Age-dependent expression of the apamin-sensitive calcium-activated K+ channel in fast and slow rat skeletal muscle.

An altered expression of the apamin-sensitive K+ channel from skeletal muscle is apparently implicated in human myotonic dystrophy (MD). We found, in rats, that the expression of this channel depends on age and the type of muscle. This result may be one of the bases of the different susceptibilities of fast and slow muscles to drug-induced myotonia.

Neural control of the expression of a Ca(2+)-activated K+ channel involved in the induction of myotonic-like characteristics.

1. Expression of the apamin-sensitive K+ channel (SK+) in rat skeletal muscle is neurally regulated. The regulatory effect of the nerve over the expression of some muscle ion channels has been attributed to the electrical activity triggered by the nerve and/or to a trophic effect of some molecules transported from the soma to the axonal endings. 2. SK+ channels apparently are involved in myotonic dystrophy (MD), therefore understanding the factors that regulate their expression may ultimately have important clinical relevance. 3. To establish if axoplasmic transport is involved in this process, we used two experimental approaches in adult rats: (a) Both sciatic nerves were severed, leaving a short or a long nerve stump attached to the anterior tibialis (AT). (b) Colchicine or vinblastine (VBL), two axonal transport blockers of different potencies, was applied on one leg to the sciatic nerve. To determine whether electrical activity affects the expression of SK+ channels, denervated AT were directly stimulated. The corresponding contralateral muscles were used as controls.

Energetic metabolism and fatigability in experimental myotonia.

Experimental myotonia was induced in rats by 2,4-dichloro-phenoxyacetic acid (2,4-D). After 4 to 24 h of treatment, the anterior tibialis muscles exhibited increased fatigue at low frequency (30 Hz) nerve stimulation, but they developed normal tension at high-frequency (100 Hz) stimulation. Glycogen content and the activities of glycogen phosphorylase, lactate dehydrogenase and malate dehydrogenase remained normal. The absence of correlation between fatigability and energetic metabolism in this experimental model of myotonia suggests a dysfunction in excitation-contraction coupling.

Intramuscular pH and endurance are abnormal in skeletal muscles from rats with experimental myotonia.

The relation between resistance to fatigue and intramuscular pH was studied in fast muscles (anterior tibialis and extensor digitorum longus) from rats treated with 2,4-dichlorophenoxyacetic acid (2,4-D) to induce myotonia. Fatiguability was studied in muscles indirectly stimulated at 30 Hz (330 ms/s; 1 train/s) for 2 min. The resistance to fatigue decreased significantly 1 h after drug treatment and remained low 24 h later. The intramuscular pH was lower than normal in resting muscles from 2,4-D-treated rats. After 2 min of stimulation the pH decreased in both control and drug-treated muscles. However, this decrement was reduced in the experimental muscles. The pH of control and of 2,4-D-treated muscles were similar after the stimulation period, but only the drug-treated muscles were fatigued. Therefore, a decrease in intramuscular pH would not be the cause of the observed decrease in muscle resistance to fatigue after 2,4-D treatment. The reduced endurance of drug-treated muscles could not be attributed to impaired neuromuscular transmission.

Sarcoplasmic reticulum ATPase in tibialis anterior muscle from rats with experimental myotonia.

The sarcoplasmic reticulum ATPase was studied (after 3 h to 14 days) in rats treated with 2,4-dichlorophenoxyacetate to induce myotonia. It was found that ATPase decreased in the treated rats after the establishment of myotonia.

Age-related responses of skeletal muscle after ectopic innervation, with particular reference to 16S acetylcholinesterase, in adult rats.

The formation of ectopic junctions between the foreign fibular nerve and the soleus muscle of young (35-day-old) and mature (200-day-old) adult rats was induced by severing the normal nerve 4 weeks after implanting the foreign nerve. The various molecular forms of acetylcholinesterase (AChE) were studied both at the implanted region and at the original denervated endplates. The velocity of contraction was also studied. In young rats the 16S form was first detected in the ectopic junctions around day 5 after reinnervation; this form rapidly increased during the following weeks, reaching a plateau by day 20. By contrast, in mature rats the appearance of the 16S AChE was dramatically delayed; in fact, it could not be observed before day 80 after reinnervation. (The 16S AChE form appeared at day 20 after reinnervation in the original denervated endplates of young rats; however, at the same time, no effect was observed in mature animals.) The original, slow muscle fibers of the soleus became faster upon reinnervation; this change occurred also much earlier in younger than in mature rats. Our results indicate a loss of plasticity in the skeletal muscle of mature rats. We suggest caution in the use of the ectopic innervation model to study development in mature adult rats.

Axonal transport blockade and denervation have qualitatively different effects upon skeletal muscle metabolism.

The activity and isoenzyme pattern of muscle lactic dehydrogenase (LDH) was measured at different times after axonal transport blockade by colchicine or after denervation. After denervation, total LDH activity decreased and the isoenzyme pattern was altered, LDH-1 being the most affected form. In contrast, after axonal transport blockade there was a decrease in LDH activity but the isoenzyme pattern was not modified. Denervation abolishes both nerve-evoked muscle activity and the release of neuro trophic substances from the nerve whereas colchicine blocks axonal transport without affecting the nerve capacity to conduct action potentials or neuromuscular transmission. It is then concluded that nerve-evoked muscle activity is the most important factor in the regulation of muscle LDH isoenzyme distribution. On the other hand, muscle metabolism can also be regulated by axonally transported molecules. The results presented here show that there is a qualitative difference between the effects of denervation and those of axonal transport blockade upon the muscle, since only denervation altered the isoenzyme pattern of muscle LDH.

Axonal transport dysfunction in dystrophia myotonica.

Axonal transport of acetylcholinesterase (AChE) was measured in the median and sural nerves of a subject who suffered from dystrophia myotonica and in a control subject. It was found that the basal activity of AChE was increased in myotonic nerves while its proximodistal transport was inhibited.

Axonal microtubules: their regulation by the electrical activity of the nerve.

The effect of nerve impulses upon the axonal microtubules was studied in the sural nerve of the cat. We showed that a sustained activity (40 impulses/sec for 2 h) determines a difference in microtubule counts: the active nerve exhibited 24% more profiles than the resting one. In addition, microtubules of the active nerve appear to be less sensitive to the depolymerizing action of colchicine. We propose that the action potential leads to a decrease of the disassembly rate constant of microtubules.

Muscle fibrillation caused by cytochalasin-B applied to the motor nerve.

Cytochalasin-B, a drug known to interfere with axoplasmic transport, evoked fibrillary potentials in the geniohyoid muscle when applied to its motor nerve. Despite this denervation-like effect, neuromuscular transmission remained normal. Some contractile characteristics of the muscle were studied. It was found that contraction time, isometric twitch tension, and half-relaxation time were not altered by the drug treatment. The present findings show that neurogenic molecular factors conveyed by axoplasmic transport to the nerve terminal are involved in the regulation of some muscle membrane characteristics but do not modify the muscle contractile features.

Neurotrophic effects: Axoplasmic transport involvement in the regulation of skeletal muscle soluble proteins.

Skeletal muscle soluble proteins were assayed by polyacrylamide gel disc-electrophoresis at different times (4-36 days) after injection of 10 mM colchicine into the motor nerve and after denervation. Results showed that effects of drug-induced axoplasmic transport blockage differ from those of denervation as to quantitative changes in the protein concentration of certain bands. Results point to the conclusion that axoplasmic transport is involved in the neural control of a number of muscle proteins. This does not invalidate the participation of acetylcholine and muscle activity in the regulation of muscle metabolism.

Influence of intermittent long-term stimulation on contractile, histochemical and metabolic properties of fibre populations in fast and slow rabbit muscles.

Slow (m.soleus) and fast (m.tibialis anterior) muscles of the rabbit were subjected to indirect long-term intermittent stimulation (3 weeks, 8 hrs daily) with a frequency pattern of 10 imp/sec. Whereas no changes were observed in case of the slow muscle, stimulation induced profound changes in the fast tibialis anterior muscle. These consisted in a rearrangement of the enzyme activity pattern of energy-supplying metabolism, e.g. decrease in glycogenolytic and glycolytic enzyme activities and severalfold increase in key enzymes of aerobic endoxidation of substrates in beta-oxidation and the citric acid cycle. Concomitant with the increase in aerobic oxidative capacity, there was an increased resistance to fatigue. Histochemical studies revealed a strong increase in mitochondria of all fibres. The bimodal distribution of fibre cross-sectional area in the normal tibialis anterior muscle was changed by stimulation into a more homogeneous population of fibres with a smaller cross-sectional area. Despite a 50% increase in time to peak of isometric twitch contraction no changes were observed in the fibre population with regard to myofibrillar ATPase reaction in quantitative evaluation of whole cross-sections of the muscles. The percentage of fibres histochemically classified as slow amounted to 2.8% and 3.1% in control and stimulated tibialis anterior muscle. Nevertheless the data suggest a transformation of the fibre population under the influence of long-term intermittent stimulation.