Coupling strength between localized Ca2+ transients and K+ channels is regulated by protein kinase C

Localized Ca2+ transients resulting from inositol trisphosphate (IP3)-depen dent Ca2+ release couple to spontaneous transient outward currents (STOCs) in murine colonic myocytes. Confocal microscopy and whole cell patch-clamp techniques were used to investigate coupling between localized Ca2+ transie nts and STOCs. Colonic myocytes were loaded with fluo 3. Reduction in exter nal Ca2+ ([ Ca2+](o)) reduced localized Ca2+ transients but increased STOC amplitude and frequency. Simultaneous recordings of Ca2+ transients and STO Cs showed increased coupling strength between Ca2+ transients and STOCs whe n [Ca2+](o) was reduced. Gd3+ (10 muM) did not affect Ca2+ transients but i ncreased STOC amplitude and frequency. Similarly, an inhibitor of Ca2+ infl ux, 1-2-(4-methoxyphenyl)-2-[3-(4-methoxyphenyl) propoxy]ethyl-1H-imidazole (SKF-96365), increased STOC amplitude and frequency. A protein kinase C (P KC) inhibitor, GF-109203X, also increased the amplitude and frequency of ST OCs but had no effect on Ca2+ transients. Phorbol 12-myristate 13-acetate ( 1 muM) reduced STOC amplitude and frequency but did not affect Ca2+ transie nts. 4 alpha -Phorbol (1 muM) had no effect on STOCs or Ca2+ transients. Si ngle channel studies indicated that large-conductance Ca2+-activated K+ (BK ) channels were inhibited by a Ca2+-dependent PKC. In summary 1) Ca2+ relea se from IP3 receptor-operated stores activates Ca2+-activated K+ channels; 2) Ca2+ influx through nonselective cation channels facilitates activation of PKC; and 3) PKC reduces the Ca2+ sensitivity of BK channels, reducing th e coupling strength between localized Ca2+ transients and BK channels.