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ABSTRACT

The effects of angiotensin II (AII), acetylcholine and vasopressin on the intracellular concentration of Ca²⁺ have been little studied in adrenocortical cells from the zona fasciculata/reticularis (ZFR).

Primary cultures of bovine ZFR cells maintained in suspension culture for 72 h produce cortisol in response to AII (0·1 μm), acetylcholine (0·1 mm) and vasopressin (1 μm). This response is accompanied by a breakdown of membrane phosphoinositides from [³H]inositol-prelabelled cells.

Using cells loaded with the Ca²⁺ indicator fura-2, the intracellular concentration of Ca²⁺ was measured in response to increasing doses of all three agonists and found to increase in a graded fashion in each case. The basal intracellular concentration of Ca²⁺ was 75±3 nm (mean±s.e.m., n=52), rising to a maximum 1·82±0·14-fold (n=6) for AII (0·1 μm), 1·35±0·05-fold (n=7) for acetylcholine (0·1 mm) and 1·27±0·10-fold (n=6) for vasopressin (1 μm).

In the case of AII and acetylcholine, agonists were added sequentially in medium of normal extracellular Ca²⁺ concentration (1·2 mm) or in medium in which the Ca²⁺ concentration was buffered to approximate to the intracellular concentration of Ca²⁺ (75–100 nm). Evidence was thereby obtained that both AII and acetylcholine mobilize a common intracellular pool of Ca²⁺.

Our findings suggest that these three agonists, all of which stimulate phospholipase C, increase intracellular Ca²⁺ through a mechanism which depends, at least in part, on the release of Ca²⁺ from a common intracellular pool.

ABSTRACT

The effects of TSH and the activation of the cyclic AMP (cAMP) and Ca²⁺-phosphatidylinositol (Ca²⁺-PI) cascades on the activity and expression of the selenoenzyme thyroidal type-I iodothyronine deiodinase (ID-I) have been studied using human thyrocytes grown in primary culture. Stimulation of ID-I activity and expression was obtained with TSH and an analogue of cAMP, 8-bromo-cAMP. In the presence or absence of TSH, the addition of the phorbol ester, phorbol 12-myristate 13-acetate (PMA) together with the calcium ionophore A23187, caused a decrease in ID-I activity; a decrease in ID-I expression was also observed as assessed by cell labelling with [⁷⁵ Se]selenite. PMA alone had no effect on ID-I activity in the presence or absence of TSH. A23187 alone produced a small but significant reduction in ID-I activity, but only in TSH-stimulated cells. These data provide evidence that the expression of thyroidal ID-I is negatively regulated by the Ca²⁺-PI cascade, and positively regulated by the cAMP cascade.

ABSTRACT

Using tritiated-thymidine incorporation as a measure of cell growth, interleukin-1β stimulated the growth of bovine zona fasciculata/reticularis adrenocortical cells after 72h in primary culture. Within the range of 10–1000pg/ml, interleukin-1β produced over 40% of angiotensin II-stimulated [³H]thymidine incorporation (P<0.005 compared with basal for 10pg/ml and 1000pg/ml; P<0.05 for 100pg/ml; two-tailed unpaired Student's t-test). Interleukin-1β did not directly stimulate cortisol secretion.

By stimulating adrenocortical growth, the increase in interleukin-1 during fever provides a potential mechanism for chronically raising glucocorticoid output. This study is the first demonstration of a long-term effect involving interleukin-1β on the adrenal cortex.

ABSTRACT

Cells isolated from the zona fasciculata/reticularis (ZFR) of the bovine adrenal cortex and maintained in culture were found to secrete cortisol in response to vasopressin stimulation. The increased cortisol secretion was dose dependent, with a threshold response at 1 nm and a maximal response (1·68-fold over basal) at 0·1 μm. In cells cultured in the presence of [³H]inositol (to prelabel the membrane phosphoinositide pool), stimulation with vasopressin in the presence of LiCl (10 mm) resulted in a similar dose-dependent increase in labelling of the phosphoinositol fraction, with a maximal response (1·45-fold over basal) at 10 nm. The increased labelling of the phosphoinositol fraction was independent of extracellular Ca²⁺ as it was not abolished in medium with [Ca²⁺ ] buffered to intracellular resting levels. This suggests that vasopressin stimulation results in the activation of a phosphoinositidase C. It is probable that cortisol secretion by bovine ZFR cells in response to vasopressin is dependent upon activation of this Ca²⁺-independent phosphoinositidase C. However, the small magnitude of the cortisol secretory response makes it unlikely that vasopressin is a primary regulator of cortisol secretion in vivo.