Administration Of GH-releasing Hormone Research Articles

Combined administration of ghrelin and GHRH synergistically stimulates GH release in Holstein preweaning calves

Ghrelin is a gut peptide which participates in growth regulation through its somatotropic, lipogenic and orexigenic effects. Synergism of ghrelin and growth hormone-releasing hormone (GHRH) on growth hormone (GH) secretion has been reported in humans and rats, but not in domestic animals in vivo. In this study, effects of a combination of ghrelin and GHRH on plasma GH and other metabolic parameters, and changes in plasma active and total ghrelin levels were studied in Holstein bull calves before and after weaning. Six calves were intravenously injected with vehicle (0.1% BSA-saline), ghrelin (1 μg/kg BW), GHRH (0.25 μg/kg BW) or a combination of ghrelin plus GHRH at the age of 5 weeks and 10 weeks (weaning at 6 weeks of age). Ghrelin stimulated GH release with similar potency as GHRH and their combined administration synergistically stimulated GH release in preweaning calves. After weaning, GH responses to ghrelin and GHRH became greater compared with the values of preweaning calves, but a synergistic effect of ghrelin and GHRH was not observed. The GH areas under the concentration curves for 2 h post-injection were greater in weaned than in preweaning calves ( P < 0.05) if ghrelin or GHRH were injected alone, but were similar if ghrelin and GHRH were injected together. Basal plasma active and total ghrelin levels did not change around weaning, but transiently increased after ghrelin injection. Basal plasma insulin, glucose and non-esterified fatty acid levels were reduced after weaning, but no changes by treatments were observed. In conclusion, ghrelin and GHRH synergistically stimulated GH release in preweaning calves, but this effect was lost after weaning.

Plasma growth hormone concentrations in female mithun (Bos frontalis) of different ages: relations with age and body weight

The role of growth hormone (GH) in postnatal somatic growth is well established. Its basal level and relation to growth performance in different age group mithun (Bos frontalis), a semiwild ruminant has not been characterized until now. To estimate the normal blood GH level and also to assess the influence of age and body weight (BW) on blood GH level in captive mithuns, a total of 65 female mithuns was divided into six age groups (group I, 0-6 months; group II, >6-12 months; group III, >1-2 years; group IV, >2-2.5 years; group V, >2.5-3.0 years and group VI, >3.0 years). Blood samples collected weekly for six consecutive weeks were assayed for GH. GH was also estimated in the samples collected from six growing mithuns at -60, -45, -30, -15, -10, -5 and 0 min prior to GH-releasing hormone (GHRH) administration for calculation of basal GH level and at 5, 10, 15, 30 min and thereafter at 15-min interval up to 8 h post-GHRH to assess blood GH response following GHRH administration in growing mithuns. For calculation of basal plasma GH in adult mithuns, GH was measured in blood samples collected at 30-min interval for 24 h from four animals. BW of all animals was recorded on two consecutive days per week and average of weekly BW was considered for growth rate calculation. It was found that both mean GH and GH per 100 kg BW between the age groups differ (p < 0.01). With increasing age and BW, GH and GH per 100 kg BW both decreased (p < 0.01). The age group with higher plasma GH and GH per 100 kg BW showed higher growth rates (r = 0.83 and 0.97 respectively). Interestingly, mean plasma GH for six consecutive weeks in all the groups showed much greater GH concentration (group I, 86.6 +/- 9.7 ng/ml to group VI 33.2 +/- 5 ng/ml) than reported in other species. Mean basal plasma GH calculated in growing and adult mithuns was 29.6 +/- 4.01 ng/ml and around 25 +/- 3.6 ng/ml respectively. The GH peak (444 +/- 21.3 ng/ml) was registered at 15 min post-GHRH administration in growing mithuns. In conclusion, age and BW influence plasma GH and GH per 100 kg BW but the latter is a better indicator of growth. The basal plasma GH and GH response to GHRH administration is six to eight and four to five times higher in mithun than in other species reported so far. An accurate assessment of the relationship between GH profiles and protein metabolism, proper receptor level study for GH action at the cellular level and the interaction of GH with other growth factors awaits better understanding of higher GH in this unique species.

Sleep in spontaneous dwarf rats

Spontaneous dwarf rats (SDRs) display growth hormone (GH) deficiency due to a mutation in the GH gene. This study investigated sleep in SDRs and their somatotropic axis and compared to Sprague–Dawley rats. SDRs had almost undetectable levels of plasma GH. Hypothalamic GH-releasing hormone (GHRH) mRNA was increased, whereas GHRH-receptor (GHRH-R) and somatostatin mRNAs were decreased in SDRs. Hypothalamic GHRH and somatostatin peptide content decreased in SDRs. Quantitative immunohistochemistry for GHRH and GHRH-R corroborated and extended these findings. In the arcuate nucleus, the number of GHRH-positive cells was significantly higher, whereas GHRH-R-positive perikarya were diminished in SDRs. Cortical GHRH and GHRH-R measurements showed similar expression characteristics as those found in the hypothalamus. SDRs had less rapid eye movement sleep (REMS) and more non-REMS (NREMS) than the control rats during the light period. The electroencephalogram (EEG) delta and theta power decreased during NREMS in the SDRs. After 4-h of sleep deprivation, SDRs had a significantly reduced REMS rebound compared to the controls, whereas NREMS rebound was normal in SDRs. The enhancement in delta power was significantly less than in the control group during recovery sleep. Intracerebroventricular (icv) administration of GHRH promoted NREMS in both strains of rats; however, increased REMS and EEG delta activity was observed only in control rats. Icv injection of insulin-like growth factor 1 increased NREMS in control rats, but not in the SDRs. These results support the ideas that GHRH is involved in NREMS regulation and that GH is involved in the regulation of REMS and in EEG slow wave activity regulation during NREMS.

Exogenous GH-releasing hormone increases GH and LH secretion in growing mithuns ( Bos frontalis)

To investigate the effects of growth hormone-releasing hormone (GHRH) administration on the patterns of GH and LH secretion in growing female mithuns, 12 mithuns within the age group of 10–12 months of age were divided into two groups (treatment and control groups) of six each in such a way that average body weight between the groups did not differ significantly ( P > 0.05). Both the groups were administered i.v. either with synthetic bGHRH [bGHRH (1-44)-NH 2] at 10 μg/100 kg body weight (treatment group) or equal volume of normal saline (control group). Blood samples collected prior to and after GHRH challenge at −60, −45, −30, −15, −10, −5 min and 5, 10, 15, 30 min, and thereafter, at an interval of 15 min up to 8 h post-injection were assayed for plasma GH and LH. Plasma progesterone was estimated in twice-a-week samples collected for six consecutive weeks preceding GHRH challenge to assess whether either group has begun ovarian cyclicity. Body weight of all animals was recorded once in a week during the period. A peak of GH was registered in all animals within 5–25 min post-GHRH administration with a mean peak of 443.5 ± 25.32 ng/ml at 15 min post-administration, which was much higher than in any other bovines reported following GHRH challenge. The patterns of LH secretion were pulsatile in nature in both the groups. Interestingly, the hormone concentrations exhibited higher pulsatility with greater amplitude after GHRH challenge in GHRH-treated than in control mithuns. The GHRH-treated mithuns averaged 0.44 pulses/h (4 pulses/9 h) and the rate was 0.20/h (2 pulses/9 h) in controls. The rate of pulse frequency and amplitude differed significantly with time of sampling. The mean plasma LH levels after GHRH administration were significantly higher in treatment group than those recorded in control mithuns. The mean plasma progesterone was similar ( P > 0.05) in both the groups and no animal from either group had begun ovarian cycle. In conclusion, exogenous GHRH significantly increases plasma GH and also LH pulse frequency and amplitude with higher mean post-GHRH LH levels in growing mithuns suggesting thereby its possible use for enhancement of maturity process in this unique meat animal.

Effects of growth hormone secretagogues on the release of adenohypophyseal hormones in young and old healthy dogs

The effects of three growth hormone secretagogues (GHSs), ghrelin, growth hormone-releasing peptide-6 (GHRP-6), and growth hormone-releasing hormone (GHRH), on the release of adenohypophyseal hormones, growth hormone (GH), adrenocorticotropic hormone (ACTH), thyroid-stimulating hormone (TSH), luteinising hormone (LH), prolactin (PRL) and on cortisol were investigated in young and old healthy Beagle dogs. Ghrelin proved to be the most potent GHS in young dogs, whereas in old dogs GHRH administration was associated with the highest plasma GH concentrations. The mean plasma GH response after administration of ghrelin was significantly lower in the old dogs compared with the young dogs. The mean plasma GH concentration after GHRH and GHRP-6 administration was lower in the old dogs compared with the young dogs, but this difference did not reach statistical significance. In both age groups, the GHSs were specific for GH release as they did not cause significant elevations in the plasma concentrations of ACTH, cortisol, TSH, LH, and PRL. It is concluded that in young dogs, ghrelin is a more powerful stimulator of GH release than either GHRH or GHRP-6. Ageing is associated with a decrease in GH-releasing capacity of ghrelin, whereas this decline is considerably lower for GHRH or GHRP-6.

Growth Hormone Response to Growth Hormone Releasing Hormone in Calves that Differ in Genetic Merit for Milk Yield

Holstein heifer, steer, and bull calves from control (CL) and select (SL) lines of cows that differed by more than 4000kg of milk during a 305-d lactation (SL>CL) were used to determine growth hormone (GH) response to 5 doses of GH releasing hormone (GHRH) and how this response was affected by gender, period (age), and genetic merit for milk yield. Doses (0, 2.5, 5, 10, and 20μg/100kg of BW) of a GHRH analog were assigned randomly to each heifer (4 CL, 4 SL), steer (4 CL, 4 SL), and bull (3 CL, 3 SL) calf and administered on consecutive days at approximately 3, 6, and 10 mo of age (periods; P1, P2, and P3). Jugular blood samples (n = 15) collected between −30 and 240min relative to GHRH administration were used to quantify area under the GH response curve (AUC) after subtracting mean prechallenge GH concentrations. Estimates of maximum response (Rmax) and sensitivity (ED50) to GHRH were obtained from the hyperbolic dose response curves (AUC vs. dose). Data were analyzed for effects of dose, line, period, gender, and their interactions with period as the repeated effect. Prechallenge GH concentrations were not affected by genetic line, gender, or period. The AUC was not affected by line, but decreased with period and increased with GHRH dose. The Rmax did not differ between lines or among genders, but decreased with period. The ED50 did not differ between lines or among periods, but heifers were more sensitive to GHRH than steers or bulls. Although GH response to GHRH has been identified as a potential indicator of genetic merit, it did not differ between these substantially different genetic lines.

Loss of Gq/11 family G proteins in the nervous system causes pituitary somatotroph hypoplasia and dwarfism in mice.

Heterotrimeric G proteins of the Gq/11 family transduce signals from a variety of neurotransmitter and hormone receptors and have therefore been implicated in various functions of the nervous system. Using the Cre/loxP system, we generated mice which lack the genes coding for the alpha subunits of the two main members of the Gq/11 family, gnaq and gna11, selectively in neuronal and glial precursor cells. Mice with defective gnaq and gna11 genes were morphologically normal, but they died shortly after birth. Mice carrying a single gna11 allele survived the early postnatal period but died within 3 to 6 weeks as anorectic dwarfs. In these mice, postnatal proliferation of pituitary somatotroph cells was strongly impaired, and plasma growth hormone (GH) levels were reduced to 15%. Hypothalamic levels of GH-releasing hormone (GHRH), an important stimulator of somatotroph proliferation, were strongly decreased, and exogenous administration of GHRH restored normal proliferation. The hypothalamic effects of ghrelin, a regulator of GHRH production and food intake, were reduced in these mice, suggesting that an impairment of ghrelin receptor signaling might contribute to GHRH deficiency and abnormal eating behavior. Taken together, our findings show that Gq/11 signaling is required for normal hypothalamic function and that impairment of this signaling pathway causes somatotroph hypoplasia, dwarfism, and anorexia.

Marked GH secretion after ghrelin alone or combined with GH-releasing hormone (GHRH) in obese patients.

Ghrelin is a 28-amino-acid peptide, predominantly produced by the stomach. It displays a strong GH-releasing activity mediated by the hypothalamus-pituitary GH secretagogue (GHS)-receptor (GHS-R). There are different studies that suggest the importance of ghrelin in feeding and weight homeostasis. In obesity there is a markedly decreased GH secretion. For both children and adults, the greater the body mass index (BMI), the lower the GH response to provocative stimuli, including the response to GHRH. However, the response to the natural GH secretaogogue ghrelin is unclear at the present time. The aim of the present study was to evaluate the GH response to ghrelin alone or combined with GHRH in a group of obese patients, in order to further understand the deranged GH secretory mechanisms in obesity and to clarify the mechanism of action of ghrelin. Six obese female patients (31 +/- 3.4 years) with a BMI of 36.1 +/- 7.7 kg/m(2) were studied. As a control group, six normal nonobese female subjects of similar age and sex were studied. Four tests were performed: placebo, GHRH [1 micro g/kg, no more than 100 micro g, intravenous (i.v.)], ghrelin (1 micro g/kg, no more than 100 micro g, i.v.) and GHRH (1 micro g/kg, no more than 100 micro g, i.v.) plus ghrelin (1 micro g/kg, no more than 100 micro g, i.v.). Blood samples were taken at appropriate intervals for determination of GH. Statistical analyses were performed by Wilcoxon and by Mann-Whitney tests. After GHRH, the median peak GH secretion in obese patients was 2.4 micro g/l (range 0.9-8.9 micro g/l). Ghrelin-induced GH secretion showed in obese patients a median peak of 24.4 micro g/l (range 7.4-85.0 micro g/l), significantly greater than the response after GHRH (P < 0.05). After the combined administration of GHRH plus ghrelin in obese patients the median peak GH secretion was 39.9 micro g/l (range 19.2-120.0 micro g/l), significantly greater than the response after GHRH (P < 0.05) or ghrelin (P < 0.05). GHRH-induced GH secretion in normal control subjects showed a median peak of 25.0 micro g/l (range 16.5-33.4 micro g/l). Ghrelin-induced GH secretion in normal showed a median peak of 68.5 micro g/l (range 22.5-119.5 micro g/l), significantly greater than the response after GHRH (P < 0.05). After the combined administration of GHRH plus ghrelin, in normal subjects the median peak GH secretion was 117.8 micro g/l (range 77.5-280.1 micro g/l), significantly greater than the response after GHRH or ghrelin alone (P < 0.05). When we compare the response of normal and obese patients, after GHRH alone, it was markedly decreased in obese people when compared with normal patients (P < 0.05) with a median GH peak of 25.0 micro g/l (range 16.5-33.4 micro g/l) and 2.4 micro g/l (range 0.9-8.9 micro g/l) for normal and obese patients, respectively. When we compare the response of normal and obese patients, after ghrelin alone or GHRH plus ghrelin, it was only blunted in obese subjects when compared with normal subjects with a median GH peak of 68.5 micro g/l (range 22.5-119.5 micro g/l) and 24.4 micro g/l (range 7.4-85 micro g/l) for normal and obese subjects, respectively, after ghrelin alone (P < 0.05) and a median GH peak of 117.8 micro g/l (range 77.5-280.1 micro g/l) and 39.9 micro g/l (range 19.2-120.0 micro g/l) for normal and obese patients, respectively, after GHRH plus ghrelin (P < 0.05). This study has demonstrated a massive GH response to ghrelin alone or combined with GHRH in obese patients, suggesting that altered ghrelin secretion could play a major role in the blunted GH secretion present in obese patients.

Chronic orchidectomy does not influence the sensitivity of the pituitary somatotropes to varying doses of GHRH administered intravenously to the adult male rhesus monkey

In the present study, the pituitary growth hormone (GH) response to graded doses of GH-releasing hormone (GHRH) was determined in intact (n = 3) and chronically orchidectomized (n = 3) adult rhesus monkeys (Mucaca mulatta). GHRH in doses of 0, 6.25, 12.5 and 25 μg/kg BW was infused through a teflon cannula implanted in the saphenous vein. Blood samples were collected 60 min before and 90 min after the injection of the neurohormone at 15 min intervals. All bleedings were carried out under ketamine hydrochloride anesthesia. The plasma levels of GH were determined by using AutoDELFIA time-resolved flouroimmunoassay, whereas plasma levels of testosterone and estradiol were determined using specific radioimmunoassay systems. The GH responses to GHRH were not significantly different between intact and chronically orchidectomized monkeys at any of the dose levels tested (p > 0.05). The administration of GHRH resulted in a significant (p < 0.05) stimulation of GH secretion at all the doses tested and in both the groups studied. In both intact and orchidectomized animals, the greatest response was observed at 6.25 μg/kg and no further increase was noted with the higher doses of GHRH. In conclusion, the present study suggests that chronic orchidectomy does not influence the sensitivity of the pituitary somatotropes to GHRH stimulation implying that the responsiveness of the pituitary somatotropes to GHRH is independent of testicular steroid modulation.

Gender differences in sleep endocrine regulation following administration of growth-hormone releasing hormone (GHRH) – relevance for the pathophysiology of major depression

Gender differences in sleep endocrine regulation following administration of growth-hormone releasing hormone (GHRH) – relevance for the pathophysiology of major depression

Effect of GHRH and GHRP-2 treatment in vitro on GH secretion and levels of GH, pituitary transcription factor-1, GHRH-receptor, GH-secretagogue-receptor and somatostatin receptor mRNAs in ovine pituitary cells.

Growth hormone (GH)-releasing hormone (GHRH) and GH-releasing peptides (GHRPs) stimulate the release of GH through their specific receptors on somatotropes. Combined GHRH and GHRP administration causes a synergistic GH release in vivo by an unknown mechanism. The current study focuses on the direct action of GHRH and GHRP on several molecular targets in somatotropes. To clarify the mechanism of action, ovine somatotropes were used to measure the expression of mRNAs encoding for GH, pituitary transcription factor-1 (Pit-1), GH-secretagogue receptor (GHS-R), GHRH-R, somatostatin receptor subtypes (sst-1 and sst-2) and GH release after GHRH and GHRP-2 treatment for 0.5, 1, 1.5 and 2 h. GHRH (10 nM), GHRP-2 (100 nM) and combined GHRH-GHRP-2 increased the levels of GH mRNA and GH release from 0.5 to 2 h in a time-dependent manner. The levels of Pit-1, GHRH-R and GHS-R mRNA were increased after 0.5 h treatment of cells with GHRH and GHRP-2. The levels of sst-1 but not sst-2 mRNA were significantly increased after 0.5 and 1 h of GHRH treatment. In contrast, both sst-1 and sst-2 mRNA expression was inhibited after 0.5-2 h of GHRP treatment. These data demonstrate a direct in vitro modification of ovine somatotropes by GHRH and GHRP-2 resulting in altered GHRH-R, GHS-R, Pit-1, sst-1, sst-2 and GH gene expression; this may underlie the regulatory action of GHRH and GHRP-2 on GH secretion.

Regulation of active ghrelin levels by various feedback mechanisms

Background: Ghrelin is a potent stimulator of growth hormone (GH) secretion. Furthermore, it is involved in energy homeostasis. Octanoylation of the peptide at serine-3 is necessary for biological activity. We investigated changes in active ghrelin plasma levels due to two different putative feedback mechanisms. Methods: Twelve healthy humans were tested by intravenous administration of the synthetic ghrelin analogue GHRP6 (1µg/kg), possibly activating a short feedback loop. On a different day, GH levels were increased by intravenous administration of growth hormone-releasing hormone (GHRH, 100µg iv) plus Arginine (30g iv over 30′) to test for a GH-dependent feedback mechanism. Blood samples were taken for measurement of ghrelin and GH. Active ghrelin-levels were determined by a specific radioimmunoassay that exclusively detects the octanoylated form of ghrelin. Results: Basal active ghrelin levels demonstrated significant inter-individual differences, but very little intra-individual variation (r=0.93, p<0,005). Administration of GHRP6 resulted in a 20-fold increase in GH levels. However, no significant changes of active ghrelin levels were observed over a 90′ period. Application of GHRH+Arginine led to an even higher 63-fold increase of GH levels, again without any significant change in levels of active ghrelin over a period of 120′. Conclusions: Acute increases in GH levels either by GHRP6 or by GHRH+Arginine did not feedback on systemic concentrations of active ghrelin. Furthermore, a ghrelin analogue had no short loop feedback effects on systemic active ghrelin levels. Due to its metabolic role, systemic ghrelin levels may be more dependent on energy homeostasis.

Effects of growth hormone-releasing peptides in healthy dogs and in dogs with pituitary-dependent hyperadrenocorticism

The aim of this study is to investigate the effects of ghrelin and GH-releasing peptide-6 (GHRP-6) on the release of growth hormone (GH), adrenocorticotrophic hormone (ACTH), and cortisol in dogs with pituitary-dependent hyperadrenocorticism (PDH) and in healthy dogs of comparable age. In eight healthy dogs, the responses to ghrelin and GHRP-6 were compared to those of GH-releasing hormone (GHRH) and NaCl 0.9% (control). In seven dogs with PDH, the effects of ghrelin and GHRP-6 were compared with their effects in healthy dogs. In the healthy dogs, GHRH, GHRP-6, and ghrelin caused a significant rise in plasma GH concentrations. GHRH administration elicited significantly higher plasma GH concentrations than administration of ghrelin and GHRP-6. In the dogs with PDH, the GHRP-6-induced release of GH was significantly lower than in healthy dogs. Administration of ghrelin elicited a GH release that did not differ significantly between dogs with PDH and healthy dogs. Ghrelin and GHRP-6 did not cause a significant rise in plasma ACTH and cortisol concentrations in either the healthy dogs or the dogs with PDH. It is concluded that in comparison with GHRH, GHRP-6 and ghrelin have a low GH-releasing potency in healthy dogs. In dogs with PDH, the GH release in response to GHRP-6 is impaired. Neither GHRP-6 nor ghrelin activates the pituitary–adrenocortical axis in healthy elderly dogs and dogs with PDH.

GH-related and extra-endocrine actions of GH secretagogues in aging.

Growth hormone (GH) secretion declines with aging in mammals, including humans. Defective pituitary function does not play a major role in this event. Rather, age-related changes involving the function of specific hypothalamic peptides for GH regulation, GH-releasing hormone (GHRH) and somatostatin (SS), appear to be the fundamental factors. Experimental evidence indicates that GHRH synthesis is impaired with aging in the rat hypothalamus, and that a relative hyperfunction of the SS-ergic system is present. In the elderly, systemic, short-term administration of GHRH enhances GH secretion and increases the formation of the GH-dependent peptide IGF-1. In old dogs, a combination of GHRH and clonidine (CLO), an alpha(2)-adrenoceptor agonist, reportedly acting via GHRH stimulation and SS inhibition, is the most effective stimulus to rejuvenate the apulsatile GH secretion. Discovery of GH secretagogues (GHSs), a series of peptydil and non-peptydil synthetic molecules endowed with a strong GH-releasing activity, the cloning of a GHS receptor (GHS-R), present in the hypothalamus and the pituitary, the isolation of the endogenous ligand of GHS-R, ghrelin, a 28-amino acid peptide whose main source is the stomach, pose the issue for the physiologic role of the GHS/ghrelin system and forces revision of our current understanding of GH regulation in different GH deficiency (GHD) states, including aging. GHSs are very effective for enhancing GH secretion in old subjects, but long-term studies are needed to assess their safety and the real biological impact of GHS replacement therapy in aging. Therapeutic use of GHSs can also be envisaged to restore, via GH-independent mechanisms, functional, and structural age-related alterations, such as anorexia, sexual dysfunction, cardiovascular damage, bone loss.

Growth hormone (GH) rebound rise following somatostatin infusion withdrawal: studies in dogs with the use of GH-releasing hormone and a GH-releasing peptide.

Evidence has been presented that in both animals and humans the rebound secretion of growth hormone (GH) following withdrawal of an infusion of somatostatin (SS) is due to the functional activation of the hypothalamic GH-releasing hormone (GHRH) neurons of the recipient organism. Based on this premise, this study has sought to assess the existence of functional interactions between endogenous GHRH released by a SS infusion withdrawal (SSIW) and growth hormone-releasing peptides (GHRPs), a class of compounds allegedly acting via GHRH. Five young dogs (3 to 4 years old, 2 male and 3 female) were administered, on different occasions, three consecutive intravenous boli of physiological saline (0.1 ml/kg), or GHRH (2 microg/kg), or EP92632 (125 microg/kg), a GHRP compound, or GHRH plus EP92632 at the end of three cycles of 1-h SS infusions (8 microg/(kg x h)) or during a 6-h infusion of saline. Under saline infusion (SALI), plasma GH levels were unaltered, whereas each SSIW cycle was followed by similar GH secretory episodes. Administration of the first GHRH bolus under SALI induced a rise in plasma GH concentrations slightly higher than that induced by the first cycle of SSIW, but the GH response to the second and third GHRH boli was similar to that after SSIW. Following SSIW, the response to the first bolus of GHRH was higher than that during SALI, but the second and third cycles of SSIW induced GH responses similar to those evoked by the GHRH bolus. During SALI, administration of the first bolus of EP92632 induced a rise in plasma GH which was higher than that induced by the first GHRH bolus, the second bolus elicited a GH peak of lesser amplitude and there was a partial restoration of the GH response to the third peptide bolus. SSIW strikingly enhanced the GH release to the first EP92632 bolus, a pattern also present, although to a lesser extent, with the second and third cycles of SSIW. Under SALI, combined administration of GHRH and EP92632 had a synergistic effect on GH release, but a progressive reduction was present in the GH response to the second and third GHRH plus EP92632 boli. SSIW increased only weakly the GH response to the first co-administration of the peptides over that present after administration of EP92632 alone, and did not induce a GH response higher than that present during SALI when the second bolus of the peptides was administered; after the third SSIW a GH rise higher than that present during SALI was elicited by the combined administration of the peptides. (i) the uniformity of the GH rebound responses to multiple cycles of SSIW may indicate that the latter activate a physiological mechanism which mimics that normally controlling GH pulse generation; (ii) EP92632 elicits, under our experimental conditions, a plasma GH rise higher than that induced by GHRH; (iii) SSIW enhances the GH response to EP92639 alone, to an extent reminiscent of that following combined administration of GHRH and EP92632. This pattern reinforces the view that SSIW elicits release of endogenous GHRH, and infers that the GHRP challenge after SSIW may be exploited in humans to distinguish between healthy and GH-deficient adults.

Involvement of Nitric Oxide in the Regulation of Growth Hormone Secretion in Dogs

Nitric oxide (NO) is a highly reactive gas that has been suggested to function as a neurotransmitter in the neuroendocrine system. In this work, we have evaluated the role of NO pathways in growth hormone (GH) secretion by assessing the effect of L-arginine infusion, a precursor of NO formation, and L-NAME, a nitric oxide synthase (NOS) inhibitor. The experiments were carried out on 7 adult beagle dogs. A saline infusion was carried out on all the dogs as a control test. L-arginine (infusion i.v. 10 g in 100 ml of saline, from t = 0 to 30 min) and L-nitro-arginine-methyl ester, L-NAME (infusion of 300 µg/kg in 120 ml of saline, from t = –30 to 45 min) were administered alone and together with growth hormone-releasing hormone (GHRH) (i.v. bolus at 0 min, at a dose of 100 µg), the synthetic GH secretagogue GHRP-6 (i.v. bolus at 0 min, at a dose of 90 µg), and the 5-HT1D serotonin receptor agonist sumatriptan, SUM (s.c. injection at the dose of 3 mg). Plasma cGH was determined by RIA. Results were evaluated by one-way analysis of variance, followed by the Newman-Keuls test for multiple comparisons. L-arginine administration resulted in a slight increase in plasma cGH in comparison with saline controls. Combined administration of L-arginine and GHRH enhanced cGH release in comparison with GHRH alone. L-NAME alone did not modify baseline cGH levels, but completely suppressed the GH release induced by GHRH or GHRP-6. It also strongly reduced, but did not abolish the effect of the two peptides (GHRH plus GHRP-6) administered together. Finally, administration of the 5-HT1D agonist SUM induced a significant cGH secretion in all dogs, a response which was not modified when L-NAME was administered in combination with SUM. In conclusion, our data show that inhibition of NO blunts both GHRH or GHRP-6-induced cGH release, and are compatible with the hypothesis that it acts by decreasing hypothalamic somatostatin release.

Changes in Immune Cell Counts and Interleukin (IL)-1β Production in Humans after a Somnogenically Active Growth Hormone-Releasing Hormone (GHRH) Administration

Growth hormone-releasing hormone (GHRH) has been shown to enhance slow-wave sleep (SWS) and non-rapid eye movement sleep in animals and humans. In animals the somnogenic effect of interleukin (IL)-1β appears to be mediated by GHRH. Neuroimmunological interactions in sleep are most frequently studied in humans by sleep deprivation or by cytokine administration. The present study, in contrast, investigates in humans the effect of enhanced sleep through GHRH administration on selected immune parameters. Results reveal that a single intravenous bolus of 50 μg GHRH which enhanced SWS stage 4 in the first half of the night suppressed circulating suppressor T cell (CD3+/CD8+) numbers, with a similar tendency for B cells (CD19+) and suppressed mitogen-stimulated IL-1β production. When the same amount of GHRH was administered distributed across five repetitive boluses of 10 μg GHRH within 1 h, neither corresponding sleep nor immune parameters were changed significantly compared to placebo. These data suggest that GHRH can modulate immune functions through brain mechanisms which are also involved in the regulation of sleep.

Effects of long-term administration of recombinant bovine tumor necrosis factor-alpha on glucose metabolism and growth hormone secretion in steers.

To investigate the effects of long-term administration of recombinant bovine tumor necrosis factor-alpha (rbTNF) on plasma glucose and growth hormone concentrations, and to determine whether treatment with rbTNF causes insulin resistance in steers. 5 steers treated with rbTNF and 5 steers treated with saline (0.9% NaCl) solution (control). In experiment 1, rbTNF (5.0 microg/kg of body weight) or saline solution (5 ml) was administered SC daily for 12 days. Blood samples were obtained before treatment, and plasma was harvested for determination of glucose, insulin, and growth hormone (GH) concentrations. In experiment 2, insulin, glucose, or growth hormone-releasing hormone (GHRH) was administered IV on days 7, 9, and 11, respectively, after initiation of rbTNF or saline treatment in experiment 1. Plasma glucose and insulin concentrations were measured before and at various times for 4 hours after insulin or glucose administration. Plasma GH concentrations were measured at various times for 3 hours after GHRH administration. In experiment 1, administration of rbTNF resulted in hyperinsulinemia without hypoglycemia and decreased plasma GH concentrations. In experiment 2, plasma glucose concentrations were higher in steers treated with rbTNF and insulin than in controls. Plasma GH concentrations were lower in steers treated with rbTNF and GHRH than in controls. Prolonged treatment with rbTNF induced insulin resistance and inhibited GHRH-stimulated release of GH in steers. Results indicate that rbTNF is a proximal mediator of insulin resistance and inhibits release of GH during periods of endotoxemia or infection.

Growth Hormone-Releasing Peptide-2 (GHRP-2) Acts Synergistically with Growth Hormone-Releasing Hormone (GHRH) to Release Growth Hormone (GH) in Swine

The aim of the present study was to examine the growth hormone (GH) response to the combined and repeated administrations of growth hormone-releasing peptide-2 (GHRP-2) plus growth hormone releasing hormone (GHRH) in swine. Six cross-bred castrated male swine, 130-135 days of age were used in this study. Animals were given a single intravenous (i.v.) injection of either GHRP-2 (30μg/kg BW), GHRH (1μg/kg BW) and GHRP-2 (30μg/kg BW) plus GHRH (1μg/kg BW) or 5ml saline (served as control) in random order with two days apart. GHRP-2 or GHRH alone and in combination with together stimulated the release of GH in swine. The peak concentrations of GH and areas under the GH response curve (GH AUC) for 120min after these treatments were significantly higher (P<0.01) than those in controls. The combined administration of these two peptides [GHRP-2 (30μg/kg BW) and GHRH (1μg/kg BW)] resulted in a GH AUC significantly greater than the arithmetic sum of those following their separate administrations (P<0.05), demonstrating the synergistic effect for GH release. GH responses to two successive i.v. injections of GHRP-2 (30μg/kg BW) plus GHRH (1μg/kg BW) at 2h interval in swine were increased after each injection; however, peak concentration of GH and GH AUC for 120min after the second injection were significantly lower (P<0.01) than those after the first injection. These results of the present study indicated that the combined administration of GHRP-2 and GHRH synergistically stimulates the release of GH in swine, and the GH responses to the co-administration of these two peptides decrease after the repeated injection. These findings suggest that GHRP-2 and GHRH act through different receptors and intracellular signal transduction and can be used together for stimulating further release of GH in swine.

GH-releasing hormone and GH-releasing peptide-6 for diagnostic testing in GH-deficient adults

The diagnosis of growth hormone (GH) deficiency in adults is based on provocative testing of GH secretion. The insulin tolerance test (ITT), currently the favoured test for this diagnosis, has been criticised for poor reproducibility and inconvenience. Since the combined administration of GH-releasing hormone (GHRH) plus GH-releasing peptide-6 (GHRP-6) is the most potent stimulus of GH secretion, we did a multicentre study comparing GH peaks elicited by ITT with those elicited by the GHRH/GHRP-6 test in healthy controls and GH-deficient individuals (cases). 125 adult patients with organic pituitary disease and 125 healthy individuals were studied. All cases and controls were given GHRH 1 microg per kg bodyweight intravenously plus GHRP-6 1 microg per kg intravenously at 0 min and blood samples were obtained during a subsequent 120 min period. 27 controls and all cases had an ITT. Inclusion criteria were severe GH deficiency--ie, a GH peak after ITT of < or = 3 microg/L. Results of the GHRH/GHRP-6 test were analysed by receiver-operating characteristic curve methodology. GH peaks seen after the GHRH/GHRP-6 test did not result in any side-effects and were not affected by age, sex, amount of adipose tissue, or by the GH assay system used. The GH mean peak after the GHRH/GHRP-6 test was 59.2 microg/L (SD 2.2) for controls and 4.1 microg/L (0.3) for cases, whereas after ITT the mean peak was 14.3 microg/L (1.7) and 0.5 microg/L (0.06), respectively. The differential peak responses of controls and cases was greater (p<0.001), for GHRH/GHRP-6 test than for ITT. When individually analysed GH peaks were a continuum, from 139.0 microg/L to 0.01 microg/L, with a cut-off point of 15.0 microg/L. The GHRH/GHRP-6 test performed well under the ROC curve analysis. For clinical utility, it is then proposed that values > or = 20.00 microg/L be considered normal and < or = 10.00 microg/L as GH deficient. The GHRH/GHRP-6 test is a convenient, safe and reliable test for adult GH deficiency and is not confounded by clinical factors known to alter GH secretory patterns. An evoked GH concentration of > or = 15.0 microg/L accurately distinguishes between healthy and GH-deficient adults.