Evidence for Na⁺/Ca²⁺exchange in intact single skeletal muscle fibers from the mouse

The myoplasmic free Ca2+ concentration ([Ca2+]i) was measured in intact single fibers from mouse skeletal muscle with the fluorescent Ca2+ indicator indo 1. Some fibers were perfused in a solution in which the concentration of Na+ was reduced from 145.4 to 0.4 mM (low-Na+ solution) in an attempt to activate reverse-mode Na+/Ca2+ exchange (Ca2+ entry in exchange for Na+ leaving the cell). Under normal resting conditions, application of low-Na+ solution only increased [Ca2+]i by 5.8 +/- 1.8 nM from a mean resting [Ca2+]i of 42 nM. In other fibers, [Ca2+]i was elevated by stimulating sarcoplasmic reticulum (SR) Ca2+ release with caffeine (10 mM) and by inhibiting SR Ca2+ uptake with 2,5-di(tert-butyl)-1,4-benzohydroquinone (TBQ; 0.5 microM) in an attempt to activate forward-mode Na+/Ca2+ exchange (Ca2+ removal from the cell in exchange for Na+ influx). These two agents caused a large increase in [Ca2+]i, which then declined to a plateau level approximately twice the baseline [Ca2+]i over 20 min. If the cell was allowed to recover between exposures to caffeine and TBQ in a solution in which Ca2+ had been removed, the increase in [Ca2+]i during the second exposure was very low, suggesting that Ca2+ had left the cell during the initial exposure. Application of caffeine and TBQ to a preparation in low-Na+ solution produced a large, sustained increase in [Ca2+]i of approximately 1 microM. However, when cells were exposed to caffeine and TBQ in a low-Na+ solution in which Ca2+ had been removed, a sustained increase in [Ca2+]i was not observed, although [Ca2+]i remained higher and declined slower than in normal Na+ solution. This suggests that forward-mode Na+/Ca2+ exchange contributed to the fall of [Ca2+]i in normal Na+ solution, but when extracellular Na+ was low, a prolonged elevation of [Ca2+]i could activate reverse-mode Na+/Ca2+ exchange. The results provide evidence that skeletal muscle fibers possess a Na+/Ca2+ exchange mechanism that becomes active in its forward mode when [Ca2+]i is increased to levels similar to that obtained during contraction.