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Endocrine Reviews 23(1):1–15
Copyright © 2002 by The Endocrine Society
Orexins in the Brain-Gut Axis
ANNETTE L. KIRCHGESSNER
Department of Physiology and Pharmacology, State University of New York Downstate Medical Center, Brooklyn,
New York 11203-2098
Orexins (hypocretins) are a novel pair of neuropeptides implicated in the regulation of energy balances and arousal.
Previous reports have indicated that orexins are produced
only in the lateral hypothalamic area, although orexincontaining nerve fibers were observed throughout the neuroaxis. Recent evidence shows that orexins and functional
orexin receptors are found in the periphery. Vagal and spinal
primary afferent neurons, enteric neurons, and endocrine
cells in both the gut and pancreas display orexin- and orexin
receptor-like immunoreactivity. Orexins excite secretomotor
neurons in the guinea pig gut and modulate gastric and in-
I.
II.
III.
IV.
V.
VI.
VII.
VIII.
IX.
testinal motility and secretion. In addition, orexins modulate
hormone release from pancreatic endocrine cells. Moreover,
fasting up-regulates the phosphorylated form of cAMP response element binding protein in orexin-immunoreactive enteric neurons, indicating a functional response to food status
in these cells. The purpose of this article is to summarize
evidence for the existence of a brain-gut network of orexincontaining cells that appears to play a role in the acute regulation of energy homeostasis. (Endocrine Reviews 23: 1–15,
2002)
Introduction
Discovery of the Orexins
Characterization of the Orexin Receptors
The Orexin System in the Brain
A. Hypothalamus
B. Dorsal vagal complex
C. Distribution of orexin receptors
Orexins and the Regulation of Feeding Behavior
A. Orexins increase food intake
B. Orexin neurons respond to metabolic signals
C. Orexins and cephalic phase reflexes
The Orexin System in the Gut
A. Characteristics of the enteric nervous system
B. Distribution of orexins and orexin receptors
C. Orexin-containing endocrine cells
D. Orexins and motility
The Orexin System in the Pancreas
Other Homeostatic Systems Affected by the Orexins
Conclusions and Future Directions
8 –10), or direct injection of orexins into the LHA (11), has
been shown to increase food intake in rodents in a dosedependent manner. Conversely, an orexin receptor antibody
inhibits food intake in fasted rats, indicating that endogenous
orexins are necessary for feeding (12). Furthermore, orexin
mRNA expression is up-regulated by fasting (1, 13), suggesting that these neurons become activated under conditions of hunger.
Orexin neurons project within the hypothalamus and
throughout the central nervous system (CNS) to nuclei
known to be important in the control of feeding (3, 4, 14, 15).
In addition, abundant orexin nerve fibers and orexin receptors are found in nuclei concerned with maintenance of
wakefulness (14, 15), and several reports implicate a dysfunction of the orexin system in human and canine narcolepsy (16 –18). Recently, genetic ablation of orexin neurons in
mice was shown to result in narcolepsy, hypophagia, and
obesity (19). This finding confirms the importance of hypothalamic orexin-containing neurons in the regulation of
sleep/wake states and further suggests a role in energy
metabolism.
Although the hypothalamus has received considerable attention regarding energy homeostasis, the gut also participates in the regulation of food intake. The presence of food
in the bowel, through activation of chemo- and mechanosensitive endings, stimulates the release of several regulatory
peptides that control gut motility and secretion (20, 21). Several of these peptides [for example, cholecystokinin (CCK)]
also act as feedback “satiety” signals responsible for termination of a meal (21–23). Satiety signals are inhibited during
fasting, and replaced by “hunger”-related signals, also from
the gut, that may be enhanced by low plasma concentrations
of glucose (21).
The enteric nervous system (ENS), which is composed of
neurons that reside within the wall of the gastrointestinal
tract and contains as many neurons as the entire spinal cord
(24), directly senses, integrates, and regulates the machinery
I. Introduction
T
HE OREXINS (1), also called hypocretins (2), were first
described just 4 yr ago; yet more than 280 papers on the
peptides have already been published. Interest in the orexins
began with the observation that these novel neuropeptides
are produced by a small group of neurons in the perifornical
and lateral hypothalamic area [LHA (1– 4)], a region classically implicated in the control of mammalian feeding behavior (5–7). Intracerebroventricular administration (1,
Abbreviations: [Ca2⫹]i, Intracellular calcium concentration; CCK,
cholecystokinin; CNS, central nervous system; DMV, dorsal motor nucleus of the vagus; EC, enterochromaffin cell; ENS, enteric nervous
system; 5-HT, 5-hydroxytryptamine; LHA, lateral hypothalamic area;
MCH, melanin concentrating hormone; NPY, neuropeptide Y; NTS,
nucleus of the solitary tract; PVN, paraventricular nucleus; VIP, vasoactive intestinal peptide; VMH, ventromedial hypothalamic nucleus.
1
2
Endocrine Reviews, February 2002, 23(1):1–15
of the gut involved in energy metabolism (24, 25). It is the
only region of the peripheral nervous system that is intrinsically capable of mediating gut-related reflex activity (25).
These reflexes, which may be secretory or peristaltic, are
made possible by the presence within the bowel of microcircuits that contain the necessary primary afferent neurons
and interneurons, as well as the excitatory and inhibitory
motor neurons that innervate gastrointestinal smooth muscle
and glands.
Recently, neurons and endocrine cells in the gut were
reported to display orexin-like immunoreactivity (26). In addition, orexins were shown to modulate the electrical properties and synaptic inputs of secretomotor neurons and stimulate colonic motility (26). We also provided evidence that
orexins were increased during periods of fasting, indicating
a functional response to food status in these cells (26). With
these observations, orexins were added to the growing list
of peptides that coexist and act in both the CNS and ENS,
giving further support to the concept of a peptidergic
brain-gut axis involved in the regulation of feeding and
energy homeostasis.
The discovery that orexins are produced in the gut is not
surprising. Many different classes of neurotransmitter have
been found in the ENS, including glutamate (27), the major
excitatory neurotransmitter of the brain (28). The gut also
contains, in neurons and/or epithelial endocrine cells, most
of the orexigenic and anorectic neuropeptides found in the
hypothalamus, including neuropeptide Y [NPY (29, 30)],
ghrelin (31), galanin (32, 33), and cocaine- and amphetamineregulated transcript (34, 35). Moreover, like the hypothalamus, the ENS contains neurons that are sensitive to glucose
(36) and are modulated by the adipocyte-derived hormone
leptin (36). Although some of these factors may lack physiological relevance in the regulation of food intake, it is likely
that many will prove to act as peripheral mediators of energy
homeostasis.
During the past 4 yr, several reviews on the orexins and
the central regulation of feeding and arousal have been published (14, 15, 37–39). The purpose of the present review is
to give an overview of the orexins in the brain-gut axis and
clarify the role of these neuropeptides in enteric and pancreatic function.
II. Discovery of the Orexins
The orexins were discovered during a search for endogenous ligands that activate orphan G protein-coupled receptors (1). Using more than 50 cell lines, each expressing a
distinct orphan G protein-coupled receptor cDNA, Sakurai et
al. (1) tested the ability of HPLC fractions of rat brain extracts,
to increase intracellular calcium concentrations ([Ca2⫹]i). The
investigators discovered several HPLC fractions that elicited
a robust increase in [Ca2⫹]i in a human embryonic kidney
(HEK293) cell line expressing a receptor originally termed
HFGAN72. This receptor was initially identified as an expressed sequence tag from human brain. When the major
peak of bioactivity in the fractions was purified to homogeneity and sequenced, a novel peptide of 33 amino acids in
length, with an N-terminal pyroglutamyl residue and an
Kirchgessner • Orexins in Brain and Gut
amidated C terminus, was discovered. The peptide, termed
orexin-A, also contained two intrachain disulfide bonds, and
sequencing of similar extracts from bovine brain revealed
exact interspecies homology.
In addition to orexin-A, the HPLC fractions contained two
minor peaks of activity, designated B and B⬘. Peak B consisted of a 28-amino acid peptide, termed orexin-B, which
also possessed an amidated C terminus and was 46% identical in sequence to orexin-A. Peak B⬘ consisted of an Nterminally truncated orexin-B, which was termed orexin-B
(3–28). Further analysis revealed that both orexin-A and
orexin-B were derived from the same 130-amino acid precursor, rat prepro-orexin, by proteolytic processing. Human
and mouse prepro-orexin sequences were determined and
found to be 83% and 95% identical to their rat counterparts,
respectively. Radiation hybrid mapping showed that the human prepro-orexin gene maps to a locus at chromosome
17q21. Thus, the prepro-orexin gene has been proposed to be
a candidate gene for a group of neurodegenerative disorders
called “chromosome 17-linked dementia” (40).
Independently, de Lecea et al. (2), using directional tag
PCR subtraction, identified a hypothalamic-specific mRNA
encoding a precursor protein that they called prepro-hypocretin and predicted that processing of this prepro-peptide
would yield two peptides, one of 39 and another of 29 amino
acids. Because the cell bodies that expressed this gene were
thought to be located exclusively in the hypothalamus, and
because of a weak homology to the gut peptide secretin, the
peptides were named hypocretin-1 and hypocretin-2. Subsequent comparisons revealed that prepro-orexin and prepro-hypocretin were the same gene, and that hypocretin-1
and hypocretin-2 had sequences in common with orexin-A
and -B, respectively (15, 37). Thus, orexin and hypocretin are
the same molecule. Nevertheless, in this article, the term
orexins will be used to denote the orexin/hypocretin peptides, since orexin immunoreactivity and orexin mRNA
expression have been found outside the hypothalamus
(1, 26, 41).
III. Characterization of the Orexin Receptors
In their remarkable paper, Sakurai et al. (1) not only reported the structure of the orexins, but also identified the
amino acid sequences of the receptors for the two peptides.
The original HFGAN72 receptor, subsequently called OX1R,
was shown to bind orexin-A with high affinity and bind
orexin-B with 100- to 1,000-fold lower affinity. However, a
related receptor, OX2R, identified by searching database entries with the OX1R sequence, was demonstrated to have
equally high affinities for both peptides. Thus, OX2R was
concluded to be a nonselective receptor for both orexin-A
and -B peptides, while OX1R was concluded to be moderately selective for orexin-A. The binding of both ligands to
either receptor was associated with changes in intracellular
calcium concentrations. Evidence from receptor-expressing
cells suggests that OX1R is coupled exclusively to the Gq
subclass of G proteins, whereas OX2R may couple to Gi/o
and/or Gq (1, 15, 42).
Kirchgessner • Orexins in Brain and Gut
IV. The Orexin System in the Brain
Immunohistochemical and in situ hybridization studies
have shown that in the CNS, orexin-producing cells are restricted to a few nuclei in the hypothalamus, including the
perifornical nucleus, the LHA, and the dorsomedial hypothalamic nucleus (Fig. 1 and Refs. 1– 4, 43, and 44). Orexin
neurons are organized bilaterally and symmetrically and
have been observed in all species investigated so far, including bovine, guinea-pig (A. L. Kirchgessner, unpublished observations), hamster, human, monkey, mouse, rat, and frog
(Fig. 1 and Refs. 1– 4 and 43– 48). Approximately 1,100 orexincontaining cell bodies have been estimated to be present in
the rat brain, using an antibody to prepro-orexin (14). Despite
their highly restricted origin, orexin nerve fibers ramify
widely throughout the CNS, with particularly abundant projections found in the olfactory bulb, cerebral cortex, thalamus, hypothalamus, brainstem, and all levels of the spinal
cord (3, 4, 13–15, 45– 49). The widespread projections of the
orexin neurons throughout the neuroaxis suggest that activation of orexin circuits probably modulates a variety of
systems (14, 15), including those involved in the regulation
of food intake. The fact that orexins can increase the release
of either excitatory or inhibitory neurotransmitters, by acting
directly on axon terminals (2, 42), indicates that the peptides
could ultimately increase or decrease the activity of innervated brain circuits.
A. Hypothalamus
Within the hypothalamus, orexin neurons project to the
arcuate nucleus (3, 4) and specifically innervate NPYcontaining cell bodies (46). Reciprocal connections from NPY
neurons to orexin neurons in the LHA have also been identified (43, 45, 46). In addition, orexin-containing nerve fibers
terminate in close apposition to NPY-immunoreactive nerve
terminals in the paraventricular nucleus [PVN (45, 46)]. NPY
Endocrine Reviews, February 2002, 23(1):1–15
3
is a potent orexigenic peptide that is released in the PVN and
surrounding sites to stimulate feeding (50, 51). Since arcuate
NPY neurons are excited by orexins (15, 42), probably
through the activation of an OX1R Gq-coupled pathway (15),
this suggests that orexin-stimulated feeding might occur
through NPY pathways (4, 52). Both NPY Y1 and Y5 receptor
antagonists reduce orexin-stimulated feeding (53, 54). Thus,
activation of NPY-containing feeding pathways is at least
partially responsible for the effects of orexin on food intake.
B. Dorsal vagal complex
In the hindbrain, orexin-immunoreactive fibers are found
in the dorsal vagal complex, comprising the nucleus of the
solitary tract (NTS) and dorsal motor nucleus of the vagus
(DMV), and the area postrema (3, 15). The NTS relays vagally
transmitted afferent signals from the gut that are related to
feeding (21, 23). The NTS also contains glucosensitive neurons that respond with altered electrical activity to changes
in blood glucose and the presence of food in the gut (55, 56).
The DMV consists of motor neurons that control gut motility
and the secretory responses that are especially important for
digestion. The DMV is also responsible for the initiation of
cephalic-phase responses that prepare the gut for the arrival
and subsequent digestion of nutrients (57). The area postrema, which is interconnected with the dorsal vagal complex, is an important circumventricular organ through which
circulating systemic factors, such as gut peptides and glucose, can gain access to the brain (58).
Although the types of neurons in the dorsal vagal complex
that are innervated by orexin-containing nerve terminals
have not been identified, it is likely that orexins alter the
activity of vagal motor neurons and/or modulate the response of NTS neurons to gastrointestinal stimuli. It has been
demonstrated that stimulation of the LHA excites neurons in
the DMV (59) and increases the activity of vagal efferents
FIG. 1. Orexin-A-immunoreactive neurons and nerve processes in the lateral hypothalamic area of the guinea pig brain. Scale bars, 50 ␮m (A),
30 ␮m (B).
4
Endocrine Reviews, February 2002, 23(1):1–15
(60). Thus, it is reasonable to expect that modulation of vagal
activity by orexins could influence cephalic phase reflexes
and/or affect gastrointestinal motility and secretion. Furthermore, since neurons in the NTS project back to the LHA
(61), orexin neurons may be regulated by satiety signals
relayed through the NTS.
Kirchgessner • Orexins in Brain and Gut
minergic projection and serves a key role in brain reward
mechanisms, which may mediate the positive reinforcing
effect of food (73).
V. Orexins and the Regulation of Feeding Behavior
A. Orexins increase food intake
C. Distribution of orexin receptors
In parallel to the diffuse orexin-containing projections
from the LHA, in situ hybridization studies with orexin receptor riboprobes demonstrate that orexin receptors are expressed in a pattern consistent with orexin nerve fibers (62–
65). However, the expression patterns for OX1R and OX2R
are strikingly different. Within the hypothalamus, OX1R
mRNA is most abundant in the dorsomedial portion of the
ventromedial hypothalamic nucleus (VMH). Dense expression of OX1R is also found in the anterior hypothalamic area
just dorsal to the suprachiasmatic nucleus. In contrast, OX2R
mRNA is expressed in many hypothalamic nuclei including
the tuberomammillary nucleus, the LHA, the arcuate nucleus, and PVN (63, 65). The tuberomammillary nucleus is
the only source of histamine in the CNS. Since histamine is
crucial for the maintenance of wakefulness (66), OX2R in this
region has been postulated to play a role in the regulation of
sleep/wake states (65).
The expression of orexin receptors in the VMH, LHA,
arcuate nucleus, and PVN is consistent with a role of hypothalamic orexin systems in regulating food intake. The VMH
is strongly implicated in the regulation of food intake since
its destruction causes obesity (67). The PVN is the site of
action of many orexigenic agents including NPY (50, 51) and
galanin (68). The PVN is also involved in the regulation of gut
functions via its projection to the dorsal vagal complex (69).
For example, stimulation of the PVN evokes an increase in
gastric acid secretion (70) and a transient increase in motility
(71).
Surprisingly, little orexin receptor mRNA has been detected in the NTS and DMV (65), although these regions
appear to receive a moderately dense orexin innervation (3,
15). Locations with dense orexin immunoreactivity but little
receptor mRNA may reflect presynaptic innervation on axon
terminals (42), whose cell bodies are located at some distances. Recent findings support a presynaptic action of
orexin-A in the DMV (72). However, a postsynaptic action on
DMV neurons via OX1R receptors has also been demonstrated and appears to mediate the stimulatory effects of
central orexin-A on gastric motility (72).
Other brain areas that display relatively dense expression
of OX1R include the CA1 and CA2 regions of the hippocampus, raphe nuclei, and the locus coeruleus (62– 65). The locus
coeruleus and dorsal/median raphe nuclei are major centers
for the noradrenergic and serotonergic neurons, respectively.
High levels of OX1R expression in these nuclei suggest a
regulatory role of orexins on the monoaminergic systems. On
the other hand, OX2R mRNA is also present in basal forebrain structures (amygdala and bed nucleus of the stria terminalis), linked to such functions as memory storage and
attention, and the nucleus accumbens (62– 65). The nucleus
accumbens is the major recipient of the mesolimbic dopa-
Sakurai et al. (1) initially examined the effects of the orexins
on feeding behavior, because the mRNA for the precursor of
these peptides was abundantly expressed in the LHA, a
region classically implicated in the regulation of both food
intake and metabolism (5–7). In fact, Anand and Brobeck (74)
called the LHA a “feeding center,” since lesions of the LHA
caused substantial reductions in food intake and body
weight leading to starvation if the animals were not force-fed
(7). The fact that electrical stimulation of the LHA produced
vigorous feeding, leading to an increase in body weight (7),
suggested that stimulation activated an orexigenic pathway
originating within or in the vicinity of the LHA.
Sakurai et al. (1) demonstrated that intracerebroventricular
administration of orexin-A or orexin-B in rats increased food
intake in a dose-dependent manner, with orexin-A significantly more effective than orexin-B, possibly due to activation of both OX1R and OX2R subtypes (1). Based on these
findings, the orexins were named after the Greek word
orexis, which means appetite (1). Subsequently, orexin-A has
been reported to increase food intake in several species (8 –10,
15, 75), including goldfish (76), and after microinjections in
several hypothalamic nuclei, including the PVN, dorsomedial hypothalamic nucleus, LHA, and perifornical area (11,
14, 15). In addition, Yamada et al. (12) reported a profound
inhibition of natural feeding in fasted rats by central injection
of an antiorexin-A antibody. Furthermore, a selective OX1R
receptor antagonist, SB-334867-A, has been shown to inhibit
spontaneous nighttime feeding over several days, as well as
orexin-A-induced feeding, and (over the first 4 h) feeding
stimulated by an overnight fast (77, 78). Thus, endogenous
orexins and stimulation of the OX1R receptor appear to be
necessary for normal feeding.
In contrast to orexin-A, the results obtained with orexin-B
have been more variable; therefore, orexin-B has been concluded to have little, if any, effect on feeding (8, 10, 11, 79).
The lack of apparent feeding disturbances in OX2R mutant
dogs further supports this conclusion (80); however, the
moderately dense expression of OX2R in the arcuate, PVN,
and LHA (65) suggests that conclusions regarding the specific receptor(s) involved in the feeding effects of orexins
cannot be made without further study.
A comparative evaluation of the potency of orexins, administered intracerebroventricularly, with other hypothalamic orexigenic peptides has demonstrated that orexin-A is
significantly less potent in stimulating food intake than NPY
(8, 51). In addition, unlike NPY, chronic administration of
orexin-A does not induce obesity in normal rats (81, 82).
However, its duration of action is longer than that of NPY (1),
and the magnitude of the effect of orexins is similar to that
of other hypothalamic appetite-stimulating peptides, such as
melanin concentrating hormone (MCH) and galanin (8). Interestingly, MCH and galanin also do not cause obesity in
Kirchgessner • Orexins in Brain and Gut
normal rats (83, 84). However, mice with targeted deletions
of the MCH gene are hypophagic (85), as are orexin knockout
mice (19). Thus, orexins appear to be involved in the shortterm regulation of feeding, rather than the long-term maintenance of body weight.
Orexin-A appears to increase food intake by delaying behavioral satiety, i.e., the normal transition from eating
through grooming to resting (75). In addition, results with
chronic intracerebroventricular infusion of orexin-A over
several days, suggest that the peptide also disrupts the normal circadian feeding pattern in rats by increasing daytime
and decreasing nighttime food intake (52, 77, 79, 81). Metabolic effects of orexins have also been shown to be dependent on circadian phase (86). Interestingly, disruptions of
normal circadian feeding patterns are well described effects
of LHA lesions (7). In addition, a role for the LHA in the
regulation of sleep-wakefulness has been established (6, 87).
This suggests that increased arousal or prolonged wakefulness may contribute to orexin-A-stimulated food intake in
continuously infused rats.
Orexin neurons innervate and activate brain areas that
promote wakefulness, such as the aminergic locus coeruleus,
which diffusely activates the cortex, and the tuberomammillary nucleus (14, 15, 88). Application of orexin-A increases
the firing rates of aminergic neurons, in vitro (88) and suppresses rapid eye movement sleep in a dose-dependent manner (89). Furthermore, orexin knockout mice exhibit a phenotype strikingly similar to human narcolepsy (90), as do
canines with a mutation of the OX2R gene (80). People with
narcolepsy have chronic, sometimes severe, daytime sleepiness that is often accompanied by episodic intrusions of
rapid eye movement sleep and sudden episodes of muscular
paralysis or weakness known as cataplexy. These findings
leave no doubt that orexin neurons play essential roles in the
control of feeding and energy balance but also regulate wakefulness. This is of significance because, during periods of
nutritional depletion, alertness may help to ensure survival.
Clearly, in such circumstances, searching for food would be
preferable to sleeping.
B. Orexin neurons respond to metabolic signals
Orexin neurons respond to several metabolic signals that
reflect the state of energy resources. Hypothalamic preproorexin mRNA levels are increased significantly after 48 h of
fasting and by acute (6 h) insulin-induced hypoglycemia (1,
13, 91), suggesting activation of these neurons under conditions of hunger. However, no changes in expression occurred
when rats with acute or chronic insulin-induced hypoglycemia were allowed to eat (91, 92) or when they were given
glucose to maintain euglycemia (93). In addition, no increases in hypothalamic prepro-orexin mRNA levels were
seen in rats with increased appetite due to insulin-deficient
diabetes, or access to palatable foods (91, 94). Thus, orexin
neurons are not stimulated under all conditions of hunger.
Based on these findings, Cai et al. (91) concluded that low
plasma glucose levels and/or absence of food from the gut
stimulates orexin neurons and postulated that orexins are
involved in short-term feeding behavior. Furthermore, they
suggested that orexin neurons might belong to a subset of
Endocrine Reviews, February 2002, 23(1):1–15
5
LHA neurons that are stimulated by falls in serum glucose
and inhibited by vagally transmitted satiety signals that are
relayed through the NTS.
It is well known that a decline of blood glucose level can
signal the initiation of food intake (95). Circulating glucose
concentrations show a dip before the onset of most meals in
human subjects and rodents. When the glucose dip is prevented, the next meal is delayed (21). Mayer’s (96) glucostatic
theory of feeding postulates that eating occurs to maintain
glucose availability. The LHA contains “glucosensitive” neurons that are activated by hypoglycemia and suppressed by
elevated blood glucose, suggesting a role in short-term nutrient sensing. Glucosensitive neurons account for approximately 25% of LHA neurons (95, 97); therefore, at least some
of the orexin-containing neurons may be glucosensitive. Recent studies support this suggestion. Approximately 30% of
orexin-immunoreactive neurons were shown to display Foslike immunoreactivity, a marker of neuronal activation, during insulin-induced hypoglycemia (92, 98). In addition, Shiraishi et al. (99) demonstrated that orexin-A caused an
increase in spike discharge in about 67% of glucosensitive
LHA neurons that they tested. Thus, glucosensitive cells
express excitatory orexin receptors. Interestingly, orexin-A
inhibited the activity of glucoresponsive neurons found in
the VMH. Glucoresponsive neurons are excited by glucose,
and stimulation of these cells has been postulated to contribute to the cessation of eating. The opposite effects of
orexins on the activity of glucosensitive and glucoresponsive
hypothalamic neurons are consistent with the antagonistic
roles of the LHA and VMH in feeding regulation (7).
The dorsal vagal complex is another important component
of orexin feeding circuits. Therefore, it is not surprising that
neuronal activation of orexin neurons was accompanied by
the appearance of Fos immunoreactivity in neurons in the
NTS and adjacent DMV (92). The NTS relays information
from vagal afferents, including glucoreceptors in the gut and
liver. In addition, the NTS, like the LHA, contains glucosensitive cells that are stimulated by hypoglycemia (5, 55, 56).
The similar pattern of Fos activity in the NTS and LHA
during insulin-induced hypoglycemia led the authors to conclude that the signals that triggered orexin neurons might be
relayed via the NTS, which has a major projection to the LHA
(61). Thus, the NTS may be an important regulator of orexin
neurons and their responses to changes in glucose availability and prandial signals.
Orexin-containing neurons may also be sensitive to leptin.
Leptin is a protein product of the ob (obese) gene (100) that
is secreted by adipocytes in proportion to fat stores. Exogenously administered leptin reduces body weight and can
inhibit the increased feeding stimulated by several orexigenic peptides (51, 101). The arcuate nucleus is a major site
of leptin-responsive neurons and is considered an important
“satiety center” on the basis of lesioning studies (51). Leptinmediated inhibition of arcuate NPY neurons, and excitation
of neurons that coexpress the anorectic peptides, cocaineand amphetamine-regulated transcript and POMC, are believed to underlie the suppression of appetite by leptin
(15, 102).
Leptin receptor immunoreactivity and signal transducer
and activator of transcription 3, a transcription factor acti-
6 Endocrine Reviews, February 2002, 23(1):1–15
vated by leptin, are found in orexin-containing cells (103).
Beck and Richy (104) showed that chronic administration of
leptin reduces orexin-A levels in the LHA. In addition, Lopez
et al. (105) found that leptin could inhibit the increase in
orexin gene expression induced by fasting. They also found
increased OX1R mRNA expression with fasting that was
suppressed by leptin treatment. Interestingly, no change in
OX2R mRNA expression was detected in either fasted or
leptin-treated conditions. Thus, it seems likely that orexin
cells are modulated by leptin, probably via OX1R, and
changes in orexin expression are involved in the response to
fasting.
C. Orexins and cephalic phase reflexes
The anatomical and experimental data described above
clearly imply that central orexins play a role in the regulation
of feeding behavior. Although several lines of evidence suggest that the hypothalamus is the primary brain site targeted
by the orexins, the projection from the NTS to the LHA also
appears to be an important regulator of orexin neurons and
their response to changes in glucose availability and prandial
signals (92). As such, the NTS may be involved in triggering
hunger and eating in response to hypoglycemia and perhaps
in terminating feeding episodes.
Orexin fibers and receptors are also found in the DMV (3,
15, 65). This raises the possibility that the orexins may control
vagal outflow to the gastrointestinal tract and modulate activities such as gastric acid secretion and/or motility.
Orexin-A dose-dependently increases gastric acid secretion
when given centrally and with an intact vagus nerve (106).
Others have found that orexin-A potently increases gastric
motility when applied to the DMV (72). These findings suggest a role for orexins in the brain-gut axis.
In 1895, Pavlov and Schumowa-Simanowskaja (107) demonstrated that sensory stimulation induced by sham feeding
evokes gastric acid secretion. It is now well known that
gastric acid secretion occurs as part of cephalic phase reflexes, which consist of simultaneous activation of gastric
acid and pancreatic enzyme secretion, antroduodenal motility, release of pancreatic polypeptide and gastrin, and gallbladder contraction. Cephalic phase responses are provoked
by the thought, sight, smell, taste, and chewing of food that
has an appetizing effect in the subjects studied (108). These
responses are important because they prime the secretory
capability of the gut, increasing the efficiency of the subsequent gastric and intestinal phases of secretion that occur in
response to a meal. The potent stimulatory effect of orexin-A
on gastric acid secretion and motility and its dependence on
vagal cholinergic outflow to the stomach suggests that orexins may mediate cephalic phase reflexes (109). Other peptides that have been shown to evoke cephalic phase responses include TRH (109), NPY (110), and ghrelin (111).
Interestingly, like orexin, both NPY (82) and ghrelin (31,
112) stimulate feeding, and both peptides are found in the
gut. Thus, orexigenic gut peptides evoke cephalic phase
responses.
Kirchgessner • Orexins in Brain and Gut
VI. The Orexin System in the Gut
A. Characteristics of the ENS
The ENS is directly responsible for controlling the motility
and secretory activity of the bowel (24, 25, 113). It consists of
groups of nerve cell bodies (ganglia) arranged in two interconnected plexuses (Fig. 2). The myenteric ganglia form a
single plexus between the longitudinal and circular smooth
muscle layers and are mainly concerned with control of
motility. In small animals, the submucosal ganglia form a
single layer and are concerned mainly with control of transmucosal water and electrolyte transport and local blood flow.
In larger mammals, the submucosal ganglia form layers that
are identified as inner, outer, and, in some cases, intermediate plexuses (114). Some data suggest that the outer submucosal plexus (closest to the circular muscle) shares control
of the circular muscle with the myenteric plexus. The musculature, mucosal epithelium, and vasculature are the gut’s
effector systems. The ENS coordinates activity of the primary
effectors to produce different patterns (digestive, interdigestive, peristaltic) of gut behavior (113).
The innervation of the bowel differs from that of other
peripheral organs because the gut is able to manifest reflex
activity in the absence of connections with the CNS (25). This
activity is made possible by the presence within the bowel of
microcircuits that contain the necessary primary afferent
neurons and interneurons, as well as the excitatory and inhibitory motor neurons that innervate gastrointestinal
smooth muscle and glands. The ENS also contains neurons
that project out of the gut to send signals to other digestive
FIG. 2. Types of neurons in the guinea pig ileum. 1, Noncholinergic
secretomotor neuron; 2, submucosal intrinsic primary afferent neuron; 3, cholinergic interneuron; 4, cholinergic secretomotor/vasodilator neuron; 5, excitatory circular muscle motor neuron; 6, inhibitory
circular muscle motor neuron; 7, myenteric intrinsic primary afferent
neuron; 8, intestinofugal neuron; 9, descending interneuron; 10, excitatory and inhibitory longitudinal muscle motor neurons. Muc, Mucosa; SM, submucosal plexus; CM, circular muscle; MP, myenteric
plexus; LM, longitudinal muscle; bv, blood vessel.
Kirchgessner • Orexins in Brain and Gut
organs, for example to the pancreas (115, 116) and gallbladder (117). The complexity of the functions controlled by the
ENS is reflected in an equally complex organization that
resembles that of the CNS more than the remainder of the
peripheral nervous system. Many different classes of neurotransmitters have been found in the ENS, including most
of those known also to be present in the CNS. On this basis,
the ENS is sometimes referred to as the “brain-in-the-gut” or
enteric “minibrain” (113, 118).
Despite the ability of the ENS to function independently
of control by the CNS, it does not normally do so. The CNS
affects the motility and secretory activity of the bowel (118 –
120). In fact, abdominal pain, diarrhea, nausea, and altered
food intake can all be manifestations of emotional or traumatic stress.
The gut receives input from the brain in efferent vagal
fibers and descending pathways in the spinal cord that connect to sympathetic preganglionic neurons in the thoracolumbar region. Efferent vagal fibers form synapses with neurons in enteric ganglia to activate circuits, which ultimately
affect the outflow of signals in motor neurons to the effector
systems (121–123). When the effector system is the musculature, its innervation consists of both inhibitory and excitatory motor neurons that participate in reciprocal control. If
the effector systems are gastric glands or digestive glands,
the secretomotor neurons are excitatory and stimulate secretory behavior.
Extrinsic primary afferent neurons carry information
about the state of the gastrointestinal tract to the CNS to
influence feeding behavior. Vagal primary afferent neurons
have cell bodies in the nodose ganglia and axons that reach
the gut via the vagus nerves, and spinal primary afferent
neurons have cell bodies in the dorsal root ganglia. The axons
of spinal primary afferent neurons in the thoracic and lumbar
regions pass through sympathetic ganglia to reach the gut via
splanchnic and mesenteric nerves. The axons of most spinal
primary afferent neurons with cell bodies in sacral ganglia
follow the pelvic nerves to reach the colon and rectum. Direct
recordings of extrinsic primary afferent neurons have revealed that they react to several types of stimuli, including
chemical changes in the intestinal lumen, distension of the
gut wall, mechanical distortion of the mucosa, and changes
in osmolarity and temperature. Hormones may also affect
afferent nerve endings; therefore, it is still not clear whether
sensory afferents are directly activated by nutrients or secondarily by hormones that are released by nutrients. In addition, spinal afferent nerves also carry nociceptive information to the CNS.
B. Distribution of orexins and orexin receptors
As the ENS contains a number of neuropeptides that have
been reported to stimulate or suppress food intake (29 –35),
we hypothesized that orexins might be present in the gut. If
orexins were present in the ENS, then, as in other sites where
these peptides exist, the enteric ganglia would be expected
to contain neurons that displayed orexin immunoreactivity.
Orexin-like immunoreactivity was found in the ENS with
a panel of antibodies raised against different sequences in
prepro-orexin, orexin-A, and orexin-B (26). In addition, RT-
Endocrine Reviews, February 2002, 23(1):1–15
7
PCR analysis revealed prepro-orexin and orexin receptor
mRNA expression in the myenteric plexus of the rat intestine.
Orexin peptides were found in the ENS of a number of
species (guinea pig, rat, mouse, and humans), suggesting
that the orexin gene has been conserved over long periods of
mammalian evolution. Orexin-immunoreactive cell bodies
and nerve fibers were observed in all regions of the gut;
however, the densest innervation was in the duodenum into
which the stomach empties after a meal.
Interestingly, there are few reports on orexins in visceral
organs. Northern blot analysis and in situ hybridization studies have detected orexin and orexin receptor mRNA expression almost exclusively in the CNS (1, 2). Nevertheless, Sakurai et al. (1) did find orexin mRNA expression in the testes,
and Lopez et al. (124) demonstrated a strong level of OX1R
and OX2R mRNA expression in the adrenal medulla of the
rat. It appears that the expression and possible effects of
orexins in the gut have just not been considered.
Orexin-immunoreactive neurons were found in both submucosal (Fig. 3A) and myenteric ganglia (Fig. 3B). Nevertheless, orexin antibodies marked distinct populations of
enteric neurons that represent only a small subset (⬃25%) of
the neurons that make up the ENS. Two neurochemically
distinct, nonoverlapping populations of orexin neurons were
identified in submucosal ganglia in the guinea pig ileum:
noncholinergic neurons that contained vasoactive intestinal
peptide (VIP), and cholinergic neurons that costored substance P.
VIP-ergic neurons are known to terminate near secretory
epithelial cells, the function of which is to secrete chloride
into the intestinal lumen (125). Chloride secretion, with accompanying fluid accumulation, is one of the ways the intestine lubricates itself, ensuring appropriate mixing and
flow of digesta along its length. Substance P is a marker of
submucosal primary afferent neurons that have been shown
to project to the mucosa and to carry sensory information
from the gut lumen to submucosal and myenteric ganglia
(25). Intrinsic primary afferent neurons are critical for orchestrating the coordination of motility and secretion (125–
127) and are essential for the initiation of peristaltic activity
(25). Orexin-like immunoreactivity was also displayed by
putative primary afferent neurons in the myenteric plexus.
In addition, orexin neurons in both plexuses displayed leptin
receptor immunoreactivity; therefore, like their CNS counterparts, orexin cells in the gut may be able to integrate this
signal from adipose tissue, along with nutrient signals from
the mucosa.
We had previously shown that leptin receptors are found
in the ENS (27). Glucoresponsive enteric neurons, like pancreatic ␤-cells and glucoresponsive VMH neurons (128), displayed leptin receptor immunoreactivity, and leptin inhibited the activity of these cells by opening ATP-sensitive
potassium channels (27). Therefore, the gut, like the endocrine pancreas and hypothalamus, contains cells that seem
specialized in the integration of messages coming directly
from extracellular nutrients (glucose) and indirectly from
hormones (leptin) that signal the nutritional state of the
organism.
To determine whether orexin synthesis in the gut was
responsive to hunger, we examined the distribution of
8 Endocrine Reviews, February 2002, 23(1):1–15
Kirchgessner • Orexins in Brain and Gut
FIG. 3. Orexin-A immunoreactivity in the guinea pig gut and pancreas. A, Submucosal ganglion. Orexin-A immunoreactivity is displayed by
a subset of cell bodies (arrow) and varicosities (arrowhead). B, Myenteric ganglion. Orexin-A immunoreactivity is displayed by a subset of cell
bodies (arrow) and varicose nerve fibers in interganglionic connectives (arrowhead) and surrounding unlabeled cell bodies (*). C, Orexin-A
immunoreactive endocrine cells (arrow) in the gastric mucosa. D, Orexin-A immunoreactive cells (arrow) in a pancreatic islet. Orexin-Acontaining cells are present throughout the islet. Scale bars, 50 ␮m (A and B), 10 ␮m (C and D).
orexin-like immunoreactivity in the ENS after a 3-d fast (26).
Strongly immunoreactive orexin-containing cell bodies, not
seen in fed controls, were observed in submucosal ganglia of
hungry guinea pigs. As a result, significantly more orexinpositive submucosal neurons were found in preparations
from fasted animals.
Fasting also activated orexin neurons, as demonstrated by
the induction of nuclear immunoreactivity to the phosphoSer (133) form of the Ca2⫹/cAMP response element binding
protein in submucosal orexin-immunoreactive cells (26).
These findings are consistent with parallel experiments in the
LHA showing that hypothalamic orexins are stimulated by
hunger (1, 13). The effects of fasting on orexin and orexin
receptor mRNA expression in the ENS have not yet been
determined; however, plasma orexin-A concentrations are
increased during fasting in humans (129) and rats (R. Oue-
draogo and A. L. Kirchgessner, unpublished observations).
Interestingly, plasma leptin concentrations are concomitantly and significantly decreased during fasting. Thus, peripheral orexin-A and leptin concentrations inversely change
during fasting and appear to be correlated with energy metabolism in humans. The source of peripheral orexins has not
yet been determined.
As in the CNS, orexin nerve fibers are observed to be
abundant in the gut (26). Orexin fibers are varicose in appearance and encircle subsets of ganglion neurons (Fig. 3, A
and B). Orexin varicosities display synaptophysin immunoreactivity; therefore, orexin appears to be in nerve terminals
making synaptic contact. Orexin fibers are found in both
myenteric and submucosal ganglia, and in the circular muscle, where they contain VIP. VIP is a marker of inhibitory
motor neurons in the myenteric plexus and mediates relax-
Kirchgessner • Orexins in Brain and Gut
ation of the circular muscle (24, 25). The presence of orexin
in VIP-ergic nerve fibers suggests that orexin might modulate intestinal motility.
An extensive network of orexin-containing fibers is also
found in the mucosa where nutrients are absorbed from the
lumen of the gut. Orexin fibers encircle mucosal crypts
(which contain secretory epithelial cells) and extend within
the lamina propria to the tips of the villi (which contain
absorptive and endocrine cells). In the guinea pig, these
fibers costore SP; therefore, at least a subset is likely to originate from submucosal primary afferent neurons.
The mucosa is also innervated by extrinsic primary afferent neurons with cell bodies in vagal and dorsal root ganglia
(130). It is possible, therefore, that some of the orexin innervation of the mucosa originates from extrinsic sources.
Orexin fibers in the NTS (15) and the dorsal horn of the spinal
cord (49), central areas associated with sensory transmission,
support this idea. In addition, Bingham et al. (41) demonstrated recently that both orexin-A and OX1R are present in
dorsal root ganglion neurons, as well as the spinal cord, in
the mouse. Furthermore, rat nodose ganglion neurons display orexin-A and OX1R immunoreactivity, and at least a
subset of these cells innervates the stomach (131). Whether
orexin-containing neurons in dorsal root ganglia innervate
the gut will need to be examined in future studies. Nevertheless, these studies demonstrate that orexin and orexin
receptors are found in both intrinsic and extrinsic primary
afferent neurons and that sensory afferents are likely to be
activated by orexins released from peripheral (enteric?)
sources. Thus, orexins probably play a role in sensory
transmission.
In parallel to the distribution of orexin-immunoreactive
axons, orexin receptors are widely distributed in the ENS. We
examined the distribution of orexin receptor protein and
mRNA in the gut, by immunocytochemistry and RT-PCR,
respectively (26). Orexin receptor-like immunoreactivity was
found in close proximity to orexin-containing nerve fibers. In
the stomach and small intestine, OX1R was found in the
circular muscle. Since OX1R expression was detected in total
RNA isolated from strips of circular muscle, this finding
suggests that smooth muscle cells may express OX1R. OX1R
immunoreactivity was also displayed by submucosal and
myenteric neurons, many of which were contacted by orexin
varicosities. OX1R mRNA expression in enteric neurons was
confirmed by RT-PCR. In addition, OX1R immunoreactivity
was displayed by nerve fibers in ganglia, muscle, and mucosa, consistent with a presynaptic localization of the receptor and/or presence of receptor on sensory afferents.
Interestingly, OX2R immunoreactivity appeared to be localized to enteroendocrine cells in the intestinal mucosa.
Thus, orexins might be able to modulate the release and/or
effects of hormones via activation of OX2R. Support for this
idea comes from a recent study, which showed that orexin
potentiates the increase in [Ca2⫹]I in response to CCK, in the
intestinal mucosa of rats (132).
To determine whether orexin receptors were functionally
active, we examined the response of submucosal neurons in
the guinea pig ileum to orexin-A by intracellular electrophysiology (26). Orexin-A increased spike activity in these
cells, similar to the increase in neuronal excitation observed
Endocrine Reviews, February 2002, 23(1):1–15
9
in CNS neurons (42). In addition, orexin-A affected synaptic
activity recorded in these cells. Bath application of the peptide reduced the amplitude of both stimulus-evoked fast and
slow excitatory postsynaptic potentials and evoked slow inhibitory postsynaptic potentials. The fact that orexins directly increased the excitability of submucosal neurons and
modulated both excitatory and inhibitory synaptic transmission, presumably by acting directly on axon terminals, suggests that the peptides could ultimately increase or decrease
the activity of innervated enteric circuits.
C. Orexin-containing endocrine cells
In addition to enteric neurons, numerous endocrine cells
in the stomach and throughout the intestine displayed
orexin-like immunoreactivity (Fig. 3C and Ref. 26). The gut,
therefore, may well be under the influence of orexins released from local neurons as well as orexins secreted as
hormones from endocrine cells. The finding that both endocrine cells and neurons in the gut contained orexin was not
surprising. CCK is released from enteroendocrine cells in the
duodenum in response to a meal. It then stimulates the
endings of vagal afferent neurons and results in reflex inhibition of gastric emptying and satiation (20 –23). In addition,
CCK is also found in the ENS where it acts as a neuromodulator and influences intestinal motility (24).
Orexin immunoreactivity colocalized with gastrin in the
antrum of the stomach and OXR1 was found on cells in the
gastric corpus (26). Gastrin is a peptide hormone that was
discovered in 1905 as an acid secretagogue (133). Gastrin is
released into the circulation after a meal and stimulates gastric acid secretion through mobilization of histamine from
enteroendocrine-like cells. Histamine, in turn, then stimulates the parietal cell to secrete acid (134). Thus, based on the
localization of orexin and OXR1 immunoreactivity in the
stomach, it seems likely that orexin influences gastric acid
secretion.
Recently it was shown that central administration of
orexin-A stimulates gastric acid secretion in rats, and that this
effect is mediated by central OXR1 activation (106). Surprisingly, no effect on gastric acid secretion was seen during
peripheral administration of the peptide.
In the intestinal mucosa, a subset of orexin-A-immunoreactive cells displays 5-hydroxytryptamine (5-HT) immunoreactivity (26). Based on this finding, orexin-immunoreactive
endocrine cells were identified as enterochromaffin (EC)
cells. Furthermore, EC cells expressed OX2R (135); therefore,
orexins might be able to modulate 5-HT release and possibly
regulate their own release in an autocrine-like fashion. 5-HT
is released from EC cells in response to mucosal stimulation
and activates both intrinsic (20, 24, 25) and extrinsic (136)
primary afferent neurons, resulting in reflex alteration of gut
function. 5-HT has also been shown to participate in the
initiation of the peristaltic reflex (137). It is not yet known
whether orexin-A is released from EC cells; however, orexins, similar to 5-HT and several other gut hormones, can
modulate gastrointestinal motility (26, 135, 138).
10
Endocrine Reviews, February 2002, 23(1):1–15
Kirchgessner • Orexins in Brain and Gut
D. Orexins and motility
To investigate whether peripheral orexins specifically regulate motility, we examined the effects of orexin-A on the
reflex-initiated rate of propulsion of an artificial fecal pellet
in the guinea pig distal colon (26). Incubation of colonic
segments with orexin-A caused a dose-dependent increase of
the rate of propulsion. Thus, enteric orexin receptors are
functional, and activation of these receptors can modulate
motility. An excitatory action of orexin-A on muscle contraction has also been observed in human colon (138). In
contrast, iv infusions of orexin-A inhibit the migrating motor
complex in the small intestine of rats, inducing a more “fedlike” motility (135). The migrating motor complex is a specific motor pattern evoked by fasting that sweeps along the
gut. Of particular interest is the fact that iv infusion of
orexin-A produced a similar inhibitory effect as VIP on the
migrating motor complex (135, 139). Since orexin nerve fibers
in the circular muscle of the rat duodenum appear to contain
VIP (135), the inhibitory effect of orexin-A on the migrating
motor complex may be mediated by VIP or, at least, involve
the same mechanism as VIP.
Convergent information suggests that orexins modulate
motility through peripheral actions of the peptides. The fact
that in both rat and mouse, orexin-A was not detected in the
brain after iv administration, although high levels were
found in the blood, further supports this idea (41). However,
we still cannot precisely locate the site of the peptide’s action.
OX1R-like immunoreactivity is displayed not only by enteric
neurons, but also nerve fibers in the mucosa and circular
muscle; therefore, orexin-A could modulate motility by acting on receptors located at nerve synapses within the enteric
plexuses, mucosa, and/or muscle. In addition, since endocrine cells of the mucosa express OX2R, the effects of orexin
may be mediated through the EC cell, which contains possible motility-regulatory peptides.
VII. The Orexin System in the Pancreas
Endocrine cells in the pancreas also display orexin-like
immunoreactivity (Fig. 3D and Ref. 26). Double-label immunocytochemistry revealed that orexin-A is present in insulinimmunoreactive cells of guinea pigs, rats, and mice (26). As
constituents of the pancreatic ␤-cells, it is conceivable that
orexins are released along with insulin and that they might
function as hormones and/or paracrine or autocrine substances. Insulin-immunoreactive islet cells also display
OX1R-like immunoreactivity, and OX1R mRNA was detected in the rat pancreas (26). Thus, orexins could potentially
modulate insulin secretion. Recent studies support this idea.
Nowak et al. (140) examined the effects of orexins on insulin
release in rats and found that sc orexin-A stimulated insulin
secretion in a dose-dependent manner (Fig. 4). In addition,
orexin-A was also effective in stimulating insulin release in
an in vitro perfusion system (140). Nevertheless, from these
studies, it is not known whether orexin-A increases insulin
release by acting directly on the ␤-cells.
Changes in insulin secretion produced by orexin-A may
contribute to the increased food intake evoked by the peptide. Insulin will decrease circulating glucose levels and this
FIG. 4. Effects of orexin-A and orexin-B on insulin secretion in perfused rat pancreas preparation. Results are expressed as means ⫾
SEM. [Reprinted with permission from K. W. Nowak et al.: Life Sci
66:449 – 454, 2000 (140). © Elsevier Science.]
could lead to feeding. In addition, insulin secretion occurs as
part of the secretory response to cephalic stimulation (57,
141). Parallel to secretion, gastrointestinal motility is enhanced. Thus, the increase in secretion and motility produced by orexins suggests that the peptides might play a role
in the integrative response of the gut and pancreas to cephalic
stimulation.
Very recently, we have shown that orexin-A immunoreactivity is also displayed by the pancreatic ␣-cells (R.
Ouedraogo and A. L. Kirchgessner, unpublished data). Low
glucose stimulates the release of orexin-A from rat-isolated
islets. Moreover, orexin-A evokes the release of glucagon.
These findings indicate that, like neurons in the hypothalamus and gut, orexin-containing pancreatic ␣-cells are glucosensitive and become activated during periods of hypoglycemia. The stimulation of orexin release by low glucose
might potentiate the effects of glucagon on glucose production during periods of hypoglycemia.
In addition to endocrine cells, orexin-like immunoreactivity is also found in nerve fibers in pancreatic ganglia, and
pancreatic neurons display OX1R-like immunoreactivity
(26). Ganglia are found throughout the pancreas, and pancreatic neurons contain most of the neuroactive substances
found in enteric neurons (142). No neuronal cell bodies contain orexin; therefore, it is likely that the orexin nerve fibers
originate from neurons located outside the pancreas. Pancreatic ganglia receive an extensive innervation from neurons located in the stomach and duodenum (115, 142). Enteropancreatic neurons reside in myenteric ganglia, which
also contain orexin neurons; however, it is not known
whether these cells project to the pancreas.
VIII. Other Homeostatic Systems Affected by the
Orexins
The location of orexin-producing cells in the perifornical
and LHA and their extensive projections in the brain and
spinal cord suggested that the peptides might have roles in
other homeostatic processes (3, 14, 15). Intracerebroventricular administration of the orexins has been shown to affect
not only feeding but also several other systems. Orexins
increase blood pressure and heart rate (143, 144), affect the
Kirchgessner • Orexins in Brain and Gut
release of LH, GH, and PRL (145–147), increase drinking
(148) and locomotor activity (149), and maintain wakefulness
(15, 88, 89). Since orexin receptors are expressed in the adrenal medulla (124), orexins may also modulate epinephrine
release. These findings indicate that orexins play a role in the
regulation of the autonomic and neuroendocrine systems,
including stimulation of sympathetic nerves (150). Interestingly, many of these effects are also associated with changes
in food intake and gastrointestinal motility.
For example, sleep has been shown to be a major determinant of interdigestive (151, 152) and digestive (153–155)
motility. Sleep is associated with diminished intestinal motility during the night and the day (154), suggesting that
intestinal motility is regulated not only by circadian rhythm
(156). Fasting alters sleep-wake cycles (151) and evokes a
specific motor activity, the migrating motor complex, in the
gut (152). Thus, sleep-wake, motility, and feeding patterns
appear to be coordinated, and orexins may play a significant
role in coordinating these complex physiological activities.
IX. Conclusions and Future Directions
In this article, I have summarized evidence for the existence of a brain-gut network of orexin-containing cells that
appears to play a role in the acute regulation of energy
homeostasis. We know that orexins are found in hypothalamic and enteric neurons that are activated by fasting (15,
26) and increase sensory arousal, wakefulness, food intake,
gastrointestinal motility, and secretion (15, 26, 106). Hypothalamic orexin-containing neurons are sensitive to nutrient
(glucose) and hormonal (leptin) messages (91, 92, 98, 104,
105) and are likely to be the glucosensitive neurons in the
LHA that are activated when blood glucose falls (91, 99).
Orexin neurons in the gut express leptin receptors (26), and,
like the hypothalamus, the ENS contains both glucosensitive
and glucoresponsive cells (36). Thus, it is likely that enteric
orexin neurons are glucosensitive, although this interesting
possibility remains to be tested in future studies. Nevertheless, these findings are consistent with the idea that glucosensitive cells in the brain and gut that are stimulated when
extracellular glucose falls below a certain threshold might
regulate the release of orexins. Such cells may be directly
activated either through their glucose sensor or indirectly
through the release of an inhibitory substance from neighboring glucoresponsive cells (128).
Vagal primary afferent neurons are sensitive to glucose
(157), and we have found that orexin-A and orexin receptor
immunoreactivity is displayed by vagal afferent neurons
innervating the gut (131). Thus, orexins are present in the
brain-gut axis and are likely to play a role in conveying
sensory information from the gut to the brain. In support of
this idea, orexins have been noted in nerve fibers/terminals
in the NTS (3, 15); however, it has been assumed that innervation of the NTS originates entirely from orexin neurons in
the LHA. The presence of orexin immunoreactivity in nodose
ganglion cells (131) indicates that orexin innervation of the
NTS also consists of vagal afferent terminals.
Hypothalamic orexin neurons receive sensory information
from the gut, via an ascending projection from the NTS, and
Endocrine Reviews, February 2002, 23(1):1–15
11
are able to modify vagal reflexes via a descending projection
to the DMV. Signals that reach the NTS are initiated by
mechanical or chemical stimulation of the gut during food
ingestion, neural input related to energy metabolism in the
liver, and humoral signals, such as CCK, that are released
upon nutrient stimulation of endocrine cells lining the intestinal lumen. The presence of OX1R immunoreactivity
within neuronal cell bodies in the nodose ganglion (131)
suggests that orexin receptors are found on gut afferent fibers
and that orexins can modulate primary afferent information
from the gut to the brainstem. Orexin receptors are also
found on nerve fibers in the lamina propria of the intestinal
mucosa (26); therefore, orexins are likely to influence neural
sensations conveyed by intrinsic primary afferent neurons
(127).
Glucose-sensitive endocrine cells in the gut and pancreas
express orexins and orexin receptors. Recent work demonstrating that luminal glucose induces the release of 5-HT
from intestinal EC cells, and that 5-HT is responsible for
eliciting the powerful vagal nodose neuronal responses
evoked by glucose (157), suggests that EC cells might act as
glucose sensors. Most gut endocrine cells have surfaces bearing microvilli that are directly apposed to the gut lumen.
These surfaces are detectors that “taste” the luminal contents,
in response to which the cells release hormones that enter the
perfusing vasculature and/or excite afferent nerve endings
beneath the mucosal epithelium (20). Orexins and OX2R are
found in EC cells (26, 135); therefore, orexins might act as
hormones and/or be able to modulate 5-HT release and
possibly regulate their own release in an autocrine-like
fashion.
Orexins are also found in gastrin-producing cells of the
stomach (26). Gastrin cells are sensitive to glucose, since
gastrin release and gastric acid secretion are reduced during
hyperglycemia (158). Interestingly, orexins are not found in
gut cells that produce the incretin hormones, glucagon-like
peptide 1 and glucose-dependent insulinotropic peptide, or
the satiety hormone CCK. Since the incretins and CCK are
released in response to a meal, it appears that orexins are
selectively produced in cells that are responsive to acute
reductions in energy substrate availability (glucose). The
presence of orexin A in pancreatic ␣-cells and the stimulation
of orexin release by low glucose in rat isolated islets (R. Ouedraogo and A. L. Kirchgessner, unpublished observations)
further supports this hypothesis.
In spite of the recent effort that has been made to define
the role of the orexins in the periphery, not much is known
about the orexin-signaling system in the gut or pancreas. For
example, we do not know the signals that trigger orexin
release or whether the peptides act as hormones and/or
neuromodulators, and we know very little about the outcome of the effects of orexins on gut motility or secretion.
Transgenic (orexin/ataxin-3) mice, depleted of orexin neurons have been generated, and these animals convincingly
demonstrate that orexins influence feeding and energy homeostasis (19). The orexin/ataxin-3 mice eat less during the
night when most feeding occurs and become obese even on
a normal diet. If orexins are expressed in the gut, enteric
neurons should also be ablated, causing gastrointestinal disorders. No defects have yet been reported in the ENS of
12
Endocrine Reviews, February 2002, 23(1):1–15
orexin/ataxin-3 mice. The absence of these reports, however,
should not be interpreted to mean that the ENS of these
animals is normal. Their ENS has not been investigated in
any detail. The possibility also exists that the orexin promoter
fragment used in the transgenic mice is not efficient in enteric
neurons. Clearly, future experiments should determine
whether orexin/ataxin-3 and/or orexin knockout mice have
any defects in gut and/or pancreatic function, especially
during conditions of hypoglycemia.
The role of orexins in sensory transmission also needs to
be carefully examined. In addition to intrinsic and vagal
primary afferent neurons, orexins and orexin receptors have
recently been found in dorsal root ganglion cells (41). Moreover, peripheral orexins were shown to have analgesic properties (41). Thus, manipulation of spinal and/or enteric
orexinergic systems may be efficacious in inflammatory conditions of the bowel in which hyperalgesia is a significant
factor.
Acknowledgments
I would like to thank Drs. Erik Näslund and Raogo Ouedraogo for
critically reading the manuscript, and Melina and Alexandro Hassiotti
for secretarial assistance. Special thanks go to my family for their patience during the preparation of this manuscript.
Address all correspondence and requests for reprints to: Annette
L. Kirchgessner, Ph.D., Department of Physiology and Pharmacology,
State University of New York Downstate Medical Center, 450 Clarkson Avenue, Box 29, Brooklyn, New York 11203-2098. E-mail:
[email protected]
This work was supported by NIH Grant NS-27645 and The American
Diabetes Association.
Kirchgessner • Orexins in Brain and Gut
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