SR Calcium Release Channel Regulation in Aging Muscle

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DESCRIPTION (provided by applicant): A decline in skeletal muscle function is a major contributor to decreased mobility and independence in the elderly. In addition, muscle weakness in a significant factor contributing to falls resulting in serious injury. Thus, although skeletal muscle weakness is a major public health concern, the molecular mechanisms underlying skeletal muscle dysfunction in aging are not well understood. The long-term objective of this work is to clarify the mechanisms underlying aging-induced skeletal muscle weakness. Muscle contraction is initiated by Ca}+ efflux from the sarcoplasmic reticulum (SR) via the SR Ca}+ release channel/ryanodane receptor (RyR1). SR Ca}+ release is depressed in muscle from aged individuals. Thus incomplete contractile activation contributes to muscle weakness. Being the major SR Ca}+ effiux pathway, RyR1 dysfunction is a likely contributor to impaired Ca}+ release in aging muscle. In vitro oxidation of RyR1 channels modifies their response to physiological effectors. In addition, aging slows the RyR1 turn-over rate potentially allowing the accumulation of detrimental protein modifications. Preliminary experiments suggest 1) an age-associated decline in ATP and caffeine activation of RyR1; and 2) modification of a reactive cysteine in the RyR1 CaM binding domain. Therefore, I hypothesize that RyR1 from aged skeletal muscle is oxidatively modified and its regulation by important endogenous effectors is impaired. To test this hypothesis, Aim 1 will determine whether aging alters the regulation of RyR1 by adenine nucleotides and effector ions; Aim 2 will determine the impact of aging on the functional interaction of RyR1 with the accessory protein calmodulin; and Aim 3 will determine the extent of age-associated oxidation of RyR1 cysteine residues. These studies will use a variety of techniques including [3H]ryanodine and [35S]calmodulin binding to SR vesicles to assess RyR1 regulation by physiological regulators, single channel analysis to determine the molecular mechanisms by which aging alters RyR1 channel function, and monobromobimane determination of the free thiol content of SR vesicles and purified RyR1 channels. This work will further our understanding of the molecular mechanisms underlying age-associated muscle weakness and aid in the development of therapies to treat and prevent skeletal muscle frailty.