Calcium currents and transients in co-cultured contracting normal and Duchenne muscular dystrophy human myotubes

1. The goal of the present study was to investigate differences in calcium movements between normal and Duchenne muscular dystrophy (DMD) human contra cting myotubes co-cultured with explants of rat spinal cord with attached d orsal root ganglia. Membrane potential, variations of intracellular calcium concentration and T- and L-type calcium currents were recorded. Further, a descriptive and quantitative study by electron microscopy of the ultrastru cture of the co-cultures was carried out. 2. The resting membrane potential was slightly less negative in DMD (-61.4 +/- 1.1 mV) than in normal myotubes (-65.5 +/- 0.9 mV). Both types of myotu be displayed spontaneous action potentials (mean firing frequency, 0.42 and 0.16 Hz, respectively), which triggered spontaneous calcium transients mea sured with Indo-1. 3. The time integral under the spontaneous Ca (2+) transients was significa ntly greater in DUD myotubes (97 +/- 8 nM s) than in normal myotubes (67 +/ - 13 nM s). 4. The L- and T-type current densities estimated from patch-clamp recording s were smaller in DMD cells (2.0 +/- 0.5 and 0.90 +/- 0.19 pA pF(-1), respe ctively) than in normal cells (3.9 +/- 0.7 and 1.39 +/- 0.30 pA pF(-1), res pectively). 5. The voltage-dependent inactivation relationships revealed a shift in the conditioning potential at which inactivation is half-maximal (V-h,V-0.5) o f the T- and L-type currents towards less negative potentials, from -72.1 /- 0.7 and -53.7 +/- 1.5 mV in normal cells to -61.9 +/- 1.4 and -29.2 +/- 1.4 mV in DMD cells, respectively, 6. Both descriptive and quantitative studies by electron microscopy suggest ed a more advanced development of DMD myotubes as compared to normal ones. This conclusion was supported by the significantly larger capacitance of th e DMD myotubes (408 +/- 45 pF) than of the normal myotubes (299 +/- 34 pF) of the same apparent size. 7. Taken together, these results show that differences in T- and L-type cal cium currents between normal and DMD myotubes cannot simply explain all obs erved alterations in calcium homeostasis in DMD myotubes, thus suggesting t hat other transmembrane calcium transport mechanisms must also be altered i n DMD myotubes compared vith normal myotubes.