Voltage-regulated calcium channels involved in the regulation of enkephalin synthesis are blocked by phorbol ester treatment

• Published in: The Journal of biological chemistry Vol. 263; no. 26; pp. 13173 - 13178
• Main Authors: Pruss, R M, Stauderman, K A
• Format: Journal Article
• Language: English
• Published: Bethesda, MD Elsevier Inc 15-09-1988; American Society for Biochemistry and Molecular Biology
• Subjects: Animals; Biological and medical sciences; calcium; Calcium - metabolism; Cattle; Cell physiology; chromaffin cells; Enkephalin, Methionine - metabolism; enkephalins; Enkephalins - biosynthesis; Fundamental and applied biological sciences. Psychology; Ion Channels - metabolism; Lanthanum - pharmacology; Membrane and intracellular transports; Molecular and cellular biology; Nicotinic Acids - pharmacology; Oxadiazoles; Potassium Chloride - pharmacology; RNA, Messenger - metabolism; Tetradecanoylphorbol Acetate - pharmacology; Chlorides Potassium; Cell culture; Bovine; Calcium; Enkephalin; Fluorescence; Ion transport; Ionic channel; Chromaffin cell; Ester; Vertebrata; Regulation(control); Mammalia; Messenger RNA; Synthesis; Artiodactyla; Intracellular; Ungulata
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)37687-7

Treatment of bovine chromaffin cells with 40 mM KCl stimulates a 3-fold increase in total methionine enkephalin immunoreactivity (medium plus cells) and a 4-fold increase in proenkephalin mRNA (mRNAenk). These effects of KCl, which are dependent on extracellular calcium, can be blocked by treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA), although release of methionine enkephalin appears less affected. Using fura-2-loaded chromaffin cells and a dual-excitation wavelength spectrofluorometer, we have examined whether the actions of KCl and TPA on methionine enkephalin synthesis and release can be explained by changes in intracellular free calcium ([Ca2+]i). KCl produced a rapid 600 nM increase in [Ca2+]i from resting levels of approximately 170 nM. Subsequently, [Ca2+]i declined to a new steady-state plateau which was approximately 275 nM higher than the original resting levels. The postdepolarization plateau of [Ca2+]i was reduced by TPA, (-)-(R)-202,791 (a dihydropyridine calcium channel antagonist), and LaCl3 (a nonselective calcium channel blocker). TPA also inhibited potentiation of the KCl-stimulated plateau of [Ca2+]i due to (+)-(S)-202,791, a calcium channel agonist. In contrast, TPA had no effect on resting [Ca2+]i and only slightly inhibited the initial rapid KCl-stimulated increase in [Ca2+]i. The inhibitory effects were maintained for 24 h in the continuous presence of TPA. We conclude 1) that TPA inhibits enkephalin synthesis by inactivating dihydropyridine-sensitive voltage-dependent calcium channels, 2) that these channels alone maintain elevated [Ca2+]i following KCl depolarization, and 3) that sustained elevation in [Ca2+]i is necessary in order to increase enkephalin synthesis in KCl-treated chromaffin cells.