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The Effect of Cyproheptadine on Plasma Growth Hormone (GH) and on Somatostatin Response to GH-Releasing Hormone in Man R. H. Rosskamp, F. Haverkamp and G. von Kalckreuth Universitats-Kinderklinik und Poliklinik, Bonn, Germany

Cyproheptadine (CPH) - a putative serotonin antagonist — is known to inhibit growth hormone (GH) response to various pharmacological stimuli, as well as during sleep. To elucidate the possible site at which this drug takes effect, we examined plasma G H and somatostatin response to i. v. G H R H 1 - 4 4 (1 ug/kg body wt.) before and after CPH treatment in 10 healthy volunteers. The oral administration of CPH (8—12 mg daily for 5 days; total dose 56 mg) significantly curbed G H response to G H R H as expressed in peak plasma G H values (32.0 ± 6.1 ug/1 vs. 12.6 ± 3 . 2 u.g/1; P < 0.01) and in integrated G H response area (2368 ± 5 1 7 ug x l _ 1 x 2 h vs. 744 ± 1 7 2 ug x 1~ x 2 h; P < 0.01). Plasma somatostatin levels did not change in response to G H R H . Key-Words Growth Hormone-Releasing Hormone — Growth Hormone-Cyproheptadine — Somatostatin

Introduction Several studies suggest that serotoninergic mechanisms are involved in regulating human growth hormone (GH) secretion. Administration of L-tryptophan (Woolf and Lee 1976; Fraser, Tucker, Grubb, Wigand and Blackard 1979) and of 5-hydroxy-tryptophan (Lancranjan, Wirz-Justice, Puhringer and Del Pozo 1977; Imura, Nakai and Yoshimi 1973) causes increase in human plasma GH. In addition, G H release can be blocked by serotonin antagonist cyproheptadine (CPH) following oral hydroxytryptophan and arginine infusion (Nakai, Imura, Sakurai, Kurahachi and Yoshimi 1974), insulin-induced hypoglycemia and physical exercise (Smythe and Lazarus 1974), and during sleep (Chihara, Kato, Maeda, Matsukura and Imura 1976). However, where action takes place precisely and at which exact level serotoninergic stimuli modulate human pituitary G H release is unknown. Therefore, to find these answers we investigated the effect of oral administration of CPH on the plasma G H and somatostatin response to GH-releasing hormone (GHRH) in man. Horm. meta. Res. 22 (1990) 295 - 297 © Georg Thieme Verlag Stuttgart • New York

10 healthy non-obese volunteers (21—32 years; 3 females) after giving their informed consent, were studied. GHRHtesting was performed after an overnight fast with the subject in a recumbent position. A cubital vein was kept open with intravenous 0.9 % saline infusion. Blood samples were drawn at —30,0,15, 30,45,60,90 and 120 min before and after bolus iv injection of GHRH1-44 (1 ug/kg body wt; Bissendorf Peptide, Wedemark, Germany). The samples for somatostatin determination were collected in prechilled tubes containing EDTA and aprotinin. They were centrifuged within 15 min at 4 ° C and aliquots frozen at — 20 ° C until assayed. After an initial GHRH test all subjects received a regimen of oral CPH (Nuran tabl., Frosst Pharma, Munich, Germany), in doses sufficient to block GH release (Smythe and Lazarus 1974). On the 1st, 4th and 5th days 4 mg of CPH three times daily. A final 4 mg dose was given 1 hour before the second GHRH-test so that cumulative dosage of CPH was 56 mg. GHRH-testing was then performed again as previously described. Plasma GH was measured in duplicate by RIA using a commercial kit (Serono, Freiburg, Germany). Intra- and inter-assay coefficients of variation were less than 8 % and 10 %, respectively. Somatostatin RIA was performed as recently described (Rosskamp, Becker, Haverkamp, Thomas, Bruhl, Klumpp and Liappis 1987). Recovery of cyclic somatostatin that had been added to human pool plasma ranged from 84% to 98%. The lower limit of detection was found to be 6.4 ±2.1 ng/1 per assay. The intra- and interassay coefficients of variation were 8.7% and 13.9%, respectively. The area under the curve was calculated by trapezoidal integration. Comparison of mean GH values before and after treatment was carried out by means of the Kruskall-Wallis-H-test. Peak GH values and integrated areas under the curve were compared by means of the Mann-Whitney-U-test. Results were calculated as mean ± SEM. Results In all subjects studied, GHRH-injection caused an increase in plasma GH. Pretreatment with CPH significantly (P < 0.01) reduced the GH-releasing effect of G H R H (Fig. 1). The oral administration of CPH significantly curved G H response to G H R H as expressed in peak plasma G H values (32.0 ±6.1 ug/1 vs. 12.6 ± 3 . 2 ug/1; P < 0.01) and in integrated G H response area (2368 ± 517 ug x 1 x 2 h vs.

Received: 26 Apr. 1989

Accepted: 1 Sept. 1989

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Materials and Methods Summary

R. H. Rosskamp, F. Haverkamp and G. von Kalckreuth

Fig. 1 Mean±SEM plasma GH levels in response to GHRH1-44 i.v. (1 ng/kg body wt.) before ( • — • ) and after ( • - — • ) CPH treatment in 10 healthy adult volunteers.

Fig. 2 Individual and mean peak GH values in response to GHRH144 i. v. (1 ng/kg body wt.) before and after CPH treatment in 10 healthy adult volunteers.

744 ± 172 u,g x 1 ' x 2 h; P < 0.01). Individual peak GH and mean peak GH levels in response to GHRH before and after CPH treatment are shown in Fig. 2. Pretreatment with CPH suppressed the plasma GH rise after GHRH in 8 of 10 subjects.

pothalamic somatostatin secretion before and after CPH treatment. However, there is some evidence that elevated gastrointestinal somatostatin levels may effect GH release in animals (Tannenbaum 1981). In children, a negative correlation was reported between the degree of the serum GH peak following GHRH injection and the basal levels of serum somatostatin (Liapi, Evain-Brion, Argente, Vaudry and Donnadieu 1986). We were not able to confirm this finding in our study group of adults.

Fasting levels of plasma somatostatin were at 44.7 ± 6.7 ng/1 before, and at 54.0 ± 6.2 ng/1 after CPH treatment. there was no significant change in plasma somatostatin levels in the posttreatment group, there was no correlation between basal plasma somatostatin levels and GHRH-induced GH increase.

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296 Horm. meta. Res. 22 (1990)

Pituitary insensitivity could be another reason for the suppressed GH response to GHRH following CPH treatment. A direct effect of CPH at the pituitary level on ACTH secretion has already been reported (Ishibashi and Discussion Yamaji 1981; Suda, Tozawa, Mouri, Sasaki, Shibasaki, In the present study, pretreatment with CPH — Demura and Shizume 1983). Relatively high concentrations of a putative serotonin antagonist with mild antihistamine and serotonin and serotoninergic nerve fibers are located in the anticholinergic effects — reduced the GH-releasing effect of pituitary (Johnston, Spinedi and Negro-Vilar 1984; MoguilevGHRH. This finding indicates, at least in part, the possible ski, Faigon, Rubio, Scacchi and Swarcfarb 1985; Westlund and role of serotoninergic mechanisms in regulating GH secretion. Childs 1982). More recently, serotonin receptors have been In addition, we suggest that modulation of hypothalamic identified and characterized in the rat pituitary gland (Souza GHRH release is not the major effect that serotoninergic in- 1986). Further support for direct serotoninergic action at the volvement has on GH regulation in man. However, in rats an pituitary level comes from in vivo studies showing direct GH injection of anti-GHRH antibody blunted a GH rise induced release in autografted rat pituitaries perifused with 5-hydroxyby a previous dose of 5-hydroxytryptophan or serotonin tryptamine (Lopez, Gonzalez and Aguilar 1986). However, 5(Murakami, Kato, Kabayama, Tojo, Inoue and Imura 1986) hydroxytryptamine did not stimulate GH release from the suggesting a possible mediation of serotoninergic mechanisms pituitary in vitro (Kato et al. 1980). by hypothalamic GHRH in this species. Alternatively, the reAlthough there is increasing evidence that duced GH response found in our study could have been caused by an increase in somatostatin secretion or by a re- serotoninergic stimuli alter the release of pituitary GH, the exact site of action is still unclear. The results of our study sugduced pituitary sensitivity to GHRH. gest that serotoninergic modulation of GHRH release seems In urethrane-anesthesized rats the injection of not to be the major effect on GH release. Modulation of soma5-hydroxytryptamine had no effect on the release of soma- tostatin release and/or direct involvement at the pituitary level tostatin from the median eminence (Chihara, Arimura and seem to be of more likely importance. Schally 1979), but pretreatment with an anti-somatostatin antibody aggravated the GH secretion induced by 5-hydroxytrypReferences tophan (Kato, Matsushita, Katakami and Imura 1980). Since the major part of somatostatin detected in the peripheral circu- Chihara, K., Y. Kato, K. Maeda, S. Matsukura, H. Imura: Suppression by cyproheptadine of human growth hormone and Cortisol lation originated in the gut (Lucey 1986; Rosskamp 1988) our secretion during sleep. J. Clin. Invest. 57:1393-1402 (1976) data can provide but little information about the hy- Chihara, K., A. Arimura, A. V. Schally: Effect of intraventricular injecpothalamic somatostatin release after CPH treatment in man. tion of dopamine, norepinephrine, acetylcholine and 5-hydroxyPhysiological TSH release is modulated by somatostatin tryptamine on immunoreactive somatostatin release into rat hysecretion. Therefore differences of TSH levels after TRH pophyseal portal blood. Endocrinology 104: 1656—1662(1979) stimulation would provide more information about hy-

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Fraser, W. M., H. St. Tucker, S. R. Grubb, J. P. Wigand, W. G. BlackNakai, Y., H. Imura, H. Sakurai, H. Kurahachi, T. Yoshimi: Effect of ard: Effect of L-tryptophan on growth hormone and prolactin recyproheptadine on human growth hormone secretion. J. Clin. Enlease in normal volunteers and patients with secretory pituitary docrinol. Metab. 38:446-449 (1974) tumors. Horm. Metab. Res. 11:149-155(1979) Rosskamp, R., M. Becker, F. Haverkamp, B. Thomas, S. Briihl, J. Imura, H., Y. Nakai, T. Yoshimi: Effect of 5-hydroxytryptophan (5Klumpp, N. Liappis: Plasma levels of growth hormone-releasing HTP) on growth hormone and ACTH release in man. J. Clin. Enhormone and somatostatin in response to a mixed meal and during docrinol. Metab. 36:204-206 (1973) sleep in children. Acta Endocrinol. (Copenh.) 116: 549—554 Ishibashi, M., T. Yamaji: Direct effects of thyrotropin-releasing hor(1987) mone, cyproheptadine, and dopamine on adrenocorticotropin Rosskamp, R.: Klinisch-physiologische Untersuchungen zum secretion from human corticotroph adenoma cells in vitro. J. Clin. Wachstumshormon-Releasing-Hormon. Habilitation, Faculty of Invest. 68:1018-1027(1981) Medicine, University of Bonn (1988) Johnston, C. A., E. Spinedi, A. Negro-Villar: Aromatic L-amino acid Smythe, G. A., L. Lazarus: Suppression of human growth hormone decarboxylase activity in the rat median eminence, neurointermesecretion by melatonin and cyproheptadine. J. Clin. Invest. 54: diate lobe and anterior lobe of the pituitary. Neuroendocrinology 116-121(1974) 39:54-59(1984) Souza, De E. B.: Serotonin and dopamine receptors in the rat pituitary Kato, Y, N. Matsushita, H. Katakami, H. Imura: Brain serotonin and gland: autoradiographic identification, characterization, and lothe secretion of prolactin and growth hormone. Clin. Endocrinol. calization. Endocrinology 119:1534-1542 (1986) (Tokyo) 26:625-628 (1980) Suda, T, F. Tozawa, T. Mouri, A. Sasaki, T. Shibasaki, H. Demura, K Lancranjan, J., A. Wirz-Justice, W. Piihringer, E. DelPozo: Effect of 1 —Shizume: Effects of cyproheptadine, reserpine and synthetic corti5 hydroxytryptophan infusion on growth hormone and prolactin cotropin-releasing factor on pituitary glands from patients with secretion in man. J. clin. Endocrinol. Metab. 45:588-593 (1977) Cushing's disease. J. Clin. Endocrinol. Metab. 56: 1094-1099 Liapi, C, D. Evain Brion, J. Argente, H. Vaudry, M. Donnadieu: Nega(1983) tive correlation between peripheral plasma somatostatin levels Tannenbaum, G. S.: Growth hormone secretory dynamics in streptoand GH responses to GH-RH stimulation tests in children. Acta zotocin diabetes: Evidence of a role for endogenous circulating soEndocrinol. (Copenh.) 113:1-4 (1986) matostatin. Endocrinology 108:76-82(1981) Lopez, F., D. Gonzalez, E. Aguilar: Serotonin stimulates GH secretion Westlund, K, G. V. Childs: Localization of serotoninergic fibers in the through a direct pituitary action: studies in hypophysectomized rat adenohypophysis. Endocrinology 111: 1761 —1763 (1982) autografted animals and in perifused pituitaries. Acta Endocrinol. Woolf, P. D., L. Lee: Effect of serotonin precursor, tryptophan, on (Copenh.) 113:317-322 (1986) pituitary hormone secretion. J. Clin. Endocrinol. Metab. 45:123— Lucey, M. R.: Endogenous somatostatin and the gut. Gut 27:457-467 133 (1977) (1986) Moguilevski, J. A., M. R. Faigon, M. C. Rubio, P. Scacchi, B. Swarcfarb: Sexual differences in the effect of serotonin on LH Requests for reprints should be addressed to: secretionin rats. Acta Endocrinol. (Copenh.) 109:320-325 (1985) Murakami, Y., Y. Kato, Y Kabayama, K. Tojo, T. Inoue, H. Imura: Priv.-Doz. Dr. R. Rosskamp Involvement of growth hormone (GH)-releasing factor in GH Universitats-Kinderklinik secretion induced by serotoninergic mechanism in conscious rats. Adenauerallee 119 Endocrinology 119:1089-1092(1986) D-5300 Bonn 1 (Germany)

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Effect of Cyproheptadine on GH Response to GHRH

The effect of cyproheptadine on plasma growth hormone (GH) and on somatostatin response to GH-releasing hormone in man.

Cyproheptadine (CPH)--a putative serotonin antagonist--is known to inhibit growth hormone (GH) response to various pharmacological stimuli, as well as...
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