Peptides 26 (2005) 17711781

Review

The Agouti-related protein and its role in energy homeostasis

Adrian M. Stutz, Christopher D. Morrison, George Argyropoulos
Pennington Biomedical Research Center, 6400 Perkins Road, Baton Rouge, LA 70808, USA
Received 14 July 2004; accepted 1 December 2004
Available online 14 June 2005

Abstract
The melanocortin system plays an important role in the regulation of energy homeostasis. The Agouti-related protein (AGRP) is a natural
antagonist of the action of alpha-melanocyte stimulating hormone (-MSH) at the melanocortin receptors (MCR). AGRP is upregulated by
fasting while intracerebroventricular injections of synthetic AGRP lead to increased appetite and food intake. Transgenic mice overexpressing
AGRP are also hyperphagic and eventually become obese. AGRP is, therefore, a significant regulator of energy balance and a candidate
gene for human fatness. Indeed, humans with common single nucleotide polymorphisms (SNPs) in the promoter or the coding region are
leaner and resistant to late-onset obesity than wild-type individuals. AGRP is also expressed in the periphery. Recent studies show that AGRP
in the adrenal gland is upregulated by fasting as much as it is in the hypothalamus. These data open up the possibility for a wider role by
AGRP not only in food intake but also in the regulation of energy balance through its actions on peripheral tissues. This review summarizes
recent advances in the biochemical and physiological properties of AGRP in an effort to enhance our understanding of the role this powerful
neuropeptide plays in mammalian energy homeostasis.
2005 Elsevier Inc. All rights reserved.
Keywords: Obesity; Adrenal; Hypothalamus; Leptin; Polymorphism

Contents
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Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
AGRP gene structure and tissue expression . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Physiological properties of AGRP action . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Molecular mechanism of AGRP action . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Hormonal and substrate regulation of AGRP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
AGRP action downstream of the ARC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
AGRP action in the periphery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Genetic models of AGRP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Human genetics of AGRP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Acknowledgement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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Abbreviations: AGRP, agouti-related protein; -MSH, alpha-melanocyte stimulating hormone; MC3R & MC4R, melanocortin receptor 3 and 4; i.c.v.,
intracerebroventricular; POMC, proopiomelanocortin; PVN, paraventricular nucleus; LHA, lateral hypothalamic area; ARC, arcuate nucleus; NPY, neuropeptide
Y; SNP, single nucleotide polymorphism; aa, amino acid; RNAi, RNA interference; SHU9119, Ac-Nle-c[Asp-His-Dnal(2 )-Arg-Trp-Lys]-Nh2
Corresponding author. Tel.: +1 225 763 2530; fax: +1 225 763 3030.
E-mail address: argyrog@pbrc.edu (G. Argyropoulos).
0196-9781/$ see front matter 2005 Elsevier Inc. All rights reserved.
doi:10.1016/j.peptides.2004.12.024

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A.M. Stutz et al. / Peptides 26 (2005) 17711781

1. Introduction
In 1992, Bultman et al. [16] discovered that the ectopic
expression of the Agouti protein was the basis for the monogenic mouse obesity model known as the Ay mouse. Ay is a
dominant mutation that leads to both yellow coat color and
morbid obesity [16,81]. Agouti is normally expressed within
the hair follicle, and promotes yellow hair color by antagonizing the action of alpha-melanocyte stimulating hormone
(-MSH) on the melanocortin receptor 1 (MC1R) [6,88].
The overexpression of Agouti within the hair follicle, thus,
explains the dominant yellow coat color, and it was also
hypothesized that the ectopic expression of Agouti within
regions of the brain controlling body weight homeostasis was
likewise responsible for the obesity and hyperphagia in this
animal model [69].
These potent anabolic effects of Agouti led to the
subsequent hypothesis that an endogenous antagonist of
melanocortin receptors might be produced locally within
the hypothalamus, and that this antagonist might participate in the hypothalamic regulation of energy balance. This
hypothesis was confirmed when Shutter et al. [110] identified a hypothalamic protein with high sequence homology to
Agouti, termed Agouti-related transcript and subsequently
Agouti-related protein (AGRP). These initial observations
are the basis for a growing body of evidence suggesting that
AGRP plays a significant role in the hypothalamic regulation of energy homeostasis [30,71,125,138], and this review
describes recent advances in our understanding of AGRP
action within the hypothalamus, as well as the potential for
AGRP-dependant effects in the periphery.

2. AGRP gene structure and tissue expression

The human AGRP is a relatively short gene, spanning
1.1 kb on chromosome 16q22. It consists of four exons and
encodes a 132 amino acid (aa) protein, with the rodent
ortholog encoding a 131 aa protein [13,76,110]. The AGRP
protein contains nine cysteine residues at conserved positions that form disulfide bridges and are important for its
function [110]. Human and murine AGRP expression profiles showed a predominant presence of both orthologs in
the hypothalamus, the subthalamic nucleus, and a shorter
transcript lacking the 5 non-coding exon in the adrenal and
at lower expression levels in the testis, lung, kidney, spinal
cord, and dorsal root ganglia [7,110]. Although, at present,
it is unclear whether these transcripts arise from differential splicing or from two independently regulated promoters,
the 5 untranslated exon demonstrated significant promoter
activity in a periphery-derived cell line only (Chinese hamster ovary) [13]. This finding suggests that this exon may play
a role in the peripheral expression of AGRP, and supports
the existence of two active promoters, each responsible for
either brain- or periphery-specific (the adrenal cortex, testis,
and lung) expression [13]. AGRP has also been sequenced or

identified in the pig [45], the sheep [1], the fish Fugu (Takifugu rubripes) [63], the zebrafish (Danio rerio) [113], the
goldfish [17], the Japanese quail [10,93] and in ring doves
(Streptopelia risoria) [115,116]. In many of these species
AGRP levels are upregulated by fasting [1,43], underscoring
a conserved role for AGRP in energy homeostasis.

3. Physiological properties of AGRP action

Perhaps the best-documented aspect of AGRP action is its
profound stimulation of food intake. Transgenic mice overexpressing AGRP exhibit severe obesity, have increased body
length, hyperinsulinemia, late-onset hyperglycemia, pancreatic islet hyperplasia, and reduced corticosterone levels [38].
These effects of AGRP are proposed to be mediated predominantly by the brain, as both murine and human AGRP
orthologs stimulate hyperphagia when administered intracerebroventricularly (i.c.v.) [42,102,103,112]. AGRP is one
of the most potent orexigenic peptides known, and is unique
in that it displays long-lasting effects on food intake, with animals displaying hyperphagia even 7 days following a single
i.c.v. injection [42]. While the mechanisms underlying this
long-lasting effect are unclear, it appears to be independent
of a persistent blockade of melanocortin receptors [42,62].
In addition to alterations in feeding behavior, recent data
indicate that AGRP also acts to suppress energy expenditure,
and that this effect contributes significantly to its overall effect
on energy homeostasis. Central administration of AGRP
decreases uncoupling protein 1 (UCP1) expression and temperature within brown adipose tissue (BAT) [111,135], likely
via an inhibition of sympathetic outflow [135]. Interestingly,
administration of N-terminal parts of AGRP increased body
and fat weight in the absence of hyperphagia [37]. Furthermore, reduction of AGRP levels by 50% using RNA interference resulted in increased energy expenditure and loss of
body weight without changes in food intake [73]. Together,
these data indicate that alterations in energy expenditure play
a significant role in the regulation of energy homeostasis by
AGRP.
AGRP also influences physiological systems beyond feeding and energy expenditure. In particular, recent evidence
suggests that AGRP influences neuroendocrine function by
regulating the hypothalamic pituitary axis. The i.c.v. injection of human AGRP into ovariectomized rhesus monkeys
resulted in elevated cortisol, adrenocorticotropin hormone
(ACTH), and prolactin (PRL) release [132]. In addition, i.c.v.
injected AGRP enhanced the ability of IL-1 to increase
ACTH, likely by affecting the function of -MSH at hypothalamic melanocortin receptors. This observation supports a
modulatory role of AGRP in the neuroendocrine responses to
inflammation, which itself may possibly promote obesity and
type 2 diabetes [22]. Evidence also suggests that AGRP has an
inhibitory influence over the hypothalamicpituitarythyroid
axis, with AGRP administration suppressing hypothalamic thyroid stimulating hormone (TSH)-releasing hormone

A.M. Stutz et al. / Peptides 26 (2005) 17711781

(TRH) expression and decreasing circulating levels of thyroid

hormones [31].

4. Molecular mechanism of AGRP action

Available evidence suggests that AGRP is a melanocortin
receptor antagonist, blocking the effect of -MSH and other
agonists to stimulate these receptors [32,83,134]. However,
several studies [47,83] indicate that AGRP may go beyond a
simple antagonist and, additionally, act as an inverse agonist
at the MC4R. Inverse agonist activity would indicate effects
of AGRP on feeding behavior and energy homeostasis that
occur independent of -MSH or other melanocortin receptor
ligands.
In addition to -MSH, other Proopiomelanocortin
(POMC) derived ligands such as -MSH, desacetyl -MSH,
ACTH, and -lipotrophic hormone regulate melanocortin
receptors. Importantly, the orthosterical, antagonistic function of AGRP (aa83132) was the same for all POMC
peptides, suggesting that AGRP (aa83132) inhibits Gscoupled accumulation of intracellular cAMP at the MC4R
in a more general way [94]. Evidence also supports a role
for the cell surface heparan sulfate proteoglycan syndecan-3
in the facilitation of AGRP action at the melanocortin receptor, again influencing the balance of agonist to antagonist
[99,100].
Most studies focusing on in vivo AGRP function have used
carboxyl-terminal AGRP peptides that mimic the effect of the
full-length protein. This approach appears legitimate, as synthetic peptides of the amino-terminus, hAGRP(aa2551), and
the mid-portion, hAGRP(aa5482), of the human AGRP protein were devoid of antagonistic action for -MSH [96,97],
whereas a synthetic variant containing the 46 C-terminal
residues, hAGRP(aa87132), was active and equipotent to
the mature mouse homolog [46,47,119,133]. Moreover, the
hAGRP(aa87132) peptide was able to bind effectively
the melanocortin receptors MC3R, MC4R, and MC5R,
thus, inhibiting binding of -MSH [32,83,134]. In depth
NMR structure analysis of this AGRP domain (aa87132)
revealed an inhibitor cysteine-knot structure [51], which
makes contact with the melanocortin receptor 3 and 4 with
two loops present in this structure [77]. Recently, elongation of the minimum core decapeptide Yc[CRFFNAFC]Y
of AGRP, which acts as an antagonist at the MC4R but
not the MC3R, by two amino acids at either the C- or
the N-terminus conferred antagonistic function also on the
MC3R, resembling the profile of full-length AGRP [53]. All
these reports underscore the importance of the C-terminus
of AGRP in the antagonistic function at the melanocortin
receptors.
Despite this clear importance of the C-terminus, a recent
study suggested that N-terminal parts of AGRP, which
are unable to bind to melanocortin receptors, have effects
on energy expenditure that are, therefore, independent of
melanocortin receptors [37]. A single i.c.v. injection of AGRP

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(aa83132) increased cumulative food intake, whereas Nterminal parts AGRP (aa2551) and AGRP (aa5482) did not
affect the amount of food consumption. However, injection of
any of the three portions of AGRP resulted in decreased oxygen consumption and colonic temperature, both readouts for
energy expenditure, and the two N-terminal parts of AGRP
increased body weight and epididymal/mesenteric fat weight,
despite the absence of hyperphagia and cross-reactivity with
MC4R [37]. Additional experiments are necessary to confirm and elucidate this interesting observation. One possible
explanation for this melanocortin independent effect is a
potential interaction with co-receptors. One such example
could be syndecan-3 [99,100], which can only bind to Nterminal AGRP. A repeat of the study, e.g. in syndecan-3
knockout mice or by altering the availability of membranebound syndecan-3, together with further mutagenesis and in
vitro binding experiments would be helpful to understand the
implications of this finding.

5. Hormonal and substrate regulation of AGRP

AGRP is produced predominately by a subpopulation of
neurons within the hypothalamic arcuate nucleus (ARC).
Considering that the ARC mediates the effects of an array
of peripheral metabolic signals, this anatomical localization
suggests that AGRP also plays a role in energy homeostasis.
Indeed, AGRP is predominately localized to a population of
neurons within the medial aspect of the ARC that co-express
neuropeptide Y (NPY). NPY has long been associated with
hypothalamic regulation of feeding behavior, and by coexpressing AGRP and NPY; these so called NPY/AGRP
neurons are considered key mediators in the response to
nutrient depletion and subsequent regulation of energy
balance.
Multiple hormonal signals influence AGRP expression.
Long acting satiety signals such as leptin or insulin act to
decrease AGRP, while maintaining physiological levels of
these hormones blocks fasting induced increases of AGRP
[12,105,128]. Loss of leptin or insulin receptors within the
brain leads to markedly elevated AGRP expression, and
AGRP is also upregulated in leptin-deficient (ob/ob) obese
mice, irrespective of fasting [89,110]. Together these observations suggest that these hormones reduce appetite, in
part, by inhibiting AGRP (and NPY) expression [15,110].
Additionally, recent work indicates that these hormones can
acutely alter membrane potential and reduce neuronal firing from neurons containing NPY, suggesting that leptin and
insulin rapidly inhibit the activity of NPY/AGRP neurons
[124]. Although progress has been made in identifying factors downstream of leptin signaling on AGRP [12,105,128],
it is not clear through which mechanism leptin regulates
AGRP. Potential signaling pathways include the AMP-kinase
pathway [78,79], the JAK-STAT pathway [9,35,44,48], and
the PI3K pathway [84,85]. It remains to be seen which
pathway, if not a combination of two or more, is respon-

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A.M. Stutz et al. / Peptides 26 (2005) 17711781

sible in mediating the leptin effect on AGRP expression

and, therefore, reducing food intake and increasing energy
expenditure.
The gut-derived protein ghrelin has also been implicated
in the regulation of AGRP neurons. Unlike leptin and insulin,
ghrelin principally acts to stimulate feeding and body weight
gain [64,121] and activates NPY/AGRP neurons. Ghrelin
is an endogenous ligand for the growth hormone secretagogue receptor (GHS-R) and has been shown to up-regulate
expression of AGRP [5456,82,131]. In addition, administration of ghrelin acutely induces c-Fos (a marker of neuronal
activation) within neurons containing NPY (and presumably
AGRP). Genetic evidence also supports a critical role for
the NPY/AGRP neurons in mediating ghrelin action, since
AGRP/NPY double knockout (k.o.) mice are resistant to
ghrelin-dependant increase in food intake [19]. However,
mice lacking only AGRP or only NPY do not display this
phenotype, whereas AGRP expression was normal in ghrelin
k.o. mice [130].
In addition to leptin, insulin, and ghrelin, other endocrine
signals also influence AGRP levels within the hypothalamus.
Glucocorticoids have been implicated in the regulation of
energy homeostasis and removal of glucocorticoid signaling (for instance, by adrenalectomy) ameliorates obesity in
a number of physiological and genetic models. Adrenalectomy decreases sensitivity to both AGRP [26] and NPY
[136] while increasing the sensitivity to -MSH [26] and
leptin [26,72]. In a different study, adrenalectomy blocked
fasting induced increases in AGRP [70]. Exogenous administration of glucocorticoids, on the other hand, increases food
intake, body weight, as well as AGRP and NPY expression [52]. Another study, supportive of an essential role of
glucocorticoids on AGRP expression, demonstrated that corticosterone secretion temporally coincided with the rising
phase of diurnal AGRP expression [70]. Depletion of corticosterone by adrenalectomy abolished this AGRP diurnal
rhythm, which was restored by exogenous corticosterone
replacement, highlighting its requirement to maintain the normal diurnal AGRP expression cycle [70]. Together, these
observations suggest that glucocorticoids have significant
effects on energy homeostasis potentially mediated by action
on hypothalamic NPY/AGRP neurons. Other hormones that
could potentially regulate AGRP expression include PYY
[30], cholecystokinin (CCK) [80], gastrin [122], glucagonlike peptide 1 [120], and vasopressin [104] that are transmitted as afferent signals by the vagus nerve to the brain [11].
More elaborate studies though are required to address the role
that these hormones play on the regulation of AGRP.
Fatty acids (FAs) may also influence the expression of
AGRP. Food intake and appetite in humans were significantly
affected by intestinal infusions of intralipid and emulsions
of oils enriched with stearic, oleic, and linoleic acids [34],
while i.c.v. administration of oleic acid in animals mimics
the anorexigenic effects of leptin and downregulates NPY
expression [87]. Furthermore, feeding mice a highly saturated
fat diet, but not polyunsaturated FAs (PUFAs), downregu-

lated AGRP expression [127]. Fat-specific satiety has been

reported in humans for fat high in linoleic acid [57], while
the degree of saturation of FAs can influence post-ingestive
satiety with the PUFAs exerting a stronger control than the
monounsaturated fatty acids (MUFAs) [66]. The exact mechanism by which FAs may influence AGRP expression is not
known at this point, but we hypothesize that it could happen through the activation of critical transcription factors
[68,109]. Indeed, FAs are involved in the activation of PPARs
[23,61] and PUFAs, in particular, are linked to the activation
of SREBP [91,92] and the suppression of other transcription
factors [86]. FAs may also be involved in the activation of
leptin [98], thus, employing multiple pathways for regulating AGRP expression.
Finally, changes in glucose concentration are known
to influence AGRP expression potentially linking appetite
to metabolic need. Central administration of 2-Deoxy-dglucose (2DG), a nonmetabolizable glucose analog that
inhibits glucose utilization, increased AGRP gene expression and elicits glucoprivic food intake [33,107]. Recently,
the effect of high glucose levels to decrease AGRP expression could be correlated with decreased phosphorylation of
AMP-kinase [67]. Further studies are needed to confirm these
findings and establish a role for glucose in AGRP regulation
in vivo.
In addition to being upregulated in response to fasting,
AGRP is also increased in other physiological situations
whereby increased food intake is desirable or necessary. For
example, during pregnancy AGRP levels, but not POMC,
MC4R or NPY, were elevated in Wistar rats, suggesting that
AGRP could play a role in pregnancy-associated hyperphagia [101]. Similarly, AGRP is upregulated in lactating sheep
[114] while ring doves express elevated AGRP levels during
the post hatching stages when parents eat more food to feed
their young [115]. Some diseases that result in insufficient
food intake correlate with reduced levels of AGRP, such as a
mouse model of PraderWilli syndrome, in which neonates
display failure-to-thrive [36]. In a rat experimental model
of anorexia nervosa, central infusion of AGRP prevented
self-starvation by counteracting physical hyperactivity and
dominantly stimulating food intake [58]. AGRP treatment in
tumor-bearing animals lead to a maintenance of lean body
mass and circadian activity patterns during tumor growth,
without negatively affecting tumor size [75]. AGRP, therefore, could modulate disease-related changes in food intake
and contribute to maintenance to overall energy balance in
response to hormonal and substrate stimuli.

6. AGRP action downstream of the ARC

The neuronal sites and signaling mechanisms mediating AGRP action within regions downstream of the ARC
are currently not well characterized. Neurons within the
paraventricular nucleus (expressing TRH, corticotrophinreleasing hormone (CRH), opioidergic) and lateral hypotha-

A.M. Stutz et al. / Peptides 26 (2005) 17711781

lamus (expressing melanin-concentrating hormone (MCH)

and orexin) are strong candidates as mediators of downstream
AGRP signaling. Within the PVN, AGRP has been shown to
specifically inhibit TRH neurons [31], in opposition to both
-MSH and leptin, whereas a synthetic melanocortin receptor antagonist (SHU9119) that mimics AGRP action was able
to block leptin induced c-Fos (a marker of neuronal activation) within paraventricular nucleus (PVN) neurons [106].
This model of opposing agonist and antagonist activity is
well documented in the PVN, and is likely replicated in
other areas receiving input from AGRP and POMC neurons.
For instance, AGRP neurons also strongly project to MC4R
expressing neurons within the lateral hypothalamic area
(LHA) [29] and where AGRP increases c-Fos. Interestingly,
both -MSH and leptin are believed to inhibit neurons within
the LHA, indicating that although the agonist/antagonist balance exists in this region, the specific effects on neuronal
activity differ [30].
AGRP neurons also project to a variety of areas outside the
hypothalamus, and MC4 receptors are also expressed in many
of these areas. Central administration of AGRP induces c-Fos
within nucleus accumbens shell, lateral septum, NTS and
central amygdala [40], and the AGRP dependant activation
of these regions was more robust 24 h after AGRP injection as compared to 2 h, suggesting that these regions may
mediate the long-lasting effects of AGRP [40]. AGRP was
also shown to specifically increase c-Fos in orexin neurons
within the LHA even 24 h after i.c.v. injection, suggesting
that an activation of LHA orexin neurons could also mediate
the long-lasting hyperphagia induced by AGRP [137].
Recent work also suggests that AGRP signals in part
via an opiodergic pathway, as combined inhibition of and the -opioid receptors signaling blocked AGRP-induced
feeding [14,41]. Interestingly, one report indicates that mice
injected with AGRP favor calorically dense chow to a palatable sucrose solution, indicating that the main function of
AGRP on food behavior may be to satisfy energy needs rather
than enhance palatability or hedonic value of food [90,129].

7. AGRP action in the periphery

The site of the most abundant hAGRP, human cocaine- and
amphetamine-regulated transcript (hCART), and hPOMC
expression is the adrenal gland [39]. Within the adrenal cortex, the short isoform of the rat AGRP is upregulated during
fasting [8], paralleling the 18-fold increase that occurs within
the hypothalamus [43]. Moreover, plasma levels of AGRP
are increased in obese men [60] and fasted rats [108]. Nevertheless, the function of AGRP in the periphery remains
unresolved. The active isoform of human AGRP (aa83132)
crosses the bloodbrain barrier [59], raising the possibility for
peripherally expressed AGRP to directly access melanocortin
receptors within the brain.
Although only a few studies have focused on peripheral
AGRP action, these studies highlight the potential for cir-

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culating or locally-produced AGRP to influence peripheral

tissues. The obvious question is what is the role of AGRP
in the periphery? In all probability, AGRP in the periphery
does the same as it does in the hypothalamus, i.e. binds to the
melanocortin or perhaps other G protein-coupled receptors.
Both AGRP and Agouti have direct effects on adipocytes,
influencing the expression of fatty acid synthase and leptin
[20,21,28], and blocking -MSH dependant effects on leptin gene expression [49]. In addition to effects in adipocytes,
AGRP may also have a paracrine role in adrenal function,
with adrenal-derived AGRP being regulated by glucocorticoids and blocking the induction of corticosterone secretion
by -MSH [24,25]. Furthermore, experiments in the chicken
have revealed that AGRP binds to MC3R in the adrenal
[117,118]. The situation is less clear if a negative feedback
loop exists in which AGRP itself acts as a negative regulator of leptin action, as was first suggested by Ebihara et al.
[28]. In other experiments, however, central administration of
AGRP in rats for 3- and 7-day periods resulted in significant
increases of plasma leptin levels, even when AGRP induced
hyperphagia was prevented [65]. In a recent study, AGRP
was found to be robustly expressed in epididymal fat and to
be upregulated in this tissue by feeding rather than by fasting
[18]. Leptin was also upregulated by feeding in the epididymal fat but unexpectedly also in the adrenal gland. This was
observed in both the obesity-susceptible C57BL/6J (C57)
and the obesity-resistant CAST/Ei mouse strain. Surprisingly, AGRP expression was higher in the fasted state in the
hypothalamus and the adrenal gland in the leaner and obesityresistant CAST/Ei strain instead of in the obesity-susceptible
C57 strain. This result was recapitulated in another obesitysusceptible mouse strain, tub/tub that also had reduced AGRP
expression in the hypothalamus [4]. These two reports present
the fresh idea that gene-specific obesity (tub/tub mutation)
or generalized fatness (C57 mouse) does not correlate positively with AGRP levels. It is also possible for a lean mouse
(CAST/Ei) to express higher than the C57 AGRP levels in the
hypothalamus without becoming obese, provided that energy
is expended through a hyperactive lifestyle. Rather unexpectedly, in vitro leptin treatment of a mouse hypothalamusderived cell line upregulated endogenous AGRP and this
result was recapitulated in adrenal-derived cells, after an
acute 6-h exposure. However, a 60-h exposure to leptin
resulted in downregulation of AGRP [18]. Treatment of
hypothalamic and adrenal cells with AGRP not only upregulated its endogenous expression but also upregulated endogenous leptin in the adrenal cells [18]. These data reveal a
novel feedback loop and reciprocal transcriptional regulation between AGRP and leptin in the hypothalamus and the
periphery. An autocrine role for AGRP in the rat adrenal cortex modulating the actions of POMC has also been proposed
[8]. Therefore, peripheral expression of AGRP, and particularly in the adrenal gland, could play a significant role in
energy homeostasis. This is even more relevant to human
physiology since AGRP (unlike its murine ortholog) is not
expressed in visceral nor subcutaneous fat [18].

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A.M. Stutz et al. / Peptides 26 (2005) 17711781

Table 1
Phenotypes of AGRP mouse models
Model

Phenotype

Reference

AGRP transgenic
i.c.v. injection of AGRP (aa87132)
AGRP k.o.
AGRP/NPY double k.o.

Food intake , body weight , body length , corticosterone levels

Food intake , body weight , energy expenditure
Normal weight gain and feeding behavior, slight difference in MCH
Normal weight gain and feeding behavior, unresponsive to ghrelin
administration
50% Inhibition of AGRP expression, metabolic rate , body weight ,
normal food intake

[38]
[37,42,62,102,103,112,116,132,137]
[19,95]
[19,95]

AGRP RNAi injection into

hypothalamus

[73]

, Increase; , decrease.

8. Genetic models of AGRP

In an effort to elucidate the functional properties of AGRP,
mouse models were created to overexpress or be AGRPdeficient (Table 1). Specifically, transgenic mice overexpressing AGRP are hyperphagic, exhibit severe obesity, show
increased body length, hyperinsulinemia, late-onset hyperglycemia, pancreatic islet hyperplasia, and reduced corticosterone levels [38]. These observations suggest that the elevation of AGRP observed following fasting or in response to
hormonal signals, promotes increased feeding behavior. Surprisingly, AGRP knockout mice (AGRP/ ) exhibit normal
feeding behavior without changes in body weight and cumulative food intake, in comparison to their wild-type littermates
[95]. AGRP/ mice are fertile and exhibit no gross histological or pathological abnormalities, and also have comparable
serum levels of glucose, leptin, triglycerides, and free fatty
acids to wild-type mice. Furthermore, AGRP/ mice do not
show a higher preference for high fat or high sucrose food
over their wild-type littermates. AGRP/ , NPY/ and double AGRP/NPY k.o. mice show no feeding or body weight
deficits and maintain a normal response to starvation [95]. In
AGRP/ mice, expression levels of MC3R, MC4R or NPY
were comparable to levels found in wild-type mice. These
data suggest that the imprinted loss of AGRP could result
in the development of compensatory mechanisms. Indeed,
a marginal increase in the orexigenic MCH expression was
observed in the AGRP/ mice [95]. Another contributing
factor to the absence of a robust phenotype of these AGRP/
mice, could be the genetic background of the mice. Passage
of the mutation to another mouse strain could address this
issue.
In another study, however, RNA interference (RNAi) in the
ARC was used to decrease AGRP expression rather than completely delete the gene, which may be a more physiologically

relevant approach. Indeed, RNAi lead to a 50% inhibition of

AGRP message within the hypothalamus, which was associated with increased metabolic rate and reduced body weight
but without changing food intake [73]. This is consistent with
conclusions from pair-fed animals treated with AGRP, which
resulted in increased BAT UCP1 and increased fat mass and
leptin levels, potentially regulating overall energy expenditure [111]. Therefore, an approach to moderately decrease
AGRP expression levels may provide a means to regulate
body weight and increase energy expenditure.

9. Human genetics of AGRP

In addition to animal models that clearly implicate AGRP
in the regulation of energy balance, studies in humans show
promising associations of AGRP single nucleotide polymorphisms (SNPs) with resistance to obesity and perhaps type 2
diabetes mellitus (T2DM) in Black Africans.
AGRP plasma levels were elevated in obese men [60] and,
consistently with data from animal models, during fasting
[108]. The former finding was recently replicated in another
cohort while a 6-day fast increased AGRP plasma levels in
lean individuals but not in obese ones [50].
Mutation screening has revealed several functional SNPs
in AGRP that are associated with various phenotypes of leanness (Table 2). Two SNPs in the 5 -UTR have been reported
in Black but not White individuals. SNP 3019G > A was in
complete linkage disequilibrium (LD) with another promoter
SNP, 38C > T [5]. Despite LD, the 3019G > A SNP exhibited functional dimorphism whereby the two linked alleles
had opposite effects on promoter activity but the net effect
was likely determined by the 38C > T SNP because of its
position in a region with 1000-fold higher promoter activity [5]. The T allele of the 38C > T SNP was associated

Table 2
Described SNPs in the human AGRP gene
SNP

Function

Phenotype

Reference

3019G > A
38
G423A
C690T
Ala67Thr

Alters promoter activity

Alters promoter activity
Silent
Silent
Alters secondary structure (algorithm prediction)

In complete LD with 38C > T

Associated with obesity, T2D
None reported
None reported
Associated with anorexia nervosa, resistance to late-onset obesity

[5]
[2,76]
[123]
[123]
[3,13,27,74,126]

A.M. Stutz et al. / Peptides 26 (2005) 17711781

with leanness in Sierra Leoneans and reduced fatness in the

Blacks of the HERITAGE Family Study [2]. In addition, all
the diabetics (T2DM) recorded in the Sierra Leonean cohort
were CC homozygous (for the 38C > T SNP) suggesting that
the T allele may not only predispose to obesity-resistance in
Blacks on both sides of the Atlantic, but may also provide
a protective mechanism against the development of T2DM
in the Black Africans. Two silent mutations in the coding
regions were not associated with any obese phenotypes [123].
Another SNP in the coding region that changed the alanine
at position 67 to a threonine was not associated with obesity
[13,27] but was associated with anorexia nervosa [126]. Interestingly, this SNP was found in Whites only, in contrast to
the two promoter SNPs (3019G > A and 38C > T) that were
found in Blacks only. Subsequent studies have shown that heterozygotes for the Ala67Thr SNP were resistant to late-onset
obesity [3] and this finding was later replicated in another
cohort for the rare homozygous individuals for the threonine
allele who, again, had reduced body fatness [74]. These data
suggest that AGRP SNPs could affect metabolic phenotypes
in humans, with individuals carrying the less frequent alleles
(either the Black- or White-specific ones) potentially protected from developing obesity in obesigenic environments.

10. Summary
AGRP is a unique neuropeptide representing a rare
endogenous receptor antagonist or inverse agonist. By acting on melanocortin receptors in brain regions controlling
food intake and energy expenditure, AGRP represents a critical component in a complex regulatory system that exists to
maintain energy balance. Its expression in peripheral tissues
and in particular the adrenal gland suggests a wider role for
AGRP in energy homeostasis, potentially involving feedback
regulatory interactions with leptin. Although genetic AGRPdeficiency does not induce a dramatic phenotype in mice,
a large body of evidence suggests that an increase in AGRP
contributes to an increase in appetite and a decrease in energy
expenditure. From the human studies we learn that common
AGRP SNPs predispose individuals to obesity-resistance and
may provide a protective mechanism against fatness under
obesigenic conditions. AGRP is an attractive target gene for
modulating pathological food intake and obesity but its exact
role in energy expenditure and overall energy homeostasis
requires further investigation. Future studies to elucidate the
peripheral actions of AGRP in the context of its binding to
melanocortin or perhaps other G-protein-coupled receptors
should be intensified. A better understanding of both the
peripheral and hypothalamic interactions between AGRP and
leptin would help to identify the best step to modulate this
complex feedback network. Furthermore, the initial observations that the N-terminal part of AGRP may influence energy
homeostasis independent of food intake and melanocortin
signaling should be investigated in more detail. This could
lead to the design of selective drugs that affect certain inter-

1777

actions of AGRP without necessarily affecting signaling of

POMC-derived peptides on melanocortin receptors.

Acknowledgement
This work was supported by the National Institute of Diabetes and Digestive and Kidney Diseases (grant DK62156 to
GA).

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