EFFECTS OF ENDOTHELIN-1 ON ITS RECEPTOR CONCENTRATION AND THYMIDINE INCORPORATION IN CALF ADRENAL ZONA GLOMERULOSA CELLS - A COMPARATIVE-STUDY WITH PHORBOL ESTERS

Endothelin (ET-1) is a 21-amino acid peptide with potent vasopressor and vasoconstrictive properties. Specific, high affinity receptors for ET-1 have been found in the adrenal gland. The stimulation by ET-1 of aldosterone secretion in cultured calf zona glomerulosa cells was shown to depend on the serum used for culturing and was not related to the growthpromoting effects of serum or the response to another secretagogue, such as angiotensin-II. In this study, binding of [125I]ET-1 to crude membrane preparations from calf adrenal cortex slices showed that ET-1 binding was greater in the outer slices, corresponding to the zona glomerulosa, than in inner slices, corresponding to the zona fasciculata. ET-1 stimulated aldosterone, but not cortisol, biosynthesis. Adrenal zona glomerulosa preincubated with ET-1 resulted in homologous down-regulation. Since ET-1 action involves activation of protein kinase-C (PKC), we studied the effect of a phorbol ester (PMA) on the down-regulation of ET-1 receptors. PMA decreased the number of cell surface receptors, and its effect was prevented by pretreatment with the PKC inhibitors H-7 and sphyngosine. Agonist-mediated down-regulation could not be blocked by pretreatment with PKC inhibitors, suggesting that PKC is involved in phorbol ester-mediated, but not agonistmediated, down-regulation of ET-1 receptors. Both effectors increased the endocytosis rate constant as well as the steady state cytosolic membrane-bound ratio for ET-1 receptors, suggesting that the decrease in the number of cell surface receptors is at least partially due to an increased internalization of the hormone-receptor complex. ET-1 and PMA decreased the incorporation of [3H]thymidine into calf zona glomerulosa cell cultures. We conclude that ET-1 and PMA have similar effects on glomerulosa cells, producing down-regulation of ET-1 receptors and an antimitogenic effect, but these actions are through different mechanisms. © 1990 by The Endocrine Society.