Abstract
This study examined the effects of 10 days of buthionine sulfoximine (BSO) treatment on in vitro contractility and sarcoplasmic reticulum calcium pump (SERCA) expression and function in adult (AD; 6–8 months old) and middle aged (MA; 14–17 months old) rat diaphragm in both the basal state and following fatiguing stimulation. BSO treatment reduced the cellular concentrations of free glutathione (GSH) by >95% and oxidized glutathione (GSSG) by >80% in both age cohorts. GSH content in AD Control diaphragm was 32% higher (P < 0.01) than in MA Control, with no differences in GSSG. The ratio of GSH:GSSG, an indicator of cellular oxidative state, was 34.6 ± 7.4 in MA Control, 52.5 ± 10.1 in AD Control, 10.6 ± 1.7 in MA BSO, and 9.5 ± 1.1 in AD BSO (BSO vs. Control, P < 0.05). Several findings suggest that the effects of BSO treatment are age dependent. AD BSO diaphragm had 26% higher twitch and 28% higher tetanic force (both P < 0.05) than AD Controls, whereas no significant difference existed between the two MA groups. In contrast to our previous work on BSO-treated AD rats, BSO treatment did not influence maximal SERCA ATPase activity in MA rat diaphragm, nor did SERCA2a expression increase in BSO-treated MA diaphragm. Biotinylated iodoacetamide binding to SERCA1a, a specific marker of free cysteine residues, was reduced by 35% (P < 0.05) in AD Control diaphragm following fatiguing stimulation, but was not reduced in any other group. Collectively, these results suggest an important role for redox regulation in both contractility and SERCA function which is influenced by aging.
Highlights
It is well established that a basal level of reactive oxidants is necessary for normal muscle function (Reid et al 1993; Reid, 2001a; Smith and Reid 2006)
We have demonstrated that buthionine sulfoximine (BSO) treatment affects Ca2+ handling properties, with higher maximum rates of ATP hydrolysis and Ca2+ uptake by the sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) in freshly homogenized diaphragm (Tupling et al 2007)
BSO treatment resulted in significant declines (P < 0.05) in the body weights of AD and middle aged (MA) BSO groups with reductions of 4.2 Æ 0.5% and 8.8 Æ 1.3%, respectively, corresponding to significantly lower food intake
Summary
It is well established that a basal level of reactive oxidants is necessary for normal muscle function (Reid et al 1993; Reid, 2001a; Smith and Reid 2006). Reid (Reid, 2001b) has developed a biphasic model describing the inotropic response of muscle to changes in the redox state, that is, the balance between the oxidizing species and the antioxidant species. In this model, the optimum point for force production occurs in the presence of low concentrations of reactive oxygen or nitrogen species (ROS, RNS), where both increases and decreases in ROS concentration decrease contraction force. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.
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