New Findings: Hindlimb Unloading Causes Nucleocytoplasmic Ca 2+ Overload and DNA Damage in Skeletal Muscle.

Disuse atrophy of skeletal muscle is associated with a severe imbalance in cellular Ca ²⁺ homeostasis and marked increase in nuclear apoptosis. Nuclear Ca ²⁺ is involved in the regulation of cellular Ca ²⁺ homeostasis. However, it remains unclear whether nuclear Ca ²⁺ levels change under skeletal muscle disuse conditions, and whether changes in nuclear Ca ²⁺ levels are associated with nuclear apoptosis. In this study, changes in Ca ²⁺ levels, Ca ²⁺ transporters, and regulatory factors in the nucleus of hindlimb unloaded rat soleus muscle were examined to investigate the effects of disuse on nuclear Ca ²⁺ homeostasis and apoptosis. Results showed that, after hindlimb unloading, the nuclear envelope Ca ²⁺ levels ([Ca ²⁺ ] _NE ) and nucleocytoplasmic Ca ²⁺ levels ([Ca ²⁺ ] _NC ) increased by 78% ( p < 0.01) and 106% ( p < 0.01), respectively. The levels of Ca ²⁺ -ATPase type 2 (Ca ²⁺ -ATPase2), Ryanodine receptor 1 (RyR1), Inositol 1,4,5-tetrakisphosphate receptor 1 (IP ₃ R1), Cyclic ADP ribose hydrolase (CD38) and Inositol 1,4,5-tetrakisphosphate (IP ₃ ) increased by 470% ( p < 0.001), 94% ( p < 0.05), 170% ( p < 0.001), 640% ( p < 0.001) and 12% ( p < 0.05), respectively, and the levels of Na ⁺ /Ca ²⁺ exchanger 3 (NCX3), Ca ²⁺ /calmodulin dependent protein kinase II (CaMK II) and Protein kinase A (PKA) decreased by 54% ( p < 0.001), 33% ( p < 0.05) and 5% ( p > 0.05), respectively. In addition, DNase X is mainly localized in the myonucleus and its activity is elevated after hindlimb unloading. Overall, our results suggest that enhanced Ca ²⁺ uptake from cytoplasm is involved in the increase in [Ca ²⁺ ] _NE after hindlimb unloading. Moreover, the increase in [Ca ²⁺ ] _NC is attributed to increased Ca ²⁺ release into nucleocytoplasm and weakened Ca ²⁺ uptake from nucleocytoplasm. DNase X is activated due to elevated [Ca ²⁺ ] _NC , leading to DNA fragmentation in myonucleus, ultimately initiating myonuclear apoptosis. Nucleocytoplasmic Ca ²⁺ overload may contribute to the increased incidence of myonuclear apoptosis in disused skeletal muscle.