Regulation of calcium influx and catecholamine secretion in chromaffin cells by a cytochrome P450 metabolite of arachidonic acid.
These studies were designed to determine the role of arachidonic acid metabolites in catecholamine secretion from adrenal chromaffin cells. Inhibitors of the cytochrome P450-dependent metabolism of arachidonic acid were shown to interfere with stimulus-secretion coupling in cultured chromaffin cells. Ketoconazole (10 microM), clotrimazole (20 microM), and piperonyl butoxide (50 microM) inhibited carbachol-dependent catecholamine secretion by 44%, 83%, and 100%, respectively; histamine-dependent secretion by 25%, 60%, and 81%, and secretion induced by 59 mM KCl depolarization by 25%, 55%, and 89%. Uptake of 45Ca2+ into the cells in response to carbachol was inhibited 63% by ketoconazole, 86% by clotrimazole, and 95% by piperonyl butoxide; KCl-dependent uptake was inhibited 7%, 56%, and 85%, respectively. However, cytochrome P450 inhibitors did not inhibit catecholamine secretion when cells were stimulated with the calcium ionophores ionomycin or lasalocid. These results indicated the involvement of a cytochrome P450 product in controlling Ca2+ influx in response to membrane depolarization. Cells prelabeled with [3H]arachidonic acid formed a 3H-labeled metabolite which comigrated with authentic 5,6-epoxyeicosatrienoic (5,6-EET) acid on reverse phase and normal phase HPLC. Pretreatment with clotrimazole inhibited the production of this 3H-labeled metabolite. Addition of synthetic 5,6-EET (1 nM) to cells pretreated with piperonyl butoxide resulted in catecholamine secretion. These data suggest a role for a cytochrome P450 metabolite of arachidonic acid in agonist-stimulated catecholamine secretion.
Hildebrandt
E
E
Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas 75235, USA.
Albanesi
J P
JP
Falck
J R
JR
Campbell
W B
WB
D015126:8,11,14-Eicosatrienoic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000311:Adrenal Glands / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002395:Catecholamines / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D002837:Chromaffin Granules / Q000187:drug effects / Q000378:metabolism*; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006632:Histamine / Q000502:physiology
D002395:Catecholamines; D065607:Cytochrome P-450 Enzyme Inhibitors; D016718:Arachidonic Acid; C040776:5,6-epoxy-8,11,14-eicosatrienoic acid; D006632:Histamine; D003577:Cytochrome P-450 Enzyme System; D015126:8,11,14-Eicosatrienoic Acid; D002118:Calcium