The effects of buthionine sulfoximine treatment on diaphragm contractility and SERCA pump function in adult and middle aged rats

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.