Residual sarcoplasmic reticulum Ca2+ concentration after Ca2+ release in skeletal myofibers from young adult and old mice

Contrasting information suggests either almost complete depletion of sarcoplasmic reticulum (SR) Ca2+ or significant residual Ca2+ concentration after prolonged depolarization of the skeletal muscle fiber. The primary obstacle to resolving this controversy is the lack of genetically encoded Ca2+ indicators targeted to the SR that exhibit low-Ca2+ affinity, a fast biosensor: Ca2+ off-rate reaction, and can be expressed in myofibers from adult and older adult mammalian species. This work used the recently designed low-affinity Ca2+ sensor (Kd = 1.66 mM in the myofiber) CatchER (calcium sensor for detecting high concentrations in the ER) targeted to the SR, to investigate whether prolonged skeletal muscle fiber depolarization significantly alters residual SR Ca2+ with aging. We found CatchER a proper tool to investigate SR Ca2+ depletion in young adult and older adult mice, consistently tracking SR luminal Ca2+ release in response to brief and repetitive stimulation. We evoked SR Ca2+ release in whole-cell voltage-clamped flexor digitorum brevis muscle fibers from young and old FVB mice and tested the maximal SR Ca2+ release by directly activating the ryanodine receptor (RyR1) with 4-chloro-m-cresol in the same myofibers. Here, we report for the first time that the Ca2+ remaining in the SR after prolonged depolarization (2 s) in myofibers from aging (similar to 220 mu M) was larger than young (similar to 132 mu M) mice. These experiments indicate that SR Ca2+ is far from fully depleted under physiological conditions throughout life, and support the concept of excitation-contraction uncoupling in functional senescent myofibers.