The Cholinergic Anti-inflammatory Pathway: A Missing Link
VALENTIN A PAVLOV,1HONG WANG,1CHRISTOPHER J CZURA,1STEVEN G FRIEDMAN,1,2
AND KEVIN J TRACEY1
This review outlines the mechanisms underlying the interaction between the nervous and immune systems of the host in
response to an immune challenge. The main focus is the cholinergic anti-inflammatory pathway, which we recently described
as a novel function of the efferent vagus nerve. This pathway plays a critical role in controlling the inflammatory response
through interaction with peripheral α7 subunit–containing nicotinic acetylcholine receptors expressed on macrophages. We
describe the modulation of systemic and local inflammation by the cholinergic anti-inflammatory pathway and its function as
an interface between the brain and the immune system. The clinical implications of this novel mechanism also are discussed.
Inflammation is a normal response to disturbed homeostasis caused
by infection, injury, and trauma. The host responds with a complex
series of immune reactions to neutralize invading pathogens, repair
injured tissues, and promote wound healing (1,2). The onset of
inflammation is characterized by release of pro-inflammatory medi-
ators including tumor necrosis factor (TNF), interleukin (IL)-1,
adhesion molecules, vasoactive mediators, and reactive oxygen
species (1–3). The early release of pro-inflammatory cytokines by
activated macrophages has a pivotal role in triggering the local
inflammatory response (2). Excessive production of cytokines, such
as TNF, IL-1β, and high mobility group B1 (HMGB1), however, can
be more injurious than the inciting event, initiating diffuse coagula-
tion, tissue injury, hypotension, and death (2,4–6). The inflamma-
tory response is balanced by anti-inflammatory factors including
the cytokines IL-10 and IL-4, soluble TNF receptors, IL-1 receptor
antagonists, and transforming growth factor (TGF)β. Although sim-
plistic (7,8), the pro-/anti- terminology is widely used in the discus-
sion of the complex cytokine network. Apart from their involve-
ment in local inflammation, TNF and IL-1β are signal molecules for
activation of brain-derived neuroendocrine immunomodulatory
responses. Neuroendocrine pathways, such as the hypothalamo-
pituitary-adrenal (HPA) axis and the sympathetic division of the
autonomic nervous system (SNS) (9–15), control inflammation as an
anti-inflammatory balancing mechanism. The host thereby mobi-
lizes the immunomodulatory resources of the nervous and
endocrine systems to regulate inflammation.
Restoration of homeostasis as a logical resolution of inflam-
mation does not always occur. Insufficient inflammatory responses
may result in increased susceptibility to infections and cancer.
On the other hand, excessive responses are associated with auto-
immune diseases, diabetes, sepsis, and other debilitating condi-
tions. When control of local inflammatory responses is lost,
pro-inflammatory mediators can spill into the circulation, result-
ing in systemic inflammation that may progress to shock, multi-
ple organ failure, and death. Effective therapies for diseases of
excessive inflammation are needed.
We recently discovered the anti-inflammatory role of the vagus
nerve (16,17) in an animal model of endotoxemia and shock. This
previously unrecognized immunomodulatory circuit termed the
“cholinergic anti-inflammatory pathway” is a mechanism for
neural inhibition of inflammation (18,19), and interfaces the brain
with the immune system. Can it be a “missing link” in
neuroimmunomodulation that will validate the notion of a mind-
This review outlines brain-derived control mechanisms of
immune function and specifically the role of cholinergic anti-
inflammatory pathway in the regulation of inflammation.
INTERACTION BETWEEN THE IMMUNE SYSTEM
AND BRAIN IN RESPONSE TO IMMUNE CHALLENGE
Communication between the immune, nervous, and endocrine
systems is essential for host defense and involves a variety of
mediators including cytokines, neurotransmitters, hormones, and
humoral factors. The influence of the brain on immune function
and the mechanisms involved in these interactions have been
elucidated over the past 3 decades (17–19). Two important ques-
tions arise when describing the brain-derived immunomodula-
tion: 1) How is the brain initially signaled by cytokines to trigger
corresponding neural and neuroendocrine responses; and 2) how
is immunomodulation achieved through these mechanisms?
The brain can monitor immune status and sense peripheral
inflammation through 2 main pathways: neural and humoral
(Figure 1; for a review, see 20, 21).
1Laboratory of Biomedical Science, North Shore LIJ-Research Institute, and 2Department of Vascular Surgery,
North Shore University Hospital, Manhasset, NY.
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Neural pathway. The neural mechanism relies upon activation of
vagus nerve afferent sensory fibers that signal the brain that
inflammation is occurring. Immunogenic stimuli activate vagal
afferents either directly by cytokines released from dendritic cells,
macrophages, and other vagal-associated immune cells, or indi-
rectly through the chemoreceptive cells located in vagal paragan-
glia (22). For instance, intraperitoneal administration of endotoxin
can induce IL-1β immunoreactivity in dendritic cells and macro-
phages within connective tissues associated with the abdominal
vagus nerve and subsequently in vagal paraganglia and afferent
fibers (23). Visceral vagus afferent fibers, residing in the nodose
ganglion, terminate primarily within the dorsal vagal complex
(DVC) of the medulla oblongata. The DVC consists of the nucleus
tractus solitarius (NTS), the dorsal motor nucleus of the vagus
(DMN), and the area postrema (AP) (24). The DMN is the major
site of origin of preganglionic vagus efferent fibers; cardiovascular
vagal efferents also originate within the medullar nucleus ambigu-
ous. The AP, which lacks a blood-brain barrier, is an important
circumventricular organ and site for humoral immune-to-brain
communication, as described below. The main portion of vagal
sensory input is received by neurons in the NTS that coordinate
autonomic function and interaction with the endocrine system
(25). Ascending projections from the NTS reach forebrain sites
including hypothalamic nuclei, amygdala, and insular cortex. One
of the hypothalamic nuclei receiving input from the NTS is the
paraventricular nucleus (PVN). The PVN is associated with the
synthesis and release of corticotropin releasing hormone (CRH),
an important substance in the HPA axis. This ascending link
between the NTS and PVN provides a pathway that can modulate
neurohormonal anti-inflammatory responses. Synaptic contacts
also exist between the neurons in the NTS and C1 neurons in the
rostral ventrolateral medulla (RVM), which occupies an important
role in control of cardiovascular homeostasis. The RVM neurons in
turn project to the locus coeruleus (LC), which is the main source
of noradrenergic innervations of higher brain sites, including the
hypothalamus and PVN. Projections emanate from the RVM and
LC to sympathetic preganglionic neurons in the spinal cord. There
are also descending pathways from the PVN to the RVM and NTS.
These ascending and descending connections provide a neuronal
substrate for interaction between HPA axis and SNS as an
The transmission of cytokine signals to the brain through the
vagal sensory neurons depends upon the magnitude of the
immune challenge. Subdiaphragmatic vagotomy inhibits the
stimulation of the HPA axis (26) and norepinephrine (NE) release
in hypothalamic nuclei (27) in response to intraperitoneal admin-
istration of endotoxin or IL-1β. Intravenous endotoxin adminis-
tration induces expression of the neural activation marker c-Fos
in the brainstem medulla, regardless of the integrity of the vagus
nerve (28). Vagotomy fails to suppress high dose endotoxin-
induced IL-1β immunoreactivity in the brain (29) and increases
blood corticosterone levels (30). It is likely that the vagal afferent
neural pathway plays a dominant role in mild to moderate
peripheral inflammatory responses, whereas acute, robust
inflammatory responses signal the brain primarily via humoral
Humoral pathway. A large body of evidence supports the
involvement of humoral mechanisms in the immune-to-brain
communication, especially in cases of systemic immune challenge
(21, 31–33). The question remains, however, of how the circulating
cytokines interact with brain structures involved in the anti-
inflammatory response, and how circulating cytokines induce
central cytokine production associated with fever and sickness.
Blood-borne IL-1β and TNF can cross the blood-brain barrier and
enter cerebrospinal fluid and the interstitial fluid spaces of the
brain and spinal cord by a saturable carrier-mediated mechanism
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Figure 1. Neural and humoral pathways in immunomodulation. During
immune challenge activated macrophages and other immune and
nonimmune cells release cytokines that signal the brain for activation of
immunomodulatory mechanisms. Central immunomodulation is
achieved by the cholinergic anti-inflammatory pathway, HPA axis, and
SNS (see text for details). AP, area postrema; NTS, nucleus tractus solitar-
ius; DMN, dorsal motor nucleus of the vagus; PVN, paraventricular
nucleus; RVM, rostral ventrolateral medulla; LC, locus coeruleus;
SNS, sympathetic nervous system; ACTH, adrenocorticotropin hormone;
GC, glucocorticoids; EN, epinephrine; NE, norepinephrine; ACh, acetyl-
choline; LPS, lipopolysaccharide (endotoxin).
(34) that may function only at very high plasma cytokine concen-
trations. Cytokines also can bind to receptors at the surface of the
endothelium of the brain capillaries and can enhance the synthe-
sis and release of soluble mediators such as prostaglandins and
nitric oxide, which diffuse into the brain parenchyma and modu-
late the activity of specific groups of neurons (21,35,36). It has
been suggested that prostaglandins mediate fever and HPA axis
Cytokine-to-brain communication also may occur via circum-
ventricular organs that lack normal blood-brain barrier function.
Among the circumventricular organs, the AP appears to represent
the best candidate for such a transduction site (for a review, see
37). The AP is located in the floor of the caudal 4th ventricle (38)
and dendrites of neurons in the NTS and DMN penetrate both the
APand floor of the 4th ventricle (39,40). The close proximity of AP
to NTS and RVM and the existing neural connections provide a
way of signaling the SNS and HPA axis. Cytokine-induced pro-
duction of prostaglandins within the AP, NTS, and RVM may acti-
vate the catecholamine projections to the PVN, resulting in subse-
quent HPA axis activation (37). This is one possible interaction
between the neural and humoral mechanisms of immune-to-
brain communication through which the brain mediates anti-
Apart from their function in signaling the brain for immuno-
modulatory responses, cytokines play a multifunctional role in
brain injury and neurodegenerative diseases (for review, see 41–43).
The brain exerts strong modulatory effects on immune function by
activation of the HPA axis and the SNS, which results in increased
synthesis and release of glucocorticoids and catecholamines (see
Figure 1). The immunomodulating properties of α-melanocyte
stimulating hormone (α-MSH) and estrogens also are known. The
HPA axis is a neurohormonal pathway; its role in the regulation
of the immune function has been widely studied (for reviews, see
14,44–46). The components of the HPA axis are the hypothalamic
PVN, the anterior pituitary gland, and the adrenal cortex. Spe-
cialized neurons in PVN synthesize CRH, which is released into
the pituitary portal blood system and stimulates the synthesis of
adrenocorticotropin hormone (ACTH) from the anterior pituitary.
ACTH is the main inducer of the synthesis and release of
immunosuppressive glucocorticoids (cortisol in humans and cor-
ticosterone in rats) from the adrenal cortex. Pro-inflammatory
cytokines trigger the HPA axis via the neural or humoral mecha-
nisms described above. At both the hypothalamic and pituitary
level, the HPA axis is subject to a classical negative feedback loop
by the final product: glucocorticoids. ACTH also inhibits the syn-
thesis of CRH from the PVN. The hypothalamo-pituitary circuit
of the HPA axis is regulated by neural mechanisms including
acetylcholine (ACh)-, catecholamine-, GABA-, serotonin-, and
Glucocorticoids exert their effects by binding to intracellular
receptors and subsequently triggering up-regulation or down-
regulation of gene expression (47). Apart from triggering the acti-
vation of the HPA axis, cytokines such as IL-1 and IL-6 also can
alter peripheral glucocorticoid effects by directly influencing the
function of corresponding glucocorticoid receptors (45). Immuno-
suppressive glucocorticoid influence is mainly linked to sup-
pression of nuclear factor-κB activity (14,48,49), which plays an
important role in regulating cytokine synthesis (50). As summa-
rized by Webster and others (14), glucocorticoids inhibit the syn-
thesis of pro-inflammatory cytokines, such as TNF, IL-1, IL-8, IL-
11, IL-12, and interferon-γ; and they activate the synthesis of the
anti-inflammatory cytokines IL-4 and IL-10. Inhibition of neu-
trophil, eosinophil, monocyte, and macrophage infiltration, and
adhesion molecule expression are attributed to glucocorticoid
suppression of local inflammation (14,45). Glucocorticoids are
also potent clinical anti-inflammatory agents (45).
The SNS plays a dual role in the regulation of inflammation,
because it mediates both pro- and anti-inflammatory activities; it
is thus an integral component of the host defense system against
injury and infection (15,51). The locus coeruleus (LC) and RVM,
brain functional components of the SNS, project to sympathetic
preganglionic cholinergic neurons in the spinal cord. Sympathetic
innervation of primary (thymus and bone marrow) and second-
ary (spleen, lymph nodes, and tissues) lymphoid organs is the
anatomic basis for modulation of immune function by the SNS
(15,51,52). Sympathetic postganglionic norepinephrine (NE)-
ergic and neuropeptide Y-ergic neurons also innervate blood
vessels, the heart, the liver, and the gastrointestinal tract (15,52).
NE, released from sympathetic postganglionic nerve endings,
exerts anti-inflammatory effects by interacting with adrenocep-
tors expressed on lymphocytes and macrophages. Adrenoceptors
belong to the G-protein coupled receptor superfamily and are
divided into α and β subtypes, which can be further subdivided.
Sympathetic control on cytokine production is achieved by
2 mechanisms: (1) synaptic-like junctions with corresponding
adrenoceptors (as in the spleen) and (2) nonsynaptic, distant
mechanisms, such as NE diffusion through the parenchyma
before interaction with the receptor. The nonsynaptic mechanism
plays a dominant physiological role (15). The release of NE is sub-
ject to complex presynaptic regulation, involving the effects of
neuropeptide Y, acetylcholine, dopamine, prostaglandins, and
other micro-environmental factors. Sympathetic immunomodula-
tion also is mediated via epinephrine, and to a lesser extent, by
NE released from the chromaffin cells of the adrenal medulla. The
chromaffin cells represent homologs of the sympathetic ganglia.
Activation of the preganglionic sympathetic neurons innervating
these cells leads to an increase in the release of catecholamines in
the bloodstream, which can act systematically as hormones. Thus,
sympathetic neural regulation is converted into “hormonal regu-
lation” within the adrenal glands. The adrenals are therefore an
important peripheral component of the CNS-controlled immuno-
regulation responsible for the synthesis of glucocorticoids (from
the cortical cells) and catecholamines (from the medullar chro-
SNS activation protects the organism from the detrimental
effects of pro-inflammatory cytokines. Activation of β-adrenocep-
tors leads to marked inhibition of endotoxin induced serum TNF,
IL-1, IL-12, interferon-γ, and nitric oxide production, and elevation
of IL-6 and IL-10. Stimulation of β-adrenoceptors also is accompa-
nied by suppression of the TNF and IL-1 expression caused by
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hemorrhagic shock. NE and epinephrine inhibit production of pro-
inflammatory cytokines through stimulation of β2-adrenoceptor-
cAMP-protein kinase A pathway, and they stimulate the synthesis
of anti-inflammatory cytokines. Healthy volunteers receiving epi-
nephrine and subsequent endotoxin treatment developed signifi-
cantly decreased TNF levels and elevated IL-10 levels, as compared
with controls exposed to endotoxin alone (53), raising the possibil-
ity that pharmacological control of cytokine production in septic
patients could be achieved by selective adrenoceptor agonists and
antagonists. Acatecholamine-based strategy for treating sepsis and
its complications is difficult, however, because of the broad effects
of these agents and the need to balance the cardiovascular and
immunomodulatory drug effects (15).
During the early stages of some cases of inflammation, stim-
ulation of the SNS can be associated with activation of the local
inflammatory responses and neutrophil accumulation (1,54). NE
stimulation of the α2–subtype adrenoceptor has been linked to an
increase in endotoxin-induced production of TNF and other
cytokines (15,55). This dual role of the SNS in immunomodulation
agrees with the general mode of SNS function, depending on the
peripheral receptors involved. A classic example of dual effects is
the “fight-or-flight” response, leading to increased perfusion of
the heart, skeletal muscles, and brain, and release of glucose from
the liver. Gastrointestinal motility and the blood supply to the
skin are simultaneously depressed.
Anti-inflammatory properties of α-MSH have been shown in
animal models of inflammation including sepsis and rheumatoid
arthritis (for a review, see 56, 57). In rodents α-MSH is mainly pro-
duced from the intermediate lobe of the pituitary gland. The cells
of the pituitary pars distalis and various extrapituitary cells, such
as monocytes, astrocytes, and keratinocytes, are the source of this
hormone production in humans. The activity of α-MSH is medi-
ated through G-protein coupled melanocortin receptors widely
distributed in peripheral tissues and in the brain. Immunomodu-
latory effects of α-MSH are associated with stimulation of its
receptors on peripheral immune target cells and on glial cells in
brain; α-MSH may also exert indirect effects via a brain-spinal
cord pathway and sympathetic neurons (58). Although the exact
molecular mechanisms of α-MSH immunosuppression and pro-
inflammatory cytokine downregulation are not well understood,
hormonal inhibition of nuclear factor κB may play an important
role. The administration of endotoxin to normal human subjects
is accompanied by an increase in α-MSH blood concentrations (in
addition to ACTH) and decreased TNF plasma levels, demon-
strating the role of α-MSH in the inflammatory response (59). It is
not clear how pro-inflammatory cytokines cause stimulation of
pituitary α-MSH secretion, but the ascending pathways from NTS
to the hypothalamus underlying the HPA axis activation may
play a role.
Female sex hormones and estrogens, in particular, also exert
immunomodulatory and anti-inflammatory effects. Because their
synthesis and blood concentrations are under control of
hypothalamo-pituitary hormones, estrogen-affected immune
functions can be imputed to the brain. Female sex hormones play
immunoregulatory roles during pregnancy and in diseases like
rheumatoid arthritis and osteoporosis (60). The classic mecha-
nism of steroid hormone action (as described above for glucocor-
ticoids) may contribute to estrogen immunomodulating activity.
Estrogens are neuroprotectors (61); they prevent cartilage degra-
dation during inflammation associated with increased production
of IL-1 (62); and they inhibit the production of pro-inflammatory
cytokines at different stages of their synthesis (63,64).
CHOLINERGIC ANTI-INFLAMMATORY PATHWAY
The sympathetic and parasympathetic parts of the autonomic
nervous system rarely operate alone; autonomic responses repre-
sent the interplay of both parts. A link between the para-
sympathetic part of the autonomic nervous system and immuno-
regulatory processes was suggested 30 years ago, when alleviation
of T-lymphocyte cytotoxicity by muscarinic cholinergic stimulation
was noted (65). Despite this observation, the role of the parasym-
pathetic/vagal efferents in immunomodulation is not completely
We recently demonstrated the existence of a parasympathetic
pathway of modulation of systemic and local inflammatory
responses (16,69), which focuses attention on neural immuno-
modulatory mechanisms via the vagus nerve.
Evidence for Parasympathetic (Vagus Nerve) Control
of Systemic And Local Inflammation
Acetylcholine is an important neurotransmitter and neuromodu-
lator in the brain. It mediates neural transmission in the ganglion
synapses of both sympathetic and parasympathetic neurons, and
is the principle neurotransmitter in the postganglionic parasym-
pathetic/vagal efferent neurons. Acetylcholine acts through
2 types of receptors: muscarinic (metabotropic) (70) and nicotinic
(ionotropic) (71). In addition to the brain and “wire-innervated”
peripheral structures, the RNA for these receptor subtypes (mus-
carinic) and subunits (nicotinic) has been detected on mixed pop-
ulations of lymphocytes and other immune and non-immune
cytokine-producing cells (72–77). Most of these cells can also pro-
duce acetylcholine (78).
We recently discovered that the α7 subunit of the nicotinic
acetylcholine receptor is expressed on macrophages (16). Acetyl-
choline significantly and concentration-dependently decreases
TNF production by endotoxin-stimulated human macrophage
cultures via a post-transcriptional mechanism. Using specific
muscarinic and nicotinic agonists and antagonists, we demon-
strated the importance of an α-bungarotoxin-sensitive nicotinic
receptor in the inhibition of TNF synthesis in vitro by acetyl-
choline. Acetylcholine also is effective in suppressing other endo-
toxin-inducible pro-inflammatory cytokines, such as IL-1β, IL-6,
and IL-18, by a post-transcriptional mechanism; release of the
anti-inflammatory cytokine IL-10 from endotoxin-stimulated
macrophages is not affected by acetylcholine (16).
Because of the immunosuppressive effects of acetylcholine
in vitro, we studied the possible immunonodulatory role of the
parasympathetic division of the autonomic nervous system in
vivo. In a rat model, vagotomy without electrical stimulation
significantly increases serum and liver TNF levels in response to
intravenously administered endotoxin (Figure 2A and 2B), sug-
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gesting a direct role of efferent vagus neurons in the regulation
of TNF production in vivo. Augmentation of efferent vagus
nerve by direct electrical stimulation significantly attenuates
endotoxin-induced serum and hepatic TNF (see Figure 2A and
2B). TNF amplifies inflammation by activating the release of
pro-inflammatory mediators such as IL-1, HMGB1, nitric oxide,
and reactive oxygen species (3,6). TNF also plays an essential
role in endotoxin-induced shock by inhibiting cardiac output,
activating microvascular thrombosis, and modulating capillary
leakage syndrome (4,79). These activities of TNF are consistent
with the finding that attenuation of serum TNF via cervical
vagus nerve stimulation prevents hypotension and shock in ani-
mals exposed to lethal doses of endotoxin (see Figure 2C) (16).
Animals subjected to vagotomy without vagus nerve stimula-
tion develop profound shock more quickly than sham-operated
animals (see Figure 2C), demonstrating a role for vagus nerve
efferent signaling in maintaining immunological homeostasis.
Importantly, the immunomodulatory effects of the efferent
vagus nerve also play a role in localized peripheral inflamma-
tion, because electrical stimulation of the distal vagus nerve also
inhibits the local inflammatory response in a standard rodent
model of carrageenan-induced paw edema (69). Pretreatment
with acetylcholine, muscarine, or nicotine localized within the
site of inflammation also prevents the development of hind paw
swelling (69). Vagal efferents are distributed throughout the
reticuloendothelial system and other peripheral organs, and the
brain-derived motor output through vagus efferent neurons is
rapid. The cholinergic anti-inflammatory pathway is therefore
uniquely positioned to modulate inflammation in real time.
Pharmacological Activation of the Cholinergic
It may be possible to activate the cholinergic anti-inflammatory
pathway with centrally acting pharmacological agents, because
the tetravalent guanylhydrazone CNI-1493 induces vagus nerve
firing (69) and confers anti-inflammatory effects through activa-
tion of the cholinergic anti-inflammatory pathway in both local
and systemic models of inflammation (69,80). Extensive research
on the anti-inflammatory activities of CNI-1493 reveals that it
confers protection in experimental models of cancer, pancreatitis,
rheumatoid arthritis, endotoxin shock, and sepsis (81–83). These
studies led to current testing of CNI-1493 in Phase II clinical trials
for Crohn’s disease. CNI-1493 was originally developed as an
inhibitor of macrophage activation that prevented phosphoryla-
tion of p38 mitogen-activated protein kinase (84–86). In models of
cerebral ischemia, CNI-1493 delivered via the intracerebroventric-
ular (ICV) route suppresses cerebral TNF synthesis and reduces
infarct volume (87). Surprisingly, CNI-1493 administered via the
same route suppresses systemic TNF production in response to
peripheral endotoxin challenge. Further studies revealed that
CNI-1493 stimulates efferent vagus nerve firing (69), suggesting
that the cholinergic anti-inflammatory pathway may mediate the
peripheral anti-inflammatory activity of this compound.
Intravenous (IV) pretreatment of endotoxemic rats with CNI-
1493 leads to a significant and dose-dependent inhibition of
serum TNF-release and prevents the development of endotoxin-
induced hypotension (80). The lowest IV dose tested in these
experiments (100 µg/kg body weight) failed to cause down-reg-
ulation of TNF release or prevent hypotension. When 1/100 of
this ineffective IV dose was injected ICV, it significantly attenu-
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Figure 2. Vagus nerve stimulation attenuates the endotoxin-induced
serum TNF response, hepatic TNF response, and development of endo-
toxemic shock. Rats were subjected to sham surgery (A and B: Sham, ?;
C: ?; n = 7), bilateral cervical vagotomy (A and B: VGX,
or vagotomy and electrical stimulation (A and B: VGX + STIM,
n = 7). A: Serum TNF response. B: Hepatic TNF response. C: Development
of endotoxemic shock. Mean arterial blood pressure data are normal-
ized to MABP at time = 0. Sham-surgery, vagotomy, and electrical stim-
ulation with vagotomy did not significantly affect MABP in vehicle-treat-
ed controls (not shown). Data are means +/- SEM. *P < 0.05; **P < 0.005
compared with SHAM + endotoxin; #P < 0.05 compared with VGX +
endotoxin. Taken from Borovikova and others (16) with Nature’s copy-
right permission <http: //www.nature.com>.
, C: ?; n = 7),
ated serum TNF release and protected against hypotension.
Much lower ICV drug doses (in the range 10 to 0.1 ng/kg) also
inhibited TNF release and protected against the development of
endotoxin-induced shock, suggesting that brain-dependent
mechanisms contribute to the systemic anti-inflammatory action
of CNI-1493. Similarly, CNI-1493 delivered into the cerebral ven-
tricles at doses significantly lower than peripherally active doses
also suppressed local inflammation in a standardized model of
carrageenan-induced paw edema (69).
The anti-inflammatory effects of CNI-1493 require the vagus
nerve, regardless of the route (ICV or IV) of drug administration,
because bilateral cervical vagotomy eliminates its effects. These
results indicate that CNI-1493 mediates protection against local
and systemic inflammation and shock via the cholinergic anti-
inflammatory pathway. Activation of the cholinergic anti-inflam-
matory pathway by centrally-acting compounds such as CNI-
1493, therefore, may hold significant clinical potential for
treatment of sepsis and other cytokine-mediated diseases.
It is plausible to consider two potential mechanisms through
which CNI-1493 activates vagus nerve efferent signaling: this
compound may either gain access to the DMN and activate
vagus efferent signals directly, or it may activate other neurons
within the DVC or higher brain structures and indirectly acti-
vate the efferent vagus nerve. A variety of receptors have been
identified within the DVC, the primary termination site of affer-
ent vagus fibers. Within the DMN, for example, studies have
revealed the presence of muscarinic receptors, but they have not
been associated with vagal efferent neurons (88,89). Muscarinic
(especially the M2subtype) and nicotinic binding sites have
been detected on the caudal and medial parts of the NTS (90).
The cholinergic system in the NTS has been identified on the
basis of the presence of choline acetyltransferase, acetylcholine-
esterase, and acetylcholine (88,91). The NTS cholinergic system
participates in the regulation of cardiovascular output and mod-
ulation of the baroreceptor reflex, which is centrally mediated
by glutamate. The presence of glutamate receptors is also well
documented within the DVC (92–94). Vagal afferents (cardiovas-
cular and abdominal visceral) terminating in the NTS are pre-
dominantly glutamatergic; they synapse on NTS neurons
through both NMDA and non-NMDA receptors (94,95). Expo-
sure to either IL-1 or endotoxin can activate the vagal gluta-
matergic system in the NTS. Both types of glutamate receptors
transmit AP neuron excitatory inputs to the NTS (96). NMDA
glutamate receptors also are present on AP and DMN neurons.
Vagus preganglionic motor neurons located in the DMN extend
dendritic fields to the NTS. Nicotinic (especially the α7 subtype),
neuropeptide Y, GABA (A and B), neurokine-1, and neurokine-3
receptors have each been localized to vagus efferent neurons in
the DMN (97–100). Ionotropic P2X purinoceptors of ATP (acting
as a neurotransmitter) are present on the DMN and AP neurons
(101). It is possible that some of these receptor mechanisms rep-
resent critical components of the pathways leading to pharma-
cological activation of the cholinergic anti-inflammatory path-
way. CNI-1493 may also activate other receptors in higher brain
regions, and intermediate neurons may relay CNI-1493 signal-
ing to the efferent vagus nerve.
Nicotinic Acetylcholine Receptor α7 Subunit Is an
Essential Component of the Cholinergic
Inhibition of peripheral TNF synthesis by the efferent vagus
nerve implicates a mechanism for signaling from the vagus nerve
to TNF-producing cells, such as macrophages. In a series of exper-
iments using specific cholinergic agonists and antagonists, as well
as antisense and gene knockout approaches, we have shown that
the α7 subunit of the nicotinic acetylcholine receptor is expressed
on macrophages and mediates the anti-inflammatory activity of
the efferent vagus nerve.
Nicotinic acetylcholine receptors are a family of ligand-gated,
pentameric ion channels. The main function of this receptor fam-
ily is to transmit acetylcholine signals at neuromuscular junctions
and in the central and peripheral nervous systems (71,102–104). In
humans, 16 different nicotinic acetylcholine receptor subunits
(α1–7, α9–10, β1–4, δ, ε, γ) have been identified (71,104). These
subunits have the potential to form a large number of homo- and
heteropentameric receptors with distinct properties and func-
tions. Among the 16 subunits, only the α1, α7, and α9 subunits
bind an antagonist derived from snake venom, α-bungarotoxin;
the α-bungarotoxin-binding activity of the α10 subunit is
Macrophages specifically bind FITC-labeled α-bungarotoxin,
an antagonist of a subset of nicotinic receptors. This surface bind-
ing can be competed with nicotine, suggesting that macrophages
express functional α1, α7, and/or α9 subunits of the nicotinic
acetylcholine receptor. RT-PCR analyses revealed mRNA expres-
sion of the α1, α7, and α10 subunits, but not α9. Further charac-
terization by western blotting analyses and pull-down methods
showed that fully differentiated primary human macrophages
specifically express the α7 subunit (105).
The functional relevance of the macrophage nicotinic receptor
α7 subunit in the cholinergic anti-inflammatory pathway was
tested using antisense oligonucleotides to the α7 subunit. Inhibi-
tion of α7 subunit expression restores the endotoxin-stimulated
TNF responses in the presence of nicotine, whereas antisense
oligonucleotides to the α1 and α10 subunits, under similar condi-
tions, fail to significantly change TNF release in the presence of
nicotine (105). The essential role of the α7 subunit in mediating
the activity of the cholinergic anti-inflammatory pathway in vivo
has been defined in α7 subunit-deficient mice. These gene knock-
out mice develop normally and show no gross anatomical defects
(106); however, these animals are more sensitive to inflammatory
stimuli, because α7 subunit-deficient mice release significantly
more TNF, IL-1, and IL-6 into the serum during endotoxemia as
compared with wild type mice (105). Electrical stimulation of the
vagus nerve inhibits serum and tissue levels of pro-inflammatory
cytokines in endotoxemic wild type mice, but is ineffective in α7-
deficient mice (Figure 3) (105). Peritoneal macrophages isolated
from mice lacking the α7 subunit do not respond to acetylcholine
and nicotine, and continue to produce TNF in the presence of
these cholinergic agonists. α7 Nicotinic receptor subunits have
been found in the superior cervical and pancreatic ganglia, but no
functional role for this subunit in ganglionic transmission has
been demonstrated in vivo. α3 Nicotinic receptor subunits medi-
1 3 0
| M O L E C U L A R M E D I C I N E | M A Y – A U G U S T 2 0 0 3V O L U M E 9 , N U M B E R 5 – 8
ate fast synaptic transmission in the autonomic ganglia. These
observations suggest that the higher sensitivity of α7 subunit-
deficient mice to inflammatory stimuli cannot be attributed to
impaired parasympathetic and/or sympathetic ganglionic trans-
mission. Taken together these data indicate that the nicotinic
acetylcholine receptor α7 subunit is a necessary component of the
cholinergic anti-inflammatory pathway. Moreover, the cholinergic
anti-inflammatory pathway represents a highly specific function
of the efferent vagus nerve, because it can signal through nicotinic
α7 receptors on macrophages, rather than “classical” muscarinic
Integration of the Cholinergic Anti-inflammatory
Pathway in Brain-Derived Immunomodulation
Involvement of vagus efferent neurons in neuroimmunomodula-
tion is supported by the protective role of the cholinergic anti-
inflammatory pathway in systemic and local inflammation. This
previously unrecognized neural-immune circuit sheds new insight
on brain regulation of immune function (19), and several aspects of
this “missing link” in neuroimmunomodulation are under investi-
gation in our laboratory.
The role of the efferent vagus nerve in minute-to-minute mod-
ulation of immune activation remains uncertain. Bilateral cervical
vagotomy renders animals more sensitive to endotoxemic shock,
and results in increased serum and organ TNF levels (see Figure 2A
and 2B), suggesting that efferent vagus activity exerts a tonic effect
on peripheral inflammatory responses. Electrical stimulation
appears to potentiate the tonic immunosuppression conferred by
the vagus nerve. These findings might be related to the intriguing
question of whether the parasympathetic part of the autonomic
nervous system is depressed during sepsis. In the systemic inflam-
matory response syndrome, the initial sympathocardiac activation
decreases during the progression to sepsis (107,108). Spectral analy-
sis of heart rate variability, or the length of time between heartbeats,
can provide important information about the status of the auto-
nomic nervous system. Analysis of instantaneous heart rate meas-
urements identifies low frequency and high frequency oscillations,
which are indicative of sympathetic and parasympathetic tone,
respectively. In comparison to survivors of critical illness, nonsur-
vivors have significantly less heart rate variability, reduced sympa-
thovagal balance, and reduced parasympathetic tone; reduced
sympathovagal balance is associated with an increased risk for
death (107,108). Interestingly, low sympathovagal balance (as indi-
cated by a low frequency/high frequency heart rate oscillation
ratio of <1) may be an early marker for sepsis in the critically ill,
since patients who went on to develop sepsis were admitted to the
intensive care unit with a lower low frequency/high frequency
ratio than those patients who remained sepsis-free (107). In total,
these findings suggest that autonomic dysfunction is an important
aspect of lethal critical illness and sepsis. The precise influence of
the parasympathetic division of the autonomic nervous system,
particularly in terms of afferent or efferent signaling, is unclear
from these studies; however, the sensitivity of animals devoid of
the nicotinic acetylcholine receptor α7 subunit to endotoxin chal-
lenge (105) implies a critical role of the cholinergic anti-inflamma-
tory pathway in regulation of peripheral inflammation.
How is the cholinergic anti-inflammatory pathway activated in
response to inflammatory stimuli? It appears that the general path-
ways of signaling the brain for triggering the HPAaxis and the SNS
described above may play a role. Proinflammatory cytokines
released upon immune challenge can activate vagal afferent signal-
ing and subsequent direct or indirect (through NTS neurons) acti-
vation of vagal efferents in the DMN. Thus, the sensory vagal affer-
ents, together with the regulatory vagus efferents, form an
inflammatory reflex that continually monitors and modulates the
inflammatory status in the periphery (18). The cholinergic anti-
inflammatory pathway also can be activated by cytokine signals
via AP. Thus, in addition to the cholinergic efferent fibers, the
cholinergic anti-inflammatory pathway may be comprised of at
least two brainstem medullar structures (such as NTS and DMN)
that may be signaled by pro-inflammatory cytokines through either
neural (vagus afferents) or humoral (AP) mechanisms. Intravenous
administration of IL-1β induces activation of the efferent vagus
fibers, innervating the thymus (67). Indeed, administration of
endotoxin induces neuronal activation in the DMN as well as the
NTS and AP (28). Absence of this neuronal surveillance results in
higher levels of inflammation as seen in vagotomized endotoxemic
mice (16). This centrally integrated vagal anti-inflammatory reflex
is similar for example to the vago-vagal reflex mechanism control-
ling the gastrointestinal tract. These observations suggest that the
cholinergic anti-inflammatory pathway is activated during the
The NTS may integrate the cholinergic anti-inflammatory
pathway with other central immunomodulatory responses,
because the NTS can transmit afferent vagus nerve signals to two
areas of the brain involved in neuroimmunomodulation. For exam-
ple, bidirectional neuronal circuits between the NTS and the hypo-
thalamic PVN can activate the HPAaxis, leading to glucocorticoid-
mediated immunosuppression. NTS neurons also project to
V O L U M E 9 , N U M B E R 5 – 8M O L E C U L A R M E D I C I N E | M A Y – A U G U S T 2 0 0 3 |
1 3 1
Figure 3. Vagus nerve stimulation does not inhibit TNF in nicotinic acetyl-
choline receptor α7 subunit–deficient mice. α7 Subunit–deficient
mice (–/–) or age- and sex-matched wild-type littermates (+/+) were
subjected to either sham operation (Sham) or vagus nerve stimulation
(VNS, left vagus, 1 volt, 2 ms, 1 Hz); blood was collected 2 h after endo-
toxin administration. Serum TNF levels were determined by ELISA. Sham
α7+/+, n = 10; VNS α7+/+, Sham α7–/–, VNS α7–/–, n = 11. *P < 0.05 com-
pared with Sham α7+/+. Taken from Wang and others (105) with Nature’s
copyright permission: <http://www.nature.com>.
brainstem nuclei, such as the RVM and the LC, which may activate
the SNS and modulate immune responses (see Figure 1). The key
structures in neuroimunomodulation are (1) the NTS, which is
associated with the reception and further transmission of the
cytokine signal to the cholinergic anti-inflammatory pathway, the
SNS, and the HPAaxis; (2) the hypothalamic PVN, which is respon-
sible for conversion of the neural signal into a hormonal; and (3) the
adrenal glands, which release epinephrine from the chromaffin
cells under activation of the SNS and complete the main neurohor-
monal route via release of glucocorticoids.
Both divisions of the autonomic nervous system are activated
by immunogenic stimuli and both contribute to modulation of
inflammation. The SNS down-regulates inflammation via
β-adrenoceptors. In some cases, however, norepinephrine may
increase TNF-α release, most likely through α-adrenoceptors (55).
Activation of the cholinergic anti-inflammatory pathway may
counteract instances of excessive TNF release. The cholinergic anti-
inflammatory pathway and the SNS also act synergistically to con-
trol inflammation. While the SNS can cause immunosupression
through β-adrenoceptors, vagus nerve downregulation of cytokine
production is mediated, at least in part, through nicotinic acetyl-
choline receptors containing the α7 subunit. In contrast to
endocrine-mediated mechanisms, neural regulation of immune
responses is rapid and more precisely localized, and thus may be
an important early response to peripheral inflammation.
Sympathetic and vagus innervation of the thymus, liver, heart,
lungs, gastrointestinal tract, pancreas, and kidneys may provide the
anatomical basis for coregulation of tissue macrophages, dendritic
cells, mast cells, Kupffer cells (in the liver), and other immune and
nonimmune cytokine-producing cells in the tissues. The liver is an
important organ in the acute phase of the inflammatory response,
supplying the necessary components for the host defense at the site
of inflammation and coordinating the activation of acute phase
plasma proteins (2). Hepatic sinusoidal macrophages (Kupffer cells)
are thought to be the major source of cytokines in endotoxemia
(109); therefore, stimulation of efferent vagus neurons may modu-
late the hepatic inflammatory response. The heart is well-innervated
by the 2 divisions of the autonomic nervous system. The autonomic
dysfunction (high sympathetic, low parasympathetic tone) that
occurs after myocardial infarction is a powerful predictor of early
mortality (110). In the heart, resident macrophages and cardiac
myocytes are the main sources of TNF, and TNF receptors have
been found in cardiac myocytes (111,112). TNF released from both
myocardial macrophages and cardiac myocytes contribute to
myocardial dysfunction and cardiomyocyte death in sepsis, chronic
heart failure, ischemia-reperfusion injury, viral myocarditis, and car-
diac allograft rejection (111). The effectiveness of vagus nerve stim-
ulation in inhibiting cardiac TNF (80) warrants further exploration
of the immunomodulatory role of the cholinergic anti-inflammatory
pathway in the heart and other organs.
Therapeutic Implications of the Cholinergic
Many current approaches for treatment of unrestrained inflam-
mation are based on direct suppression of pro-inflammatory
cytokines or cytokine activity. The identification of the cholinergic
anti-inflammatory pathway now suggests several new
approaches to modulate cytokines and inflammatory responses to
therapeutic advantage. For example, stimulation of the vagus
nerve may represent a novel approach to inhibit TNF production
and protect against pathological inflammation. Permanently
implanted vagus nerve stimulators are clinically approved
devices for treatment of epilepsy and depression (113–115). Vagus
nerve stimulation prevents seizures by stimulating sensory vagal
afferents associated with limbic and cortical function. Although
vagal efferents also may be activated as a result of vagus nerve
stimulation, no cardiac, gastric, or pulmonary complications have
been observed (116). In animals, vagus nerve stimulation causes
neural activation (assessed by c-Fos technique) in NTS, LC, DMN,
and hypothalamic nuclei, including PVN (117,118). Vagus nerve
stimulation enhances the activity of key components of the brain-
derived anti-inflammatory response. In addition to the vagal
efferents, the HPA axis and the SNS may also be activated as a
“side effect” of vagus nerve stimulation. The occurrence of such
activation remains to be evaluated by testing the fluctuations in
serum glucocorticoid levels in patients with vagus nerve stimula-
tors. If so, it may be possible to use currently existing, approved
devices to control inflammatory responses.
The discovery of the cholinergic anti-inflammatory pathway
identifies at least 1 receptor type that may be pharmacologically
targeted to modulate cytokine activity. The nicotinic acetylcholine
receptor α7 subunit is essential for regulation of peripheral
inflammatory responses; activation of this receptor via vagus
nerve stimulation (105) or cholinergic agonists (119,120) sup-
presses cytokine release and protects against lethal murine endo-
toxemia and sepsis. CNI-1493, the tetravalent guanylhydrazone
that was instrumental in the discovery of the cholinergic anti-
inflammatory pathway, exerts its anti-inflammatory effects in
vivo by a central mechanism involving activation of the vagus
nerve (69,80). It is possible that other experimental and clinically
approved therapeutics function through the unanticipated mech-
anism of activating neural pathways. For example, low doses of
centrally administered α-MSH or salicylates elicit specific periph-
eral anti-inflammatory responses. Likewise, the cardiac anti-
arrhythmic drug amiodarone, as well as aspirin, indomethacin,
and ibuprofen substantially increase vagus nerve activity
(reviewed in 18). The precise identification of the brain receptor(s)
that mediate these effects will facilitate the development of effec-
tive, specific receptor agonists to pharmacologically activate the
cholinergic anti-inflammatory pathway.
It also is interesting to reconsider alternative therapeutic
approaches in light of the cholinergic anti-inflammatory pathway.
For instance, hypnosis, meditation, prayer, biofeedback, acupunc-
ture, and even Pavlovian conditioning of immunological
responses are believed to involve central mechanisms that modu-
late experimental systemic or peripheral inflammatory responses.
Moreover, autonomic dysfunction occurs not only in association
with lethal critical illness and sepsis but also is considered a com-
plication of diabetes, rheumatoid arthritis, and other autoimmune
diseases (reviewed in 18). Whether physiological augmentation of
vagus nerve activity through any of these methods can modulate
disease activity remains an open question.
1 3 2
| M O L E C U L A R M E D I C I N E | M A Y – A U G U S T 2 0 0 3 V O L U M E 9 , N U M B E R 5 – 8
We have described the cholinergic anti-inflammatory pathway, a
previously unknown neural circuit that provides a new physio-
logical mechanism for immunomodulation. This specific function
of the motor branch of the vagus nerve provides new insight into
CNS control of peripheral inflammatory responses. Better under-
standing of the receptor mechanisms and neuronal circuits
involved in the cholinergic anti-inflammatory pathway, and eval-
uation of its stimulation for treatment of inflammatory disorders,
may improve the lives of patients with diseases characterized by
Grant support came from the Dept. of Defense (DARPA) and NIH
Address correspondence and reprint inquiries to Kevin J Tracey,
Head, Laboratory of Biomedical Science, North Shore LIJ-Research
Inst., 350 Community Drive, Manhasset, NY 11030. Phone: 516-562-
2813; fax 516-562-2356; e-mail: firstname.lastname@example.org.
Submitted March 7, 2003; accepted for publication June 27, 2003.
1.Sell S. (2001) Immunology, immunopathology, and immunity (6th ed). ASM
Press, Washington, D.C.
Baumann H, Gauldie J. (1994) The acute phase response. Immunol. Today
Koj A. (1997) Initiation of acute phase response and synthesis of cytokines.
Biochim. Biophys. Acta. 1317:84-94.
Tracey KJ et al. (1986) Shock and tissues injury induced by recombinant human
cachectin. Science 234:470-4.
Wang H et al. (1999) HMG-1 as a late mediator of endotoxin lethality in mice.
Wang H, Yang H, Czura CJ, Sama AE, Tracey KJ. (2001) HMGB1 as a late medi-
ator of lethal systemic inflammation. Am. J. Respir. Crit. Care Med. 164:1768-73.
Sporn MB. (1997) The importance of context in cytokine action. Kidney Int.
Kushner I. (1998) Semantics, inflammation, cytokines and common sense. Cyto-
kine Growth Factor Rev. 9:191-6.
Reichlin S. (1993) Neuroendocrine-immune interactions. New Engl. J. Med.
10. Peristein RS, Whitnall MH, Abrams JS, Mougey EH, Neta R. (1993) Synergistic roles
of interleukin-6, interleukin-1 and tumor necrosis factor in adrenocorticotropin
response to bacterial lipopolysaccharide in vivo. Endocrinology 132:946-52.
11. Gaillard RC. (1995) Neuroendocrine-immune system interactions. Trends Endo-
crinol. Metab. 7:303-9.
12. Mulla A, Buchingham JC. (1999) Regulation of the hypothalamo-pituitary-
adrenal axis by cytokines. Baillieres Best. Prac. Res. Clin. Endocriol. Metab.
13. Rivest S. (2001) How circulating cytokines trigger the neural circuits that control
the hypothalamic-pituitary-adrenal axis. Psychoneuroendocrinology 26:761-88.
14. Webster JI, Tonelli L, Sternberg EM. (2002) Neuroendocrine regulation of immu-
nity. Annu. Rev. Immunol. 20:125-63.
15. Elenkov IJ, Wilder RL, Chrousos GP, Vizi ES. (2000) The sympathetic nerve—an
integrative interface between two supersystems: the brain and the immune
system. Pharmacol. Rev. 52:595-638.
16. Borovikova LV et al. (2000) Vagus nerve stimulation attenuates the systemic
inflammatory response to endotoxin. Nature 405:458-61.
17. Tracey KJ, Czura CJ, Ivanova S. (2001) Mind over immunity. FASEB J. 15:1575-6.
18. Tracey KJ. (2002) The inflammatory reflex. Nature 420:853-9.
19. Blalock JE. (2002) Harnessing a neural-immune circuit to control inflammation
and shock. J. Exp. Med. 195:F25-8.
20. Watkins LR, Maier SF, Goehler LE. (1995) Cytokine-to-brain communication: a
review and analysis of alternative mechanisms. Life Sci. 57:1011-26.
21. Elmquist JK, Scammell TE, Saper CB. (1997) Mechanisms of CNS response to sys-
temic immune challenge: the febrile response. Trends Neurosci. 20:565-9.
22. Goehler LE, Gaykema RPA, Hansen MK, Anderson K, Maier SF, Watkins L. (2000)
Vagal immune-to-brain communication: a visceral chemosensory pathway.
Auton. Neurosci. 85:49-59.
23. Goehler LE et al. (1999) Interleukin-1 in immune cells of the abdominal vagus
nerve: a link between the immune and nervous system? J. Neurosci. 19:2799-
24. Berthhoud HR, Neuhuber WL. (2000) Functional anatomy of afferent vagal sys-
tem. Auton. Neurosci. 85:1-17.
25. Iversen S, Iversen L, Saper CB. (2000) The autonomic nervous system and the
hypothalamus. In: Principles in Neural Science. 4th ed. Kendel ER, Schwartz JH,
Jessel TM (eds.) McGraw Hill, New York. pp. 960–81.
26. Gaykema RPA, Dijkstra I, Tilders FJH. (1995) Subdiaphragmic vagotomy sup-
presses endotoxin-induced activation of hypothalamic corticotropin-releasing
hormones neurons and ACTH secretion. Endocrinology 136:4717-20.
27. Ishizuka Y et al. (1997) Effects of area postrema lesion and abdominal vagoto-
my on interleukin-1β–induced norepinephrine release in the hypothalamic par-
aventricular nucleus region in the rat. Neurosci. Lett. 223:57-60.
28. Hermann GE, Emch GS, Tovar CA, Rogers RC. (2001) C-Fos generation in the
dorsal vagal complex after systemic endotoxin is not dependent on the vagus
nerve. Am. J. Physiol. Regulatory Integrative Comp. Physiol. 280:R289-99.
29. Van Dam AM et al. (2000) Vagotomy does not inhibit high dose LPS-indiced
interleukin-1β immunoreactivity in the rat brain and pituitary gland. Neurosci.
30. Hansen MK et al. (2000) Effects of vagotomy on serum endotoxin, cytokines
and corticosterone after intraperitoneal lipopolysaccharide. Am. J. Physiol.
31. Hopkins SJ, Rothwell N. (1995) Cytokines and nervous system I: expression and
recognition. Trends Neurosci. 18:83-8.
32. Szelényi J. (2001) Cytokines and the central nervous system. Brain Res. Bull.
33. Rivest S et al. (2000) How the blood talks to the brain parenchyma and the par-
aventricular nucleus of the hypothalamus during systemic inflammatory and
infectious stimuli. Proc. Soc. Exp. Biol. Med. 223:22-38.
34. Banks WA, Kastin AJ, Broadwell RD. (1995) Passage of cytokines across the
blood-brain barrier. Neuroimmunomodulation 2:241-8.
35. Nadeau S, Rivest S. (1999) Effects of circulating tumor necrosing factor on the
neuronal activity and expression on the genes encoding the tumor necrosis
factor receptors (p55 and p75) in the rat brain: a view from the blood-brain
barrier. Neuroscience 93:1449-64.
36. Ek M, Engblom D, Saha S, Blomqvist A, Jacobsson P-J, Ericsson-Dahlstrand A.
(2001) Inflammatory response: pathway across the blood-brain barrier. Nature
37. Buller KM. (2001) Circumventricular organs: gateways to the brain. Role of cir-
cumventricular organs in pro-inflammatory cytokine-induced activation of the
hypothalamic-pituitary-adrenal axis. Clin. Experiment. Pharmacol. Physiol.
38. Afifi AK, Bergmann RA. (1998) Medulla oblongata. In: Functional neuroanatomy.
McGraw-Hill, New York, p. 117.
39. Rogers RC, McCann MJ. (1993) Intramedullary connections of the gastric
region in the solitary nucleus: a biocytin histochemical tracing study in the rat.
J. Auton Nerv. Syst. 42:119-30.
40. Rogers RC, Hermann GE, Travagli RA. (1999) Brainstem pathways responsible
for the oesophageal control of gastric motility and tone in the rat. J. Physiol.
41. Rothwell NJ, Hopkins SL. (1995) Cytokines and the nervous system II: actions and
mechanism of action. Trends Neurosci. 18:130-6.
42. Benveniste EN. (1998) Cytokine actions in the central nervous system. Cytokine
Growth Factor Rev. 9:259-75.
43. Hallenbeck JM. (2002) The many faces of tumor necrosis factor in stroke.
Nature Med. 8:1363-8.
44. Lilly MP, Gann DS. (1992) The hypothalamic-pituitary-adrenal-immune axis: a
critical assessment. Arch. Surg. 127:1463-74.
45. McEwen BS et al. (1997) The role of adrenocorticoides as modulators of
immune function in health and disease: neural, endocrine and immune inter-
actions. Brain Res. Rev. 23:79-133.
46. Karalis K, Muglia LJ, Bae D, Hildebrand H, Majzoub JA. (1997) CRH and the
immune system. J. Immunology 72:131-6.
47. Adcock IM. (2000) Molecular mechanisms of glucocorticoid action. Pulmon.
Pharmacol. Ther. 13:115-26.
48. Scheinman RI, Cogswell PC, Lofquist AK, Baldwin AS. (1995) Role of transcrip-
tional activation of I κB alpha in mediation of immunosuppression by gluco-
corticoids. Science 270:283-6.
49. McKay LI, Cidlowski JA. (1999) Molecular control of immune/inflammatory
responses: interactions between nuclear factor κB and steroid receptor–
signaling pathways. Endocrine Rev. 20:435-59.
50. Ghosh S, May MJ, Kopp EB. (1998) NF-κB and Rel proteins: evolutionary con-
served mediators of immune responses. Annu. Rev. Immunol. 16:225-60.
51. Hasko G, Szabo C. (1998) Regulation of cytokine and chemokine production
by transmitters and co-transmitters of the autonomic nervous system. Biochem.
52. Bellinger DL, Lorton CL, Felton DL. (2001) Innervation of lymphoid organs: asso-
ciation of nerves with cells of the immune system and their implications in dis-
ease. In: Psychoneuroimmunology. Ader R, Felten DI, Cohen N (eds.). 3rd ed.
Academic Press, New York, pp. 55.
V O L U M E 9 , N U M B E R 5 – 8M O L E C U L A R M E D I C I N E | M A Y – A U G U S T 2 0 0 3 |
1 3 3
IN OVERVIEW Download full-text
53. van der Poll T, Coyle SM, Barbosa K, Braxton CC, Lowry SF. (1996) Epinephrine
inhibits tumor necrosis factor-α and potentiates interleukin-10 production dur-
ing human endotoxemia. J. Clin. Invest. 97:713-9.
54. Madden KS, Sanders VM, Felten DL. (1995) Catecholamine influences and sym-
pathetic neuronal modulation of immune responsiveness. Annu. Rev. Pharmacol.
55. Zhou M, Yang S, Koo DJ, Ornan DA, Chaudry IH, Wang P. (2001) The role of
Kupffer cell α2-adrenoceptors in norepinephrine-induced TNF-α production.
Biochim. Biophys. Acta. 1537:49-57.
56. Lipton JM, Catania A. (1997) Anti-inflammatory actions of the neuroim-
munomodulator α-MSH. Immunol. Today 140:140-5.
57. Catania A, Airaghi L, Colombo G, Lipton JM. (2000) α-Melanocyte-stimulating
hormone in normal human physiology and disease states. Trends Exp. Med.
58. Macaluso A et al. (1994) Anti-inflammatory influences of α-MSH molecules:
central neurogenic and peripheral action. J. Neurosci. 14:2377-82.
59. Catania A, Suffredini AF, Lipton JM. (1995) Endotoxin causes release of α-
melanocyte-stimulating hormone in normal human subjects. Neuroimmuno-
60. Janson L, Holmdahl R. (1998) Estrogen-mediated immunosupression in autoim-
mune diseases. Inflamm. Res. 47:290-301.
61. Behl C. (2002) Oestrogen as a neuroprotective hormone. Nature Rev. Neurosci.
62. Da Silva JA, Colville-Nash P, Spector TD, Scott DL, Willoughby DA. (1993)
Inflammation-induced cartilage degradation in female rodents. Protective
role of sex hormones. Arthritis Rhem. 36:1007-15.
63. Deshpande R, Khalili H, Pergolizzi RG, Michael SD, Chang MD. (1997) Estradiol
down-regulates LPS-induced cytokine production and NF-κB activation in
murine macrophages. Am. J. Reprod. Immunol. 38:46-54.
64. Srivastava S, Weitzmann MN, Cenci S, Ross FP, Adler S, Pacifici R. (1999) Estrogen
decreases TNF gene expression by blocking JNK activity and the resulting pro-
duction of c-Jun and Jun D. J. Clin. Invest. 104:503-13.
65. Strom TB, Deisseroth A, Morganroth J, Carpenter CB, Merrill JP. (1972) Alteration
of the cytotoxic action of sensitized lymphocytes by cholinergic agents and
activators of adenylate cyclase. Proc. Natl. Acad. Sci. U.S.A. 69:2995-9.
66. Antonica A, Magni F, Mearini L, Paolocci N. (1994) Vagal control of lymphocyte
release from rat thymus. J. Auton. Nerv. Syst. 48:187-97.
67. Niijima A, Hori T, Katafuchi T, Ichijo T. (1995) The effect of interleukin-β on the effer-
ent activity of the vagus nerve to the thymus. J. Auton. Nerv. Syst. 54: 137-44.
68. Hori T, Katafuchi T, Take S, Shimizu N, Niijima A. (1995) The autonomic nervous
system as a communication channel between the brain and the immune sys-
tem. Neuroimmunomodulation 2:203-15.
69. Borovikova LV et al. (2000) Role of vagus nerve signaling in CNI-1493-mediated
suppression of acute inflammation. Auton. Neurosci. 85:141-7.
70. Caulfield P, Birdsall NJM. (1998) Intl. union of pharmacology. XVII. Classification
of muscarinic acetylcholine receptors. Pharmacol. Reviews 50:279-90.
71. Lindstrom J. (1997) Nicotinic acetylcholine receptors in health and disease.
Mol. Neurobiol. 15:193-222.
72. Hiemke C et al. (1996) Expression of alpha subunit genes of nicotinic acetyl-
choline receptors in human lymphocytes. Neurosci. Lett. 214:171-4.
73. Mita Y, Dobashi K, Suzuki K, Mori M, Nakazawa T. (1996) Induction of muscarinic
receptor subtypes in monocytic/macrophagic cells differentiated from EoL-1
cells. Eur. J. Pharmacol. 297:121-7.
74. Toyabe S et al. (1997) Identification o as thymus in mice. Immunol. 92:201-5.
75. Sato KZ et al. (1999) Diversity of mRNA expression for muscarine acetylcholine
receptor subtypes and neuronal nicotinic acetylcholine receptor subunits in
human mononuclear leukocytes and leukemic cell lines. Neurosci. Lett. 266:
76. Walch L, Brink C, Norel X. (2001) The muscarinic receptor subtypes in human
blood vessels. Therapie 56:223-6.
77. Tayebati SK, El-Assouad D, Ricci A, Amenta F. (2002) Immunochemical and
immunocytochemical characterization of cholinergic markers in human
peripheral blood lymphocytes. J. Neuroimmunol. 132:147-55.
78. Kawashima K, Fujii T. (2000) Extraneural cholinergic system in lymphocytes.
Pharmacol. Ther. 86:29-48.
79. Tracey KJ et al. (1987) Anti-cachectin/TNF-monoclonal antibodies prevent sep-
tic shock during lethal bacteraemia. Nature 330:662-4.
80. Bernik TR et al. (2002) Pharmacological stimulation of the cholinergic anti-
inflammatory pathway. J. Exp. Med. 195:1-9.
81. Villa P et al. (1997) Inhibition of multiple pro-inflammatory mediators (TNF, IL-6,
and NO) abrogate lethality in a murine mode of polymicrobial sepsis. J. Endot.
82. Martiney JA et al. (1998) Prevention and treatment of experimental autoimmune
encephalomyelitis by CNI-1493, a macrophage deactivating agent. J. Immunol.
83. Akerlund K et al. (1999) Anti-inflammatory effect of a new TNFα inhibitor (CNI-
1493) in collagen-induced arthritis in rats. J. Clin. Exp. Immunol. 115:32-41.
84. Bianchi M et al. (1996) Suppression of pro-inflammatory cytokines in monocytes
by tetravalent guanylhidrazone. J. Exp. Med. 83:927-36.
85. Wang H, Zhang M, Bianchi M, Sherry B, Sama A, Tracey KJ. (1998) Fetuin (α-2-
HS-Glycoprotein) opsonizes cationic macrophage-deactivating molecules.
Proc. Natl. Acad. Sci. U.S.A. 95:14429-34.
86. Tracey KJ. (1998) Suppression of TNF and other pro-inflammatory cytokines by
tetravalent guanylhydrazone CNI-1492. Prog. Clin. Biol. Res. 397:335-43.
87. Meistrell ME III et al. (1997) TNF is a brain-damaging cytokine in cerebral
ischemia. Shock 8:341-8.
88. Hoover DB, Hancock JC, DePorter TE. (1985) Effect of vagotomy on cholinergic
parameters in nuclei of rat medulla oblongata. Brain Res. Bull. 15:5-11.
89. Hyde TM, Gibbs M, Peroutka SJ. (1988) Distribution of muscarinic cholinergic
receptors in the dorsal vagal complex and other selected nuclei in the human
medulla. Brain Res. 447:287-92.
90. Lawrence AJ, Jarrot B. (1996) Neurochemical modulation of cardiovascular
control in the nucleus tractus solitarius. Prog. Neurobiol. 48:21-53.
91. Shihara M, Hori N, Hirooka Y, Eshima K, Akaike N, Takeshita A. (1999) Cholinergic
system in the nucleus of the solitary tract of rats. Amer. J. Physiol.-Regulat.
Integrat. Compar. Physiol. 276:R1141-8.
92. Sykes RM, Spyer KM, Izzo PN. (1997) Demonstration of glutamate immunoreac-
tivity in vagal sensory afferents in the nucleus tractus solitarius of the rat. Brain
93. Mascarucci P, Perego C, Terrazzino S, De Simoni MG. (1998) Glutamate release
in the nucleus tractus solitarius induced by peripheral lipopolysaccharide and
interleukin-1β. Neuroscience 86:1285-90.
94. Hornby P. (2001) Receptors and transmission in the brain-gut axis. II. Excitatory
amino acid receptors in the brain-gut axis. Am. J. Physiol.-Gastrointest. Liver
95. Lin LH, Talman WT. (2000) N-methyl-D-aspartate receptors on neurons that syn-
thesize nitric oxide in rat nucleus tractus solitarii. Neuroscience 100:581-8.
96. Chen C-Y, Bonham AC. (1998) Non-NMDA and NMDA receptors transmit area
postrema input to aortic baroreceptor neurons in NTS. Am. J. Physiol.-Heart
Circulat. Physiol. 275:H1695-706.
97. Ferreira M et al. (2001) Evidence for functional alpha-7 neuronal nicotinic
receptor subtype located on motoneurones of the dorsal motor nucleus of the
vagus. J. Pharmacol. Exper. Therap. 296:260-9.
98. Browning KN, Travagli RA. (2001) Mechanism of action of baclofen in rat dorsal
motor nucleus of the vagus. Am. J. Physiol.-Gastrointest. Liver Physiol. 280:
99. Lewis MW, Travagli RA. (2001) Effects of substance P on identified neurons of the
rat dorsal motor nucleus of the vagus. Am. J. Physiol.-Gastrointestin. Liver
100. Blondeau C, Clerc N, Baude A. (2002) Neurokinin-1 and neurokinin-3 receptors
are expressed in vagal efferent neurons that innervate different parts of the
gastro-intestinal tract. Neuroscience 110:339-49.
101. Atkinson L, Batten TF, Deuchars J. (2000) P2X(2) receptor immunoreactivity in the
dorsal vagal complex and area postrema of the rat. Neuroscience 99:683-96.
102. Steinlein O. (1998) New functions for nicotine acetylcholine receptors? Behav.
Brain Res. 95:31-5.
103. Marubio LM, Changeux J-P. (2000) Nicotinic acetylcholine receptor knockout
mice as animal models for studying receptor function. Eur. J. Pharmacol. 393:
104. Leonard S, Bertrand D. (2001) Neuronal nicotinic receptors: from structure to
function. Nicotine Tob. Res. 3:203-23.
105. Wang H, Yu M, Ochani M, et al. (2003) Nicotinic acetylcholine receptor α7 sub-
unit is an essential regulator of inflammation. Nature 421:384-8.
106. Orr-Urtreger A et al. (1997) Mice deficient in the α7 neuronal nicotinic acetyl-
choline receptor lack α-bungarotoxin binding sites and hippocampal fast nico-
tinic currents. J. Neurosci. 17:9165-71.
107. Korach M et al. (2001) Cardiac variability in critically ill adults: Influence of sep-
sis. Crit. Care Med. 29:1380-5.
108. Winchell RJ, Hoyt DB. (1996) Spectral analysis of heart rate variability in the ICU:
a measure of autonomic function. J. Surg. Res. 63:11-6.
109. Chensue SW, Terebuh PD, Remick DG, Scales WE, Kunkel SL. (1991) In vivo bio-
logical and immunohistochemical analysis of interleukin-1 α, β and tumor
necrosis factor during experimental endotoxemia. Am. J. Pathol. 138:395-402.
110. Honzikova N, Semrad B, Fiser B, Labrova R. (2000) Baroreflex sensitivity deter-
mined by spectral method and heart rate variability, and two-years mortality
in patients after myocardial infarction. Physiol. Res. 49:643-50.
111. Meldrum DR. (1998) Tumor necrosis factor in the heart. Am. J. Physiol. 274:R577-95.
112. Torre-Amione G, Kapadia S, Lee J, Bies RD, Lebovitz R, Mann DL. (1995)
Expression and functional significance of tumor necrosis factor receptors in
human myocardium. Circulation 92:1487-93.
113. George MS et al. (2000) Vagus nerve stimulation: a new tool for brain research
and therapy. Biol. Psychiatry 47:287-95.
114. Valencia I, Holder DL, Helmers SL, Madsen JR, Riviello JJ. (2001) Vagus nerve
stimulation in pediatric epilepsy: a review. Pediatr. Neurol. 25:368-76.
115. George MS et al. (2000) Vagus nerve stimulation. A potential therapy for resist-
ant depression? Psychiatr. Clin. North Am. 23:757-83.
116. Handforth A et al. (1998) Vagus nerve stimulation therapy for partial-onset
seizures: A randomized active-control trial. Neurology 51:48-55.
117. Krahl SE, Clark KB, Smith DC, Browning RA. (1998) Locus coeroleus lesions sup-
press the seizure attenuating effects of vagus nerve stimulation. Epilepsia
118. Naritoku DK, Terry WJ, Helfert RH. (1995) Regional induction of Fos immunore-
activity in the brain by anticonvulsant stimulation of the vagus nerve. Epilepsy
119. Wang H et al. (2002) Nicotine inhibits the release of HMGB1 through post-tran-
scriptional mechanism. Shock 17(S1):62.
120. Han JL et al. (2003) Cholinergic suppression of cytokine release from human
macrophages. Shock 19(S1):22.
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