Acute clonidine administration potentiates spontaneous diurnal, but not nocturnal, growth hormone secretion in normal short children.

It has been shown that alpha 2-adrenoreceptor activation induced by clonidine (CLON) increases plasma GH levels in both adults and children. In this study the effects of CLON (150 micrograms/m2, orally) on GH secretion were studied both in the morning (from 0800-1100 h) and at night (from 2300-0200 h) in nine short children previously shown to have normal spontaneous nocturnal GH secretion. In the morning, CLON induced a GH increase higher than placebo [peak (mean +/- SEM), 23.8 +/- 4.3 vs. 3.4 +/- 1.4 micrograms/L; P = 0.0001; area under curve (AUC), 624.4 +/- 62.7 vs. 135.6 +/- 33.3 micrograms/L.h; P less than 0.00001]. In the night, no difference was observed between GH secretion after CLON (peak, 15.4 +/- 3.2 micrograms/L; AUC, 562.2 +/- 57.5 micrograms/L.h) and placebo (peak, 13.1 +/- 4.7 micrograms/L; AUC, 497.2 +/- 83.5 micrograms/L.h). Spontaneous GH secretion was higher during the night than in the morning (P = 0.0001), whereas nocturnal GH secretion overlapped with that in the morning after CLON. The data presented show that alpha 2-adrenoreceptor activation is probably mediated by increased endogenous GHRH release; our results suggest that the endogenous GHRH secretion is maximally stimulated at night.

Enhancement of cholinergic tone by pyridostigmine promotes both basal and growth hormone (GH)-releasing hormone-induced GH secretion in children of short stature.

Increased cholinergic tone induced by pyridostigmine (PD) increases basal plasma GH levels and potentiates the GH response to GHRH in normal adults. In this study the effects of PD (60 mg, orally) on both basal and GHRH (1 microgram/kg)-induced GH secretion in seven children with familial short stature (FSS), six with GH deficiency (GHD) and 10 with constitutional growth delay (CGD) were studied and compared with results obtained by stimulation with insulin-induced hypoglycemia (IH) and GHRH alone. The mean peak plasma GH levels were variable, but individual values were frequently low in all groups after both IH [FSS, 9.7 +/- 1.3 (+/- SEM) ng/mL; GHD, 1.6 +/- 0.4 ng/mL; CGD, 7.0 +/- 0.8 ng/mL] and GHRH (FSS, 23.8 +/- 6.6 ng/mL; GHD, 11.1 +/- 5.8 ng/mL; CGD, 15.1 +/- 4.5 ng/mL) administration. PD induced GH responses (FSS, 14.5 +/- 1.6 ng/mL; GHD, 3.8 +/- 0.8 ng/mL; CGD, 18.3 +/- 3.2 ng/mL) that in many children in the FSS and CGD groups were higher than those after IH and GHRH treatment. PD clearly increased the GH response to GHRH in all children [FSS, 69.5 +/- 9.4 ng/mL (P less than 0.01 vs. other stimuli); GHD, 18.0 +/- 7.5 ng/mL; CGD, 50.0 +/- 8.5 ng/mL (P less than 0.01 vs. other stimuli)]. We conclude that in children with short stature, as in adults, enhancement of cholinergic tone increases both basal and GHRH-induced GH secretion, and that PD plus GHRH is the best provocative stimulus for evaluating the somatotroph response.

Alpha 1-blocker doxazosin improves peripheral insulin sensitivity in diabetic hypertensive patients.

The antihypertensive doxazosin is a selective alpha 1-adrenoceptor-blocking drug whose favorable impact on lipid metabolism is well known. A single-blind placebo-controlled crossover study was designed to determine whether antihypertensive treatment with doxazosin affects insulin sensitivity in diabetic, mildly hypertensive, non-obese patients. Twelve subjects (diastolic blood pressure, 98 +/- 1.5 mm Hg; body mass index, 25 +/- 0.6 kg/m2; hemoglobin A1c [HbA1c], 7.6% +/- 0.4%) who were not taking drugs and were treating diabetes only by diet were randomly assigned to placebo treatment for 6 weeks and then to doxazosin for the same period, or vice versa. The doxazosin dose (maximum, 12 mg/d) was increased to achieve a normotensive blood pressure (final diastolic pressure, 85 +/- 2 mm Hg, P < .05). A euglycemic (100 +/- 4 mg/dL) hyperinsulinemic (61 +/- 6 microU/mL) glucose clamp was performed at baseline and at the end of both placebo and doxazosin administration. Hepatic glucose production was measured by the isotope dilution technique using 3H-glucose. Body weights and HbA1c did not vary during the entire study. The basal mean glucose uptake and the insulin sensitivity index (2.3 +/- 0.3 mg/kg/min and 4 +/- 0.5 mg/kg/min per U/L x 100) remained unchanged during placebo administration (2.5 +/- 0.4 and 4 +/- 0.6, NS), but significantly increased during doxazosin treatment (3.3 +/- 0.4 and 5.6 +/- 0.7, P < .05). Hepatic glucose production showed no modification during both placebo and doxazosin. These data provide evidence that doxazosin improves insulin sensitivity in diabetic hypertensive patients, mainly through peripheral effects.

Effect of cholinergic enhancement by pyridostigmine on growth hormone secretion in obese adults and children.

In obesity the reduced growth hormone (GH) responses to several provocative stimuli including growth hormone-releasing hormone (GHRH) indicate a diminished somatotroph responsiveness but do not distinguish between primary pituitary and hypothalamic pathogenesis. However, it has been shown that the cholinergic system positively influences Gh secretion likely by modulating somatostatin release in a negative way. Thus, the effect of cholinergic activity enhancement by pyridostigmine (PD), an acetylcholinesterase inhibitor, on both basal and GHRH-induced GH secretion was studied in 14 obese subjects (eight adults and six children). Eighteen nonobese subjects (seven adults and 11 children) were studied as controls. In obese subjects the GHRH-induced GH increase was lower than in controls (peak, mean +/- SEM, adults, 9.2 +/- 2.7 v 16.8 +/- 5.7 ng/mL; children, 8.0 +/- 0.8 v 20.3 +/- 4.6 ng/mL) attaining statistical significance only in children group (P less than .02). The PD-induced GH response in the two obese groups was similar to that observed in relative controls (adults, 5.3 +/- 1.0 v 7.4 +/- 1.7 ng/mL; children, 9.6 +/- 1.6 v 13.3 +/- 1.4 ng/mL). PD clearly potentiated the GH response to GHRH in obese subjects, both adults (P less than .05 v GHRH alone) and children (P less than .0005 v GHRH alone). However, the GH responses to PD + GHRH was significantly reduced in obese subjects compared with controls (adults, 18.1 +/- 2.2 v 42.7 +/- 10.7 ng/mL, P less than .05; children, 28.3 +/- 4.5 v 58.2 +/- 7.7 ng/mL, P less than .01). In conclusion, PD is able to potentiate the blunted GH responses to GHRH in obese adults and children, inducing a GH increase similar to that observed after GHRH alone in normal subjects. This finding suggests that an alteration of somatostatinergic tone could be involved in the reduced GH secretion in obesity.

Growth hormone responses to pyridostigmine in normal adults and in normal and short children.

There is evidence indicating that the cholinergic system positively modulates GH release probably by inhibiting somatostatinergic tone. In the present study, the effects of cholinergic enhancement by pyridostigmine, (PD), a cholinesterases inhibitor, on GH release in normal adults (n = 14) (NA) and in both normal (n = 5) (NC) and short children (n = 19) (SC) with familial short stature (n = 7) or constitutional growth delay (n = 12) were studied. In SC the insulin hypoglycaemia (IH)-induced GH increase was also studied. In both NC and SC 60 mg orally PD induced a significant GH increase with mean peak at 90 min (mean +/- SEM 11.0 +/- 2.2 ng/ml in NC and 11.2 +/- 2.3 ng/ml in SC). The GH areas under response curve (AUC) were 379.3 +/- 76.6 and 327.8 +/- 43.2 ng/ml/h in NC and SC respectively. In NA 120 mg orally PD induced a significant GH increase with mean peak at 120 min (5.1 +/- 1.1 ng/ml) which was significantly lower (P less than 0.05) than that observed in both NC and SC. This statistical difference was strengthened by evaluating AUC (NA:205.6 +/- 33.7 ng/ml/h, P less than 0.05 vs NC and SC). The correlation of drug dosage with body area ruled out that this difference could be related to the different PD dose in adults and children. In SC, IH induced a GH increase significantly lower than that observed after PD (GH peak 7.8 +/- 0.6 vs 16.4 +/- 1.9 ng/ml P less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Cholinergic enhancement of pyridostigmine potentiates spontaneous diurnal but not nocturnal growth hormone secretion in short children.

It has been shown that enhanced cholinergic tone induced by pyridostigmine (PD) increases both basal and GHRH-stimulated GH levels in both adults and children. In this study the effects of PD (60 mg orally) on GH secretion were studied both in the morning (from 8.00 to 12.00) and in the night (from 23.00 to 3.00) in 7 short children previously shown as having a normal spontaneous nocturnal GH secretion. In the morning, PD induced a GH increase higher than saline (peak, mean +/- SEM: 17.4 +/- 3.4 vs. 5.5 +/- 3.0 ng/ml, p less than 0.02; area under curve (AUC): 360.8 +/- 71.4 vs. 109.4 +/- 44.7 ng/ml/h, p less than 0.01). In the night, no difference was observed between GH secretion after PD (peak: 16.7 +/- 2.4 ng/ml; AUC: 468.2 +/- 95.5 ng/ml/h) and saline (peak: 16.0 +/- 2.7 ng/ml; AUC: 409.1 +/- 97.7 ng/ml/h). Spontaneous GH secretion was higher during the night than in the morning (p less than 0.02) whereas nocturnal GH secretion overlapped with that in the morning after PD. The ability of PD to increase GH secretion during the morning but not GH hypersecretion occurring at night implies that the cholinergic tone in the central nervous system areas controlling GH secretion is already maximally stimulated at night. Since, reportedly, the cholinergic system negatively modulates somatostatin secretion, presence of a physiologically reduced somatostatinergic tone may be envisaged at night.

A neuroendocrinological approach to evidence an impairment of central cholinergic function in aging.

A hypothalamic pathogenesis for the reduced GH secretion in aging has been reported for both animal and man. To further address this issue we studied in 31 elderly normal subjects (6 males and 25 females, aged 66-90 yr) and in 22 young healthy controls (13 males and 9 females, aged 20-35 yr) the GH responses to GHRH test (GHRH29, 1 microgram/kg i.v. as a bolus at 0 min) alone and combined with pyridostigmine, a cholinesterase inhibitor (PD, 120 mg po 60 min before GHRH), or with arginine (ARG, 30 g in 100 ml infused from 0 to 30 min). Serum IGF-I levels were lower in elderly than in young subjects (mean +/- SE: 86.9 +/- 7.2 vs 288.7 +/- 22.1 micrograms/L, p < 0.01). The GHRH-induced GH increase was lower in elderly than in young subjects (p < 0.01). PD increased the GH response to GHRH in both groups (p < 0.001), but in elderly subjects this response persisted lower (p < 0.0001) than that observed in young adults. Also ARG coadministration potentiated the GHRH-induced GH release in both groups (p < 0.0001) but in this case the elderly's responses overlapped with the young's. The GH increase observed after combined administration of ARG and GHRH was higher (p < 0.0001) than that elicited by PD plus GHRH in elderly but not in young subjects. Analyzing individual GH responses, a GH peak below the limit of normality for young adults was observed in 19 (61.3%) elderly subjects after PD plus GHRH administration while ARG plus GHRH test elicited a normal GH peak in all but one.(ABSTRACT TRUNCATED AT 250 WORDS)

Pyridostigmine potentiates L-dopa- but not arginine- and galanin-induced growth hormone secretion in children.

The coadministration of growth hormone (GH) secretagogues can provide insight into the neuroregulation of GH secretion. The GH response to L-dopa (125, 250 and 500 mg orally for body weights less than 15 kg, between 15 and 30 kg and greater than 30 kg, respectively), arginine (Arg; 0.5 g/kg infused intravenously over 30 min) and galanin (GAL; 15 micrograms/kg infused intravenously over 60 min) when administered alone or combined with pyridostigmine (PD; 60 mg orally), a cholinergic agonist that likely acts via inhibition of endogenous somatostatin secretion, was studied in children with familial short stature. The GH-releasing effect of PD was also evaluated. In 8 children, PD and L-dopa when administered alone induced an equivalent GH rise (area under the response curve, mean +/- SEM: 241.4 +/- 31.1 vs. 202.9 +/- 38.6 micrograms/l/h) while their coadministration had an additive effect (435.4 +/- 41.4 micrograms/l/h; p less than 0.02 vs. PD and L-dopa alone). On the contrary, in other 8 children, PD and Arg induced similar GH increases either when administered alone (394.2 +/- 68.5 vs. 405.8 +/- 103.9 micrograms/l/h) or in combination (535.8 +/- 97.3 micrograms/l/h). GH increases almost superimposable were also observed when PD and GAL were administered alone (405.2 +/- 72.3 vs. 412.6 +/- 94.1 micrograms/l/h) or in combination (537.9 +/- 139.0 micrograms/l/h) in other 7 children. These data show that the enhancement of the cholinergic activity by PD increases the L-dopa-induced GH release but fails to modify both Arg- and GAL-induced GH release in short children.(ABSTRACT TRUNCATED AT 250 WORDS)

Arginine potentiates the GHRH- but not the pyridostigmine-induced GH secretion in normal short children. Further evidence for a somatostatin suppressing effect of arginine.

To investigate the mechanism underlying the GH-releasing effect of arginine (ARG), we studied the interactions of ARG (0.5 g/kg infused i.v. over 30 min) with GHRH (1 microgram/kg i.v.) and with pyridostigmine (PD, 60 mg orally) on GH secretion in 15 children and adolescents with familial short stature (5.1-15.4 years). In a group of eight subjects ARG induced a GH increase not statistically different to that observed after GHRH (peak, mean +/- SEM: 38.0 +/- 10.4 vs 64.0 +/- 14.4 mU/l). The combined administration of ARG and GHRH led to GH levels (101 +/- 15.2 mU/l) higher than those observed after GHRH (P less than 0.025) or ARG alone (P less than 0.001) and overlapping with those recorded after combined PD and GHRH administration (111 +/- 22.4 mU/l). In the other seven subjects, ARG and PD administration induced a similar GH response either when administered alone (25.2 +/- 13.6 and 27.8 +/- 4.0 mU/l, respectively) or in combination (33.8 +/- 5.4 mU/l). In conclusion, our results show that in children ARG administration potentiates GHRH- but not PD-induced GH increase. These findings agree with the hypothesis that the GH-releasing effect of both ARG and PD is mediated via the same mechanism, namely, by suppression of endogeneous somatostatin release. Combined administration of either ARG or PD with GHRH has a similar striking GH-releasing effect which is clearly higher than that of GHRH alone.

Acute administration of pyridostigmine and clonidine has an additive stimulatory effect on GH release in normal children.

It has been shown in humans that both alpha 2-adrenoceptor activation by clonidine (CLON) and cholinergic enhancement by pyridostigmine (PD) have a clear-cut stimulatory effect on GH release. As this effect is probably mediated by two different mechanisms, i.e. via increased endogenous GHRH for CLON and via inhibition of endogenous somatostatin for PD, in 8 normal children we studied the effect of both single and combined acute oral administration of CLON (150 micrograms/m2) and PD (60 mg). When administered alone, CLON and PD induced a similar GH increase (peak, mean +/- SE: 14.6 +/- 2.4 vs 14.2 +/- 3.1 ng/ml; area under curve, AUC: 376.9 +/- 57.6 vs 390.0 +/- 74.3 ng/ml/h). Combined administration of CLON and PD had an additive effect on GH release (peak: 27.5 +/- 4.5 ng/ml; AUC: 920.8 +/- 153.3 ng/ml/h; p less than 0.005 vs CLON and PD alone). In conclusion, presented data show that: i) CLON and PD have similar GH-releasing effect in normal children; ii) The additive stimulatory effect on GH release exerted by acute combined administration of CLON and PD agrees with the hypothesized different mechanism of action of these two drugs; iii) A therapeutic association of CLON and PD may be envisaged in the treatment of some children of short stature.

A new test for the diagnosis of growth hormone deficiency due to primary pituitary impairment: combined administration of pyridostigmine and growth hormone-releasing hormone.

The diagnosis of growth hormone (GH) deficiency (GHD) is currently based on failure to increase plasma GH levels to an arbitrary cutoff point of 7 or 10 micrograms/l in response to two provocative stimuli. False negative responses to these tests, however, frequently occur thus reducing their diagnostic reliability. The aim of this study was to assess a combination of pyridostigmine (PD) and GH-releasing hormone (GHRH) (60 mg oral PD 60 min before 1 microgram/Kg GHRH iv) as a reliable test probing pituitary somatotropic function. In fact PD, an acetylcholinesterase inhibitor, strikingly potentiates GH response to GHRH likely by inhibiting somatostatin release. The combination PD + GHRH was tested in normal children and adolescents (NS, n = 27) and in a large group of short children classified as having familial short stature (FSS, n = 24), constitutional growth delay (CGD, n = 34) and GH deficiency (organic, oGHD, n = 6; idiopathic, iGHD, n = 10). In all groups results obtained by PD + GHRH were compared with those obtained by testing with GHRH, clonidine (CLON) and PD alone and by studying spontaneous nocturnal GH secretion over 8 hours. Assuming 7 micrograms/l as minimum normal GH peak, a positive response occurred in only 18/24, 11/12 and 12/13 NS for GHRH, CLON, and PD, respectively. In contrast even assuming a minimum normal GH peak as high as 20 micrograms/l, PD + GHRH induced a positive response in 27/27 NS all having a nocturnal GH mean concentration (MC) greater than or equal to 3 micrograms/l. Therefore PD + GHRH test gave no false negative responses and this was true not only in NS but even in all FSS and CGD having a GH MC greater than or equal to 3 micrograms/l. On the other hand, PD + GHRH induced a negative GH response in all oGHD and in 8/10 iGHD patients. In the remaining two iGHD patients, PD + GHRH demonstrated a normal pituitary GH reserve in spite of a GH MC less than 3 micrograms/l and low IGF-I level, thus pointing to a hypothalamic pathogenesis for the GHD. Considering FSS and CGD children having a GH MC less than 3 micrograms/l, PD + GHRH showed a primary pituitary GH deficiency in 3/12 CGD with low plasma IGF-I levels. In conclusion, in slowly growing children PD + GHRH test is the most reliable provocative test for the diagnosis of primary pituitary GH deficiency being capable to discriminate between an unequivocally normal and impaired somatotropic function.(ABSTRACT TRUNCATED AT 400 WORDS)

Pyridostigmine partially restores the GH responsiveness to GHRH in normal aging.

In 11 elderly normal subjects and in 17 young healthy subjects we studied the response of plasma growth hormone to GH-releasing hormone (GHRH(29), 1 microgram/kg iv) alone and preceded by pyridostigmine (120 mg orally 60 min before GHRH), a cholinesterase inhibitor likely able to suppress somatostatin release. The GH response to pyridostigmine alone was also examined. Basal plasma GH levels were similar in elderly and young subjects. In the elderly, GHRH induced a GH rise (AUC, median and range: 207.5, 43.5-444.0 micrograms.l-1.h-1) which was lower (p = 0.006) than that observed in young subjects (548.0, 112.5-2313.5 micrograms.l-1.h-1). The pyridostigmine-induced GH rise in the elderly was similar to that in young subjects (300.5, 163.0-470.0 vs 265.0, 33.0-514.5 micrograms.l-1.h-1). Pyridostigmine potentiated the GH responsiveness to GHRH in both elderly (437.5, 152.0-1815.5 micrograms.l-1.h-1; p = 0.01 vs GHRH alone) and young subjects (2140.0, 681.5-4429.5 micrograms.l-1.h-1; p = 0.0001 vs GHRH alone). However, the GH response to pyridostigmine + GHRH was significantly lower (p = 0.0001) in elderly than in young subjects. In conclusion, the cholinergic enhancement by pyridostigmine is able to potentiate the blunted GH response to GHRH in elderly subjects, inducing a GH increase similar to that observed after GHRH alone in young adults. This finding suggests that an alteration of somatostatinergic tone could be involved in the reduced GH secretion in normal aging. However, a decreased GH response to combined administration of pyridostigmine and GHRH in elderly subjects suggests that other abnormalities may coexist, leading to the secretory hypoactivity of somatotropes.

Glucagon stimulates GH secretion after intramuscular but not intravenous administration. Evidence against the assumption that glucagon per se has a GH-releasing activity.

In order to verify the true GH-releasing effect of glucagon and to explain the mechanism underlying this effect, we studied the effect of glucagon (GLU, 1 mg) administered either iv or im on both basal and GHRH (1 microgram/kg)-induced GH rise in 48 normal short children and adolescents. Moreover, the in vitro effect of GLU on rat anterior pituitary cells was studied. Intravenous administration of GLU induced no significant GH rise. On the other hand, im GLU administration induced a clear-cut GH increase (mean +/- SE GH peak after GLU vs placebo = 25.7 +/- 3.9 vs 10.1 +/- 3.6 micrograms/L, p < 0.01). Intravenous administration of GLU failed to modify the GHRH-induced GH rise either when coadministered with the neurohormone (35.2 +/- 4.1 vs 34.1 +/- 6.0 micrograms/L) or when given 60 min earlier (20.2 +/- 5.8 vs 21.1 +/- 8.3 micrograms/L). Differently from iv GLU, im GLU strikingly potentiated the GH response to GHRH given 90 min later (57.5 +/- 6.3 vs 24.7 +/- 9.1 micrograms/L, p < 0.01). Mean plasma glucose levels increased 30 min after GLU, administered either iv or im, and returned to basal levels 60 min later. GH secretion from dispersed rat pituitary cells was unaffected by incubation with GLU (10(-10)-10(-4) mol/L). Incubation of the cells with 10(-7) mol/L GHRH induced instead a clear-cut stimulation of GH release. In conclusion, our data demonstrate that glucagon per se has not GH-releasing activity as indicated by its uneffectiveness to release GH in vitro and after intravenous administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of the potentiation of cholinergic activity on the variability in individual GH response to GH-releasing hormone.

In man the GH response to GHRH is highly variable and some normal subjects may be completely unresponsive to the neuropeptide. On the other hand, the potentiation of cholinergic activity by pyridostigmine (PD), a cholinesterase inhibitor, increases the GH response to GHRH, probably by inhibiting somatostatin release. The aim of this study was to assess the existence of intraindividual variability in the GH response to GHRH and verify the effects of PD treatment on inter- and intraindividual variability. Twenty normal adults (17 M and 3 F) and 10 normal prepubertal children (9 M and 1 F) underwent 2-5 administrations of 1 micrograms/kg GHRH on different days. Seven adults and all children also underwent 1-5 other tests in which GHRH was preceded (60 min before) by oral PD (120 mg in adults and 60 mg in children). The GH responses to GHRH were highly variable, not only within subjects but also in the same subject on different occasions (peak range; adults: 0.4-49.0 ng/ml; children: 2.4-50.0 ng/ml). PD always markedly increased the GH response to GHRH, even unmasking this response in 3 adults and 4 children hyporesponsive to the neuropeptide alone. However, the variability in the GH response was still present (adults: 27.2-108.5 ng/ml; children: 25.0-144.0 ng/ml), though reduced (adults: p = 0.0005; children: p = 0.0204). These data indicate that: i. A great inter- and intraindividual variability in the GH response to GHRH is present.(ABSTRACT TRUNCATED AT 250 WORDS)