Intracellular Ca2+ transients were recorded from frog twitch muscle fibres in response to voltage-clamp depolarizing pulses, using arsenazo III as an intracellular Ca2+ indicator. The effect of the activation of protein kinase C (PKC) on the Ca2+ transients was studied. With 1 microM phorbol 12,13-dibutyrate (PDBu), a PKC activator, the peak of the Ca2+ transients increased to about 120% of control during the first 0.5 h, and then decreased gradually to a plateau of 44% of control within the following 2 h. This effect of PDBu could be alleviated significantly by PKC inhibitors, 10 microM polymyxin B (PMB) or 30 microM 1-(5-isoquinolinylsulphonyl)-2-methyl-piperazine (H-7). Moreover, PDBu caused an upward shift of the strength/duration curve. In Li(+)-loaded muscle fibres the Ca2+ transients could not fully recover after 80 mM K+ exposure for 15 min, while the post-K+ Ca2- transients could be completely restored in the fibres not loaded with Li+. In the presence of 10 microM PMB or 30 microM H-7, a full restoration of the post-K+ Ca2+ transients was seen in Li(+)-loaded fibres. PMB supplemented after high-K+ exposure also could result in a complete recovery of the post-K+ Ca2+ transients in Li(+)-loaded fibres. The role of PKC in modulating excitation-contraction coupling in frog twitch muscle fibres is clearly indicated, but the mechanism(s) and physiological significance remain to be established.