PRELIMINARY OBSERVATIONS OF BOVINE ADRENAL FASCICULATA-RETICULARIS CELLS IN MONOLAYER CULTURE: STEROIDOGENESIS, EFFECT OF ACTH AND CYCLIC AMP

Abstract

ABSTRACT Bovine adrenocortical cells dispersed by trypsin digestion of fasciculata-reticularis minces were maintained in monolayer culture for up to 6 weeks. During the first week cells grown in medium containing ACTH (1 mU/ml) secreted steroids at a rate 10 to 20-fold greater than control cultures, cortisol accounting for 80–90% of the corticotrophic response. Using tracer amounts of [3H]progesterone and [3H]pregnenolone, the major products were cortisol, corticosterone, 11-deoxycortisol and 11-deoxycorticosterone in decreasing order of magnitude. After 10 to 15 days in culture steroidogenesis was no longer enhanced by ACTH. This was concomitant with an apparent loss of 11β-hydroxylase activity which was mainly manifested by a sharp increase in the formation of 11-deoxycortisol. Short-term incubations of these cells during the first week in culture provided evidence that steroidogenesis was related to ACTH concentrations (from 0.1 to 100 μU/ml) and stimulated by dibutyryl cyclic AMP, the corticotrophic responses being further enhanced by theophylline (0.5 to 50 μmoles/5 ml). Exposure of the cells to ACTH (50 μU/ml) resulted in a rapid increase in intracellular cyclic AMP concentrations concomitant with a progressive increase in the corticosteroids released into the medium. The data are consistent with the conclusion that during the first week in culture these cells provide a valuable model for the study of factors regulating steroidogenesis in the zona fasciculata-reticularis.