PROPERTIES OF A SIMPLIFIED BIOASSAY FOR ADRENOCORTICOTROPHIC ACTIVITY USING THE STEROIDOGENIC RESPONSE OF ISOLATED ADRENAL CELLS

Abstract

SUMMARY A reproducible method for isolating intact cells from the adrenal glands of a small number of rats (four to six) is described. The incubation conditions and fluorometric corticosteroid assay were modified to enable a large number of incubations to be carried out with each batch of cells. Linear regression analysis of log-dose—response curves for adrenocorticotrophin (ACTH) analogues yields potency ratios with 95% confidence limits which are usually less than ± 20%. The ED50 of the assay is about 50 pg/ml for natural ACTH and 7 pg/ml for corticotrophin-(1–24)-tetracosapeptide. Insulin and prostaglandin E2 had no effect on the steroidogenic response to ACTH while cyclic AMP at high doses was steroidogenic but gave a much steeper log-dose—response curve than ACTH. α- and β-melanocytestimulating hormone (MSH) were both agonists but only at concentrations 1 × 106 times higher than those of ACTH. Cells isolated from rabbit adrenals were similar to those from the rat in their responsiveness to ACTH but were unresponsive to amounts of MSH sufficient to stimulate rat adrenal cells.