Inhibition by nickel of the L-type Ca channel in guinea pig ventricular myocytes and effect of internal cAMP

The characteristics of nickel (Ni) block of L-type Ca current (I-Ca,I-L) we re studied in whole cell patch-clamped guinea pig cardiac myocytes at 37 de grees C in the absence and presence of 100 mu M cAMP in the pipette solutio n. Ni block of peak I-Ca,I-L had a dissociation constant (K-d) of 0.33 +/- 0.03 mM in the absence of cAMP, whereas in the presence of cAMP, the K-d wa s 0.53 +/- 0.05 mM (P = 0.006). Ni blocked Ca entry via Ca channels (measur ed as I-Ca,I-L integral over 50 ms) with similar kinetics (K-d of 0.35 +/- 0.03 mM in cAMP-free solution and 0.30 +/- 0.02 mM in solution with cAMP, P = not significant). Under both conditions, 5 mM Ni produced a maximal bloc k that was complete for the first pulse after application. Ni block of I-Ca ,I-L was largely use independent. Ni (0.5 mM) induced a positive shift (4 t o 6 mV) in the activation curve of I-Ca,I-L. The block of I-Ca,I-L by 0.5 m M Ni was independent of prepulse membrane potential (over the range of -120 to -40 mV). Ni (0.5 mM) also induced a significant shift in I-Ca,I-L inact ivation: by 6 mV negative in cAMP-free solution and by 4 mV positive in cel ls dialyzed with 100 mM cAMP. These data suggest that, in addition to block ing channel conductance by binding to a site in the channel pore, Ni may bi nd to a second site that influences the voltage-dependent gating of the L-t ype Ca channel. They also suggest that Ca channel phosphorylation causes a conformational change that alters some effects of Ni. The results may be re levant to excitation-contraction coupling studies, which have employed inte rnal cAMP dialysis, and where Ni has been used to block I-Ca,I-L and Ca ent ry into cardiac cells.