Abstract
Ca2+-activated Cl− currents were studied in isolated cells from rat portal vein smooth muscle in short-term primary culture using the whole-cell patch-clamp technique. Cl− currents can be activated separately by Ca2+ release from intracellular stores (in response to external applications of caffeine or noradrenaline) and by Ca2+ influx through voltage-dependent Ca2+ channels. The effects of several Cl− channel blockers and of spironolactone (a substance known to reduce internal Ca2+ loading) on both Cl− and Ca2+ currents were examined. Diisothiocyanostilbene-2,2′-disulfonic acid (DIDS), anthracene-9-carboxylic acid (9-AC) and diphenylamine-2,2′-dicarboxylic acid (DPC) inhibited the Ca2+-activated Cl− current (IC50 values between 16.5 and 306 μM) with no effects on the inward Ca2+ current and on internal Ca2+ loading (tested by measuring the Ca2+-activated K+ current). These results indicate that the inhibition of Cl− current by these compounds is due to a direct interaction with the Cl− channel. In contrast, spironolactone inhibited both K+ and Cl− currents (IC50=7.6 μM) by reducing the amount of Ca2+ located in the internal stores, whereas the Cl− current activated by Ca2+ current through T-type Ca2+ channels was unchanged. This preparation and the protocols developed in this study appears to be appropriate for analysis of substances interfering with Cl− channels or intracellular Ca2+ stores.
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Baron, A., Pacaud, P., Loirand, G. et al. Pharmacological block of Ca2+-activated Cl− current in rat vascular smooth muscle cells in short-term primary culture. Pflügers Arch 419, 553–558 (1991). https://doi.org/10.1007/BF00370294
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DOI: https://doi.org/10.1007/BF00370294