MINIREVIEW
Roles of prolactin-releasing peptide and RFamide related
peptides in the control of stress and food intake
Yuki Takayanagi and Tatsushi Onaka
Division of Brain and Neurophysiology, Department of Physiology, Jichi Medical University, Japan
Introduction
RFamide peptides, defined by their carboxy-terminal
arginine (R) and amidated phenylalanine (F) residues
(hence RFamide), were originally discovered in inverte-
brates [1] and have recently been identified in verte-
brates [2]. The first reported RFamide peptides in
mammals were neuropeptide FF (NPFF) and neuro-
peptide AF, which were later confirmed to be encoded
on a single gene [3]. By applying a reverse pharmaco-
logical approach, in which orphan G protein-coupled
receptor ligands were identified by detecting signal
transduction induced in cells expressing a targeted
orphan G protein-coupled receptor after stimulation,
prolactin-releasing peptide (PrRP) was identified to be
a ligand of an orphan G protein-coupled receptor,
GPR10 (hGR3 ⁄ UHR-1 ⁄ PRLHR) and to belong to the
RFamide peptide [4]. Subsequently, by utilizing DNA
databases, another gene for RFamide peptides was
identified in mammals [5,6]. The RFamide peptides
encoded by the gene were named RFamide related
peptide (RFRP)-1 ⁄ NPSF and RFRP-3 ⁄ NPVF, which
were found to be orthologs of avian peptide LPLRFa-
mide. Thus, RFRPs are allocated into the LPXRFa-
mide peptide family (X = L or Q). Several other
RFamide peptides also have been discovered by a
reverse pharmacological method and by DNA

the control of energy metabolism and stress responses. Both food intake
and stressful stimuli activate PrRP neurons. The administration of PrRP
affects energy metabolism and neuroendocrine systems. PrRP-deficient or
PrRP receptor-deficient mice show abnormal energy metabolism and ⁄ or
stress responses. On the other hand, RFRP neurons are activated by stress-
ful stimuli and the administration of RFRP induces neuroendocrine
and behavioral stress responses. Taken together, these data suggests that
PrRP and RFRP neurons play a role in the control of energy metabolism
and/or stress responses.
Abbreviations
ACTH, adrenocorticotropic hormone; CCK, cholecystokinin-8; CRH, corticotropin-releasing hormone; NPFF, neuropeptide FF;
NTS, nucleus tractus solitarii; PrRP, prolactin-releasing peptide; QFRP, pyroglutamylated RFamide peptide; RFRP, RFamide related peptide.
4998 FEBS Journal 277 (2010) 4998–5005 ª 2010 The Authors Journal compilation ª 2010 FEBS
(previously known as metastin) and pyroglutamylated
RFamide peptide (QFRP) (Table 1). RFamide peptides
show diversity in their N-terminal sequence and, as a
result, a broad pattern of biological activities, includ-
ing the control of energy metabolism and stress, as
well as effects in the neuroendocrine and cardiovascu-
lar systems. In the present minireview, we focus on
PrRP and RFRP (Table 2) and review recent progress
in research investigating the roles of these two peptides
in the control of energy metabolism and stress.
PrRP
PrRP was considered to serve as a hypothalamic-
releasing factor and to act on the anterior pituitary
to stimulate prolactin release from the pituitary.
However, no PrRP immunoreactivity was found in the
external layer of the median eminence, from where
classic hypothalamic hormones are released into the

LPXRF RFRP-1 (NPSF)
RFRP-3 (NPVF)
GPR147 (NPFF-1, OT7T022, RFRPR) Increase in the release
of ACTH, oxytocin and
prolactin
Increase
?
?
Kisspeptin Kiss1 (metastin) GPR54 (OT7T175, AXOR12, KISS-1R) ? No effects ?
QFRP QRFP (26 Rfa, P513) GPR103 (AQ27, SP9155) ? Increase [52,53] ?
Table 2. Amino acid sequences of PrRP and RFRP in human, rats and mice are shown. *Deduced from the cDNA sequence.
Peptide Species
Number of
of amino acids Sequence Reference
PrRP Human 20
31
TPDINPAWYASRGIRPVGRF-NH
2
SRTHRHSMEIRTPDINPAWYASRGIRPVGRF-NH
2
[4]*
[4]*
Rat 20
31
TPDINPAWYTGRGIRPVGRF-NH
2
SRAHQHSMETRTPDINPAWYTGRGIRPVGRF-NH
2
[4]*
[4]*

[5]*
[5]*
RFRP-3 (NPVF) Human 8
17
VPNLPQRF-NH
2
NMEVSLVRRVPNLPQRF-NH
2
[5]*, [54]
[5]*
Rat 18 ANMEAGTMSHFPSLPQRF-NH
2
[56]
Mouse 17 or 18 (V)NMEAGTRSHFPSLPQRF-NH
2
[5]*
Y. Takayanagi and T. Onaka PrRP and RFRPs in metabolism and stress
FEBS Journal 277 (2010) 4998–5005 ª 2010 The Authors Journal compilation ª 2010 FEBS 4999
caudal A2 and A1 noradrenergic neurons. Immunohis-
tochemical studies have shown that PrRP-immunoreac-
tive fibers are widely distributed in the brain [8]. The
main receptor for PrRP, GPR10, is also widely
expressed in the brain (especially in the reticular
nucleus of the thalamus, bed nucleus of the stria termi-
nalis, preoptic areas, hypothalamic paraventricular
nucleus, periventricular nucleus, dorsomedial hypothal-
amus, NTS and area postrema) [9]. In addition to
GPR10, PrRP has a high affinity for NPFF receptor 2
(also known as GPR74 ⁄ NPGPR ⁄ HLWAR77). NPFF
receptor 2 is expressed in the dorsal horn of the spinal

pin-releasing hormone (CRH) or oxytocin may relay
PrRP signaling to reduce food intake. Anatomical
studies have shown that oxytocin neurons in the hypo-
thalamus receive direct projections from PrRP neurons
in the medulla oblongata. The administration of PrRP
activates neurons expressing CRH or oxytocin in the
hypothalamus, both of which are anorexic peptides.
PrRP-induced anorexia is attenuated by a CRH recep-
tor antagonist or oxytocin receptor antagonist [16].
Furthermore, an oxytocin receptor antagonist reduces
the anorexic actions of CCK [17,18], and increases
meal size [19]. Oxytocin receptor-deficient mice show
an increased meal size [20]. Taken together, these
results suggest that the PrRP-oxytocin system plays a
pivotal role in relaying the satiety signaling of CCK.
PrRP neurons in the brainstem and hypothalamus
express leptin receptors [21]. Leptin regulates long-term
energy metabolism. Leptin induces the expression of
phosphorylated signal transducer and activator of
transcription protein 3 in PrRP neurons, especially in
the dorsomedial hypothalamus [13] (Fig. 1) and the
anorectic effects of leptin are impaired in PrRP-defi-
cient mice [13]. These data suggest that the anorectic
effects of leptin signaling are mediated, at least in part,
by PrRP.
Role of PrRP in the control of energy metabolism:
energy expenditure
PrRP has also been associated with energy expendi-
ture. An intracerebroventricular injection of PrRP
Fig. 1. Activation of PrRP and RFRP neurons in the dorsomedial

deficient mice. It is interesting to note that PrRP neu-
rons in the brainstem express estrogen receptors and
that PrRP expression in the brainstem is higher in
female than in male rats [24]. Thus, the function of
PrRP-GPR10 system in the control of energy con-
sumption might differ between sexes.
PrRP has been suggested to be involved in energy
consumption under stressful conditions. Stressful stim-
uli increase oxygen consumption. This increase in oxy-
gen consumption after stressful stimuli is lower in
PrRP-deficient mice [25]. Stressful stimuli activate
PrRP neurons, and thus it is possible that PrRP
increases energy consumption under the conditions in
which PrRP neurons are activated.
Roles of PrRP in the control of stress responses
PrRP neurons in the medulla oblongata and ⁄ or in the
dorsomedial hypothalamus are activated by a variety
of stressful stimuli [26], including restraint of body
movement, conditioned fear [27] (Fig. 1), footshocks,
hemorrhage [28], exercise [29] and inflammatory stress
(e.g. lipopolysaccharide injection) [30]. PrRP neurons
have been suggested to be involved in neuroendocrine
responses to stress. PrRP neurons project directly to
CRH neurons and oxytocin neurons in the hypothala-
mus [31]. An intracerebroventricular injection of PrRP
activates CRH neurons and oxytocin neurons in the
hypothalamus, and facilitates adrenocorticotropic hor-
mone (ACTH) and oxytocin release into the systemic
circulation. Blockade of endogenous PrRP signaling by
the administration of PrRP-neutralizing antibodies

family are RFRP-1 and RFRP-3. RFRP-1 and RFRP-
3 are derived from a single precursor protein. Immu-
nohistochemical studies have shown that single cells
contain both RFRP-1 and RFRP-3. RFRP-1 and
RFRP-3 bind with high affinity to a G protein-coupled
receptor, GPR147 (also known as OT7T022, NPFF
receptor 1 or RFRP receptor).
In birds, peptides of the LPXRFamide family are
termed gonadotropin inhibitory hormones. RFRPs
have also been reported to serve a similar function in
mammals [34]. The administration of RFRP-3 sup-
presses plasma luteinizing hormone or follicle-stimulat-
ing hormone concentrations in mammals. However,
the mechanisms are not fully understood. RFRP-3
inhibits the activity of gonadotropin-releasing hormone
neurons in the hypothalamus [35–37]. RFRP-3 has
been also reported to act on pituitary gonadotrophs to
inhibit luteinizing hormone or follicle-stimulating hor-
mone release [38–40], although it is currently a matter
of controversy as to whether RFRP-3 exerts a hypo-
physiotropic role in mammals [37,41]. RFRP mRNA
expression is not affected either by estrogen [42] or tes-
tosterone [43], whereas RFRP neurons have been
reported to express estrogen receptors [35]. The precise
physiological roles of RFRP in the mammalian repro-
ductive system need further investigation.
Y. Takayanagi and T. Onaka PrRP and RFRPs in metabolism and stress
FEBS Journal 277 (2010) 4998–5005 ª 2010 The Authors Journal compilation ª 2010 FEBS 5001
Localization of RFRPs and their receptors
The RFRP gene is expressed in the caudal hypothala-

responses has also been reported. The dorsomedial
hypothalamus where RFRP neurons exist plays an
important role in the control of stress responses as well
as food intake [45]. Subsequent to stressful stimuli, the
percentage of RFRP neurons expressing Fos protein
[48] (Fig. 1) and the expression of RFRP mRNA in
the hypothalamus are increased [49]. RFRP fibers are
observed in the hypothalamic paraventricular nucleus
and appear to project directly to cells containing CRH
or oxytocin [46] in the hypothalamus. The administra-
tion of RFRP increases Fos expression in the hypotha-
lamic paraventricular nucleus and in hypothalamic
oxytocin neurons, and facilitates the release of ACTH
and oxytocin into the peripheral circulation. Similar
patterns of Fos expression and hormone release are
observed after stressful stimuli. Furthermore, the cen-
tral application of RFRP-1 or RFRP-3 induces anxi-
ety-related behavior [48]. Taken together, these data
are consistent with the view that stressful stimuli acti-
vate RFRP neurons and that RFRP-1 and RFRP-3
are involved in neuroendocrine and behavioral
responses to stressful stimuli.
RFRP neurons express glucocorticoid receptors and
the administration of glucocorticoid increases the
expression of RFRP mRNA in the hypothalamus [49].
Fig. 2. Possible neural pathways controlling stress and food intake by PrRP and RFRP. The PrRP and RFRP systems influence energy
homeostasis and stress responses. The dorsomedial hypothalamus has direct connections to and from the limbic areas, other hypothalamic
nuclei and the brainstem, which are involved in stress and energy balance. AMY, amygdala; ARC, arcuate nucleus; BST, bed nucleus of the
stria terminalis; DMH, dorsomedial hypothalamus; LHA, lateral hypothalamic area; NTS, nucleus tractus solitarii (A2 noradrenergic region);
PBN, parabrachial nucleus; POA, preoptic area; PVN, paraventricular nucleus; SCN, suprachiasmatic nucleus; VLM, ventrolateral medulla (A1

Other neuropeptides, including GALP [57–59] and
relaxin-3 [60], also may play a role in these interac-
tions. The detailed mechanisms underlying the interac-
tions between stress and energy metabolism remain to
be clarified.
Acknowledgements
The present work was supported by KAKENHI
(20590237, 20020023, 20790194, 22120512, 22659050)
from MEXT and JSPS.
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