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.