Purinergic activation of spontaneous transient outward currents in guinea pig taenia colonic myocytes

Abstract

Spontaneous transient outward currents (STOCs) were recorded from smooth muscle cells of the guinea pig taenia coli using the whole cell patch-clamp technique. STOCs were resolved at potentials positive to −50 mV. Treating cells with caffeine (1 mM) caused a burst of outward currents followed by inhibition of STOCs. Replacing extracellular Ca²⁺with equimolar Mn²⁺caused STOCs to “run down.” Iberiotoxin (200 nM) or charybdotoxin (ChTX; 200 nM) inhibited large-amplitude STOCs, but small-amplitude “mini-STOCs” remained in the presence of these drugs. Mini-STOCs were reduced by apamin (500 nM), an inhibitor of small-conductance Ca²⁺-activated K⁺channels (SK channels). Application of ATP or 2-methylthioadenosine 5′-triphosphate (2-MeS-ATP) increased the frequency of STOCs. The effects of 2-MeS-ATP persisted in the presence of charybdotoxin but were blocked by combination of ChTX (200 nM) and apamin (500 nM). 2-MeS-ATP did not increase STOCs in the presence of pyridoxal phosphate 6-azophenyl-2′,4′-disulfonic acid, a P₂receptor blocker. Similarly, pretreatment of cells with U-73122 (1 μM), an inhibitor of phospholipase C (PLC), abolished the effects of 2-MeS-ATP. Xestospongin C, an inositol 1,4,5-trisphosphate (IP₃) receptor blocker, attenuated STOCs, but these events were not affected by ryanodine. The data suggest that purinergic activation through P_2Yreceptors results in localized Ca²⁺release via PLC- and IP₃-dependent mechanisms. Release of Ca²⁺is coupled to STOCs, which are composed of currents mediated by large-conductance Ca²⁺-activated K⁺channels and SK channels. The latter are thought to mediate hyperpolarization and relaxation responses of gastrointestinal muscles to inhibitory purinergic stimulation.