Effects of Alprazolam,a Benzodiazepine,on the ACTH-, GH- and PRL-Releasing Activity of Hexarelin,a Synthetic Peptidyl GH Secretagogue (GHS), in Patients with Simple Obesity and in Patients with Cushing's Disease

GH secretagogues (GHS) possess potent GH-releasing activity but also stimulate PRL, ACTH and cortisol (F) secretion. To further clarify the endocrine activities of GHS, in 9 obese patients, 9 patients with Cushing's Disease and 14 controls we studied the ACTH, F, GH and PRL responses to hexarelin (HEX, 2.0 µg/kg i.v.), a peptidyl GHS, alone and preceeded by alprazolam (ALP, 0.02 mg/kg p.o.), a benzodiazepine. The HEX-induced ACTH response in controls was similar to that in obese patients (peak: 9.9 ± 1.9 and 24.7 ± 7.6 ng/L, respectively) and both were lower (p < 0.002) than that in Cushing's patients (peak: 210.7 ± 58.4 ng/L). The GH response to HEX in controls (peak: 58.1 ± 10.3 g/L) was higher (p < 0.001) than those in obese and Cushing's patients (18.2 ± 3.8 and 12.6 ± 5.4 /L, respectively) which, in turn, were similar. The PRL responses to HEX in controls, obese and Cushing's patients (peak: 11.9 ± 1.6, 18.0 ± 4.5 and 12.4 ± 1.4 /L, respectively) were similar. In controls the HEX-induced ACTH response was abolished by ALP (peak: 8.6 ± 2.4 vs 28.0 ± 6.7 ng/L, p < 0.03) which, on the other hand, only blunted that in obese (peak: 12.7 ± 2.1 vs 42.4 ± 8.4 ng/L, p < 0.02) and did not modify that in Cushing's patients (205.6 ± 55.4 vs 175.9 ± 47.6 ng/L). ALP blunted the GH response to HEX in controls (peak: 31.0 ± 7.1 /L, p < 0.03) while did not modify those in obese and in Cushing's patients (14.5 ± 5.3 and 13.3 ± 11.1 /L, respectively). ALP did not modify the HEX-induced PRL response in controls, obese and Cushing's patients (peak: 13.8 ± 0.9, 16.3 ± 2.4 and 19.2 ± 1.1 /L, respectively). In conclusion, alprazolam inhibits the ACTH response to hexarelin in normal and obese subjects while fails to modify the exaggerated ACTH response in Cushing's Disease suggesting that GHS activate the HPA axis via the hypothalamus in normal and obese subjects but not in patients with Cushing's disease. Alprazolam is also able to blunt the GH-releasing activity of hexarelin in normal subjects but not the low GH response to the hexapeptide in obese and Cushing's patients. The PRL-releasing activity of hexarelin in controls, obese and hypercortisolemic patients is similar and is not modified by alprazolam pretreatment.     

Recombinant human IGF-I does not modify the ACTH and cortisol responses to hCRH and hexarelin, a peptidyl GH secretagogue, in humans

An inhibitory influence of insulin-like growth factor-I (IGF-I) on hypothalamus-pituitary-adrenal (HPA) axis has been hypothesized. In fact, it has been reported that the rhGH (recombinant human GH)-induced IGF-I increase inhibits both cortisol and GH response to MK-0677, a non-peptidyl GH secretagogue in animals. The aim of this study was to further clarify the inhibitory role, if any, of IGF-I on corticotroph function. We studied the effect of rhIGF-I (recombinant human IGF-I; 20 microg/kg s.c. at -180 min) or placebo on the ACTH and cortisol responses to hCRH (human CRH; 2.0 microg/kg i.v. at 0 min) or hexarelin (HEX; 2.0 microg/kg i.v. at 0 min), a peptidyl GHS, in normal young women. The effect of rhIGF-I on the GH response to HEX was also studied. The subjects were six normal young women [age: 26-35 yr; body mass index (BMI): 19-23 kg/m2] in their early follicular phase. The results showed that after s.c. rhIGF-I administration, circulating IGF-I levels increased approximately 77%, peaking at -60 min and persisting similar up to +120 min. The mean ACTH, cortisol and GH concentrations did not change from -180 to 0 min when evaluated after both placebo or rhIGF-I. CRH and HEX induced similar ACTH (peak vs baseline, mean+/-SE: 47.5+/-10.9 vs 21.3+/-3.0 pg/ml and 30.3+/-6.9 vs 19.2+/-3.8 pg/ml, respectively; p<0.04) and cortisol responses (177.5+/-5.4 vs 109.3+/-10.3 microg/l and 149.4+/-12.3 vs 119.8+/-16.4 microg/l, respectively, p<0.04). RhIGF-I pretreatment did not modify the ACTH and cortisol responses to hCRH (46.0+/-13.8 pg/ml and 181.1+/-16.9 microg/l, respectively) as well as those to HEX (28.8+/-5.0 pg/ml and 144.1+/-16.2 microg/l, respectively). On the other hand, the GH response to HEX was clearly reduced by rhIGF-I (23.9+/-4.7 vs 64.7+/-14.8 microg/l, p<0.05). Our findings show that rhIGF-I-induced increase of circulating IGF-I levels exerts negative feedback action on somatotroph secretion, while it does not modify the corticotroph and the adrenal responsiveness to CRH or hexarelin.     

Effects of the combined administration of hexarelin, a synthetic peptidyl GH secretagogue, and hCRH on ACTH, cortisol and GH secretion in patients with Cushing's disease

Hexarelin (HEX) is a peptidyl GH secretagogue (GHS) which markedly stimulates GH release but, like other GHS, possesses also CNS-mediated ACTH- and cortisol-releasing activity. Interestingly, the stimulatory effect of HEX on ACTH and cortisol release is exaggerated and higher than that of hCRH in patients with Cushing's disease (CD). To further clarify the mechanisms by which HEX stimulates the activity of hypothalamo-pituitary-adrenal (HPA) axis in man, in 6 patients with CD (6 women, 38-68 yr old) and in 7 control subjects (CS, 7 women, 22-29 yr old) we studied the effects of HEX (2.0 microg/kg i.v.) and/or hCRH (2.0 microg/kg i.v.) on ACTH and cortisol (F) secretion. The GH responses to HEX alone and combined with hCRH were also studied in all subjects. Basal ACTH and F levels in CD were higher than in CS (66.3+/-5.1 vs 16.5+/-0.6 pg/ml and 217.8+/-18.5 vs 134.4+/-4.6 microg/l, respectively; p<0.02). In CS, the ACTH and F responses to HEX, evaluated as deltaAUC (mean+/-SE: 128.7+/-39.2 pg x min/ml and 328.5+/-93.2 microg x min/l, respectively) were lower, though not significantly, than those after hCRH (375.8+/-128.4 pg x min/ml and 1714.2+/-598.0 microg x min/l, respectively), though the peak ACTH and F responses to both stimuli were similar. The co-administration of HEX and hCRH had an additive effect on both ACTH (1189.6+/-237.2 pg x min/ml) and F secretion (3452.9+/-648.6 microg x min/l). In fact, the ACTH and F responses to HEX+/-hCRH were significantly higher (p<0.01) than those elicited by single stimuli. In CD, HEX induced ACTH and F responses (3603.8+/-970.7 pg x min/ml and 10955.9+/-6184.6 microg x min/l, respectively) clearly higher (p<0.002) than those in CS. The HEX-induced ACTH and F responses in CD were higher, though not significantly, than those recorded after hCRH (1432.7+/-793.5 pg x min/ml and 4832.7+/-2146.5 microg x min/l, respectively). On the other hand, the hCRH-induced ACTH and F responses in CD were similar to those in CS. In CD, the coadministration of HEX and hCRH had an additive effect on ACTH (8035.7+/-1191.1 pg x min/ml) but not on F (10985.4+/-3900.8 microg x min/l) secretion. In fact, the ACTH, but not the F response to HEX+hCRH was significantly higher (p<0.02) than that elicited by single stimuli. In conclusion, the present study demonstrates that in patients with Cushing's disease as well as in subjects control Hexarelin and hCRH have an additive effect on ACTH secretion. Considering that, at least in humans, differently from hCRH, GHS have no interaction with AVP, our present findings further agree with the hypothesis that the ACTH-releasing activity of GHS is, at least partially, independent of CRH-mediated mechanisms.     

Endocrine activities of ghrelin, a natural growth hormone secretagogue (GHS), in humans: comparison and interactions with hexarelin, a nonnatural peptidyl GHS, and GH-releasing hormone

An endogenous ligand for the GH secretagogue-receptor (GHS-receptor) has recently been isolated, from both the rat and the human stomach, and named ghrelin. It is a 28-amino-acid peptide showing a unique structure with an n-octanoyl ester at its third serine residue, which is essential for its potent stimulatory activity on somatotroph secretion. In fact, it has been demonstrated that ghrelin specifically stimulates GH secretion from both rat pituitary cells in culture and rats in vivo. The aim of the present study was to test the GH-releasing activity of ghrelin in humans and to compare it with that of GHRH and hexarelin (HEX), a nonnatural peptidyl GHS, which possesses strong GH-releasing activity but also significantly stimulates PRL, ACTH, and cortisol secretion. To clarify the mechanisms of action underlying the GH-releasing activity of ghrelin in humans, its interaction with GHRH and HEX was also studied. Seven normal young volunteers (7 men; 24-32 yr old; body mass index, 20-24 kg/m(2)) were studied. All subjects underwent the administration of ghrelin, HEX, and GHRH-29 (1.0 microg/kg i.v. at 0 min) as well as placebo (2 mL isotonic saline i.v. at 0 min). Six subjects also underwent the combined administration of ghrelin and GHRH or HEX. Blood samples were taken every 15 min from -15 up to +180 min. GH levels were assayed at each time point in all sessions; PRL, ACTH, cortisol, and aldosterone levels were also assayed after administration of ghrelin and/or HEX. Ghrelin administration induced a prompt and marked increase in circulating GH levels (Cmax, mean +/- SEM, 92.1 +/- 16.7 microg/L; area under the curve, 1894.9 +/- 347.8 microg/L.h). The GH response to ghrelin was clearly higher (P < 0.01) than the one recorded after GHRH (26.7 +/- 8.7 microg/L; 619.6 +/- 174.4 microg/L.h) and even significantly higher (P < 0.05) than after HEX (68.4 +/- 14.7 microg/L; 1546.9 +/- 380.0 microg/L x h). Ghrelin administration also induced an increase in PRL, ACTH, and cortisol levels; these responses were higher (P < 0.05) than those elicited by HEX. A significant increase in aldosterone levels was recorded after ghrelin but not after HEX. The endocrine responses to ghrelin were not modified by the coadministration of HEX. On the other hand, the coadministration of ghrelin and GHRH had a real synergistical effect (P < 0.05) on GH secretion (133.6 +/- 22.5 microg/L; 3374.3 +/- 617.3 microg/L x h). In conclusion, ghrelin, a natural ligand of GHS-receptor, exerts a strong stimulatory effect on GH secretion in humans, releasing more GH than GHRH and even more than a nonnatural GHS such as HEX. Ghrelin, as well as HEX, also stimulates lactotroph and corticotroph secretion. Ghrelin shows no interaction with HEX, whereas it has a synergistical effect with GHRH on GH secretion. Thus, ghrelin is a new hormone playing a major role in the control of somatotroph secretion in humans, and its effects are imitated by nonnatural GHS.     

Interaction between glucagon and hexarelin, a peptidyl GH secretagogue, on somatotroph and corticotroph secretion in humans

OBJECTIVE: Glucagon administration stimulates both somatotroph and corticotroph secretion in humans, although this happens only if glucagon is administered by the intramuscular route and not by the intravenous route. On the other hand, GH secretagogues (GHS) strongly stimulate GH and also possess ACTH-releasing activity. DESIGN AND METHODS: To clarify the mechanisms underlying the stimulatory effects of both glucagon and GHS on somatotroph and corticotroph secretion, we studied the GH, ACTH and cortisol responses to glucagon (GLU, 0.017 mg/kg i.m.) and Hexarelin, a peptidyl GHS (HEX, 2.0 microg/kg i.v.) given alone or in combination in 6 normal young volunteers (females, aged 26-32 years, body mass index 19.7-22.5 kg/m). RESULTS: GLU administration elicited a clear increase in GH (peak vs baseline, mean+/-S.E.M.: 11.6+/-3.4 vs 3. 3+/-0.7 microg/l, P<0.02), ACTH (11.6+/-3.3 vs 4.1+/-0.3 pmol/l, P<0. 02) and cortisol (613.5+/-65.6 vs 436.9+/-19.3 nmol/l, P<0.05) levels. HEX induced a marked increase in GH levels (55.7+/-19.8 vs 3. 7+/-1.9 microg/l, P<0.005) and also significant ACTH (5.7+/-1.1 vs 3. 4+/-0.6 pmol/l, P<0.01) and cortisol (400.2+/-31.4 vs 363.4+/-32.2 nmol/l, P<0.05) responses. The GH area under the curve (AUC) after HEX was clearly higher than after GLU (1637.3+/-494.0 vs 479.1+/-115. 7 microg/l/120 min, P<0.04) while HEX and GLU coadministration had a true synergistic effect on GH release (3243.8+/-687.5 microg/l/120 min, P<0.02). The ACTH and cortisol AUCs after HEX were lower (P<0. 02) than those after GLU (208.3+/-41.3 vs 426.3+/-80.9 pmol/l/120 min and 18 874.5+/-1626.1 vs 28 338.5+/-2430.7 nmol/l/120 min respectively). The combined administration of HEX and GLU had an effect which was less than additive on both ACTH (564.02+/-76.5 pmol/l/120 min) and cortisol (35 424.6+/-5548.1 nmol/l/120 min) secretion. CONCLUSIONS: These results show that the intramuscular administration of glucagon releases less GH but more ACTH and cortisol than Hexarelin. The combined administration of glucagon and Hexarelin has a true synergistic effect on somatotroph secretion but a less than additive effect on corticotroph secretion; these findings suggest that these stimuli act via different mechanisms to stimulate somatotrophs while they could have a common action on the hypothalamo-pituitary-adrenal axis.     

Impact of two or three daily subcutaneous injections of hexarelin, a synthetic growth hormone (GH) secretagogue, on 24-h GH, prolactin, adrenocorticotropin and cortisol secretion in humans

OBJECTIVE: To extend the insights on the action of GH secretagogues (GHS) on pituitary function, we studied the impact of intermittent daily s.c. administration of a peptidyl GHS, hexarelin (HEX), on 24-h GH, PRL, ACTH and cortisol release in healthy volunteers. DESIGN: We investigated the impact of two or three times daily s.c. administration of a short-acting peptidyl GHS, the hexapeptide HEX (1.5 microg/kg) on 24-h GH, PRL, ACTH and cortisol secretion (sampling every 20 min) in six normal young men. To monitor possible down-regulation, the effect of 1 microg/kg i.v. HEX at the end of each 24-h sampling period was studied. METHODS: Multi-parameter deconvolution analysis was used to quantitate pulsatile GH, PRL, ACTH and cortisol secretion and estimate the corresponding hormone half-lives. Complementary to deconvolution analysis, approximate entropy was used as a scale- and model-independent statistic to quantify the serial orderliness or pattern regularity of hormone measurements. RESULTS: Mean and integrated (24-h) serum GH concentrations were increased from baseline values to the same extent by two and three HEX injections. Both HEX schedules equally increased GH secretory burst mass (but not burst frequency), mean daily GH production rate, GH half-life and irregularity of GH release patterns. No change occurred in the secretion of IGF-I, PRL, ACTH and cortisol. Intravenous HEX at the end of each spontaneous 24-h profile induced a significant rise in GH, PRL, ACTH and cortisol. Prior HEX administration blunted the GH response, abolished that of ACTH and cortisol and did not modify the PRL increase. CONCLUSIONS: The study showed that two or three daily s.c. injections of HEX augmented 24-h GH secretion equally, amplifying selectively GH secretory pulse mass without altering lactotroph and corticotroph secretion. IGF-I levels were not modified by these 1-day HEX treatment schedules.     

Glucagon administration elicits blunted GH but exaggerated ACTH response in obesity

Reduction in both spontaneous and stimulated GH secretion in obesity has been clearly demonstrated. Mild hyperactivity of hypothalamus-pituitary-adrenal (HPA) axis has been also reported. Glucagon, at least after im administration, induces clear increase in either GH or ACTH and F levels but its effect on somatotroph and corticotroph secretion in obesity has never been studied. In 7 patients with abdominal obesity (OB, aged 24-42 yr, BMI: 29.1-43.9 kg/m2, waist/hip ratio [WHR]: 0.86-1.00) we studied the GH, ACTH and F responses to the im administration of glucagon (0.017 mg/kg at 0 min). The results in OB were compared with those in a group of 6 age-matched controls normal subjects (Ns aged 26-32 yr, BMI 19.7-22.5 kg/m2). In Ns glucagon administration induced clear increase in GH (peak vs baseline, mean+/-SE: 11.6+/-3.4 vs 3.3+/-0.7 microg/l, p<0.02), and ACTH (52.9+/-15.2 vs 19.0+/-1.5 pg/ml, p<0.02) levels which peaked at +150 and +165 min, respectively. Increase in F levels (222.3+/-23.8 vs 158.3+/-7.0 ng/ml, p<0.05) was also recorded but peaked at +180 min. In OB glucagon administration induced GH response (7.4+/-2.3 vs 0.8+/-0.6 microg/l) lower (p<0.05) than that recorded in Ns; when the GH responses were evaluated by co-variance analysis, a significant difference between the 2 groups was recorded in term of peaks but not of AUCs. On the other hand, the ACTH response to glucagon in OB was higher than that in Ns (11452.6+/-2447.7 vs 4892.2+/-719.4 pg/ml x min, p<0.05). The F response to glucagon in OB and Ns was, however, similar (24057.9+/-4109.1 vs 29835.9+/-1566.0 ng/ml x min). In conclusion, this study demonstrates that in obese patients the im administration of glucagon elicits blunted GH response but exaggerated ACTH increase which is uncoupled with the adrenal response. These findings agree with the existence of concomitant GH insufficiency and altered corticotroph function in obesity.     

Effects of cortistatin-14 and somatostatin-14 on the endocrine response to hexarelin in humans

Cortistatin (CST)-14, a neuropeptide with high structural homology with somatostatine (SS)-14, binds all SS receptor subtypes but also shows activities not shared by SS. CST and SS are often co-expressed in the same neurons but are regulated by different stimuli. Moreover, CST, but not SS, also binds the GH secretagogue (GHS) receptor. We compared the effects of CST-14 and SS-14 (2.0 microg/kg/h i.v. from -30 to +90 min) on the endocrine response to hexarelin (HEX, 1.0 microg/kg i.v. at 0 min), a synthetic GHS, in 6 normal volunteers [age (mean+/-SEM): 28.7+/-2.9 yr; body mass index: 23.4+/-0.8 kg/m2]. GH, PRL, ACTH, cortisol, insulin and glucose levels were measured at each time point. CST-14 inhibited spontaneous GH secretion [delta-areas under curves (-AUC): -83.57+/-44.8 vs 2.3+/-2.7 microg/l/h, p<0.01] to the same extent of SS-14 (-186.1+/-162.9 microg/l/h, p<0.01). CST-14 as well as SS-14 also inhibited insulin secretion (p<0.05). The GH response to HEX was similarly inhibited by either CST-14 (AUC: 3814.1+/-924.2 vs 1212.9+/-379.8 microg/l/h, p<0.05) or SS-14 (720.9+/-158.6 microg/l/h, p<0.05). HEX significantly increased PRL, ACTH and cortisol levels but these responses were not modified by either CST-14 or SS-14. The effects of CST-14 and SS-14 on insulin and glucose levels were not modified by HEX. In conclusion, this study shows that CST-14 inhibits the GH response to HEX to the same extent of SS-14. Like SS-14, CST-14 also inhibits insulin secretion but both do not modify the stimulatory effects of HEX on lactotroph and corticotroph secretion. Thus, CST-14 exerts full SS-14 activity in humans.     

Corticotropin-releasing effect of hexarelin, a peptidyl GH secretagogue, in normal subjects pretreated with metyrapone or RU-486, a glucocorticoid receptor antagonist, and in patients with Addison's disease.

GH secretagogues (GHS) are peptidyl and nonpeptidyl molecules which possess strong GH-releasing activity but also stimulatory effect on hypothalamo-pituitary-adrenal axis. The ACTH and cortisol responses to Hexarelin (HEX), a peptidyl GHS, are abolished by low-dose dexamethasone pretreatment in normal subjects but are exaggerated and higher than those after hCRH in patients with pituitary ACTH-dependent Cushing's disease, in spite of their hypercortisolism. Based on the foregoing, we studied the ACTH, cortisol and GH responses to HEX (2.0 microgram/kg i.v. at 0 min) alone and after metyrapone (2 g p.o. at 23:00 h the night before) or RU-486 (400 mg p.o. at 02:00 h), a glucocorticoid receptor antagonist, in 6 normal women (NS, age 26-34 years). The endocrine responses (mean +/- SEM) to HEX alone were also studied in 8 patients with Addison's disease (AD, 6 males, 2 females, age 30-77 years; last hydrocortisone administration the day before testing). In NS, HEX stimulated basal ACTH (peak, mean +/- SEM: 26.0 +/- 7.8 vs. 10.7 +/- 2.0 pg/ml, p < 0. 05), cortisol (163.2 +/- 18.3 vs. 137.4 +/- 15.4 microgram/l, p < 0.05) and GH (72.6 +/- 23.5 vs. 3.7 +/- 1.3 microgram/l, p < 0.01) levels. Metyrapone markedly increased basal ACTH (294.4 +/- 61.6 pg/ml, p < 0.05), reduced basal cortisol (19.6 +/- 7.2 microgram/l, p < 0.05), while it did not modify GH levels. After metyrapone pretreatment the ACTH response to HEX was clearly increased (DeltaAUC: 2,857.4 +/- 901.9 vs. 367.3 +/- 274.0 pg/ml/h, p < 0.05), while the GH response was not modified. HEX did not stimulate the low cortisol levels after metyrapone pretreatment. RU-486 significantly increased basal ACTH (76.6 +/- 12.5 pg/ml, p < 0.05) and cortisol (312.7 +/- 22.2 microgram/l, p < 0.05), while it did not modify basal GH levels. RU-486 pretreatment did not modify the ACTH, cortisol and GH responses to HEX. In AD, HEX elicited a marked ACTH response (6,619.4 +/- 3,365.8 pg/ml/h; p < 0.01), which was clearly higher (p < 0.01) than that in NS after HEX alone but not significantly different from that after HEX+MET. The GH response to HEX in AD (1,325.6 +/- 284.1 microgram/l/h) was similar to that in NS (1,519.7 +/- 483.8 microgram/l/h). In conclusion, our present data demonstrate that the ACTH-releasing activity of HEX is increased in primary hypoadrenalism as well as in normal subjects after metyrapone but not after RU-486 pretreatment. These findings indicate that in normal subjects as well as in hypocortisolemic patients the ACTH-releasing activity of GHS is enhanced by the lack of negative glucocorticoid feedback.     

Endocrine activities of alexamorelin (Ala-His-d-2-methyl-Trp-Ala-Trp-d-Phe-Lys-NH2), a synthetic GH secretagogue, in humans

OBJECTIVE: Peptidyl and non-peptidyl synthetic GH secretagogues (GHS) possess significant GH-, prolactin (PRL)- and ACTH/cortisol-releasing activity after i.v. and even p.o. administration, acting via specific hypothalamo-pituitary receptors in both animals and humans. The hexapeptide hexarelin (HEX) is a paradigmatic GHS whose activities have been widely studied in humans. The heptapeptide Ala-His-d-2-methyl-Trp-Ala-Trp-d-Phe-Lys-NH(2) (alexamorelin, ALEX) is a new synthetic molecule which inhibits GHS binding in vitro, but its endocrine activity has never been studied in humans. DESIGN: In six young adults we studied the effects of 1.0 and 2.0 microgram/kg i.v. ALEX or HEX on GH, PRL, ACTH, cortisol and aldosterone levels and those of 20mg p.o. ( approximately 300 microgram/kg) on GH levels. RESULTS: Basal GH, PRL, ACTH, cortisol and aldosterone levels in all testing sessions were similar. ALEX and HEX (1.0 and 2.0 microgram/kg i.v.) induced the same dose-dependent increase of GH and PRL levels. Both ALEX and HEX induced a dose-dependent increase of ACTH and cortisol levels. The ACTH and cortisol responses to the highest ALEX dose were significantly higher than those after HEX. Aldosterone levels significantly increased after both i.v. ALEX doses, but not after HEX. The GH response to 20mg p.o. ALEX was higher, though not significantly, than that to the same HEX dose. CONCLUSION: ALEX, a new GHS, shows the same GH-releasing activity as HEX. On the other hand, ALEX seems endowed with an ACTH-releasing activity more marked than that of HEX; this evidence could explain the significant increase of aldosterone levels after its i.v. administration.     

Tyr-Ala-Hexarelin, a synthetic octapeptide, possesses the same endocrine activities of Hexarelin and GHRP-2 in humans

Hexarelin (HEX) and GHRP-2 are two synthetic hexapeptides, superanalogs of GHRP-6, belonging to GH secretagogue (GHS) family. GHS act via specific receptors at both the pituitary and the hypothalamic level to stimulate GH release both in animals and in humans. However, GHS also possess significant PRL- and ACTH/cortisol-releasing activity. Tyr-Ala-HEX as well as Tyr-Ala-GHRP-6 are, in turn, synthetic octapeptides generally used to perform binding studies because of their easy iodination. However, their endocrine activities have never been studied in humans. To clarify the endocrine activities of Tyr-Ala-HEX, in 7 young adult volunteers we compared the effects of the maximal effective dose of HEX (2.0 microg/kg i.v.) or GHRP-2 (2.0 microg/kg i.v.) with the same one of Tyr-Ala-HEX on GH, PRL, ACTH and cortisol levels. Basal GH, PRL, ACTH and cortisol levels in all testing sessions were similar. The administration of placebo did not modify hormonal levels. HEX and GHRP-2 administration induced the well known strong GH response (Cmax, mean+/-SE: 77.3+/-6.0 and 74.1+/-12.1 microg/l; AUC, mean+/-SE: 2596.7+/-251.1 and 2480.0+/-343.6 microg*min/l). These responses were similar to that induced by Tyr-Ala-HEX (63.7+/-18.5 microg/l; 1986.6+/-622.4 microg*min/l). Moreover, HEX, GHRP-2 and Tyr-Ala-HEX had the same significant stimulatory effect on PRL (14.9+/-2.5, 12.3+/-2.0 and 10.0+/-1.5 microg/l; 497.8+/-61.8, 480.4+/-66.9 and 415.8+/-58.5 microg*min/l), ACTH (48.0+/-10.1, 51.4+/-10.6 and 44.9+/-12.2 pg/ml; 1531.6+/-235.7, 1586.7+/-277.0 and 1338.1+/-164.5 pg*min/ml) and cortisol (179.9+/-10.0, 181.2+/-14.1 and 149.7+/-20.1 microg/l; 8465.0+/-406.6, 8689.2+/-788.1 and 6295.2+/-797.0 microg*min/l). Also the mean Tmax of the endocrine responses to HEX, GHRP-2 and Tyr-Ala-HEX were similar. In conclusion, the present results demonstrate that in humans Tyr-Ala-HEX is a GH secretagogue as potent as HEX and GHRP-2, two GHRP-6 superanalogs. Tyr-Ala-HEX also shares with HEX and GHRP-2 the same PRL- ACTH- and cortisol-releasing activity.     

Acute cardiovascular and hormonal effects of GH and hexarelin, a synthetic GH-releasing peptide, in humans.

Reduced cardiac mass and performances are present in GH deficiency and are counteracted by rhGH replacement. GH and IGF-I possess specific myocardial receptors and have been reported able to exert an acute inotropic effect. Synthetic GH secretagogues (GHS) possess specific pituitary and hypothalamic but even myocardial receptors. In 7 male volunteers, we studied cardiac performance by radionuclide angiocardiography after iv administration of rhGH or hexarelin (HEX), a peptidyl GHS. The administration of rhGH or HEX increased circulating GH levels to the same extent (AUC: 1594.6+/-88.1 vs 1739.3+/-262.2 microg/l/min for 90 min) while aldosterone and catecholamine levels did not change; HEX, but not rhGH, significantly increased cortisol levels. Left ventricular ejection fraction (LVEF), mean blood pressure (MBP) and heart rate (HR) were unaffected by rhGH (62.4+/-2.1 vs 62.1+/-2.3%, 90.6+/-3.4 vs 92.0+/-2.5 mm Hg, 62.3+/-1.8 vs 66.7+/-2.7 bpm). HEX increased LVEF (70.7+/-3.0 vs 64.0+/-1.5%, p<0.03) without significant changes in MBP and HR (92.8+/-4.7 vs 92.4+/-3.2 mm Hg, 63.1+/-2.1 vs 67.0+/-2.9 bpm). LVEF significantly raised at 15 min, peaked at 30 min and lasted up to 60 min after HEX. These findings suggest that in man, the acute administration of Hexarelin exerts a short-lasting, positive inotropic effect. This effect seems GH-independent and might be mediated by specific GHS myocardial receptors.     

Alprazolam (a benzodiazepine activating GABA receptor) reduces the neuroendocrine responses to insulin-induced hypoglycaemia in humans

OBJECTIVE: Alprazolam (ALP), a benzodiazepine-activating GABAergic receptor, possesses clear centrally mediated inhibitory effects on ACTH and cortisol secretion that could reflect an inhibitory influence on CRH- and/or AVP-secreting neurones. An inhibitory effect of ALP on catecholamine release has also been shown while its effect on GH secretion is unclear. To further clarify the neuroendocrine actions of ALP, we studied the ALP effects on the neurohormonal responses to hypoglycaemia in a group of normal subjects. DESIGN: In eight normal subjects [four women and four men, 22-34 years old, body mass index (BMI) 20-25 kg/m2] the ACTH, cortisol, GH, adrenaline (A) and noradrenaline (NA) responses to insulin-induced hypoglycaemia [ITT, 0.1 UI/kg regular insulin intravenously (i.v.) at 0 min] preceded by placebo or ALP (0.02 mg/kg orally at -90 min) were studied in two sessions at least 10 days apart. MEASUREMENTS: Blood samples were taken basely at -90 and 0 min and every 15 min up to +120 min. ACTH, cortisol, GH, A and NA level were assayed at each time point in both sessions. RESULTS: All subjects experienced hypoglycaemia (plasma glucose levels below 2.2 mmol/l). After placebo ITT induced clear-cut increases in ACTH (peak vs. baseline, mean +/- SEM: 27.9 +/- 3.9 vs. 7.1 +/- 1.5 pmol/l), cortisol (438.1 +/- 32.0 vs. 237.7 +/- 19.3 nmol/l) and GH (38.1 +/- 9.7 vs. 5.7 +/- 2.0 micro g/l) levels (P < 0.05). Marked increase in A (6627.2 +/- 116.7 vs. 263.7 +/- 71.4 pmol/l) and NA (3.8 +/- 1.5 vs. 1.6 +/- 1.0 nmol/l) levels were also recorded (P < 0.05). Pretreatment with ALP significantly inhibited the ACTH peak response to ITT (17.8 +/- 5.0 pmol/l, P < 0.05), while the cortisol response showed a non significant reduction (342.1 +/- 38.7 nmol/l). ALP also significantly reduced the GH (21.7 +/- 4.7 micro g/l, P < 0.02) and A (3828.0 +/- 1400.7 pmol/l, P < 0.02) responses to ITT. On the contrary, ALP lowered basal NA levels (P < 0.05) but did not significantly affect its response to ITT (2.2 +/- 1.2 nmol/l). Glucose changes induced by ITT were not modified by ALP. CONCLUSIONS: This study shows that GABAergic activation by alprazolam significantly inhibits the neuroendocrine and adrenomedullary responses to hypoglycaemia.     

Obese patients with obstructive sleep apnoea syndrome show a peculiar alteration of the corticotroph but not of the thyrotroph and lactotroph function

OBJECTIVE: Obstructive sleep apnoea syndrome (OSAS) is strongly associated with obesity (OB) and is characterized by several changes in endocrine functions, e.g. GH/IGF-I axis, adrenal and thyroid activity. It is still unclear whether these alterations simply reflect overweight or include peculiar hypoxia-induced hormonal alterations. Hormonal evaluations have been generally performed in basal conditions but we have recently reported that OSAS is characterized by a more severe reduction of the GH releasable pool in comparison to simple obesity. We aimed to extend our evaluation of anterior pituitary function to corticotroph, thyrotroph and lactotroph secretion under dynamic testing in OSAS in comparison with simply obese and normal subjects. SUBJECTS AND METHODS: In 15 male patients with OSAS [age, mean +/- SEM 43.5 +/- 1.6 years; body mass index (BMI) 39.2 +/- 3.1 kg/m2; apnoea/hypopnoea index, (AHI) 53.4 +/- 8.7], 15 male patients with simple obesity (OB, age 39.7 +/- 1.2 years; BMI 41.2 +/- 2.0 kg/m2; AHI 3.1 +/- 1.2 events/h of sleep) and in 15 normal lean male subjects (NS, age 38.2 +/- 1.4 years; BMI 21.2 +/- 0.8 kg/m2; AHI 1.9 +/- 0.8 events/h of sleep) we evaluated: (a) the ACTH and cortisol responses to CRH [2 microg/kg intravenously (i.v.)] and basal 24 h UFC levels; (b) the TSH and PRL responses to TRH (5 microg/kg iv) as well as FT3 and FT4 levels. RESULTS: Twenty-four-hour UFC levels in OSAS and OB were similar and within the normal range. Basal ACTH and cortisol levels were similar in all groups. However, the ACTH response to CRH in OSAS (Deltapeak: 30.3 +/- 3.8 pmol/l; DeltaAUC: 682.8 +/- 128.4 pmol*h/l) was markedly higher (P < 0.001) than in OB (Deltapeak: 9.3 +/- 1.4 pmol/l; DeltaAUC 471.5 +/- 97.3 pmol*h/l), which, in turn, was higher (P < 0.05) than in NS (Deltapeak: 3.3 +/- 0.9 pmol/l; DeltaAUC 94.7 +/- 76.7 pmol*h/l). On the other hand, the cortisol response to CRH was not significantly different in the three groups. Basal FT3 and FT4 levels as well as the TSH response to TRH were similar in all groups. Similarly, both basal PRL levels and the PRL response to TRH were similar in the three groups. CONCLUSIONS: With respect to patients with simple abdominal obesity, obese patients with OSAS show a more remarkable enhancement of the ACTH response to CRH but a preserved TSH and PRL responsiveness to TRH. These findings indicate the existence of a peculiarly exaggerated ACTH hyper-responsiveness to CRH that would reflect hypoxia- and/or sleep-induced alterations of the neural control of corticotroph function; this further alteration is coupled to the previously described, peculiar reduction of somatotroph function.     

The GH-releasing effect of ghrelin,a natural GH secretagogue,is only blunted by the infusion of exogenous somatostatin in humans

OBJECTIVE: Ghrelin, a 28-amino-acid peptide purified from the stomach and showing a unique structure with an n-octanoyl ester at the serine 3 residue, is a natural ligand of the GH secretagogue (GHS) receptor (GHS-R). Ghrelin strongly stimulates GH secretion in both animals and humans, showing a synergistic effect with GH-releasing hormone (GHRH) but no interaction with synthetic GHS. However, the activity of ghrelin as well as that of non-natural GHS is not fully specific for GH; ghrelin also induces a stimulatory effect on lactotroph and corticotroph secretion, at least in humans. DESIGN: To further clarify the mechanisms underlying the GH-releasing activity of this natural GHS, we studied the effects of somatostatin (SS, 2.0 microg/kg/h from -30 to +90 min) on the endocrine responses to ghrelin (1.0 microg/kg i.v. at 0 min) in seven normal young male volunteers [age (mean +/- SEM) 28.6 +/- 2.9 years; body mass index (BMI) 22.1 +/- 0.8 kg/m2]. In the same subjects, the effect of SS on the GH response to GHRH (1.0 microm/kg i.v. at 0 min) was also studied. MEASUREMENTS: Blood samples were taken every 15 min from -30 up to +120 min. GH levels were assayed at each time point in all sessions; PRL, ACTH and cortisol levels were assayed after ghrelin administration alone and during SS infusion. RESULTS: The GH response to ghrelin (hAUC0'-->120' 2695.0 +/- 492.6 microg min/l) was higher (P < 0.01) than that after GHRH (757.1 +/- 44.1 microg min/l). SS infusion almost abolished the GH response to GHRH (177.0 +/- 37.7 microg min/l, P < 0.01); the GH response to ghrelin was inhibited by SS (993.8 +/- 248.5 microg min/l, P < 0.01) but GH levels remained higher (P < 0.05) than with GHRH. Ghrelin induced significant increases in PRL, ACTH and cortisol levels and these responses were not modified by SS. CONCLUSIONS: Ghrelin, a natural GHS-R ligand, exerts a strong stimulatory effect on GH secretion in humans and this effect is only blunted by an exogenous somatostatin dose which almost abolishes the GH response to GHRH. The stimulatory effect of ghrelin on lactotroph and corticotroph secretion is refractory to exogenous somatostatin, indicating that these effects occur through pathways independent of somatostatinergic influence.     

The inhibitory effect of alprazolam, a benzodiazepine, overrides the stimulatory effect of metyrapone-induced lack of negative cortisol feedback on corticotroph secretion in humans.

Alprazolam (ALP), a benzodiazepine that activates gamma-aminobutyric acid-ergic receptors, inhibits the activity of hypothalamo-pituitary-adrenal (HPA) axis, probably via inhibition of hypothalamic CRH and/or arginine vasopressin release. To further clarify the effects of ALP on the HPA axis in humans, in six normal young women (26-34 yr old) we studied the effects of 0.02 mg/kg ALP (administered orally at 0700 h) or placebo on ACTH, cortisol (F), and 11-deoxycortisol (S) levels assayed after placebo or metyrapone (MET; 0.04 g/kg administered orally at 2300 h the night before). After placebo administration, ACTH, F, and S levels showed a progressive decrease from 0700-1200 h (P < 0.03). At 0700 h, ACTH, F, and S levels before ALP overlapped with those after placebo. At 1200 h, ACTH, F, and S levels after ALP were lower than those after placebo (P < 0.03). MET pretreatment strongly increased ACTH (P < 0.03) and S (P < 0.02) while clearly inhibiting F (P < 0.03) levels at 0700 h. After MET, ACTH levels did not show any decrease up to 1200 h; similarly, S levels persisted similar up to 1200 h, whereas F levels at 1200 h were significantly increased (P < 0.03). At 0700 h, MET-induced ACTH and F levels before ALP overlapped with those after MET alone. The MET-induced ACTH levels at 1200 h were markedly inhibited by ALP (P < 0.05). At 1200 h after MET and ALP, a clear reduction of S levels (P < 0.02) and an insignificant F reduction were also found. In conclusion, our present data show that ALP inhibits basal and, much more, metyrapone-induced corticotroph secretion. These findings indicate that the inhibitory effect of central gamma-aminobutyric acid-ergic activation by ALP overrides the stimulatory effect of the MET-induced lack of negative F feedback on corticotroph secretion. These results also point toward potential contraindication of ALP administration in patients with suspected hypoadrenalism.     

Hexarelin as a test of pituitary reserve in patients with pituitary disease

BACKGROUND: The insulin tolerance test (ITT) is the reference standard for the diagnosis of cortisol and growth hormone (GH) deficiency, but problems have occurred in small children in inexperienced hands and it is contraindicated in patients with cardiac disease and epilepsy. Hexarelin is a growth hormone-releasing peptide with GH-, ACTH/cortisol- and prolactin-releasing effects which involve both hypothalamic and direct pituitary mechanisms. We therefore investigated whether it could be used to test GH and ACTH/cortisol reserve in patients with pituitary disease. METHODS AND SUBJECTS: The changes in GH and cortisol in response to insulin-induced hypoglycaemia (intravenous human Actrapid 0.15 IU/kg) and hexarelin (2 microg/kg) in 19 patients with possible pituitary disease (5 males, mean age 39 years, range 21-70) were compared. The patients' responses during the hexarelin test were also compared to normal ranges of GH and cortisol responses established in healthy volunteers following hexarelin administration. RESULTS AND DISCUSSION: GH peak levels were significantly higher after hexarelin than after hypoglycaemia (mean +/- SEM; 67.1 +/- 16 vs. 26.9 +/- 6.8 mU/l respectively; P < 0. 001), while cortisol levels were significantly lower (420 +/- 34 vs. 605 +/- 50 nmol/l; P < 0.001). The peak responses of both hormones correlated significantly between the hexarelin and insulin-induced hypoglycaemia tests (r = 0.80, P < 0.001 for cortisol). Peak GH levels after hexarelin and ITT showed a significant positive correlation with IGF-I levels (r = 0.84 and r = 0.77, P < 0.001 for both). All patients with a subnormal GH response to hexarelin (<41.4 mU/l) had a peak GH response to ITT of <9 mU/l, and only one patient had a normal (although borderline) response to hexarelin with a subnormal GH response to the ITT. Although 17 of the 19 patients had corresponding cortisol responses to hexarelin and the ITT test (either failing or passing both), two patients had normal cortisol responses to hexarelin but subnormal responses to the ITT. A peak serum cortisol level following hypoglycaemia of >580 nmol/l is indicative of normal cortisol reserve, as established in patients undergoing surgery; only five of the normal volunteers and one of the thirteen patients with a normal ACTH/cortisol reserve on ITT had a peak cortisol >580 nmol/l in response to hexarelin. CONCLUSION: Adult patients who have a subnormal peak GH response to hexarelin are likely to be GH deficient on an insulin tolerance test. However, our data suggest that the hexarelin test is not a useful test of ACTH/cortisol reserve. The hexarelin test could be a useful first/screening test to diagnose adult GH deficiency, particularly in patients in whom an insulin tolerance test is contraindicated or who are already ACTH deficient and in whom the GH reserve alone is of interest.     

Neuroendocrine and metabolic effects of acute ghrelin administration in human obesity

Ghrelin stimulates appetite and plays a role in the neuroendocrine response to energy balance variations. Ghrelin levels are inversely associated with body mass index (BMI), increased by fasting and decreased by food intake, glucose load, insulin, and somatostatin. Ghrelin levels are reduced in obesity, a condition of hyperinsulinism, reduced GH secretion, and hypothalamus-pituitary-adrenal axis hyperactivity. We studied the endocrine and metabolic response to acute ghrelin administration (1.0 microg/kg i.v.) in nine obese women [OB; BMI (mean +/- SD) 36.3 +/- 2.3 kg/m(2)] and seven normal women (NW; BMI 20.3 +/- 1.7 kg/m(2)). Basal ghrelin levels in NW were higher than in OB (P < 0.05). In NW, ghrelin increased (P < 0.05) GH, prolactin (PRL), ACTH, cortisol, and glucose levels but did not modify insulin. In OB, ghrelin increased (P < 0.01) GH, PRL, ACTH, and cortisol levels. The GH response to ghrelin in OB was 55% lower (P < 0.02) than in NW, whereas the PRL, ACTH, and cortisol responses were similar. In OB, ghrelin increased glucose and reduced insulin (P < 0.05). Thus, obesity shows remarkable reduction of the somatotroph responsiveness to ghrelin, suggesting that ghrelin hyposecretion unlikely explains the impairment of somatotroph function in obesity. On the other hand, in obesity ghrelin shows preserved influence on PRL, ACTH, and insulin secretion as well as in glucose levels.     

Low dose hexarelin and growth hormone (GH)-releasing hormone as a diagnostic tool for the diagnosis of GH deficiency in adults: comparison with insulin-induced hypoglycemia test.

GH deficiency (GHD) in adults must be shown by provocative testing of GH secretion. Insulin-induced hypoglycemia (ITT) is the test of choice, and severe GHD, treated with recombinant human GH replacement, is defined by a GH peak response to ITT of less than 3 microg/L. GHRH plus arginine (ARG) is a more provocative test and is as sensitive as ITT provided that appropriate cut-off limits are assumed. GH secretagogues are a family of peptidyl and nonpeptidyl GH-releasing molecules that strongly stimulate GH secretion and, even at low doses, truly synergize with GHRH. Our aim was to verify the diagnostic reliability of the hexarelin (HEX; 0.25 microg/kg, iv) and GHRH (1 microg/kg, iv) test for the diagnosis of adult GHD. To this goal, in the present study we 1) defined the normal ranges of the GH response to GHRH+HEX in a group of normal young adult volunteers (NS; n = 25; 18 men and 7 women; age, 28.5+/-0.6 yr) and in 11 of them verified its reproducibility in a second session, and 2) compared the GH response to GHRH+HEXwith that to ITT in a group of normal subjects (n = 33; 12 men and 21 women; age, 34.1+/-1.5 yr) and hypopituitaric adults with GHD (n = 19; 10 men and 9 women; age, 39.9+/-2.2 yr; GH peak <5 microg/L after ITT). The GH response to GHRH+ARG was also evaluated in all GHD and in 77 normal subjects (40 men and 37 women; age, 28.1+/-0.6 yr). The mean GH peak after GHRH+HEX in NS was 83.6+/-4.5 microg/L; the third and first percentile limits of the normal GH response were 55.5 and 51.2 microg/L, respectively). The GH response to GHRH+HEX in NS showed good intraindividual reproducibility. In GHD the mean GH peak after GHRH+HEX (2.6+/-0.7 microg/L) was similar to that after GHRH+ARG (3.6+/-1.0 microg/L), and both were higher (P < 0.001) than that after ITT (0.6+/-0.1 microg/L); the GH responses to GHRH+HEX were positively associated with those to ITT and GHRH+ARG. Analyzing individual GH responses, 100% had severe GHD after ITT (GH peak, <3 microg/L). After GHRH+HEX all GHD had GH peaks below the third percentile limit of normality appropriate for this test (i.e. 55.5 microg/L). Thirteen of 19 (68.4%) GHD subjects had GH peaks below 3 microg/L after GHRH+HEX but all 19 (100%) had GH peaks below the first percentile limit of normality (i.e. 51.2 microg/L). The GH responses to GHRH+HEX were highly concordant with those after GHRH+ARG. In conclusion, the present results define normal limits of the GH response to stimulation with low dose HEX+GHRH in normal adults and show that this test is as sensitive as ITT for the diagnosis of adult GHD provided that appropriate cut-off limits are considered.     

Endocrine responses to ghrelin in adult patients with isolated childhood-onset growth hormone deficiency

OBJECTIVE: Ghrelin, a 28 amino acid acylated peptide, is a natural ligand of the GH secretagogues (GHS) receptor (GHS-R), which is specific for synthetic GHS. Similar to synthetic GHS, ghrelin strongly stimulates GH secretion but also displays significant stimulatory effects on lactotroph and corticotroph secretion. It has been hypothesized that isolated GH deficiency (GHD) could reflect hypothalamic impairment that would theoretically involve defect in ghrelin activity. PATIENTS: In the present study, we verified the effects of ghrelin (1 microg/kg i.v.) on GH, PRL, ACTH and cortisol levels in adult patients with isolated severe GHD [five males and one female, age (mean +/- SEM) 24.7 +/- 2.6 years, BMI 25.7 +/- 2.7 kg/m2]. In all patients, the GH response to insulin-induced hypoglycaemia (ITT, 0.1 IU regular insulin i.v.) and GH releasing hormone (GHRH) (1 microg/kg i.v.) + arginine (ARG, 0.5 g/kg i.v.) was also studied. The hormonal responses in GHD were compared with those in age-matched normal subjects (NS, seven males, age 28.6 +/- 2.9 years, BMI 22.1 +/- 0.8 kg/m2). RESULTS: IGF-I levels in GHD were markedly lower than in NS (69.8 +/- 11.3 vs. 167.9 +/- 19.2 microg/l, P < 0.003). Ghrelin administration induced significant increase in GH, PRL, ACTH and cortisol levels in all GHD. In GHD, the GH response to ghrelin was higher (P < 0.05) than that to GHRH + ARG, which, in turn, was higher (P < 0.05) than that to ITT (9.2 +/- 4.1 vs. 5.3 +/- 1.7 vs. 1.4 +/- 0.4 microg/l). These GH (1 microg/l = 2 mU/l) responses in GHD were markedly lower (P < 0.0001) than those in NS (ghrelin vs. GHRH + ARG vs. ITT 92.1 +/- 16.7 vs. 65.3 +/- 8.9 vs. 17.7 +/- 3.5 microg/l). In GHD, the highest individual peak GH response to ghrelin was markedly lower than the lowest peak GH response in NS (28.5 vs. 42.9 microg/l). GHD and NS showed overlapping PRL (1 microg/l = 32 mU/l) (10.0 +/- 1.4 vs. 14.9 +/- 2.2 microg/l), ACTH (22.3 +/- 5.3 vs. 18.7 +/- 4.6 pmol/l) and cortisol responses (598.1 +/- 52.4 vs. 486.9 +/- 38.9 nmol/l). CONCLUSIONS: This study shows that ghrelin is one of the most powerful provocative stimuli of GH secretion, even in those patients with isolated severe GHD. In this condition, however, the somatotroph response is markedly reduced while the lactotroph and corticotroph responsiveness to ghrelin is fully preserved, indicating that this endocrine activity is fully independent of mechanisms underlying the GH-releasing effect. These results do not support the hypothesis that ghrelin deficiency is a major cause of isolated GH deficiency but suggest that ghrelin might represent a reliable provocative test to evaluate the maximal GH secretory capacity provided that appropriate cut-off limits are assumed.