Satellite cell proliferation was assessed in low-frequency-stimulated hypothyroid rat fast-twitch muscle by 5-bromo-2'-deoxyuridine (BrdU) labeling and subsequent staining of labeled muscle nuclei, and by staining for proliferating cell nuclear antigen (PCNA). BrdU labeling and PCNA staining were highly correlated and increased approximately fourfold at 5 days of stimulation, decayed thereafter, but remained elevated over control in 10- and 20-day stimulated muscles. Myogenin mRNA was approximately 4-fold elevated at 5 days and 1.5-fold at 10 days. Staining for myogenin protein yielded results similar to that for PCNA and BrdU. Furthermore, a detailed examination of the pattern of myogenin staining revealed that the number of myogenin-positive nuclei was elevated in the fast pure IIB fiber population at 5 and 10 days of chronic low-frequency stimulation. By 20 days, myogenin staining was observed in transforming fast fibers that coexpressed embryonic and adult myosin heavy chain isoforms. In the slower fiber populations (i.e., IIA and I), myogenin-positive transforming fibers that coexpressed embryonic myosin heavy chain, appeared already at 5 days. Thus the satellite cell progeny on slower fibers seemed to proliferate less and to fuse earlier to their associated fibers than the satellite cell progeny on fast fibers. We suggest that the increase in muscle nuclei of the fast fibers might be a prerequisite for fast-to-slow fiber type transitions.

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Chronic low-frequency stimulation was used to study the effects of enhanced contractile activity on satellite cell content and myosin isoform expression in extensor digitorum longus muscles from hypothyroid rats. As verified by immunohistochemical staining for desmin, vimentin, and myosin heavy chain (MHC) isoforms and by histological analysis, stimulation induced a transformation of existing fast fibers toward slower fibers without signs of fiber deterioration or regeneration. Immunohistochemically detected increases in MHC I and MHC IIa isoforms, as well as reduced numbers of fibers expressing the faster MHC isoforms, mirrored the rearrangement of the thick-filament composition. These changes, especially the upregulation of MHC IIa, were accompanied by an induction of developmental MHC isoforms in the transforming adult fibers. Satellite cell content rose 2.6-, 3.0-, and 3.7-fold over that of corresponding controls (P < 0.05 in all cases) in 5-, 10-, and 20-day-stimulated muscles, respectively. Hypothyroidism alone had no effect on satellite cell content but resulted in a significant reduction in fiber size. The relative satellite cell contents increased (P < 0.05) from 3.8% in euthyroid control muscles to 7.9, 11.5, and 13.8% in the 5-, 10-, and 20-day-stimulated hypothyroid muscles, respectively. In 20-day-stimulated muscles, the relative satellite cell content reached an almost twofold higher level than that of normal slow-twitch soleus muscle. This increase occurred concomitantly with a rise in myonuclear density, most probably because of the fusion of satellite cells with existing fibers.

Rabbit fast-twitch tibialis anterior muscle was subjected to chronic low-frequency stimulation (10 Hz, 24 h/day). Measurements of the time course of changes in the concentration of metabolites of energy metabolism were performed in order to test the hypothesis whether or not alterations in the metabolite profile might represent possible signals for triggering muscle fibre type transformation. Most of the investigated metabolites displayed triphasic changes in response to persistently increased contractile activity. During the first 15 min of stimulation, drastic reductions were observed for adenosine triphosphate (ATP, 56%), phosphocreatine (PCr, 60%) and glycogen (76%), as well as 3- to 4-fold and 10-fold increases for glucose and lactate, respectively. This early metabolic perturbance coincided with a rapid reduction of isometric force. The next phase, extending to 4 days of stimulation, was characterized by a nearly complete recovery of ATP and PCr, and an overshoot in glycogen. The first signs of metabolic recovery were already detectable in 60-min-stimulated muscle when isometric force was still markedly depressed. These results demonstrated an impressive capability of the muscle to recover with ongoing stimulation from an initial, dramatic disturbance in energy metabolism. During the final phase, extending to 50 days, the metabolite profile approached that of a slow-twitch muscle with moderate reductions in total adenine nucleotides, ATP, total creatine, PCr and glycogen. A conspicuous result was the finding that, contrary to the recovery of most metabolites, the ratio of ATP to the product of free adenosine diphosphate and resting free inorganic phosphate was persistently depressed with ongoing stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)