GHRH Bolus Research Articles

Effects of propranolol on GH responsiveness to repeated GH-releasing hormone stimulations in normal subjects

The influence of beta-adrenergic blockade by oral propranolol on the variability of GH responses to GHRH and on GH responsiveness to repeated GHRH administrations was investigated. Eight normal volunteers underwent three tests on three separate occasions. Each test consisted of two administrations of 80 micrograms GHRH at 2-h intervals without other medication (test 1) or combined with oral administration of 80 mg propranolol 90 min before the first (test 2) or the second GHRH injection (test 3). In test 1 GH levels increased significantly after the first, but not the second GHRH bolus (net incremental area under the curve [nAUC], mean +/- SD: 1453 +/- 974 and 178 +/- 309 micrograms.l-1.(120 min)-1, respectively). In test 2 basal GH secretion was not influenced by propranolol administration, whereas the GH response to the first GHRH injection was significantly greater than in test 1 (2327 +/- 1814 micrograms.l-1.(120 min)-1; p less than 0.05). However, individual subjects showed the same variability of GH response as in test 1. The GH response to the second GHRH bolus remained negligible. In test 3 administration of propranolol 90 min before the second GHRH bolus led to a clear GH increase (690 +/- 1002 micrograms.l-1.(120 min)-1), not significantly different from the GH response to the first bolus (1796 +/- 1375 micrograms.l-1.(120 min)-1). However, only 4 subjects showed a marked restoration of the GH responsiveness to the second GHRH administration. In conclusion, oral administration of propranolol is able to increase GH responsiveness to GHRH without changing the great individual variability. The response to a repeated GHRH stimulation is only partially restored by propranolol.

GHRH-test in short children with “non classic” GH deficiency. A comparison with “classic” GH deficiency and short normal stature

In this study GHRH-test has been performed (2 micrograms/Kg of an iv bolus of GHRH 1-44) sampling for GH measurement every 15 min over 2 hours in three groups of short children. Group 1 consisted of 10 subjects with classic GH deficiency (CGHD): GH response less than 10 ng/ml to two conventional tests and 24-h mean GH concentration (MGHC) less than 3 ng/ml; group 2 consisted of 16 subjects with non-classic GH deficiency (NCGHD): response greater than 10 ng/ml to at least one conventional test and MGHC less than 3 ng/ml; group 3 consisted of 18 subjects with short normal stature: GH response greater than 10 ng/ml to at least one conventional test and MGHC greater than 3 ng/ml. GH peak and area under the curve (AUC) values were significantly lower in group 1 than groups 2 and 3 and in group 2 than group 3. GH peak and AUC values statistically correlated with height, height velocity, bone age/chronological age ratio and MGHC. Six children in group 1, 14 children in group 2 and all 18 children in group 3 showed after GHRH a GH peak greater than 10 ng/ml and were considered as 'responders'. Considering only the responders, GH peak and AUC values were significantly lower in group 1 than groups 2 and 3 and in group 2 than group 3. In conclusion, our data have shown that 87% of children with NCGHD responded to a single bolus of GHRH with an increase in GH levels greater than 10 ng/ml and that their responses were intermediate compared to those of CGHD and short normal subjects.

EFFECT OF ORAL GLUCOSE ON THE LATE GROWTH HORMONE RISE AND GROWTH HORMONE RESPONSES TO GHRH IN NORMAL SUBJECTS

A late rise in serum GH occurs 3-5 h following oral glucose in man. In order to investigate the mechanisms through which this occurs we have studied the late GH rise after oral glucose during administration of a supramaximal dose of GHRH. In eight normal subjects, oral glucose (100 g) greatly enhanced the GH responses to a supramaximal dose of GHRH (50 micrograms bolus, followed immediately by 100 micrograms/h infusion for 3 h) given 3.5 h after the glucose. GH peak (mean +/- SEM) elicited by GHRH (bolus + infusion) rose from 55.2 +/- 20.4 to 133.4 +/- 29.6 mU/l (P less than 0.02) after glucose pretreatment. In conclusion, it is likely that the late rise in GH secretion induced by oral glucose occurs via a non-GHRH-dependent mechanism. These data are consistent with the hypothesis that the delayed GH response to glucose is a consequence of reduced release of somatostatin from the hypothalamus.

CONTINUOUS SUBCUTANEOUS GHRH(1‐29)NH2 PROMOTES GROWTH OVER 1 YEAR IN SHORT, SLOWLY GROWING CHILDREN

We have treated eight pre-pubertal children with partial GH insufficiency with continuous subcutaneous infusions of GHRH(1-29)NH2 at a dose of 60 ng/kg/min for periods of up to 1 year. In five children treated for 1 year, mean growth velocity increased from 4.6 cm/year (range 4.4-5.2) to 7.0 cm/year (5.7-8.7) (P = 0.04). Three children treated for 3-6 months showed similar height velocity increases. A return to pretreatment growth rates was seen after cessation of treatment in all children. Twenty-four-hour GH profiles performed at intervals of 3 months showed sustained augmentation of pulsatile GH secretion without evidence of desensitization. The presence of pulsatile GH secretion during continuous GHRH administration provides strong evidence in man for the role of somatostatin in determining GH pulse frequency. The ability of the pituitary to respond to a supramaximal bolus of GHRH remained constant during the treatment. Continuous administration of GHRH(1-29)NH2 will become a practicable treatment when formulated into a sustained release or depot preparation. We have shown this to be an effective therapy for some short, slowly growing children. Further studies are required to establish the optimal dosage regimen.

Repeated GH-releasing hormone administration unravels different GH secretory patterns in normal adults and children

In normal adults, repeated GHRH administration leads to progressively decreasing somatotrope responses. To verify whether this GH secretory pattern also connotes normal growing children, we have studied the effects of two consecutive (every 120 min) 1 microgram/kg iv GHRH boluses on GH release in normal adults (N = 7, age 23.2-30.6 years) children (N = 6, age 10.4-13.2 years). In the adults, the GH response to the second GHRH bolus (peak, mean +/- SEM; 2.9 +/- 0.8 micrograms/l) was lower (P less than 0.02) than that to the first bolus (15.9 +/- 2.4 micrograms/l). Conversely, in children the GH response to the second GHRH bolus (25.6 +/- 6.3 micrograms/l) overrode the first one (13.6 +/- 6.5 micrograms/l), but this difference did not attain statistical significance. In adults cholinergic enhancement by pyridostigmine, a cholinesterase inhibitor, was previously shown to re-instate, even to potentiate somatotrope responsiveness to consecutive GHRH boluses. Thus, in 5 children GH response to repeated GHRH boluses was retested administering pyridostigmine (60 mg orally) 30 min before the second GHRH bolus. In these subjects, pyridostigmine failed significantly to potentiate the GH responsiveness to the second GHRH bolus (30.3 +/- 4.6 vs 25.0 +/- 7.6 micrograms/l). These data indicate that differently from in adults, in children repeated GHRH administration does not reduce somatotrope responsiveness and that cholinergic enhancement fails to potentiate GH responsiveness to the second GHRH bolus.

INTERACTION OF CLONIDINE AND GHRH ON GH SECRETION IN VIVO AND IN VITRO

To investigate the mechanism by which clonidine stimulates GH-secretion in vivo and in vitro, we studied its interaction with GHRH. In vivo: eight or six normal male subjects were submitted to five protocols: (1) 150 micrograms clonidine orally followed by 50 micrograms GHRH 1-44 i.v. 2 h later, (2) 50 micrograms GHRH 1-44 i.v. followed by 150 micrograms clonidine orally 2 h later, (3) 150 micrograms clonidine orally followed by GHRH i.v. 30 min later, (4) 300 micrograms clonidine orally followed by 50 micrograms GHRH i.v. 3 h later and (5) 50 micrograms GHRH i.v. followed by 300 micrograms clonidine orally 90 min later. In vitro: Rat anterior pituitary cells were coincubated with clonidine (10(-11), 10(-9), 10(-7) and 10(-5) M) and GHRH (0.005, 0.05, 10 nM) for 4 h. 150 micrograms clonidine alone does not stimulate GH-secretion. Furthermore, the GH-increase was not significantly different when GHRH bolus was given before, after or together with clonidine. When 300 micrograms clonidine was given before GHRH GH-levels were significantly higher (max 28.6 +/- 8.0 mU/l) at 90 min, compared to when clonidine was given after GHRH (max 7.8 +/- 3.6 mU/l). The GHRH bolus after clonidine led to a significantly lower GH-increase (max 31.6 +/- 17.0 mU/l) compared to the GHRH-induced GH-increase (max 47.2 +/- 13.0 mU/l) before clonidine. In vitro, clonidine had no stimulatory effect on GHRH-stimulated GH secretion. These findings are compatible with clonidine leading to stimulation of GH by inducing endogenous GHRH release.

Different GH responsiveness to repeated GHRH administration in normal children and adults.

It has been previously reported that repeated GHRH administration leads to a reduction in growth hormone responses in adults. In order to verify if this regulatory mechanism is also operating in children, we compared GH secretory responses to repeated GHRH in normal children and adults. The administration of the first GHRH bolus caused a marked GH rise (expressed as net incremental area under the curve, nAUC +/- SE) both in children (GH nAUC: 1184 +/- 216 ng/ml/h) and in adults (GH nAUC: 701.2 +/- 90 ng/ml/h). The second GHRH bolus induced a different pattern of GH secretion in the two groups. In children the peptide determined a clear GH rise (GH nAUC: 518 +/- 147 ng/ml/h), while in adults the second GHRH injection was uneffective (GH nAUC: 21 +/- 20 ng/ml/h). These preliminary results indicate that in children repeated GHRH administration does not induce refractoriness to GHRH action, suggesting different regulatory mechanism of GH secretion in normal children and adults.

HGH RESPONSE TO REPEATED GH-RH 1-44 ADMINISTRATION IN HYPOPITUITARY CHILDREN

The aim of the present investigation was to determine the effectiveness of 1 daily s.c. injection for 5 consecutive days of GHRH 1-44 (1 mcg/Kg) on the plasma hGH response to an i.v. bolus of GH-RH 1-44 (1 mcg/Kg) in hypopituitary patients who had a subnormal response of plasma hGH to previously performed tests: ITT, clonidine, sleep. The i.v. GH-RH test was performed before (I) and day after (II) the last i.m. injection.Eleven children (7 IGHD, 4 HPHD) were studied. According to the results the patients were sub-divided into 3 groups:In GR I 3 girls were in puberty and thepeak response of hGH to other tests was 5.1±1.9. In GR II 4 boys were prepubertal, had a lower peak of hGH in other tests 2.3±0.9, whereas in GR III there was no rise in any test. This investigation indicates a) that repeated administration of GH-RH even in small doses is useful in the differentiation of patients who may benefit from prolonged therapy; b) that hGH responsiveness to GH-RH may be influenced by sex hormones.

Continuous infusion of growth hormone releasing hormone does not alter the growth hormone response to different stress stimuli.

Continuous infusion of GHRH, as well as a bolus injection of GHRH, specifically stimulates the release of GH by the anterior pituitary. However, repetitive bolus injections of GHRH result in diminished responses of GH, while a constant infusion of GHRH cannot maintain high serum GH levels. To investigate whether different GH-releasing stimuli are able further to challenge the somatotroph being exposed to continuous infusion of GHRH, we determined the GH response to insulin-induced hypoglycaemia and bicycle exercise. Six normal volunteers received a bolus of 50 micrograms GHRH or vehicle followed by a continuous infusions of GHRH 1-29 amide (1 microgram/kg/h) or vehicle for 2.5 h. GHRH bolus and infusion resulted in elevated GH levels, but GH levels fell to values not significantly different from baseline levels after 150 min, GH plasma levels rose again, however, in response to insulin hypoglycaemia and bicycle exercise after both GHRH or vehicle infusion. Thus, the somatotroph's responsiveness to GHRH remains intact to respond to stress stimuli after continuous GHRH infusion.

Growth Hormone Responses to Continuous Infusions of Growth Hormone-Releasing Hormone*

The pattern of GH secretion during a continuous 4-h iv infusion of 1 microgram/kg.h GH-releasing hormone (1-44)-NH2 (GHRH-44) or saline was examined in 15 adult men. There was prompt release of GH beginning within 20 min of starting the GHRH-44 infusions, reaching peak GH levels of 43 +/- 11 (+/- SE) ng/ml within 60-90 min. This is similar to the peak GH level reached in men after a single 1 microgram/kg GHRH iv bolus dose (34 +/- 8 ng/ml). GH levels then fell progressively, but did not return to baseline during the GHRH infusions. After GHRH infusions, the response (delta) to a 1 microgram/kg GHRH bolus dose was markedly attenuated (delta GH, 2.7 +/- 0.9 ng/ml) compared to the response (delta GH, 23 +/- 3 ng/ml) after saline infusion. Dispersed rat pituicytes perifused with medium containing 10 nM GHRH-44 responded with an initial rapid rise in GH secretion, followed by a progressive decline, and after 150 min of continuous GHRH exposure, the response to pulses of an equal or higher (100 nM) GHRH concentration was blunted. These results indicate that the peak response to GHRH infusions is similar to that of maximally effective bolus doses; during infusions, the GH response is not sustained; and immediately after GHRH infusions, the response to previously effective bolus doses is reduced. These phenomena could reflect either receptor-mediated desensitization, the depletion of rapidly releasable GH stores, or both. A counterregulatory rise in hypothalamic somatostatin secretion is not necessary to produce these effects, since the same phenomenon occurs in vitro and in vivo.

Classification of patients with GH deficiency (GHD) according to the response to GH-RH

Synthetic GH-RH-1-44 was administered in a dose of 1 mcg/1 Kg to 3 normal adolescents with short stature and to 17 patients with GHD (hGH response to ITT and clonidine < 2 ng/ml). In the normal subjects the peak response of plasma hGH to GH-RH was 100, 20 and 19 ng/ml. In the GHD patients 3 patterns of response were observed: a) Good response (n=5), 18±1.9 ng/ml, i.e. GHD of hypothalamic origin; b) Partial response (n=4), 4.25±0.4 ng/ml, i.e. GHD of hypothalamic origin, possibly with a pituitary component; c) No response (n=8), 1.3±0.3 ng/ml, i.e. GHD of pituitary origin. It remains to be established whether a partial response of GH to one bolus of GH-RH may in certain patients be due to a longstanding GH-RH deficiency, with the possibility of its unmasking following repeated injections and the consequent synthesis of sufficient GH for its release.