Cortisol production by dispersed guinea-pig adrenal cells; a specific, sensitive and reproducible response to acth and it's fragments

Abstract

Naturally occurring steroids and peptide hormones, tested at supraphysiological concentrations, were without effect on basal and human (h) 1–39 ACTH (NIBSC code 74/555, 25ng/l (5.5 × 10 −12 mol/1)) stimulated cortisol production. Further, low concentrations of angiotensin II, N-pro-opiocortin ( N terminal fragment 1–76) and γ-MSH all of which have been reported to synergise with ACTH with regard to cortisol production, were without significant effect alone or in combination with ACTH over the range 2.2 × 10 −13 to 5.5 × 10 −12 mol/1. The activity of h 1–39 was compared with that of the ACTH related peptides 1–24, 1–18, 1–17, 1–16, 1–13-NH 2 (α MSH), 1–10 and 4–10. The dose responses were parallel and the same maximal cortisol output was observed with all the peptides except the 1–10 fragment. Half maximal stimulation occurred at 3.1 × 10 −12 (1–24), 4.4 × 10 −12 (h 1–39), 1.5 × 10 −11 (1–39), 3.3 × 10 −10 (1–18), 5 × 10 −9 (1–13-NH 2), 8× 10 −9 (1–17), 2 × 10 −7 (1–16) and 1 × 10 −5 (4–10) mol/1 respectively. Interference by the above ACTH-derived peptides in cortisol secretion by the cells in response to 5.5 × 10 −12 mol/1 h 1–39 ACTH was minimal over the range 5.2 × 10 −12–2.2 × 10 −6 mol/1. The sensitivity of the adrenal cells to h 1–39 ACTH was such that 2ng/l (4.4 × 10 −13 mol/1) provoked cortisol secretion over the control ( P < 0.05, n = 17). The coefficient of variation within assay for each dose on the full standard curve (2.2 × 10 −13–1.1 × 10 −10mol/1) was < 10% n = 6). Half maximal stimulation was given by 14.5 ng/1 (3.2 × 10 −12 mol/1). Between control and 1.1 × 10 −10mol/l ACTH there was a 32 ± 8 (mean ± SD, n = 9) fold change in cortisol production.