Adaptation of skeletal muscle to increased neuromuscular activity as induced by chronic low frequency stimulation.
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Adaptation, Physiological↗
Biomedical subjects
D Pette
Publications and source records attributed to D Pette.
Electrophoretically defined myosin heavy chain patterns of single human muscle spindles.
At least four myosin heavy chain (MHC) isoforms were separated by SDS-PAGE in extracts of intrafusal fibers isolated by microdissection from human lumbrical muscles. The fastest migrating MHC represents a slow isoform. The slowest migrating MHC was identified as the embryonic MHCemb. A faint band, moving slightly faster than MHCemb, most likely represents a neonatal/fetal MHC isoform. A prominent band, moving between the latter and the slow isoform is suggested to represent a hitherto unidentified, spindle-specific MHC isoform, MHCif.
Adult↗
Electrophoretic analysis of myosin heavy chain isoform patterns in extraocular muscles of the rat.
Six oculorotatory muscles and the levator palpebrae muscle of the rat were analysed by SDS-PAGE for their myosin heavy chain (MHC) isoform patterns. Oculorotatory muscles display a marked predominance of fast MHC isoforms. They contain, in addition to the slow (MHCI) and fast (MHCIIb, MHCIId, MHCIIa) skeletal MHCs, the neonatal MHCneo and the extraocular MHCeom. The levator palpebrae, generally assumed to be a member of the extraocular muscle group because of its innervation by the oculomotor nerve, does not contain MHCneo and MHCeom. It resembles a fast-twitch skeletal muscle with a predominance of MHCIId.
Animals↗
PCR-based assignment of two myosin heavy chain cDNA clones to biochemically and histochemically defined single type IIB and IID fibers of rabbit muscle.
The present study assigns two as yet unidentified fast myosin cDNA clones to specific myosin heavy chain (MHC) isoforms and their mRNAs in different fiber types of rabbit skeletal muscle. Specific oligonucleotide primers were used for reverse transcription and polymerase chain reaction (PCR) to yield products of defined lengths. The method was sensitive enough to detect specific mRNA sequences in total RNA extracts from microdissected, freeze-dried, single-fiber fragments down to 16 ng dry weight. The fibers were typed histochemically and biochemically by their electrophoretically assessed MHC complement. The following results were obtained: clone pMHC20-40 was assigned to type IIB fibers and clone pMHC24-79 to type IID fibers.
Animals↗
Relationships between alkali light-chain complement and myosin heavy-chain isoforms in single fast-twitch fibers of rat and rabbit.
The present study compares the alkali myosin light chain (LC) complement of the fast fiber types IIB, IID and IIA in single fibers from rat muscle, as well as in type IID and type IIA fibers from rabbit muscle. Single fibers were classified according to their electrophoretically determined myosin heavy chain (HC) isoforms, HCIIb, HCIId, and HCIIa. Alkali myosin light chains were analysed by densitometric evaluation of two-dimensional electrophoresis performed on extracts from the same fibers. On the average, the fraction of LC3f, i.e. LC3f/(LC1f+LC3f), was highest in type IIB fibers and lowest in type IIA fibers. Type IID fibers occupied an intermediate position. Also in the rabbit, type IID fibers displayed a higher fraction of LC3f than type IIA fibers. Large scattering of the LC3f fraction in IIB, IID, and IIA fibers indicated that each fiber type is composed of fibers identical with regard to their specific myosin heavy chain complement, but heterogeneous with regard to their fast alkali light chain composition and the resulting light-chain-based isomyosins. It is suggested that the variable proportions of the two alkali light chains in the three fast fiber populations serve as a fine tuning of contractile velocities within the ranges determined by the three fast myosin heavy-chain isoforms.
Animals↗
Coordinate changes in the expression of troponin subunit and myosin heavy-chain isoforms during fast-to-slow transition of low-frequency-stimulated rabbit muscle.
The purpose of this study was to follow the time course of changes in the expression of myosin heavy chain (HC) and troponin (Tn) subunit isoforms during fast-to-slow transition as induced in rabbit fast-twitch muscle by low-frequency stimulation. The evaluation of changes in the relative concentrations of myosin and troponin subunit isoforms were supplemented by measurements of relative protein synthesis rates using an in situ labeling technique. Changes in the amounts of mRNA encoding fast troponin C (TnC) were followed by Northern blot analysis, those for fast and slow troponin I (TnI) by in vitro translation of total RNA. The various fast myosin heavy chain (HC) and fast troponin T (TnT) isoforms were exchanged sequentially. Myosin HCIId which is the predominant fast isoform in rabbit tibialis anterior, was exchanged with HCIIa and, finally, the latter was replaced by the slow myosin HCI. The replacement of HCIId by HCIIa was accompanied by an exchange of TnT1f and TnT2f with TnT3f. The expression of HCI was accompanied by an exchange of TnT3f with the slow TnT isoforms, TnT1s and TnT2s. The changes in the relative concentrations of the TnT isoforms were preceded by similar changes of their relative synthesis rates. Pronounced decreases in the fast TnI and TnC isoforms occurred only with prolonged stimulation and were preceded by changes of the specific mRNAs and decreases in relative synthesis rates. The parallel time courses of the sequential transitions in both the myosin heavy chain and troponin T isoforms suggest the existence of coordinate programs of expression serving specific functional requirements.
Animals↗
Non-radioactive reverse transcriptase/polymerase chain reaction for quantification of myosin heavy chain mRNA isoforms in various rabbit muscles.
A method was established for measuring molecule numbers of three different myosin heavy chain (MHC) mRNA isoforms in total RNA preparations. The quantification was based on a combination of primer-directed reverse transcriptase and polymerase chain reactions with 5'-digoxigenin-labeled oligonucleotides, using external standards. The sensitivity of the method allowed the quantitation of mRNA amounts down to the range of 1,000 molecules (detection limit 50 molecules). The numbers determined for eight different rabbit muscles are in the range of 10(3)-10(9)/micrograms total RNA. In soleus muscle, the value of 1.11 x 10(9) MHCI mRNA molecules corresponds to approximately 8% of the total mRNA. With reference to myonuclei, this amount corresponds to 1-2 x 10(4) molecules/nucleus. A quantitative comparison of the two fast MHC mRNA isoforms with the distribution of different MHC isoforms at the protein level indicates that one of these two fast sequences is specific to MHCIIb and the other to MHCIId. However, our data point to the existence of additional MHCIId mRNA subtypes.
Animals↗
Fast myosin heavy chain diversity in skeletal muscles of the rabbit: heavy chain IId, not IIb predominates.
The myosin heavy chain (HC) composition of various rabbit muscles was analysed at both the mRNA and the protein level. S1-nuclease mapping was performed with a cDNA probe specific for myosin HCIIa, yielding a fully protected sequence for HCIIa, a partially protected sequence for HCIIb, and an additional signal putatively assigned to HCIId. At the protein level, three fast myosin HC isoforms, HCIIa, HCIIb and HCIId, were separated by gradient PAGE. The results obtained at the protein level were in agreement with the findings at the mRNA level. The expression of appreciable amounts of myosin HCIIb, the predominating isoform of fast-twitch muscles in rat and mouse, was restricted in the rabbit to only a few muscles, i.e. adductor magnus, gastrocnemius, latissimus dorsi and vastus lateralis. Typical fast-twitch muscles such as extensor digitorum longus, tibialis anterior and psoas contained only minute amounts of HCIIb. The HCIId isoform, demonstrated in the present study for the first time in rabbit, is the predominating fast myosin HC isoform in this species. Electrophoretic analyses of myosin HC in histochemically defined single fibers confirmed the lack of fibers expressing only HCIIb in tibialis anterior, whereas such fibers were found in the adductor magnus. In addition to fiber types IIB, IID, and IIA expressing HCIIb, HCIId, and HCIIa, respectively, an appreciable amount of hybrid fibers coexpressing two HC isoforms at various ratios were found: HCIIb > HCIId; HCIId > HCIIb; HCIId > HCIIa; HCIIa > HCIId; HCIIa > HCI; HCI > HCIIa. This fiber-type spectrum indicates possible fiber-type transitions in the order IIB<==> IIB<==>IIDB<==>IID<==>IIDA<==>IIAD<==>IIA<==>IIC<==>IC <==>I.
Animals↗
Responses of fatigable and fatigue-resistant fibres of rabbit muscle to low-frequency stimulation.
This study investigates early adaptive responses of fast-twitch muscle to increased contractile activity by low-frequency stimulation. Changes in metabolite levels and activities of regulatory enzymes of carbohydrate metabolism were investigated in rabbit tibialis anterior muscle after 24 h of stimulation. In addition, changes elicited during a 5-min lasting acute stimulation experiment were compared between 24-h-prestimulated and contralateral control muscles. Stimulation for 5 min reduced energy-rich phosphates and glycogen, and increased lactate in the control muscle. A transient elevation of fructose 2,6-bisphosphate demonstrated that activation of phosphofructokinase 2 was an immediate response to contractile activity. Prestimulated muscles displayed nearly normal values for ATP, phosphocreatine and glycogen, and did not augment lactate. Increased activities of hexokinase and phosphofructokinase 2 and permanently elevated levels of fructose 2,6-bisphosphate pointed to enhanced glycolysis with glucose as the main fuel in the prestimulated muscle. Isometric tension of the control muscle decreased rapidly a few minutes after the onset of stimulation. In the prestimulated muscles, tension was almost stable, but amounted to only 30%-40% of the initial tension of the control muscle. In view of the fibre type distribution of rabbit tibialis anterior, these findings suggested that a large fibre fraction of the prestimulated muscle, possibly the glycolytic type IID fibres, did not contract. Therefore, the possibility must be considered that the metabolite pattern of the 24-h-stimulated muscle primarily reflected metabolic activities of the contracting, less fatigable fibres, most likely type IIA and type I fibres.(ABSTRACT TRUNCATED AT 250 WORDS)
Adenosine Triphosphate↗
The continuum of pure and hybrid myosin heavy chain-based fibre types in rat skeletal muscle.
The myosin heavy chain (MHC)-based fibre composition of adult rat adductor magnus (AM) and tibialis anterior (TA) muscles was investigated using single fibre analysis. Microelectrophoresis performed on single fibre fragments demonstrated a predominance of pure fast MHC-based fibre types (expressing only one fast MHC). Most of the fibres analysed from both the AM (72%) and TA (50%) were pure type IIB (expressing only MHCIId). Pure type IID fibres (expressing only MHCIId) were also abundant in AM (20%) and TA (18%). In addition, hybrid fibres coexpressing MHCIIb and MHCIId in varying proportions (fibre types IIBD and IIDB) were found, as well as fibres coexpressing MHCIId and MHCIIa with a predominance of MHCIId (type IIDA) and some C fibres (coexpressing MHCI and MHCIIa in varying proportions). Considered altogether, these data reflect the dynamic nature of adult skeletal muscle fibres and indicate a continuum of MHC-based fibre types in normal rat muscle with transitions in the order IIB<==>IIBD<==>IIDB<==>IID<==>IIDA<==>IIAD<==>II A<==>IIC<==>IC<==>I.
Animals↗
Satellite cells from slow rat muscle express slow myosin under appropriate culture conditions.
Satellite cells were isolated at high yields from slow-twitch soleus and fast-twitch tibialis anterior (TA) muscles of adult male Wistar rats. The number of satellite cells isolated from soleus muscle exceeded that from TA muscles by a factor of three. A comparison of satellite cells grown on gelatin- or Matrigel-coated dishes revealed that Matrigel greatly enhances the maturation of the satellite-cell-derived myotubes. As judged from immunohistochemistry, myosin heavy chain electrophoresis and immunoblot analyses, only cells grown on Matrigel, but not on gelatin, expressed adult myosin isoforms. Slow myosin expression was only detected in Matrigel cultures. Soleus cultures contained, in addition to the majority of myotubes expressing fast myosin, a small fraction (maximally 10%) of myotubes coexpressing fast and slow myosins. The number of fast/slow myosin-containing myotubes was negligible in TA cultures. The expression of slow myosin increased with age. Slow myosin was nonuniformly distributed along the length of specific myotubes and accumulated around some myonuclei. These results point to the existence of myotubes with a heterogeneous population of myonuclei, probably resulting from fusion of differently preprogrammed satellite cells. We suggest that the patch-like expression of slow myosin results from local accumulation of myonuclei of slow-type satellite cells.
Animals↗
Asynchronous increases in oxidative capacity and resistance to fatigue of electrostimulated muscles of rat and rabbit.
1. The present study investigates to what extent increases in resistance to fatigue and aerobic oxidative capacity of energy metabolism are correlated in fast-twitch tibialis anterior muscles of rat and rabbit subjected to chronic low-frequency stimulation. 2. Changes in the aerobic oxidative capacity of the stimulated muscles were judged from increases in citrate synthase activity, representing the constant-proportion enzyme group of the citric acid cycle. 3. Resistance to fatigue reached maximal values in both rat and rabbit tibialis anterior muscles after stimulation periods of 14 days, whereas citrate synthase activity continued to increase with longer stimulation periods. 4. Different time courses of the changes in resistance to fatigue and citrate synthase activity were observed not only with prolonged stimulation periods but also during the first week, when pronounced increases in resistance to fatigue were accompanied by only moderate elevations in citrate synthase activity. 5. The dissociation between the changes of the two parameters studied suggests that factors other than elevated aerobic oxidative capacity contribute to enhanced resistance to fatigue.
Adaptation, Physiological↗
The histochemical profiles of fast fiber types IIB, IID, and IIA in skeletal muscles of mouse, rat, and rabbit.
This study characterized histochemically three fast fiber types (IIB, IID, IIA) in skeletal muscles of mouse, rat, and rabbit, with special reference to fiber types IIB and IID. The results are complemented by biochemical analyses of myosin heavy chain composition in these muscles. Fiber type delineation is based on various methods for mATPase staining with pre-incubations and assays under different conditions. In rat and mouse, IIB and IID fibers can be best distinguished according to their different mATPase stabilities towards formaldehyde and alkaline pH. In rabbit, the method of Matoba and Gollnick using acid pre-incubation provided best and most reproducible results. In addition to their different mATPase stabilities, the three fast fiber types differ with regard to their oxidative capacities and cross-sectional fiber areas in the three species. In general, Type IIB fibers are the largest and least oxidative, Type IIA fibers the smallest and most oxidative, and Type IID fibers intermediate. In rabbit, Type IID fibers are the predominant fast fiber population in extensor digitorum longus, psoas, and tibialis anterior muscles. As judged from histochemistry, these muscles of rabbit do not contain pure Type IIB fibers. This is in accordance with biochemical results that show the HCIId to form the majority of the myosin heavy chain complement expressed in these muscles. On the other hand, IIB fibers are numerous in rabbit adductor magnus, gastrocnemius, and vastus lateralis muscles. Similarly, appreciable amounts of myosin heavy chain HCIIb are found in the three latter muscles of rabbit.
Animals↗
Time-dependent increases in Na+,K(+)-ATPase content of low-frequency-stimulated rabbit muscle.
Chronic low-frequency stimulation of rabbit fast-twitch muscle induced time-dependent increases in the concentration of the sarcolemmal Na+,K(+)-ATPase and in mitochondrial citrate synthase activity. The almost twofold increase in Na+,K(+)-ATPase preceded the rise in citrate synthase and was complete after 10 days of stimulation. We suggest that the increase in Na+,K(+)-ATPase enhances resistance to fatigue of low-frequency-stimulated muscle prior to elevations in aerobic-oxidative capacity.
Animals↗
Inactivation of sarcoplasmic-reticulum Ca(2+)-ATPase in low-frequency-stimulated muscle results from a modification of the active site.
Molecular changes underlying the partial inactivation of the sarcoplasmic-reticulum (SR) Ca(2+-) ATPase in low-frequency-stimulated fast-twitch muscle were investigated in the present study. The specific Ca(2+)-ATPase activity, as well as the ATP- and acetyl phosphate-driven Ca2+ uptakes by the SR, were reduced by approx. 30% in 4-day-stimulated muscle. Phosphoprotein formation of the enzyme in the presence of ATP or Pi was also decreased to the same extent. Measurements of ATP binding revealed a 30% decrease in binding to the enzyme. These changes were accompanied by similar decreases in the ligand-induced (ATP, ADP, Pi) intrinsic tryptophan fluorescence. A decreased binding of fluorescein isothiocyanate (FITC) corresponded to the lower ATP binding and phosphorylation of the enzyme. Moreover, Pi-induced changes in fluorescence of the FITC-labelled enzyme did not differ between SR from stimulated and contralateral muscles, indicating that Ca(2+)- ATPase molecules which did not bind FITC were responsible for the decreased Pi-dependent phosphorylation, and therefore represented the inactive form of the enzyme. No differences existed between the Ca(2+)-induced changes in the intrinsic fluorescence of SR from stimulated and contralateral muscles which fit their similar Ca(2+)-binding characteristics. Taking the proposed architecture of the Ca2(+)-ATPase into consideration, our results suggest that the inactivation relates to a circumscribed structural alteration of the enzyme in sections of the active site consisting of the nucleotide-binding and phosphorylation domains.
Adenosine Triphosphate↗
Changes in myosin heavy-chain isoform synthesis of chronically stimulated rat fast-twitch muscle.
Chronic low-frequency stimulation was used for studying the adaptive potential of rat fast-twitch muscle to increased neuromuscular activity. The sequential exchange of myosin heavy chain isoforms HCIIb with HCIId and HCIIa was studied at the translational level using an in-vivo-labeling technique with [35S]methionine. Alterations in heavy chain isoform synthesis, i.e. a decrease in the labeling of HCIIb concomitant with an enhanced labeling of HCIId/IIa, were detectable already two days after the onset of stimulation. This time course corresponds to the previously observed alterations in the amounts of HCIIb and HCIIa mRNAs. However, significant changes in the relative protein amounts of HCIIb and HCIId/IIa were recorded only after an 8-day stimulation period. This delay at the protein level was interpreted to relate to the slow turnover of HCIIb which was estimated from its decay in long-term stimulated muscles with an approximate value of 14.7 days. Therefore, protein degradation seems to be an important post-translational regulatory step in the remodeling process of the thick filament.
Animals↗
Metabolite patterns related to exhaustion, recovery and transformation of chronically stimulated rabbit fast-twitch muscle.
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)
Adenine Nucleotides↗
Adaptation of mammalian skeletal muscle fibers to chronic electrical stimulation.
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Adaptation, Physiological↗