In vivo intracellular Ca²⁺ profiles after eccentric rat muscle contractions: addressing the mechanistic bases for repeated bout protection

Eccentric contractions (ECC) are accompanied by the accumulation of intracellular calcium ions ([Ca²⁺]_i) and induce skeletal muscle damage. Suppressed muscle damage in repeated bouts of ECC is well characterized; however, whether it is mediated by altered Ca²⁺ profiles remains unknown. We tested the hypothesis that repeated ECC suppresses Ca²⁺ accumulation via adaptations in Ca²⁺ regulation. Male Wistar rats were divided into two groups: ECC single bout (ECC-SB) and repeated bout (ECC-RB). Tibialis anterior (TA) muscles were subjected to ECC (40 times, 5 sets) once (ECC-SB) or twice 14 days apart (ECC-RB). Under anesthesia, the TA muscle was loaded with Ca²⁺ indicator Fura 2-AM, and the 340/380 nm ratio was evaluated as [Ca²⁺]_i. Ca²⁺ handling proteins were measured by Western blots. ECC induced [Ca²⁺]_i increase in both groups, but ECC-RB evinced a markedly suppressed [Ca²⁺]_i (Time: <i>P</i> < 0.01, Group: <i>P</i> = 0.0357). Five hours post-ECC, in contrast to the localized [Ca²⁺]_i accumulation in ECC-SB, ECC-RB exhibited lower and more uniform [Ca²⁺]_i (<i>P</i> < 0.01). In ECC-RB, mitochondria Ca²⁺ uniporter complex (MCU) components MCU and MICU2 were significantly increased pre-second ECC bout (<i>P</i> < 0.01), and both SERCA1 and MICU1 were better preserved after contractions (<i>P</i> < 0.01). Fourteen days after novel ECC, skeletal muscle mitochondrial Ca²⁺ regulating proteins were elevated. Following subsequent ECC, [Ca²⁺]_i accumulation and muscle damage were suppressed and SERCA1 and MICU1 preserved. These findings suggest that tolerance to a subsequent ECC bout is driven, at least in part, by enhanced mitochondrial and sarcoplasmic reticulum Ca²⁺ regulation.<b>NEW & NOTEWORTHY</b> We demonstrated a reduced [Ca²⁺]_i profile with suppressed muscle damage after a repeated bout of ECC in vivo: the ECC-induced immediate [Ca²⁺]_i increase was suppressed and the persistence of increased [Ca²⁺]_i with localized accumulation was diminished after repeated ECC. This effect occurred consonant with the upregulation of the mitochondrial Ca²⁺ uniporter complex and better preservation of SERCA1 and MICU1. These findings suggest that the mechanistic bases for repeated bout protection involve adaptation of Ca²⁺ regulation.