Endogenous and maximal sarcoplasmic reticulum calcium content and calsequestrin expression in type I and type II
C R Lamboley1, R M Murphy, M J McKenna
1G. D. Lamb: Department of Zoology, La Trobe University, Melbourne, Victoria 3086, Australia. g.lamb@latrobe.edu.au.
Abstract:
The relationship between sarcoplasmic reticulum (SR) Ca(2+) content and calsequestrin (CSQ) isoforms was investigated in human skeletal muscle. A fibre-lysing assay was used to quantify the endogenous Ca(2+) content and maximal Ca(2+) capacity of the SR in skinned segments of type I and type II fibres from vastus lateralis muscles of young healthy adults. Western blotting of individual fibres showed the great majority contained either all fast or all slow isoforms of myosin heavy chain (MHC), troponins C and I, tropomyosin and SERCA, and that the strontium sensitivity of the force response was closely indicative of the troponin C isoform present. The endogenous SR Ca(2+) content was slightly lower in type I compared to type II fibres (0.76 ± 0.03 and 0.85 ± 0.02 mmol Ca(2+) per litre of fibre, respectively), with virtually all of this Ca(2+) evidently being in the SR, as it could be rapidly released with a caffeine-low [Mg(2+)] solution (only 0.08 ± 0.01 and <0.07 mmol l(-1), respectively, remaining). The maximal Ca(2+) content that could be reached with SR Ca(2+) loading was 1.45 ± 0.04 and 1.79 ± 0.03 mmol l(-1) in type I and type II fibres, respectively (P < 0.05). In non-lysed skinned fibres, where the SR remained functional, repeated cycles of caffeine-induced Ca(2+) release and subsequent Ca(2+) reloading similarly indicated that (i) maximal SR Ca(2+) content was lower in type I fibres than in type II fibres (P < 0.05), and (ii) the endogenous Ca(2+) content represented a greater percentage of maximal content in type I fibres compared to type II fibres (∼59% and 41%, respectively, P < 0.05). Type II fibres were found on average to contain ∼3-fold more CSQ1 and ∼5-fold less CSQ2 than type I fibres (P < 0.001). The findings are consistent with the SR Ca(2+) content characteristics in human type II fibres being primarily determined by the CSQ1 abundance, and in type I fibres by the combined amounts of both CSQ1 and CSQ2.
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