Menotti Midrio

The denervated muscle: facts and hypotheses. A historical review

Denervation changes in skeletal muscle (atrophy; alterations of myofibrillar expression, muscle membrane electrical properties, ACh sensitivity and excitation–contraction coupling process; fibrillation), and their possible causes are reviewed. All changes can be counteracted by muscle electrostimulation, while denervation-like effects can be caused by the complete conduction block in muscle nerve. These results do not support the hypothesis that the lack of neurotrophic, non-motor factors plays a role in denervation phenomena. Instead they support the view that the lack of neuromotor discharge is the only cause of the phenomena and that neuromotor activity is an essential factor in regulating muscle properties. However, some experimental results cannot apparently be explained by the lack of neuromotor impulses, and may still suggest that neurotrophic influences exist. A hypothesis is that neurotrophic factors, too feeble to maintain a role in completely differentiated, adult muscles, can concur with neuromotor activity in the differentiation of immature, developing muscles.

Early effects of denervation on sarcoplasmic reticulum properties of slow-twitch rat muscle fibres

Abstract The Ca2+ release activity of the sarcoplasmic reticulum (SR) in chemically skinned single slow-twitch fibres from control, 2-day and 7-day denervated rat soleus muscle was studied. Histochemical fibre type composition of the whole muscle, electrophysiological properties and the Ca2+ sensitivity of tension development by single muscle fibres were also studied. All the data were correlated with contractile properties of the in vitro muscle. In the 2-day denervated muscle the SR Ca2+ capacity and the rate of Ca2+ uptake decreased from the control values of 0.384 ± 0.030 μmol (mg fibre protein)–1 and 19.8 ± 1.9 nmol min–1 (mg fibre protein)–1, respectively, to 0.210 ± 0.016 μmol (mg fibre protein)–1 and 13.5 ± 0.9 nmol min–1 (mg fibre protein)–1; the calculated amount of Ca2+ released upon stimulation by caffeine decreased from the control value of 0.148 to 0.078 μmol (mg fibre protein)–1. In the 7-day denervated muscle, the SR Ca2+ capacity and the rate of Ca2+ uptake increased to 0.517 ± 0.06 μmol (mg fibre protein)–1 and 21.6 ± 2.3 nmol min–1 (mg fibre protein)–1, respectively; the calculated amount of Ca2+ released increased to 0.217 μmol (mg fibre protein)–1. Both contraction time and tension of the isometric twitch decreased in 2-day denervated and increased in 7-day denervated muscles. Electrophysiological and histochemical changes, as well as changes in the Ca2+ sensitivity of the muscle fibres did not show any apparent correlation with mechanical changes. It is therefore concluded that the SR plays a prominent role in the early changes of contraction time and tension following denervation.

Expression of Sarco(endo)Plasmic Reticulum Ca2+‐Atpase Slow (SERCA2) Isoform in Regenerating Rat Soleus Skeletal Muscle Depends on Nerve Impulses

We have examined the influence of innervation on the expression of different isoforms of sarco(endo)plasmic reticulum Ca2+‐ATPase (SERCA) in regenerating rat slow twitch muscle. The process of degeneration/ regeneration was induced by injection of bupivacaine into rat soleus muscle under four different conditions: (1) in the presence of intact motor nerves, (2) after surgical denervation, (3) with nerve impulse conduction blocked by the Na+‐channel blocker tetrodotoxin (TTX), and (4) with the axoplasmic flow blocked by vinblastine. Expression of SERCA isoforms was visualized by immunohistochemical and Western blot analysis. In regenerating innervated muscle, SERCA1, the isoform normally expressed in fast twitch fibres, was present after 5 days and was then progressively replaced by SERCA2, the isoform typical of slow twitch fibres. The maximum Ca2+ uptake, measured in single skinned fibres regenerating for 10‐21 days, was similar to that of slow adult fibres and significantly lower than that of fast adult fibres. Denervation or TTX treatment prevented the expression of the SERCA2 isoform. Conversely, vinblastine did not affect the expression of SERCA isoforms. These data indicate that nerve impulses play a determinant role in the expression of the SERCA2 isoform.

High-frequency fatigue of skeletal muscle: role of extracellular Ca2+

The present study evaluated whether Ca2+ entry operates during fatigue of skeletal muscle. The involvement of different skeletal muscle membrane calcium channels and of the Na+/Ca2+ exchanger (NCX) has been examined. The decline of force was analysed in vitro in mouse soleus and EDL muscles submitted to 60 and 110 Hz continuous stimulation, respectively. Stimulation with this high-frequency fatigue (HFF) protocol, in Ca2+-free conditions, caused in soleus muscle a dramatic increase of fatigue, while in the presence of high Ca2+ fatigue was reduced. In EDL muscle, HFF was not affected by external Ca2+ levels either way, suggesting that external Ca2+ plays a general protective role only in soleus. Calciseptine, a specific antagonist of the cardiac isoform (α1C) of the dihydropyridine receptor, gadolinium, a blocker of both stretch-activated and store-operated Ca2+ channels, as well as inhibitors of P2X receptors did not affect the development of HFF. Conversely, the Ca2+ ionophore A23187 increased the protective action of extracellular Ca2+. KB-R7943, a selective inhibitor of the reverse mode of NCX, produced an effect similar to that of Ca2+-free solution. These results indicate that a transmembrane Ca2+ influx, mainly through NCX, may play a protective role during HFF development in soleus muscle.

Nerve control of type 2A MHC isoform expression in regenerating slow skeletal muscle

Bupivacaine‐induced regeneration was studied in rat soleus muscle under several conditions, with the focus on type 2A and type 1 myosin heavy chain (MHC) isoform expression. In denervated muscles, type 1 was absent, whereas type 2A was widely expressed, a pattern of regeneration which appeared to be independent of fibrillation activity of the muscle. Both type 1 and type 2A isoforms were absent in muscles regenerated during tetrodotoxin (TTX) block of impulse conduction in the sciatic nerve, but type 2A was still present when the TTX block was associated with the vinblastine block of axoplasmic flow; vinblastine block alone caused the coexpression of type 1 and type 2A isoforms in the majority of fibers. These results suggest that axoplasmic flow carries some chemical factor that inhibits 2A MHC isoform expression. The results are also of clinical interest, contributing to the understanding of factors controlling muscle differentiation and adaptation.