Studies on the Control of Adrenal Steroidogenesis: Inter-Relationships Between the Effects of Benzodiazepines, Calcium Ions and Lipoproteins

Thomson, Ian (1992) Studies on the Control of Adrenal Steroidogenesis: Inter-Relationships Between the Effects of Benzodiazepines, Calcium Ions and Lipoproteins. PhD thesis, University of Glasgow.

Abstract

Three aspects of steroidogenic control in bovine adrenocortical cells have been considered: (i) Cholesterol supply, (ii) Calcium metabolism. (iii) Activity of steroidogenic enzymes. The effects of dantrolene, high density lipoprotein cholesterol isoquinoline carboxamides and benzodiazepines, agents reported to act specifically on each control element, have been assessed. (a) Dantrolene Dantrolene is a drug which reputedly blocks specifically hormone-stimulated release of Ca2+ from intracellular stores. Dantrolene abolished the aldosterone response to angiotensin-II (A-II). However, aldosterone synthesis, from added hydroxycholesterol, was more sensitive to inhibition by dantrolene (IC50 0.02mmol/l) than A-II-stimulated synthesis although high concentrations of the drug (0.2mmol/l) did not totally abolish aldosterone synthesis in hydroxycholesterol-treated cells. Aldosterone synthesis in cells incubated with hydroxycholesterol, pregnenolone, progesterone, 11-deoxycorticosterone (DOC), corticosterone and 18- hydroxy corticosterone was significantly inhibited (p<0.05) by 89%, 69%, 59%, 31%, 62% and 28% respectively in the presence of 0.2mmol/l dantrolene. In zona fasiculata/reticularis cells, dantrolene (0.2mmol/l) significantly reduced cortisol production in cells treated with hydroxycholesterol (p<0.001) but not in those treated with pregnenolone or progesterone. In isolated mitochondria, dantrolene significantly reduced pregnenolone production from added hydroxycholesterol (92%; p<0.001), aldosterone production from added corticosterone (31%; p<0.001) and cortisol production from added 11-deoxycortisol (27%; p<0.01). Dantrolene appears not only to inhibit steroidogenesis by preventing A-II- stimulated release of intracellular calcium, but also to directly inhibit steroidogenic enzymes, in particular cholesterol side-chain cleavage activity. (b) High Density Lipoproteins Previous studies have established that HDL is readily utilised as a substrate for steroidogenesis. However, studies in platelets suggest that HDL subfractions may differentially interfere with signal-response coupling with variable consequences on platelet aggregation. In freshly isolated cells HDL-2 and HDL-3 increased basal aldosterone (1.5-3.5 fold) and cortisol synthesis (1.3-6.5 fold) in a dose-dependent manner. Both subfractions potentiated A-II stimulated aldosterone synthesis at low concentrations of peptide (10-10 mol/1: 1.5 fold) but had little effect at maximal concentrations of A-II (10-7mol/1). Cortisol synthesis at both low (10-10 mol/l: 1.8-2.3 fold) and high (10-7mol/1: 2.2-2.6 fold) concentrations of A-II was further increased by HDL. In cultured cells treated with HDL-cholesterol, basal aldosterone (3 and 3.1 fold) and cortisol synthesis (12 and 15 fold) was increased more than in cultured cells stimulated with A-II (10-8mol/1). The response to A-II in the presence of HDL- cholesterol was a 10-14 fold increase in aldosterone synthesis and a 6.5-7.6 fold increase in cortisol synthesis. This compared with a 30 fold increase in aldosterone and a 42 fold increase in cortisol in cells not treated with HDL. Neither HDL-2 nor HDL-3 (0.4mmol/l cholesterol) had any effect on acute 45Ca uptake in isolated zona glomerulosa cells. This was measured thirty seconds after addition of label. Under steady-state conditions (after thirty minutes), both subfractions significantly decreased 45Ca uptake when compared with control (15%; p<0.01). No significant effects were observed on concentrations of basal and A-II- stimulated intracellular free calcium in zona glomerulosa cells pre-incubated with HDL-2 or HDL-3 for 15 mins. In conclusion, HDL-2 and HDL-3 are similarly effective in providing substrate for aldosterone and cortisol synthesis. Neither subfraction has any acute effect on cell calcium. In the longer term, HDL-cholesterol may lower steady-state calcium content in adrenal cells as it does in platelets. The significance of this in signal-response coupling is not known. (Abstract shortened by ProQuest.).

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Additional Information: Adviser: C J Kenyon
• Keywords: Medicine, Pharmacology
• Date of Award: 1992
• Depositing User: Enlighten Team
• Unique ID: glathesis:1992-75454
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 19 Nov 2019 20:04
• Last Modified: 19 Nov 2019 20:04
• URI: https://theses.gla.ac.uk/id/eprint/75454