Abstract: | TheCl secretory response ofcolonic cells to Ca2+-mediatedagonists is transient despite a sustained elevation of intracellular Ca2+. We evaluated the effects ofsecond messengers proposed to limit Ca2+-mediatedCl secretion on thebasolateral membrane,Ca2+-dependentK+ channel(KCa) in colonic secretorycells, T84. Neither protein kinase C (PKC) nor inositoltetrakisphosphate (1,3,4,5 or 3,4,5,6 form) affectedKCa in excised inside-out patches.In contrast, arachidonic acid (AA; 3 µM) potently inhibitedKCa, reducingNPo, the productof number of channels and channel open probability, by 95%. Theapparent inhibition constant for this AA effect was 425 nM. AAinhibited KCa in the presence ofboth indomethacin and nordihydroguaiaretic acid, blockers of thecyclooxygenase and lipoxygenase pathways. In the presence of albumin,the effect of AA on KCa wasreversed. A similar effect of AA was observed onKCa during outside-out recording.We determined also the effect of thecis-unsaturated fatty acid linoleate,the trans-unsaturated fatty acidelaidate, and the saturated fatty acid myristate. At 3 µM, all ofthese fatty acids inhibited KCa,reducing NPo by 72-86%. Finally, the effect of the cytosolic phospholipaseA2 inhibitorarachidonyltrifluoromethyl ketone(AACOCF3) on thecarbachol-induced short-circuit current(Isc) responsewas determined. In the presence ofAACOCF3, the peakcarbachol-inducedIsc response wasincreased ~2.5-fold. Our results suggest that AA generation inducedby Ca2+-mediated agonists maycontribute to the dissociation observed between the rise inintracellular Ca2+ evoked by theseagonists and the associatedCl secretory response. |