Jr. Stimers et al., EFFECT OF NA(I) ON ACTIVITY AND VOLTAGE-DEPENDENCE OF THE NA K PUMP IN ADULT-RAT CARDIAC MYOCYTES/, The Journal of membrane biology, 135(1), 1993, pp. 39-47
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.