EFFECT OF NA(I) ON ACTIVITY AND VOLTAGE-DEPENDENCE OF THE NA K PUMP IN ADULT-RAT CARDIAC MYOCYTES/

We have measured the voltage dependence of the Na/K pump in isolated a dult rat cardiac myocytes using the whole-cell patch-clamp technique. In the presence of 1-2 mm Ba and 0.1 mm Cd and nominally Ca-free, Na/K pump current (I(p)) was measured as the change in current due to 1 mm ouabain. Voltage dependence of I(p) was measured between -140 and +40 or +60 mV using square voltage-pulse and voltage-ramp protocols, resp ectively. With 150 mm extracellular Na (Na(o)) and 5.4 mm extracellula r K (K(o)), we found that the Na/K pump shows a strong positive voltag e dependence between -140 and 0 mV and is voltage independent at posit ive potentials. Removing Na(o) reduced the voltage dependence at negat ive potentials with no effect at positive potentials. When K(o) was re duced, a negative slope appeared in the current-voltage (I-V) curve at positive potentials. We have investigated whether Na(i) (intracellula r Na) might also affect the voltage dependence of I(p) by varying Na i n the patch pipette (Na(pip)) between 20 and 85 mm. We found, as expec ted, that I(p) increased markedly as Na(pip) was raised, saturating at about 70 mm Na(pip) under these conditions. In contast, while I(p) sa turated near +20 mV and declined to about 40% of maximum at -120 mV, t here was no effect of Na(pip) under these conditions. In contrast, whi le I(p) saturated near +20 mV and declined to about 40% of maximum at -120 mV, there was no effect of Na(pip) on the voltage dependence of I (p). This suggests that neither Na(i) binding to the Na/K pump nor the conformational changes dependent on Na(i) binding are voltage depende nt. These results are consistent with extracellular ion binding within the field of the membrane but do not rule out the possibility that ot her steps, such as Na translocation, are also voltage dependent.