0021-972X/90/7102-0433$02.00/0 Journal of Clinical Endocrinology and Metabolism Copyright© 1990 by The Endocrine Society
Vol. 71, No. 2 Printed in U.S.A.
Acute Clonidine Administration Potentiates Spontaneous Diurnal, but not Nocturnal, Growth Hormone Secretion in Normal Short Children EZIO GHIGO, EMANUELA ARVAT, MARIO NICOLOSI, JAELE BELLONE, MARIA ROSA VALETTO, ENRICO MAZZA, ELISEO IMPERIALE, MASSIMO PROCOPIO, MARIA CRISTINA GHIGO, AND FRANCO CAMANNI Department of Clinical Physiopathology, Division of Endocrinology, University of Turin, Turin, Italy
ABSTRACT. It has been shown that «2-adrenoreceptor activation induced by clonidine (CLON) increases plasma GH levels in both adults and children. In this study the effects of CLON (150 Mg/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 ng/L; P = 0.0001; area under curve (AUC), 624.4 ± 62.7 us. 135.6 ± 33.3 Mg/Lh; P < 0.00001]. In the night, no
I
difference was observed between GH secretion after CLON (peak, 15.4 ± 3.2 Mg/L; AUC, 562.2 ± 57.5 fig/L-h) and placebo (peak, 13.1 ± 4.7 Mg/L; AUC, 497.2 ± 83.5 Mg/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 a2-adrenoreceptor activation is probably mediated by increased endogenous GHRH release; our results suggest that the endogenous GHRH secretion is maximally stimulated at night. (J Clin Endocrinol Metab 7 1 : 433-435, 1990)
Pubertal stage ranged between 1-3. None of the subjects had evidence of malnourishment, organic disease, thyroid or adrenal
T IS WELL known that clonidine (CLON), an a2adrenoreceptor agonist, induces a clear-cut rise in
plasma GH in both animals (1) and man (2). Many data
impairment, or psychosocial dwarfism.
suggest that the stimulatory effect of a2-adrenoreceptor activation is mediated by increased endogenous GHRH release (1). As the GH-releasing effect of clonidine was always shown during the morning, the aim of this study was to verify whether this drug is able to potentiate nocturnal GH secretion in normal short children. Previous observations of our group showed that pyridostigmine is able to increase GH secretion during the morning, but not GH hypersecretion occurring at night (3).
After informing and receiving consent from parents and children, the subjects were studied as in-patients. All tests were performed in random order on different days, at least 3 days apart. During nocturnal tests all children slept. Lights were turned off at 2300 h just after CLON or placebo administration. Electroencephalogram recording was not performed. Thirty minutes after an indwelling catheter was inserted into an antecubital vein, blood samples were taken every 30 min. The effects of oral CLON (Catapresan, Boehringer Ingelheim) on GH secretion were studied both in the morning (from 08001100 h; 150 Mg/m2 CLON at 0800 h) and at night (from 23000200 h; 150 Mg/m2 at 2300 h). In all subjects the effects of placebo on GH secretion were studied under the same experimental conditions both in the morning and during the night. Plasma GH levels were measured in duplicate by RIA, using reagents produced by Sorin (Saluggia, Italy). All samples from an individual child were analyzed at the same time. The sensitivity of the assay was 0.2 Mg/L. The intra- and interassay coefficients of variation were 4.5% and 7.9%, respectively. The GH secretory responses to both CLON and placebo were expressed either as absolute values (micrograms per L) or area under the response curve (AUC; micrograms per L/h), calculated by trapezoidal integration. Statistical analysis of the data was performed using Kruskal-Wallis nonparametric analysis of variance.
Subjects and Methods Nine children (six boys and three girls; age, 10.1-15.2 yr) with short stature were studied. Based on auxological criteria (4, 5) all children were classified as having familial short stature. They had 1) predicted adult height within parental target zone, 2) normal height velocity, and 3) bone age similar to chronological age (±12 months). In all children studied normal spontaneous nocturnal (from 2300-0700 h) GH secretion (mean GH concentration, >3 Mg/L) as well as normal insulin-like growth factor-I levels (>100 Mg/L) were shown. Received October 17,1989. Address requests for reprints to: Prof. F. Camanni, Divisione Endocrinologia, Ospedale Molinette C.so Polonia 14,10126 Torino, Italy. 433
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GHIGO ET AL.
434
Results During the morning, CLON induced a significant GH increase, which was higher than that observed after placebo [peak (mean ± SEM), 23.8 ± 4.3 vs. 3.4 ± 1.4 L; P = 0.0001; AUC, 624.4 ± 62.7 vs. 135.6 ± 33.3 h; P < 0.00001; Figs. 1 and 2, left panels]. In the night, GH secretion after CLON administration (peak, 15.4 ± 3.2 Mg/L; AUC, 562.2 ± 57.5 ng/L-h) overlapped with that observed after placebo (peak, 13.1 ± 4.7 /ig/L; AUC, 497.2 ± 83.5 /ig/L-h; Figs. 1 and 2, right panels). Spontaneous GH secretion was higher during the night than in the morning (P = 0.0001 for peak and P = 0.0005 for AUC). GH secretion at night, both spontaneously and after drug treatment, overlapped with that observed in the morning after CLON (Fig. 2). Clonidine administration induced mild sleepness in all children. Discussion The present study in normal short children clearly shows that «2-adrenoreceptor activation by CLON induces a clear-cut increase in plasma GH levels in the morning, but fails to potentiate the spontaneous nocturnal GH secretion, which, as previously evidenced by several researchers (6-8), greatly overrides the diurnal secretion. To our knowledge the effect of acute CLON administration on nocturnal GH secretion has not been studied in children. It has been recently reported that chronic short term CLON treatment augmented 24-h integrated GH secretion in slowly growing children with constitutional growth delay (9). Thirty-six hours after the last CLON administration, an increase in GH pulse amplitude was present, but this increase was significant only in the waking hours. This finding fits well with our results. Evidence has been presented that the GH-releasing effect of CLON is specifically due to a2-adrenoreceptor
JCE & M • 1990 Vol 71 • No 2
activation (10) and is probably mediated by stimulation of endogenous GHRH. In fact, the ability of CLON to increase GH release is preserved in reserpinized rats pretreated with somatostatin antiserum (11), while it is abolished in rats pretreated with GH-releasing hormone (GHRH) antiserum (12) or with monosodium glutamateinduced destruction of GHRH-secreting neurons (13). Additionally, CLON is able to release GHRH from rat hypothalami perfused in vitro (14), and acute or short term administration of CLON to 5-day-old rats induced changes in GH secretion overlapping with those elicited by GHRH (15, 16). In man acute CLON administration fails to enhance the GHRH-induced GH secretion (17) as well as to elicit a GH rise after a previous administration of GHRH (17). However, the latter findings were not observed by others (18). Moreover, the combined
administration of CLON with pyridostigmine, a cholinergic agonist probably acting via somatostatin inhibition (1), has an additive effect on GH secretion (19). Therefore, our results, by showing failure of CLON to increase nocturnal GH secretion, suggest that the endogenous GHRH secretion is maximally stimulated at night. Nocturnal GH secretion in short normal children was enhanced by neither pyridostigmine (3) nor the /?i-receptor blocker atenolol (20), two drugs that probably act via inhibition of somatostatin release. These findings suggested that somatostatin tone is spontaneously reduced at night. Therefore, a maximal activation of GHRHsecreting neurons and a reduced somatostatin tone may coexist at night. This hypothesis is supported by the finding that in rats, pretreatment with a somatostatin antiserum triggers the release of endogenous GHRH (21). On the other hand, to explain the ineffectiveness of CLON to enhance nocturnal GH secretion, a maximal activation of a2-adrenoreceptor might be hypothesized. This seems unlikely based on the assumption that noradrenergic activity is reduced during sleep (22). However, a selective hyperactivation of «2-adrenoreceptors in some Night
Morning 28 24
O) 20
FIG. 1. Plasma GH levels (mean ± SEM) in nine patients given 150 ng/m2 CLON, orally (A), or placebo (•) at 0800 (left) and 2300 (right) h.
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The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 14 November 2015. at 23:13 For personal use only. No other uses without permission. . All rights reserved.
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CLON ON DIURNAL AND NOCTURNAL GH SECRETION 800 r
O) 3,
O 3
Vol. 71, No. 2 Printed in U.S.A.
Acute Clonidine Administration Potentiates Spontaneous Diurnal, but not Nocturnal, Growth Hormone Secretion in Normal Short Children EZIO GHIGO, EMANUELA ARVAT, MARIO NICOLOSI, JAELE BELLONE, MARIA ROSA VALETTO, ENRICO MAZZA, ELISEO IMPERIALE, MASSIMO PROCOPIO, MARIA CRISTINA GHIGO, AND FRANCO CAMANNI Department of Clinical Physiopathology, Division of Endocrinology, University of Turin, Turin, Italy
ABSTRACT. It has been shown that «2-adrenoreceptor activation induced by clonidine (CLON) increases plasma GH levels in both adults and children. In this study the effects of CLON (150 Mg/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 ng/L; P = 0.0001; area under curve (AUC), 624.4 ± 62.7 us. 135.6 ± 33.3 Mg/Lh; P < 0.00001]. In the night, no
I
difference was observed between GH secretion after CLON (peak, 15.4 ± 3.2 Mg/L; AUC, 562.2 ± 57.5 fig/L-h) and placebo (peak, 13.1 ± 4.7 Mg/L; AUC, 497.2 ± 83.5 Mg/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 a2-adrenoreceptor activation is probably mediated by increased endogenous GHRH release; our results suggest that the endogenous GHRH secretion is maximally stimulated at night. (J Clin Endocrinol Metab 7 1 : 433-435, 1990)
Pubertal stage ranged between 1-3. None of the subjects had evidence of malnourishment, organic disease, thyroid or adrenal
T IS WELL known that clonidine (CLON), an a2adrenoreceptor agonist, induces a clear-cut rise in
plasma GH in both animals (1) and man (2). Many data
impairment, or psychosocial dwarfism.
suggest that the stimulatory effect of a2-adrenoreceptor activation is mediated by increased endogenous GHRH release (1). As the GH-releasing effect of clonidine was always shown during the morning, the aim of this study was to verify whether this drug is able to potentiate nocturnal GH secretion in normal short children. Previous observations of our group showed that pyridostigmine is able to increase GH secretion during the morning, but not GH hypersecretion occurring at night (3).
After informing and receiving consent from parents and children, the subjects were studied as in-patients. All tests were performed in random order on different days, at least 3 days apart. During nocturnal tests all children slept. Lights were turned off at 2300 h just after CLON or placebo administration. Electroencephalogram recording was not performed. Thirty minutes after an indwelling catheter was inserted into an antecubital vein, blood samples were taken every 30 min. The effects of oral CLON (Catapresan, Boehringer Ingelheim) on GH secretion were studied both in the morning (from 08001100 h; 150 Mg/m2 CLON at 0800 h) and at night (from 23000200 h; 150 Mg/m2 at 2300 h). In all subjects the effects of placebo on GH secretion were studied under the same experimental conditions both in the morning and during the night. Plasma GH levels were measured in duplicate by RIA, using reagents produced by Sorin (Saluggia, Italy). All samples from an individual child were analyzed at the same time. The sensitivity of the assay was 0.2 Mg/L. The intra- and interassay coefficients of variation were 4.5% and 7.9%, respectively. The GH secretory responses to both CLON and placebo were expressed either as absolute values (micrograms per L) or area under the response curve (AUC; micrograms per L/h), calculated by trapezoidal integration. Statistical analysis of the data was performed using Kruskal-Wallis nonparametric analysis of variance.
Subjects and Methods Nine children (six boys and three girls; age, 10.1-15.2 yr) with short stature were studied. Based on auxological criteria (4, 5) all children were classified as having familial short stature. They had 1) predicted adult height within parental target zone, 2) normal height velocity, and 3) bone age similar to chronological age (±12 months). In all children studied normal spontaneous nocturnal (from 2300-0700 h) GH secretion (mean GH concentration, >3 Mg/L) as well as normal insulin-like growth factor-I levels (>100 Mg/L) were shown. Received October 17,1989. Address requests for reprints to: Prof. F. Camanni, Divisione Endocrinologia, Ospedale Molinette C.so Polonia 14,10126 Torino, Italy. 433
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 14 November 2015. at 23:13 For personal use only. No other uses without permission. . All rights reserved.
GHIGO ET AL.
434
Results During the morning, CLON induced a significant GH increase, which was higher than that observed after placebo [peak (mean ± SEM), 23.8 ± 4.3 vs. 3.4 ± 1.4 L; P = 0.0001; AUC, 624.4 ± 62.7 vs. 135.6 ± 33.3 h; P < 0.00001; Figs. 1 and 2, left panels]. In the night, GH secretion after CLON administration (peak, 15.4 ± 3.2 Mg/L; AUC, 562.2 ± 57.5 ng/L-h) overlapped with that observed after placebo (peak, 13.1 ± 4.7 /ig/L; AUC, 497.2 ± 83.5 /ig/L-h; Figs. 1 and 2, right panels). Spontaneous GH secretion was higher during the night than in the morning (P = 0.0001 for peak and P = 0.0005 for AUC). GH secretion at night, both spontaneously and after drug treatment, overlapped with that observed in the morning after CLON (Fig. 2). Clonidine administration induced mild sleepness in all children. Discussion The present study in normal short children clearly shows that «2-adrenoreceptor activation by CLON induces a clear-cut increase in plasma GH levels in the morning, but fails to potentiate the spontaneous nocturnal GH secretion, which, as previously evidenced by several researchers (6-8), greatly overrides the diurnal secretion. To our knowledge the effect of acute CLON administration on nocturnal GH secretion has not been studied in children. It has been recently reported that chronic short term CLON treatment augmented 24-h integrated GH secretion in slowly growing children with constitutional growth delay (9). Thirty-six hours after the last CLON administration, an increase in GH pulse amplitude was present, but this increase was significant only in the waking hours. This finding fits well with our results. Evidence has been presented that the GH-releasing effect of CLON is specifically due to a2-adrenoreceptor
JCE & M • 1990 Vol 71 • No 2
activation (10) and is probably mediated by stimulation of endogenous GHRH. In fact, the ability of CLON to increase GH release is preserved in reserpinized rats pretreated with somatostatin antiserum (11), while it is abolished in rats pretreated with GH-releasing hormone (GHRH) antiserum (12) or with monosodium glutamateinduced destruction of GHRH-secreting neurons (13). Additionally, CLON is able to release GHRH from rat hypothalami perfused in vitro (14), and acute or short term administration of CLON to 5-day-old rats induced changes in GH secretion overlapping with those elicited by GHRH (15, 16). In man acute CLON administration fails to enhance the GHRH-induced GH secretion (17) as well as to elicit a GH rise after a previous administration of GHRH (17). However, the latter findings were not observed by others (18). Moreover, the combined
administration of CLON with pyridostigmine, a cholinergic agonist probably acting via somatostatin inhibition (1), has an additive effect on GH secretion (19). Therefore, our results, by showing failure of CLON to increase nocturnal GH secretion, suggest that the endogenous GHRH secretion is maximally stimulated at night. Nocturnal GH secretion in short normal children was enhanced by neither pyridostigmine (3) nor the /?i-receptor blocker atenolol (20), two drugs that probably act via inhibition of somatostatin release. These findings suggested that somatostatin tone is spontaneously reduced at night. Therefore, a maximal activation of GHRHsecreting neurons and a reduced somatostatin tone may coexist at night. This hypothesis is supported by the finding that in rats, pretreatment with a somatostatin antiserum triggers the release of endogenous GHRH (21). On the other hand, to explain the ineffectiveness of CLON to enhance nocturnal GH secretion, a maximal activation of a2-adrenoreceptor might be hypothesized. This seems unlikely based on the assumption that noradrenergic activity is reduced during sleep (22). However, a selective hyperactivation of «2-adrenoreceptors in some Night
Morning 28 24
O) 20
FIG. 1. Plasma GH levels (mean ± SEM) in nine patients given 150 ng/m2 CLON, orally (A), or placebo (•) at 0800 (left) and 2300 (right) h.
I
16
o E » (A re Q.
8
0800
08.30
09.00
09.30
10.00
1030
11.00 23.00
23.30
24.00
24.30
01.00
time (h)
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 14 November 2015. at 23:13 For personal use only. No other uses without permission. . All rights reserved.
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CLON ON DIURNAL AND NOCTURNAL GH SECRETION 800 r
O) 3,
O 3
