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We have used patch-clamp techniques to study the effects of arachidonic acid (AA) on the activity of the 70-pS K+ channel, the predominant type of the two apical K+ channels operating under physiological conditions in the thick ascending limb (TAL) of the rat kidney. Addition of 5-10 microM AA blocked the activity of the 70-pS K+ channel in both cell-attached and inside-out patches. The inhibitory effect of AA was specific, because application of 10 microM linoleic acid, oleic acid, or palmitic acid failed to mimic the effect of AA. The effect of AA could not be blocked by pretreatment of the TAL tubules with either 5 microM indomethacin (inhibitor of cyclooxygenase) or 4 microM cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate (CDC) (inhibitor of lipooxygenase). In contrast, addition of 5 microM 17-octadecynoic acid (17-ODYA), an inhibitor of P450 monooxygenases, abolised the effect of AA on the channel activity, indicating that the effect was mediated by cytochrome P450 metabolites of AA. Addition of 10 nM 20-hydroxyeicosatetraenoic acid (20-HETE), the main metabolite of the cytochrome P450 metabolic pathway in the medullary TAL, mimicked the inhibitory effect of 10 microM AA. However, addition of 100 nM 19-HETE or 17-HETE had no significant effects and 100 nM 20-carboxy AA (20-COOH) reduced the channel activity by only 20%, indicating that the inhibitory effect of 20-HETE was specific and responsible for the action of AA. Inhibition of the P450 metabolic pathway by either 5 microM 17-ODYA or 12, 12-dibromododec-11-enoic acid (DBDD) dramatically increased the channel activity by 280% in cell-attached patches. The stimulatory effect of 17-ODYA or DBDD was not observed in inside-out patches. The results strongly indicate that 20-HETE is a specific inhibitor for the 70-pS K+ channel and may play an important role in the regulation of the K+ channel activity in the TAL.

Effect of arachidonic acid on activity of the apical K+ channel in the thick ascending limb of the rat kidney.