Dual roles of phorbol 12, 13-dibutyrate in the regulation of guinea-pig gastric contraction.

Abstract

We performed experiments to investigate the actions of protein kinase C(PKC) on mechanical contraction during agonist stimulation in guinea pig stomach. We used carbachol and high K condition to enhance mechanical contraction by mobilizing intracellular Ca2+ and increasing Ca2+ influx through voltage-dependent Ca(2+)-channel, respectively. Phorbol 12, 13-dibutyrate (PDBu) increased spontaneous contractions sensitive to verapamil (438 +/- 82.2%, n = 7) and potentiated high K-induced contraction (189 +/- 22.5%, n = 5). However, carbachol (CCh)-induced contractions in PDBu-treated condition depended on extracellular Ca2+. In the presence of extracellular Ca2+, CCh-induced contraction was potentiated, while it was suppressed in the absence of extracellular Ca(2+)-preloaded muscle strips. To prove the hypothesis that such phenomena might be related with changes of myoplasmic Ca2+ concentration, we investigated the effect of PDBu on voltage-dependent Ca2+ current(ICa) and CCh-induced Ca2+ activated K current(IK(Ca)) transient using whole-cell voltage clamp technique. For recording voltage-dependent Ca2+ current(ICa), 10 mM Ba2+, instead of Ca2+, was used to enhance the current size. Voltage-dependent Ba2+ current(IBa) was increased by PDBu (212 +/- 32.2% of steady state currents, n = 5), while CCh-induced increase of IK(Ca) transient was inhibited by PDBu (n = 5), the changes of which were similar to those of muscle contractions. To analyze the steps involved in the inhibition of CCh-induced IK(Ca) transient by PDBu, we investigated the effect of PDBu on IK(Ca) in the cells perfused with Ins (1, 4, 5)P3. However, Ins (1, 4, 5)P3 induced IK(Ca) was not inhibited by the treatment with phorbol ester. From these results, it is concluded that inhibition of phosphatidylinositol phospholipase C(PI-PLC) system and potentiation of ICa by PKC are important regulatory mechanisms in agonist-induced muscle contraction.