Increased calcium entry into dystrophin-deficient muscle fibres of MDX andADR-MDX mice is reduced by ion channel blockers

1. Single fibres were enzymatically isolated from interosseus muscles of dy strophic MDX mice, myotonic-dystrophic double mutant ADR-MDX, mice and C57B L/10 controls. The fibres were kept in cell culture for up to 2 weeks for t he study of Ca2+ homeostasis and sarcolemmal Ca2+ permeability. 2. Resting levels of intracellular free Ca2+, determined with the fluoresce nt Ca2+ indicator fura-2, were slightly higher in MDX (63 +/- 20 nm; means +/- S.D.; n = 454 analysed fibres) and ADR-MDX (65 +/- 12 nM; n = 87) fibre s than in controls (51 +/- 20 nM; n = 265). 3.The amplitudes of electrically induced Ca2+ transients did not differ bet ween MDS fibres and controls. Decay time constants of Ca2+ transients range d between 10 and 55 ms in both genotypes. In 50% of MDX fibres (n = 68), bu t in only 20% of controls (n = 54), the decay time constants were > 35 ms. 4. Bath application of Mn2+ resulted in a progressive quench of fura-2 fluo rescence emitted from the fibres. The quench rate was about 2 times higher in MDX fibres (3.98 +/- 1.9% min(-1); n = 275) than in controls (2.03 +/- 1 .4% min(-1); n = 204). The quench rate in ADR-MDX fibres (2.49 +/- 1 4% min (-1); n = 87) was closer to that of controls. 5. The Mn2+ influx into MDX fibres was reduced to 10% by Gd3+, to 19% by La 3+ and to 47% by Ni2+ (all at 50 mu M). Bath application of 50 mu M amilori de inhibited the Mn2+ influx to 37%. 6. We conclude that in isolated, resting MDX muscle fibres the membrane per meability fur divalent cations is increased. The presumed additional influx of Ca2+ occurs through ion channels, but is well compensated for by effect ive cellular Ca2+ transport systems. The milder dystrophic phenotype of ADR -MDX mice is correlated with a smaller increase of their sarcolemmal Ca2+ p ermeability.