Na,K-ATPase α2 activity in mammalian skeletal muscle T-tubules is acutely stimulated by extracellular K(+)

The Na,K-ATPase α2 isoform is the predominant Na,K-ATPase in adult skeletal muscle and the sole Na,K-ATPase in the transverse tubules (T-tubules). In quiescent muscles, the α2 isozyme operates substantially below its maximal transport capacity. Unlike the α1 isoform, the α2 isoform is not required for maintaining resting ion gradients or the resting membrane potential, canonical roles of the Na,K-ATPase in most other cells. However, α2 activity is stimulated immediately upon the start of contraction and, in working muscles, its contribution is crucial to maintaining excitation and resisting fatigue. Here, we show that α2 activity is determined in part by the K(+) concentration in the T-tubules, through its K(+) substrate affinity. Apparent K(+) affinity was determined from measurements of the K(1/2) for K(+) activation of pump current in intact, voltage-clamped mouse flexor digitorum brevis muscle fibers. Pump current generated by the α2 Na,K-ATPase, Ip, was identified as the outward current activated by K(+) and inhibited by micromolar ouabain. Ip was outward at all potentials studied (−90 to −30 mV) and increased with depolarization in the subthreshold range, −90 to −50 mV. The Q(10) was 2.1 over the range of 22–37°C. The K(1/2,K) of Ip was 4.3 ± 0.3 mM at −90 mV and was relatively voltage independent. This K(+) affinity is lower than that reported for other cell types but closely matches the dynamic range of extracellular K(+) concentrations in the T-tubules. During muscle contraction, T-tubule luminal K(+) increases in proportion to the frequency and duration of action potential firing. This K(1/2,K) predicts a low fractional occupancy of K(+) substrate sites at the resting extracellular K(+) concentration, with occupancy increasing in proportion to the frequency of membrane excitation. The stimulation of preexisting pumps by greater K(+) site occupancy thus provides a rapid mechanism for increasing α2 activity in working muscles.