Developmental changes in the calcium currents in embryonic chick ventricular myocytes

Abstract

Using the patch-clamp technique, we recorded whole-cell calcium current from isolated cardiac myocytes dissociated from the apical ventricles of 7-day and 14-day chick embryos. In 70% of 14-day cells after 24 hr in culture, two component currents could be separated from total ICa activated from a holding potential (Vh) of -80 mV. L-type current (IL) was activated by depolarizing steps from Vh -30 or -40 mV. The difference current (IT) was obtained by subtracting IL from ICa. IT could also be distinguished pharmacologically from IL in these cells. IT was selectively blocked by 40-160 microM Ni2+, whereas IL was suppressed by 1 microM D600 or 2 microM nifedipine. The Ni2(+)-resistant and D600-resistant currents had activation thresholds and peak voltages that were near those of IT and IL defined by voltage threshold, and resembled those in adult mammalian heart. In 7-day cells, IT and IL could be distinguished by voltage threshold in 45% (S cells), while an additional 45% of 7-day cells were nonseparable (NS) by activation voltage threshold. Nonetheless, in most NS cells, ICa was partly blocked by Ni2+ and by D600 given separately, and the effects were additive when these agents were given together. Differences among the cells in the ability to separate IT and IL by voltage threshold resulted largely from differences in the position of the steady-state inactivation and activation curves along the voltage axis. In all cells at both ages in which the steady-state inactivation relation was determined with a double-pulse protocol, the half-inactivation potential (V1/2) of the Ni2(+)-resistant current IL averaged -18 mV. In contrast, V1/2 of the Ni2(+)-sensitive IT was -60 mV in 14-day cells, -52 mV in 7-day S cells, and -43 mV in 7-day NS cells. The half-activation potential was near -2 mV for IL at both ages, but that of IT was -38 mV in 14-day and -29 mV in 7-day cells. Maximal current density was highly variable from cell to cell, but showed no systematic differences between 7-day and 14-day cells. These results indicate that the main developmental change that occurs in the components of ICa is a negative shift with embryonic age in the activation and inactivation relationships of IT along the voltage axis.