Based on the finding of gene expression of adrenomedullin (Adm) and its receptor components in the rat testis, a paracrine effect of ADM on testicular steroidogenesis has been suggested by our group. The present study demonstrates the gene expression of Adm and the effect of ADM on testosterone production in the Leydig cell. The regulation of ADM by hCG and its interaction with endothelin 1 (EDN1) in the rat Leydig cells are also observed. Primary culture of Leydig cells produced Adm mRNA and secreted 275 ± 19 pg immunoreactive ADM per 10⁶ cells in 24 h. In addition, the Leydig cell was shown to coexpress mRNAs encoding for the calcitonin receptor-like receptor (CALCRL) and receptor activity-modifying protein (RAMP1, RAMP2, and RAMP3). These may account for the specific binding of ADM to the Leydig cells. Administration of ADM to Leydig cells resulted in an inhibition of hCG- and EDN1-stimulated testosterone production. Correlated with this, ADM reduced EDN1 production, whereas its production was increased by EDN1. Furthermore, the production of ADM and the mRNA levels of Calcrl and Ramp2 were suppressed by hCG. Our results suggest that ADM has an autocrine effect on Leydig cell steroidogenesis, possibly by interacting with EDN1 and under the control of gonadotropin. We propose that there is an ADM/EDN1 local regulatory mechanism that may be important in modulating the control of testicular functions by gonadotropins.