Membrane properties and calcium ion activity in skeletal muscle fibres of the dystrophic mouse
The ReJI29 murine model of muscular dystrophy was employed to investigate the properties of skeletal muscle plasmalemma and calcium ion movements during muscle wastage, with the intention of determining the route of calcium influx, and the efficacy of calcium ion blockers in preventing this influx. Electrophysiological parameters (Resting membrane potential [RMP] and input resistance) reached adult magnitude in normal soleus and extensor digitorum longus (EDL) by 4 weeks and 3 weeks respectively. Electrophysiological parameters in dystrophic soleus developed in a similar maimer to normal muscle, but RMP in dystrophic EDL was reduced and input resistance was elevated suggesting the presence of a population of regenerating fibres. Twitch and contracture tension development reached mature levels by 4 weeks in normal soleus muscle, but development was prolonged to 10 weeks in EDL, due to the slower development of type II fibres. Contracture tension was markedly smaller in EDL possibly due to fibre type-related differences in the concentrations of calcium sequestering proteins.Twitch and contracture tension in dystrophic soleus was identical to normal. In dystrophic EDL, 4 week old muscle did not generate tension from either electrical or chemical stimulation. Older muscles generated tension but smaller than their normal counterparts. Twitch and contracture tomography revealed similarities between dystrophic EDL and immature normal EDL corroborating with the conclusions made from the electrophysiological experiments. Caffeine contractures generated in glycinerated normal soleus muscle, incubated in low calcium saline, and then bathed in high calcium saline reached 115-185% the magnitude of caffeine contractures generated prior to incubation. 4 week dystrophic soleus attained similar sized contractures, but contractures were reduced to 77-90% in older muscles. This technique was found to be inappropriate in the study of EDL. Caffeine contractures generated by normal soleus in normal saline, after incubation in low calcium saline, reached the magnitude of contractures generated in high calcium saline after 30 minutes. Contractures generated by dystrophic soleus remained identical to those generated in high calcium saline suggesting that the sarcoplasmic reticulum in these fibres was unable to cope with a high calcium load. This result gave the first indication that soleus muscle was affected by muscular dystrophy. Membrane bound calcium derived from 45Calcium influx studies was 2nmoles/mg tissue/30 minutes in soleus and 1 Snmoles/mg tissue weight/30 minutes in EDL. Intracellular influx was 0.5-0.6nmoles/mg tissue weight/ 30 minutes in both muscles. Membrane bound calcium was elevated in dystrophic EDL to 2.5-3.8nmoles/mg tissue weight/30 minutes, but influx was normal in both dystrophic soleus and EDL, casting doubt on whether the plasmalemma is more permeable to calcium. The increase in membrane bound calcium may be artefactual. Cadmium blocked influx in both normal and dystrophic muscles. Blot weights showed continual growth in normal muscles although the rate decreased after 10 weeks. In dystrophic muscles growth ceased after 10 weeks. NCP data was limited but did show a decrease in dystrophic muscle, followed by an increase. Tension as a function of tissue mass, revealed that the contractile apparatus in normal soleus matured within 2 weeks, and by 10 weeks in EDL due to the differences in myosin isoforms present in each fibre type. These ratios were elevated in dystrophic muscle indicating calcium accumulation in the sarcoplasmic reticulum.. The results showed that EDL muscle was more severely affected by muscular dystrophy. Between 2 and 4 weeks, the muscle underwent degeneration via an unidentified process. There was regeneration, and the regenerating fibres appeared to be normal. Soleus muscle appeared resistant to the disease but succumbed under unusual chemical stresses. Some aspects of development of normal muscle were also considered.
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