Ghrelin
(1-5), Trp3, Arg5 (GSWFR) A Potent GHS Receptor Agonist
[Trp(3), Arg(5)]-ghrelin(1-5) stimulates
growth hormone secretion and food intake via growth hormone
secretagogue (GHS) receptor
Ghrelin, a
28 amino acid peptide identified as an endogenous ligand for
growth hormone secretagogue (GHS) receptor, stimulates food
intake and growth hormone (GH) secretion. We designed low
molecular weight peptides with affinity for the GHS receptor
based on the primary structure of ghrelin. We found that [Trp(3),
Arg(5)]-ghrelin(1-5) (GSWFR), a novel pentapeptide composed
of all l-amino acids, had affinity for the GHS receptor (IC(50)=10muM).
GSWFR stimulated GH secretion after intravenous or oral administration.
Centrally administered GSWFR increased food intake in non-fasted
mice. The orexigenic action of GSWFR was inhibited by a GHS
receptor antagonist, [d-Lys(3)]-GH-releasing peptide-6, suggesting
that GSWFR stimulated food intake through the GHS receptor.
The orexigenic action of GSWFR was also inhibited by a neuropeptide
Y (NPY) Y(1) receptor antagonist, BIBO3304. These results
suggest that the GSWFR-induced feeding is mediated by the
NPY Y(1) receptor.
Ohinata K, Kobayashi
K, Yoshikawa M. Peptides. 2006 Mar 10; [Epub ahead of print]
031-25
[D-Lys3]-GHRP-6's antagonist activity in Ghrelin induced growth-hormone
secretion in prepubertal rats
Ghrelin is a 28-amino-acid peptide, with an essential n-octanoyl
modification at Ser3, that elicits growth-hormone (GH) secretion
in rats and humans. At present, the mechanisms of ghrelin action
and its interactions with other systems controlling GH secretion
remain poorly characterized. In this context, the present study
was undertaken to obtain information about ontogeny and possible
gender differences in the GH-releasing activity of ghrelin,
and to delineate its primary site(s) of action at the hypothalamus
and/or pituitary. In addition, the interactions between ghrelin
and other relevant signals in the control of GH secretion, such
as excitatory amino acids (EAAs), nitric oxide (NO) and serotonin,
were assessed. Experiments were carried out in infantile-prepubertal
animals, when GH pulsatility is not yet established. Systemic
administration of ghrelin (25 nmol/rat, i.p.) to 5-, 10- and
23-day-old male and female rats increased plasma GH levels from
day 10 onwards. This action was NO dependent, since it disappeared
in 23-day-old males after pretreatment with an inhibitor of
NO synthase (NAME). Similarly, central infusion of ghrelin (3
nmol/rat, i.c.v.) elicited GH responses in 10- and 23-day-old
animals significantly higher than after systemic administration.
By contrast, in vitro challenge of pituitary tissue with increasing
doses of ghrelin (10(-9)-10(-7) M) failed to enhance GH release
into the incubation medium, whereas stimulation with GH-releasing
hormone (GHRH; 10(-7) M) or GHRP-6 (10(-7) M) was effective.
Finally, effects of ghrelin were blocked by pretreatment with
MK-801 and NBQX antagonists of EAA ionotropic receptors and
after manipulation of endogenous serotoninergic tone. In addition,
the potent releasing activity of EAA agonists NMDA and AMPA
was blunted by pretreatment with D-Lys3-GHRP-6, a selective
antagonist of the cognate ghrelin receptor, i.e. the GH-secretagogue
receptor. In conclusion, our results demonstrate that GH-releasing
activity of ghrelin appears early in the infantile period, is
NO dependent and involves a primary hypothalamic site of action.
The data also demonstrate for the first time the existence of
a cross-talk between ghrelin and other neurotransmitter systems,
such as EAAs and serotonin, in precise control of GH secretion.
Pinilla L, et al.
Neuroendocrinology 2003 Feb;77(2):83-90
Antagonism of ghrelin
receptor reduces food intake and body weight gain in mice
BACKGROUND AND AIMS: Ghrelin, an endogenous ligand
for growth hormone secretagogue receptor (GHS-R), is an appetite
stimulatory signal from the stomach with structural resemblance
to motilin. We examined the effects of the gastric peptide ghrelin
and GHS-R antagonists on energy balance and glycaemic control
in mice. MATERIALS AND METHODS: Body weight, fat mass, glucose,
insulin, and gene expression of leptin, adiponectin, and resistin
in white adipose tissue (WAT) were measured after repeated administrations
of ghrelin under a high fat diet. Gastric ghrelin gene expression
was assessed by northern blot analysis. Energy intake and gastric
emptying were measured after administration of GHS-R antagonists.
Repeated administration of GHS-R antagonist was continued for
six days in ob/ob obese mice. RESULTS: Ghrelin induced remarkable
adiposity and worsened glycaemic control under a high fat diet.
Pair feeding inhibited this effect. Ghrelin elevated leptin
mRNA expression and reduced resistin mRNA expression. Gastric
ghrelin mRNA expression during fasting was increased by a high
fat diet. GHS-R antagonists decreased energy intake in lean
mice, in mice with diet induced obesity, and in ob/ob obese
mice; it also reduced the rate of gastric emptying. Repeated
administration of GHS-R antagonist decreased body weight gain
and improved glycaemic control in ob/ob obese mice. CONCLUSIONS:
Ghrelin appears to be closely related to excess weight gain,
adiposity, and insulin resistance, particularly under a high
fat diet and in the dynamic stage. Gastric peptide ghrelin and
GHS-R may be promising therapeutic targets not only for anorexia-cachexia
but also for obesity and type 2 diabetes, which are becoming
increasingly prevalent worldwide.
Asakawa A, et al. Gut.
2003 Jul;52(7):947-52
Effects of ghrelin administered
intraperitoneally (3 nmol/mouse every 12
hours for five days) on calorie intake,
epididymal fat mass, gastrocnemius muscle,
and blood glucose, insulin, cholesterol,
triglycerides, and free fatty acid concentrations
in lean mice under a high fat diet
LF, saline
HF, saline
HF, ghrelin
Calorie intake (kcal/day)
18.83 (1.055)
23.22 (1.329)
25.94 (2.562)*
Fat pad mass(g)
0.533 (0.049)
0.797 (0.095)
1.202 (0.175)**
Skeletal musde (g)
0.337 (0.016)
0.353 (0.010)
0.340 (0.005)
Glucose (mmol/l)
7.899 (0.476)
8.393 (0.825)
8.909 (0.498)
Insulin (pmol/l)
135.0 (20.10)
177.5 (13.05)
378.0 (141.8)*
Cholesterol (mmol/l)
3.742 (0.337)
5.568 (0.497)
5.813 (0.509)**
Triglycerides (mmol/l)
0.347 (0.032)
0.305 (0.039)
0.371 (0.090)
Free fatty acids (meq/l)
1.467 (0.050)
1.623 (0.100)
1.636 (0.047)
Results are
expressed as mean (SEM).
LF and HF
indicate a standard diet and a high fat
diet, respectively.
*p<0.05,
**p<0.01 between saline treated mice
fed a standard diet and ghrelin treated
mice fed a high fat diet.
p<0.05
between saline treated mice fed a high fat
diet and ghrelin treated mice fed a high
fat diet.
Chronic effects of ghrelin administered intraperitoneally
(3 nmol/mouse every 12 hours for five days) on resistin gene
expression in the epididymal fat mass under a high fat (HF)
diet, as assessed by northern blot analysis, expressed as a
percentage of physiological saline treated controls under a
standard (LF) diet. Results are expressed as mean (SEM): n indicates
the number of mice used, *p<0.05 between saline treated mice
fed a high fat diet and ghrelin treated mice fed a high fat
diet. (B) Stimulatory effects of a high fat diet for two weeks
on ghrelin gene expression in the stomach of food deprived mice,
as assessed by northern blot analysis, expressed as a percentage
of standard diet fed controls. (Top) A representative northern
blot analysis showing gastric ghrelin mRNA during fasting after
a high fat diet for two weeks; *p<0.05 compared with the
control group by Bonferroni’/8s t test. G3PDH,
glyceraldehyde 3-phosphate dehydrogenase.
Effects of intraperitoneally administered [D-Lys-3]-GHRP-6
(2–/(200 nmol/mouse) on cumulative food intake in food
deprived lean mice: *p<0.05, **p<0.01 compared with the
control group by Bonferroni’/8s t test. (B) Effects
of intracerebroventricularly administered [D-Lys-3]-GHRP-6 (0.2–/(20
nmol/mouse) on cumulative food intake in food deprived lean
mice. ACSF, artificial cerebrospinal fluid. (C) Antagonistic
effects of [D-Lys-3]-GHRP-6 administered intracerebroventricularly
(20 nmol/mouse) on feeding induced by intraperitoneal injection
of ghrelin (3 nmol/mouse) in non-food deprived lean mice. (D)
Inhibitory effects of intraperitoneally administered [D-Lys-
3]-GHRP-6 (20–/(200 nmol/mouse) on the gastric emptying
rate one and two hours after injection in lean mice. (E) Effects
of intraperitoneally administered [D-Arg-1, D-Phe-5, D- Trp-7,
9, Leu-11] substance P (10–/(100 nmol/mouse) on cumulative
food intake in food deprived lean mice. (F) Effects of intraperitoneally
administered [D-Lys-3]-GHRP-6 (20–/(200 nmol/mouse) on
cumulative food intake in mice with diet induced obesity who
received a high fat diet for two weeks.
Acute effects of intraperitoneally administered
[D-Lys-3]-GHRP-6 (200 nmol/mouse) on cumulative food intake
in food deprived ob/ob obese mice: *p<0.05, **p<0.01
compared with physiological saline treated controls. (B) Chronic
effects of [D-Lys- 3]-GHRP-6 administered intraperitoneally
(20–/(200 nmol/mouse every 12 hours for six days) on
body weight gain in non-food deprived ob/ob obese mice.
Effects of [D-Lys-3]-GHRP-6 administered
intraperitoneally (20–/(200 nmol/mouse every
12 hours for six days) on food intake, epididymal
fat mass, gastrocnemius muscle, and blood glucose,
insulin, cholesterol, triglyceride, and free fatty
acid concentrations in ob/ob obese mice
Saline
20 nmol
200 nmol
Food intake (g/day)
4.845 (0.160)
4.527 (0.261)
4.285 (0.298)
Fat pad mass (g)
0.974 (0.066)
0.897 (0.169)
0.860 (0.086)
Skeletal muscle (g)
0.300 (0.012)
0.314 (0.009)
0.326 (0.013)
Glucose (mmolli)
13.01 (1.538)
12.06 (1.549)
7.489 (1.081)*
Insulin (pmol/1)
8294 (1676)
6242 (1628)
5481 (1304)
Cholesterol (mmol/1)
6.649 (0.346)
5.663 (0.291)
5.906 (0.565)
Triglycerides (mmol/l)
0.518 (0.049)
0.436 (0.040)
0.465 (0.068)
Free fatty acids (meq/l)
2.164 (0.075)
2.036 (0.121)
1.646 (0.078)**
*p<0.05,
**p<0.01 compared with physiological
saline treated controls.
GHRF-6, An Agonist for
Ghrelin receptor (GHS-R)
Interaction of the growth hormone releasing
peptides, ghrelin and GHRP-6, with the motilin receptor in the
rabbit gastric antrum
The structural relationship between the motilin and the growth
hormone secretagogue receptor (GHS-R), and between their respective
ligands, motilin and ghrelin, prompted us to investigate whether
ghrelin and the GHS-R agonist, growth hormone releasing peptide-6
(GHRP-6), could interact with the motilin receptor. The interaction
was evaluated in the rabbit gastric antrum with binding studies
on membrane preparations and with contraction studies on muscle
strips in the presence of selective antagonists under conditions
of electrical field stimulation (EFS) or not. Binding studies
indicated that the affinity (pKd) for the motilin receptor was
in the order of: ghrelin (4.230.07) < GHRP-6 (5.540.08) <
motilin (9.130.03). The interaction of ghrelin with the motilin
receptor requires the octanoyl group. Motilin induced smooth
muscle contractile responses but ghrelin and GHRP-6 were ineffective.
EFS elicited on- and off-responses that were increased by motilin
already at 10(-9) M, but not by 10(-5) M ghrelin. In contrast,
GHRP-6 also enhanced the on- and off-responses. The motilin
antagonist, GM-109, blocked the effect of GHRP-6 on the off-
but not on the on-responses. Under NANC conditions the effects
of motilin and GHRP-6 on the on-responses were abolished, those
on the off-responses were preserved. All responses were blocked
by NK1 and NK2 antagonists. In conclusion, ghrelin is unable
to induce contractions via the motilin receptor. However, GHRP-6
enhances neural contractile responses, partially via interaction
with the motilin receptor on non-cholinergic tachykininergic
nerves, partially via another receptor that may be a GHS-R subtype
on cholinergic nerves that corelease tachykinins.
Depoortere I, et al. J
Pharmacol Exp Ther 2003 Feb 11; [epub ahead of print]
Differential transport
of mouse ghrelin, des-octanoyl mouse ghrelin, and human
ghrelin across the blood-brain barrier in mice. Although
octanoylated (bioactive) mouse ghrelin crosses the mouse
BBB predominantly in the brain-to-blood direction, passage
for des-octanoyl mouse ghrelin was observed only in
the blood-to-brain direction. Human ghrelin, which differs
from mouse ghrelin by two amino residues only, was transported
in both directions in mice. The extent and direction
in which the ghrelin can cross the BBB is therefore
influenced by at least two features of its primary structure,
its post-translationally added fatty acid side chain
and its amino acid sequence.
William A. Banks,
Matthias Tschöp, Sandra M. Robinson and
Mark L. Heiman. THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL
THERAPEUTICS. Vol. 302, Issue 2, 822-827, August 2002
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