostasis through scavenging bacteria and foreign Ags (56). In the gastrointestinal tract, which is continuously exposed to microbial products and dietary Ags, macrophage activation is tightly controlled to prevent chronic inflammation and tissue damage. Three recent studies provide evidence that enteric neurons reflexively modulate functional phenotypes of the intestinal macrophages under normal physiological conditions and in bacterial infection. In a murine model of postoperative ileus, electrical stimulation of the vagus nerve attenuated inflammation and improved postoperative intestinal transit (57). Anterograde tracing of the vagus nerve to gut demonstrated a dense network of cholinergic vagus fibers in close proximity to muscularis macrophages. Acetylcholine released by cholinergic myenteric neurons directly modulated inflammatory responses by muscularis macrophages, and activation of α7 nicotinic receptors suppressed inflammatory responses in the muscle layer (57). Enteric neurons regulate homeostasis of muscularis macrophages through the production of CSF-1, a growth factor necessary for macrophage survival (58). Muscularis macrophages, in turn, secrete BMP-2, which activates the BMP receptor expressed by enteric neurons. BMPR-mediated activation of enteric neurons is essential for maintaining proper tissue function. Selective ablation of the muscularis macrophage population is correlated with reduced neuronal activation of SMAD1, SMAD5, and SMAD8, the downstream signaling mediators of BMP receptors, resulting in impaired contractility (58). Interestingly, antibiotic-treated mice expressed significantly less BMP-2 as compared to controls, indicating that luminal microbiota regulate the neuronal interaction with macrophages (58).

Recent studies identified a novel reflex mechanism in the gut activated by the gut microbiome that forces macrophages into a tissue protective phenotype (59). Intestinal resident macrophages exhibit distinct cell dynamics, morphological features, and functional phenotypes depending on their proximity to the gut lumen. Lamina propria macrophages reside closer to the gut mucosa, and preferentially express a proinflammatory M1 phenotype. In contrast, muscularis macrophages are located more distally from the luminal surface in the submucosal region between the circular and longitudinal muscle layers, and primarily express an M2 tissue-protective gene profile. Muscularis macrophages share a niche with a dense network of neurons. Luminal bacterial infection results in the activation of extrinsic adrenergic neurons innervating the intestine. Norepinephrine released by these neurons in turn activates development of a tissue-protective phenotype in muscularis macrophages by signaling through β2-adrenergic receptors. Electron microscopy and immune staining indicate that muscularis macrophages form direct synapses with enteric neurons (60, 61). Thus, the bacterial Ag-mediated tissue protective program in muscularis macrophages is regulated by adrenergic neural circuits.

Bacterial-derived N-formylated peptides or toxins interact with receptors expressed on nociceptor neurons (62). Immune cell activation is not necessary for bacteria-induced hyperalgesia during infection. Moreover, direct binding of bacterial products to nociceptor neurons induces an axonal-axonal reflex mechanism that modulates innate immune responses. Activated nociceptor neurons release neuropeptides, including galanin, somatostatin and calcitonin gene-related peptide (CGRP) that directly suppress innate immune activation in the region of infection (62). Accordingly, ablation of nociceptor neurons increases immune cell influx and lymphadenopathy. It is plausible that pathogenic bacteria use this neural mechanism to enhance virulence (62). Gram-negative bacteria induce hyperalgesia by directly activating TRPA1 expressing nociceptors. LPS, a cell wall component of gram-negative bacteria, activates and depolarizes TRPA1-expressing nociceptors in a TLR4-independent manner, and induces hyperalgesia (63). LPS-induced hyperalgesia may also play a major role in chronic pain and neuropathy observed in diabetes. Pattern recognition receptors, including TLR4 and CD14, are expressed on sensory neurons (6466). LPS binds directly to sensory neurons in a TLR4-dependent manner, and induces production of nociceptin (67), an opioid-related peptide that is upregulated during inflammation and associated with hyperalgesia (6769). LPS binding also triggers activation of sensory neurons to enhance production of the vasodilator CGRP from the dorsal root ganglion (70, 71). Together, these studies indicate that direct activation of nociceptors by bacterial products participate in the molecular mechanisms of pain and stimulate reflex regulation of innate immunity.

The lung is innervated by a rich network of sensory neurons that express nociceptive receptors (7274). Activation of nociceptors enhances the release of neuropeptides, including CGRP and substance P. These mediators increase vascular permeability and vasodilation, and activate innate and adaptive immune cells. Recent evidence indicates that by-products of allergic inflammatory responses activate lung nociceptors to modulate allergic lung inflammatory responses (75). Specifically, IL-5 released by activated immune cells induces the release of vasoactive peptide (VIP) by Nav1.8 positive nociceptors. VIP in turn activates CD4+ T cells and type 2 innate lymphoid cells in a VIP receptor(VPAC2)–dependent manner to produce elevated levels of IL-5, IL-13, and other cytokines. VIP signaling through VPAC2 activates and recruits immune cells to amplify inflammatory responses in the lung. Silencing of nociceptors by either genetic ablation or chemical blocking significantly inhibits cytokine production, and increases resolution of allergic lung inflammation. These studies delineate neural circuits that drive a feed-forward amplification loop to exacerbate allergic lung inflammation opening new avenues for developing therapeutic strategies.

The blood-brain barrier strictly limits the flow of proteins and cells from the blood stream to the CNS. In a mouse model of multiple sclerosis, Arima et al. (76) identified a specific neural circuit that modulates trafficking of pathogenic T cells into the CNS. In this adoptive transfer model of experimental allergic encephalomyelitis, myelinated nerve specific T cells were introduced into naive recipient mice. Autoreactive T cells preferentially accumulated at specific sites in the spinal cord near the fifth lumbar level, rather than in the brain or at other levels of the spinal cord. In an attempt to elucidate the mechanism for this specificity, they discovered that the expression of CCL20, a chemokine important for T cell accumulation, is increased in the dorsal blood vessels of the fifth lumbar cord. As the fifth lumbar vertebra lies close to the dorsal root ganglia of sensory neurons that innervate the soleus and other leg muscles, they reasoned that weight-bearing stimulation of the soleus muscles might contribute to the sensory nerve activation resulting in high CCL20 expression in the blood vessels of the fifth lumbar cord. They studied this possibility by silencing the muscle-sensitive neuronal pathway by suspending mice from their tail, so that forelimbs, and not the hind limbs, were subjected to weight bearing. This significantly decreased CCL20 expression and suppressed pathogenic T cell accumulation in the fifth lumbar cord level. Instead, these pathogenic T cells accumulated at the cervical cord, indicating that stimulation of the forelimbs opened a new gateway for the pathogenic cells. Moreover, pharmacological inhibition of adrenergic receptors significantly suppressed accumulation of pathogenic T cells, and improved clinical signs of the disease. Together, these studies reveal a reflex circuit that controls a critical step in the initiation and progression of autoimmune disease by specifically regulating blood-brain barrier permeability to pathogenic T cells.

Neural circuits play a critical role in both the development and spread of tumors. In an experimental model of prostate cancer, adrenergic nerves promoted tumor growth by producing norepinephrine, which binds to and stimulates adrenergic receptors (β2 and β3) on the stromal cells in the tumor microenvironment (77). In addition, cholinergic nerves within the tumor tissue enhanced tumor metastasis by releasing acetylcholine, which activates the stromal type 1 muscarinic receptor, and enhances prostate cancer metastasis to distant lymph nodes (77). Recent observations in patients with prostate cancer support the preclinical data, and suggest that neural circuits are critical for tumor development and metastasis (77). Higher neuronal density is observed within tumors and in normal prostate tissue surrounding tumors in patients with aggressive prostate cancers as compared with patients who had less aggressive tumors. Specifically, normal prostate tissue surrounding the tumor is densely innervated with adrenergic fibers, whereas cholinergic fibers are restricted inside the tumor. High neural densities of adrenergic fibers are associated with a higher tumor proliferative index, shorter recurrence time, and tumor metastasis, whereas cholinergic fiber density is significantly associated with extraprostatic extension. These observations correlate with the epidemiological data showing that β-blockers are associated with improved survival in prostate cancer (78, 79).

Neural reflex circuits have also been implicated in development and progression of other cancers. Denervation of the stomach reduced tumor incidence and progression in preclinical models of gastric cancer (80). In rodent models of melanoma, lung cancer, and ovarian cancer, sustained adrenergic activation promotes cancer growth and metastasis (8184). In addition to their effect on tumor cells, norepinephrine and epinephrine also stimulate endothelial cells and fibroblasts, and modulate functions of immune cells, leaving open a broader potential impact of neurotransmitters on the tumor microenvironment (85). Several retrospective clinical studies have revealed that β-blockers may have antitumor activity, reducing metastasis, tumor recurrence, and cancer-specific mortality, and increasing survival time for patients with melanoma, breast, lung, ovarian, and prostate cancer (78, 8690).

The discovery that neurons operating reflexively are functional components of the immune system represents a fundamentally new insight into understanding immunity. Cells of hematopoietic origin share the frontline of host defense with neurons. Monocytes and neurons both detect changes in the environment. Macrophages can respond by producing cytokines and other mediators, and neurons respond by producing action potentials. Neurons and macrophages indeed share the responsibility for mobilizing host responses to infection and injury, and for producing a coordinated and balanced immune response. Highly conserved neural circuits act reflexively to maintain homeostasis during the development of immunity, and inflammation. This discovery has raised many more questions that future students of immunology will likely approach, expanding their research focus from cells of hematopoietic origin to include neurons. These new studies of immunology that exploit essentials from neuroscience will continue to enlighten our understanding of basic principles governing immunity.

This work was supported by a grant from the National Institute of General Medical Sciences, National Institutes of Health (1R35 GM118182-01 to K.J.T.).

Abbreviations used in this article:

β2AR

β2-adrenergic receptor

CGRP

calcitonin gene-related peptide

VIP

vasoactive peptide.

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The authors have no financial conflicts of interest.