CORTICOSTEROIDOGENESIS IN ISOLATED ADRENAL CELLS: EFFECT OF ADRENOCORTICOTROPHIC HORMONE, ADENOSINE 3′,5′-MONOPHOSPHATE AND β1–24ADRENOCORTICOTROPHIC HORMONE DIAZOTIZED TO POLYACRYLAMIDE

Abstract

SUMMARY A cell suspension was prepared from normal rat adrenal tissue by collagenase digestion of decapsulated adrenal glands. Corticosterone produced by these cells was assayed both by competitive protein binding and by radioimmunoassay (range: 0–1 ng; sensitivity: 30 pg). Before incubation, the medium and cells equivalent to one gland contained 27·7 ± 3·2 ng corticosterone which increased to 36·3 ± 7·0 ng corticosterone after 1 h of incubation. Although a normal sigmoid log dose—response curve was obtained using adrenocorticotrophin (ACTH) (ED 50 :1·2 × 10 −4 i.u./ml), it was observed that log (ACTH concentration) was directly proportional to log (corticosterone produced) between 1 × 10 −7 and 1 × 10 −3 i.u. ACTH/ml. A similar linear relationship was demonstrated for the effect of adenosine 3′,5′-monophosphate on steroidogenesis. Time-course studies revealed that after addition of ACTH there was a definite lag (3 min) before onset of increased steroidogenesis; within 5 min a constant corticosterone production rate was achieved. β 1–24 Adrenocorticotrophin was shown to retain its ability to stimulate steroidogenesis when diazotized to beads of polyacrylamide too large to enter cells. Evidence is provided that the activity of the β 1–24 ACTH—polyacrylamide beads is not due to biologically active peptides cleaved from the complex. It is concluded that ACTH can stimulate steroidogenesis without entering its target cells.