UBC Theses and Dissertations
Histochemical and contractile properties following neonatal denervation in the fast-twitch extensor digitorum longus muscle of the mouse Redenbach, Darlene M.
Mature fast-twitch skeletal muscle depends on innervation for complete differentiation and maintenance of its fast-twitch histochemical and contractile properties. The motorneuron plays a dominant role in the development of many of these characteristics, and therefore developing muscle is even more dependent on its innervation. This study was done to look at some of the changes in histochemical and contractile properties brought about by denervation, and to examine the effects of neonatal denervation on the pattern of development of these properties in the extensor digitorum longus muscle (EDL) of the C57/BL6 mouse, at 7, 14 and 21 days of age. At 1 day of age, a unilateral sciatic neurectomy was performed. Silver and acetylcholinesterase staining was used to confirm that reinnervation does not occur by this method. Extrafusal fiber types were examined histochemically for oxidative enzyme and myosin ATPase activities (pH 4.2 and 9.4). Isometric contractile properties, including time-to-peak twitch tension, time from peak twitch tension to one-half peak twitch tension, twitch tension, tetanus tension, post-tetanic twitch potentiation, maximum velocity of unloaded shortening and resistance to fatigue were measured, in vitro, at 20°C. Denervated and control muscles were examined at 7, 14 and 21 days of age, and normal muscles were also examined at 1 day of age. At 1 day of age, all fibers stained for myosin ATPase following alkaline and acid preincubation, and all fibers were uniformly oxidative, according to NADH staining, typical of immature fibers. Over the next 21 days, normal muscles showed an increase in twitch and tetanus tension, post-tetanic twitch potentiation and velocity of unloaded shortening, with a reduction in time-to-peak twitch tension, time to one-half relaxation and in resistance to fatigue. Histochemically, the extrafusal fibers differentiated into mature fast fibers with H6% of fibers being type IIA (anaerobic in the mouse), 40% type IIB (aerobic), with the remainder being type I. In the denervated muscle there was a significant prolongation of time-to-peak twitch tension and half-relaxation compared with controls. In addition, post-tetanic twitch potentiation was absent, the maximum velocity of unloaded shortening remained low, and there was marked resistance to fatigue, at all ages studied. All denervated muscles showed significant atrophy. Histochemically, there was evidence of some continued maturation at 7 days of age, but by 14 days only two fiber groups could be distinguished. Of these, 70% were atypical fibers exhibiting dual staining for myosin ATPase and were oxidative, as seen in immature fibers. The second group stained as type IIA but, unlike the control muscles, they were oxidative. The denervated muscles returned toward slow properties of immature muscles, but the changes in the physiological properties preceeded changes in fiber type. These results suggest that removal of neural influence neonatally to extensor digitorum longus results in loss of control over the differentiation into fast-twitch muscle. In addition, there is an immediate and significant slowing of the contractile properties and although denervated muscle continues to mimick the pattern of development of normal muscle, it becomes stalled at 14 days of age, preventing further maturation. Further studies are suggested that may identify the factors contributing to the changes seen in this study.
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