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Sodium pump; Sodium ion flux; Voltage clamp; ATPase, Na+/K+-; mRNA injection; (T. californica)
(Xenopus oocyte)
Abstract:
The Na+/K+-ATPase from electroplax of Torpedo californica was incorporated into the plasma membrane of Xenopus oocytes by microinjection of mRNA coding for the α- and β-subunit of the enzyme; the mRNAs were obtained by in vitro translation of cloned cDNAs (Noguchi et al. (1988) FEBS Lett. 225, 27–32). (1) Measurements of ouabain-sensitive membrane current revealed that the Na+/K+-ATPase of Torpedo is less sensitive to ouabain than the endogenous enzyme. (2) The ouabain-sensitive membrane currents in mRNA-injected oocytes exhibit similar voltage dependence as the currents generated by the endogenous ATPase of Xenopus oocytes; in particular, the current-voltage relation exhibits a maximum and a negative slope at potentials more positive than + 20 mV. (3) A maximum can also be detected if the rate of 22Na+ efflux is determined under different voltage-clamp conditions. If membrane current and rate of Na2+ efflux are determined simultaneously, a voltage-independent ratio between current and flux is obtained suggesting voltage-independent Na+-K+ stoichiometry. The data are compatible with a 3Na+–2K+ stoichiometry.
