Pergamon J. Steroid Biochem. Molec. Biol. Vol. 57, No. 1/2, pp. 89--93, 1996 Copyright © 1996 Elsevier Science Ltd. All rights reserved 0960-0760(95)00248-0 Printed in Great Britain 0960-0760/96 $15.00 + 0.00 In Vitro Studies of the Subtypes of Endothelin (ET) Receptors Present in the Rat Testis, and of their Involvement in the Secretory Response of Leydig Cells to ET-1 Anna S. Belloni, 1 Paola G. Andreis, 1 Giuliano Neri, 1 Gian Paolo Rossifl Anna Markowska, 3 Giuseppe Gottardo, 1 Ludwik K. Malendowicz 3 and Gastone G. Nussdorfer ~* Departments of IAnatomy and 2Clinical Medicine, University of Padua, 35121 Padua, Italy and 3Department of Histology and Embryology, School of Medicine, 60781 Poznan, Poland The distribution of the endothelin (ET)-receptor subtypes ET A and ET B in the rat testis and their involvement in the secretory response of Leydig cells to ET-1 have been investigated by the use of specific ligands. Autoradiography showed that [125I]ET-1 binding was intense in the interstitial area of the testis, contairdng Leydig cells, and virtually absent in the walls of seminiferous tubules. Labelling was almost completely displaced by BQ-123, a selective ETA receptor antagonist, while sarafotoxin-6C and BQ-788, two specific ETB ligands, were ineffective. ET-1 concentration-depen- dently enhanced testosterone secretion of dispersed rat Leydig cells, and the response was suppressed by BQ-123, but not by BQ-788. Both antagonists per se did not affect either basal and hCG stimulated secretion of Leydig cells. Taken together our findings indicate that rat Leydig cells are mainly provided with ETA, and that this ET-receptor subtype mediates their secretory response to ET-1. Copyright © 1996 Elsevier Science Ltd. J. Steroid Biochem. Malec. Biol., Vol. 57, No. 1/2, pp. 89-93, 1996 INTR ODUCTION Endothelins (ET) are a family of peptides secreted by vascular endothelium, with a potent v---asoconstrictor and pressor activity (see [1] for review). The wide- spread distribution of ET in endocrine organs suggests that these peptides may be involved in the paracrine regulation of hormone ,;ecretion (see [1, 2] for review). ET-1 and its mRNA ]~ave been found in the rat testis [3, 4]. Evidence indicates that ET-1 is produced by Sertoli cells of rat [5] and humans [6], and acts on Leydig cells, which are provided with specific ET receptors [5, 7]. Accordingly, it has been demonstrated *Correspondence to G. G. Nussdorfer. Received 12 Jun. 1995; accepted 12 Sep. 1995. Abbreviations: ANOVA, analysis of variance; BSA, bovine serum albumin; ET, endothelins; ET A and ET a, endothelin-receptor sub- types A and B; hCG, human chorionic gonadotropin; HSA, human serum albumin; $6C, sarafotoxin-6C, T, testosterone. that ET-1 enhances both basal and hCG-stimulated testosterone (T) production by dispersed rat Leydig cells [8]. Parenthetically, it must be recalled that mouse Leydig tumour cells (MA-10) also possess ET recep- tors and secrete progesterone in response to ET-1 [9]. ET act via at least two main subtypes of receptors, named ETA and ETB, which are mainly expressed in vascular smooth-muscle ceils and endothelial cells, respectively (see [1, 10, 11] for review). Indirect proof seems to suggest that rat Leydig ceils mainly express ETA [5, 7]. However, Conte et al. [8] reported that dispersed rat Leydig cells also display a moderate secretory response to ET-3, an ET isoform that preferentially binds ET a receptor subtypes [1, 12]. It, therefore, seemed worthwhile to study, by the use of selective ligands, which subtypes of ET recep- tors are present in the rat testis and are involved in the mediation of the secretagogue effect of ET-1 on Leydig cells. 89