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Quantitative analysis of the cytosolic-free-Ca2+-dependency of aldosterone production in bovine adrenal glomerulosa cells. Different requirements for angiotensin II and K+.

ABSTRACT: Angiotensin II (AII) and K+ raise the cytosolic free Ca2+ concentration [( Ca2+]i) and stimulate aldosterone production in isolated bovine adrenal glomerulosa cells. The mechanisms leading to an elevation of [Ca2+]i were analysed with the fluorescent Ca2+ probe quin 2. (1) Whereas [Ca2+]i rose transiently and returned to basal values within 5 min in response to AII, the effect of K+ was sustained for at least 15 min. (2) AII released Ca2+ from intracellular stores, whereas the [Ca2+]i response to K+ depended solely on extracellular [Ca2+]. (3) When added after K+ stimulation, AII provoked a dramatic decrease in [Ca2+]i to below the resting value. The role of [Ca2+]i in stimulating steroidogenesis was determined by manipulating the concentration of this cation. (4) In a cell superfusion system, the aldosterone response to AII is biphasic. Suppressing the transient [Ca2+]i elevation triggered by AII resulted in the disappearance of the initial secretory peak, but the final production rate was similar to that of control cells. (5) Normal basal [Ca2+]i levels were, however, necessary to maintain continuous AII-induced steroidogenesis. (6) When added after AII, the antagonist analogue [Sar1,Ala8]AII suppressed steroidogenesis without affecting [Ca2+]i levels. (7) In contrast, continuously elevated [Ca2+]i values were required for the initiation and the maintenance of K+-stimulated aldosterone production. These results demonstrate important differences in the mechanisms through which AII and K+ activate the Ca2+ messenger system. Moreover, functional correlations have shown that K+, but not AII, depends solely on a sustained [Ca2+]i response for its steroidogenic effect. However, the AII-induced effect is also a Ca2+-requiring process: the initial [Ca2+]i transient accelerates the onset of steroidogenesis, which is subsequently extremely sensitive to [Ca2+]i decreases below normal basal levels.

SUBMITTER: Capponi AM

PROVIDER: S-EPMC1148413 | biostudies-other | 1987 Oct

REPOSITORIES: biostudies-other

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Project description:The mineralocorticoid aldosterone mainly produced by the adrenal gland is essential for life but an abnormal excessive secretion causes severe pathological effects including hypertension and target organ injury in the heart and kidney. The aim of this study was to determine the gene regulatory network triggered by aldosterone secretagogues in a non transformed cell system. Freshly isolated rat adrenal zona glomerulosa cells were stimulated with the two main aldosterone secretagogues, angiotensin II and potassium, for two hours and subjected to whole genome expression studies using multiple biological and bioinformatics tools. Several genes were differentially expressed by Ang II (n=133) or potassium (n=216). Genes belonging to the nucleic acid binding and transcription factor activity categories were significantly enriched. A subset of the most regulated genes were confirmed by real-time RT-PCR and then their expression analyzed in time curve studies. Differentially expressed genes were grouped according to their time-response expression pattern and their promoter regions analyzed for common regulatory transcription factors binding sites. Finally, data mining with gene promoters, transcription factors and literature databases were performed to generate gene interaction networks for either Ang II or potassium. This study provides for the first time a complete study of the genes that are regulated, and the interaction between them, by aldosterone secretagogues in rat adrenal cells. Increasing our knowledge of adrenal physiology and gene regulation in non transformed cell systems would lead us to a better approach for discovery of candidate genes involved pathological conditions of the adrenal cortex. Experiment Overall Design: Freshly isolated rat adrenal cells were treated with Ang II (100 nM) or potassium (16 mM) for 2 h at 37C. Experiment Overall Design: Total RNA from three independent experiments were isolated, labeled and hybridized to whole genome high-density oligonucleotide microarrays (Affymetrix Rat Genome 230 2.0). Experiment Overall Design: 3 independent cell preparations. Experiment Overall Design: In each experiement there is one tube control, one Ang II (100 nM) and potassium (16 mM).

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Quantitative analysis of the cytosolic-free-Ca2+-dependency of aldosterone production in bovine adrenal glomerulosa cells. Different requirements for angiotensin II and K+.

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