In vivo Ca²⁺ dynamics during cooling after eccentric contractions in rat skeletal muscle

The effect of cooling on in vivo intracellular calcium ion concentration [Ca²⁺]_i after eccentric contractions (ECs) remains to be determined. We tested the hypothesis that cryotherapy following ECs promotes an increased [Ca²⁺]_i and induces greater muscle damage in two muscles with substantial IIb and IIx fiber populations. The thin spinotrapezius (SPINO) muscles of Wistar rats were used for in vivo [Ca²⁺]_i imaging, and tibialis anterior (TA) muscles provided greater fidelity and repeatability of contractile function measurements. SPINO [Ca²⁺]_i was estimated using fura 2-AM and the magnitude, location, and temporal profile of [Ca²⁺]_i determined as the temperature near the muscle surface post-ECs was decreased from 30°C (control) to 20°C or 10°C. Subsequently, in the TA, the effect of post-ECs cooling to 10°C on muscle contractile performance was determined at 1 and 2 days after ECs. TA muscle samples were examined by hematoxylin and eosin staining to assess damage. In SPINO, reducing the muscle temperature from 30°C to 10°C post-ECs resulted in a 3.7-fold increase in the spread of high [Ca²⁺]_i sites generated by ECs (P < 0.05). These high [Ca²⁺]_i sites demonstrated partial reversibility when rewarmed to 30°C. Dantrolene, a ryanodine receptor Ca²⁺ release inhibitor, reduced the presence of high [Ca²⁺] sites at 10°C. In the TA, cooling exacerbated ECs-induced muscle strength deficits via enhanced muscle fiber damage (P < 0.05). By demonstrating that cooling post-ECs potentiates [Ca²⁺]_i derangements, this in vivo approach supports a putative mechanistic basis for how postexercise cryotherapy might augment muscle fiber damage and decrease subsequent exercise performance.