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1.
J Physiol Sci ; 61(5): 411-9, 2011 Sep.
Article in English | MEDLINE | ID: mdl-21656343

ABSTRACT

We have investigated the appropriate joint angle for detecting frequent alternating activity in synergistic muscles and the relationship between muscle activation patterns and endurance during static low-level contractions. Eleven healthy men performed prolonged static plantar flexion of the ankle at 10% of the maximal voluntary contraction, with the ankle flexed at 100°, 110°, or 120°, while seated with the right leg in full extension. The onset of muscle activation and/or inactivation was detected using quantitative analysis, and alternate activity among muscles was detected using a threshold criterion of ×e or ×1/e multiplied by the levels of mean electromyograms (EMG) calculated at 1-min intervals. Surface EMG revealed frequent alternations of activity among the lateral and medial gastrocnemius and soleus muscles at an ankle flexion of 110°. The first alternation occurred after approximately 15 min of exercise. The number of alternations per hour was four- to sevenfold higher at 110° than at 100° or 120°. Endurance was longest and shortest at 110° and 120°, respectively. These findings suggest that synergistic motor pools activated at a specific joint angle (110°) affect muscle endurance during static low-level fatiguing tasks.


Subject(s)
Ankle/physiology , Muscle Contraction/physiology , Muscle, Skeletal/physiology , Physical Endurance/physiology , Adult , Electromyography , Exercise/physiology , Humans , Leg/physiology , Male , Muscle Fatigue/physiology , Young Adult
2.
J Physiol Sci ; 61(2): 83-91, 2011 Mar.
Article in English | MEDLINE | ID: mdl-21181322

ABSTRACT

The objective of this study was to examine the effects of exercise training in hypoxia on arterial stiffness and flow-mediated vasodilation (FMD) in postmenopausal women. Sixteen postmenopausal women (56±1 years) were assigned to a normoxic exercise group (Normoxic group, n=8) or a hypoxic exercise group (Hypoxic group, n=8). The Hypoxic group performed exercise under hypobaric hypoxic conditions corresponding to 2000 m above sea level, and was exposed to these conditions for 2 h per session. Aquatic exercise was performed at an intensity of around 50% peak oxygen uptake for 30min, 4days per week, for 8 weeks. Arterial stiffness was assessed by brachial-ankle pulse wave velocity (baPWV), and FMD was evaluated by peak diameter of the popliteal artery during reactive hyperemia. After the 8 weeks of training, the Normoxic group showed no significant changes. In contrast, baPWV (P < 0.05) was significantly reduced and peak diameter (P<0.05) and %FMD (P<0.01) were significantly increased in the Hypoxic group after training. These results suggest that exercise training under mild intermittent hypoxic conditions could more effectively reduce arterial stiffness in postmenopausal women, compared with exercise training performed at the same relative intensity under normoxic conditions. Our data also indicate that hypoxic exercise training may induce vascular functional adaptation, for example an increase in FMD response. These findings therefore could have important implications for the development of a new effective exercise prescription program.


Subject(s)
Adaptation, Physiological/physiology , Arteries/physiology , Exercise/physiology , Hypoxia/physiopathology , Postmenopause/physiology , Ankle Brachial Index/methods , Female , Humans , Hyperemia/physiopathology , Middle Aged , Oxygen Consumption/physiology , Vasodilation/physiology
3.
J Muscle Res Cell Motil ; 31(3): 195-205, 2010 Sep.
Article in English | MEDLINE | ID: mdl-20706864

ABSTRACT

We determine the effects of direct electrical stimulation (ES) on the histological profiles in atrophied skeletal muscle fibers after denervation caused by nerve freezing. Direct ES was performed on the tibialis anterior (TA) muscle after denervation in 7-week-old rats divided into groups as follows: control (CON), denervation (DN), or denervation with direct ES (subdivided into a 4 mA (ES4), an 8 mA (ES8), or a 16 mA stimulus (ES16). The stimulation frequency was set at 10 Hz, and the voltage was set at 40 V (30 min/day, 6 days/week, for 3 weeks). Ultrastructural profiles of the membrane systems involved in excitation-contraction coupling, and four kinds of mRNA expression profiles were evaluated. Morphological disruptions occurred in transverse (t)-tubule networks following denervation: an apparent disruption of the transverse networks, and an increase in the longitudinal t-tubules spanning the gap between the two transverse networks, with the appearance of pentads and heptads. These membrane disruptions seemed to be ameliorated by relatively low intensity ES (4 mA and 8 mA), and the area of longitudinally oriented t-tubules and the number of pentads and heptads decreased significantly (P < 0.01) in ES4 and ES8 compared to the DN. The highest intensity (16 mA) did not improve the disruption of membrane systems. There were no significant differences in the (alpha1s)DHPR and RyR1 mRNA expression among CON, DN, and all ES groups. After 3 weeks of denervation all nerve terminals had disappeared from the neuromuscular junctions (NMJs) in the CON and ES16 groups. However, in the ES4 and ES8 groups, modified nerve terminals were seen in the NMJs. The relatively low-intensity ES ameliorates disruption of membrane system architecture in denervated skeletal muscle fibers, but that it is necessary to select the optimal stimulus intensities to preserve the structural integrity of denervated muscle fibers.


Subject(s)
Denervation , Membrane Potentials , Muscle Fibers, Skeletal/metabolism , Muscle Fibers, Skeletal/pathology , Neuromuscular Junction/metabolism , Neuromuscular Junction/pathology , Animals , Electric Stimulation Therapy , Male , Muscle Proteins/biosynthesis , Neuromuscular Junction/physiopathology , Rats , Rats, Wistar
4.
Eur J Appl Physiol ; 109(4): 641-50, 2010 Jul.
Article in English | MEDLINE | ID: mdl-20191288

ABSTRACT

Jump training is a high-impact training regimen that increases bone volume in young bones. The aim of our study was to determine whether downregulation of adipogenesis that is associated with upregulation of osteogenesis is detected after jump training in growing rat tibiae. Four-week-old rats were jump trained for 1, 2, or 4 weeks for 5 days/week, and the height of jumping progressively increased to 35 cm. We performed morphometry to directly quantitate changes in bone volume and marrow adipocyte distribution in tibiae after the jump training. We also examined changes in the expression of osteogenic and adipogenic transcription factor proteins and mRNAs after the jump training. Four weeks of jump training induced an increase in trabecular bone volume, which was associated with the recruitment of runt-related transcription factor 2 expressing cells, as well as a decrease in marrow fat volume. However, peroxisome proliferator-activated receptor-gamma2 protein and mRNA expression levels did not change after high-impact jump training. The mRNA expression levels of the adipocyte differentiation genes CCAAT/enhancer-binding proteins (C/EBPs)alpha, C/EBPbeta, and C/EBPdelta also showed no change during the training period in jump-trained rats. We suggest that the levels of osteogenic factors that were upregulated by mechanical loading from high-impact jumping suppress adipogenesis in marrow rather than adipogenic transcription factors.


Subject(s)
Adipocytes/metabolism , Adipogenesis , Bone Marrow Cells/metabolism , Osteoblasts/metabolism , Osteogenesis , Physical Exertion , Tibia/metabolism , Transcription Factors/metabolism , Weight-Bearing , Adipogenesis/genetics , Animals , Body Weight , CCAAT-Enhancer-Binding Proteins/metabolism , Core Binding Factor Alpha 1 Subunit/metabolism , Female , Gene Expression Regulation , Hindlimb , Immunohistochemistry , Muscle, Skeletal/anatomy & histology , Organ Size , Osteogenesis/genetics , PPAR gamma/metabolism , RNA, Messenger/metabolism , Rats , Rats, Inbred F344 , Reverse Transcriptase Polymerase Chain Reaction , Tibia/cytology , Time Factors , Transcription Factors/genetics
5.
Biochim Biophys Acta ; 1800(1): 31-9, 2010 Jan.
Article in English | MEDLINE | ID: mdl-19819302

ABSTRACT

BACKGROUND: It is not known if the dihydrolipoamide succinyltransferase (DLST) gene, a mitochondrial protein, undergoes alternative splicing. We identified an uncharacterized protein reacting with an anti-DLST antibody in the I bands of myofibrils in rat skeletal muscle. METHODS: Immunocytochemical staining with an anti-DLST antibody, the purification and amino acid sequence analysis of the protein, and the isolation and sequencing of the protein's cDNA were carried out to clarify the properties of the protein and its relationship to the DLST gene. RESULTS: A pyrophosphate concentration >10 mM was necessary to extract the protein from myofibrils in the presence of salt with a higher concentration than 0.6 M, at an alkaline pH of 7.5-8.0. The protein corresponded to the amino acid sequence of the C-terminal side of DLST. The cDNAs for this protein were splicing variants of the DLST gene, with deletions of both exons 2 and 3, or only exon 2 or 3. These variants possessed an open reading frame from an initiation codon in exon 8 of the DLST gene to a termination codon in exon 15, generating a protein with a molecular weight of 30 kDa. CONCLUSIONS: The DLST gene undergoes alternative splicing, generating the protein isolated from the I bands of myofibrils. GENERAL SIGNIFICANCE: The DLST gene produces two different proteins with quite different functions via alternative splicing.


Subject(s)
Acyltransferases/genetics , Alternative Splicing , Myofibrils/metabolism , Sarcomeres/metabolism , Acyltransferases/metabolism , Amino Acid Sequence , Animals , Base Sequence , Blotting, Western , DNA, Complementary/chemistry , DNA, Complementary/genetics , Electrophoresis, Polyacrylamide Gel , Gene Expression Regulation, Enzymologic , Humans , Immunohistochemistry , Isoenzymes/genetics , Isoenzymes/metabolism , Mice , Mice, Inbred C57BL , Molecular Sequence Data , Muscle, Skeletal/enzymology , Muscle, Skeletal/metabolism , Myofibrils/enzymology , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Reverse Transcriptase Polymerase Chain Reaction , Sarcomeres/enzymology , Sequence Analysis, DNA
6.
J Muscle Res Cell Motil ; 30(5-6): 225-31, 2009.
Article in English | MEDLINE | ID: mdl-19924547

ABSTRACT

Metabolic and circulatory disorders such as diabetes and hypertension are associated with cardiac dysfunction. Research on these types of experimental animals has observed abnormal calcium (Ca(2+)) sparks and waves in cells; a potential mechanism altering excitation-contraction (e-c) coupling in the myocardium. The e-c coupling depends on the intricate spatial relationship between the sarcolemma and sarcoplasmic reticulum calcium release units (CRU's). The objective of this study was to assess for a presence or absence of abnormalities in CRU's from type II diabetic and hypertensive rat models. Myocardial tissue underwent perfusion fixation followed by selective staining of the CRU's and the features observed using a high voltage electron microscope. Results revealed both diabetic groups had significant increases in body weight, a tendency toward an enlarged heart, and a significant disruption of the CRU's and displacement of transverse (t)-tubules in a longitudinal direction. The hypertensive model characteristically showed a dramatic increase in heart size, a significant increase in disrupted CRU's and a tendency towards longitudinal t-tubule orientation. We propose the two disorders of diabetes and hypertension have a similar etiology of cardiomyopathy resulting, in part, from an increase in the number of incomplete CRU's, due to a morphological change in the architecture and orientation of the t-tubules. These architectural changes could potentially explain the impaired calcium signaling previously observed in diabetic and hypertensive cardiomyopathy.


Subject(s)
Cardiomyopathies/etiology , Cardiomyopathies/pathology , Diabetes Complications/pathology , Hypertension/complications , Myocardium/pathology , Sarcoplasmic Reticulum/pathology , Animals , Body Weight/genetics , Calcium Signaling/physiology , Cardiomegaly/etiology , Cardiomegaly/pathology , Cardiomyopathies/physiopathology , Diabetes Complications/physiopathology , Disease Models, Animal , Male , Muscle Contraction/physiology , Myocardium/metabolism , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , Rats , Rats, Inbred SHR , Sarcolemma/metabolism , Sarcolemma/pathology , Sarcoplasmic Reticulum/metabolism
7.
J Appl Physiol (1985) ; 107(2): 599-604, 2009 Aug.
Article in English | MEDLINE | ID: mdl-19541734

ABSTRACT

Skeletal muscle glucose uptake closely reflects muscle activity at exercise intensity levels <55% of maximal oxygen consumption (VO2max). Our purpose was to evaluate individual skeletal muscle activity from glucose uptake in humans during pedaling exercise at different workloads by using [18F]fluorodeoxyglucose (FDG) and positron emission tomography (PET). Twenty healthy male subjects were divided into two groups (7 exercise subjects and 13 control subjects). Exercise subjects were studied during 35 min of pedaling exercise at 40 and 55% VO2max exercise intensities. FDG was injected 10 min after the start of exercise or after 20 min of rest. PET scanning of the whole body was conducted after completion of the exercise or rest period. In exercise subjects, mean FDG uptake [standardized uptake ratio (SUR)] of the iliacus muscle and muscles of the anterior part of the thigh was significantly greater than uptake in muscles of control subjects. At 55% VO2max exercise, SURs of the iliacus muscle and thigh muscles, except for the rectus femoris, increased significantly compared with SURs at 40% VO2max exercise. Our results are the first to clarify that the iliacus muscle, as well as the muscles of the anterior thigh, is the prime muscle used during pedaling exercise. In addition, the iliacus muscle and all muscles in the thigh, except for the rectus femoris, contribute when the workload of the pedaling exercise increases from 40 to 55% VO2max.


Subject(s)
Bicycling , Exercise , Fluorodeoxyglucose F18 , Glucose/metabolism , Muscle Contraction , Muscle, Skeletal/diagnostic imaging , Muscle, Skeletal/metabolism , Positron-Emission Tomography , Radiopharmaceuticals , Adult , Biological Transport , Case-Control Studies , Humans , Male , Oxygen Consumption , Thigh , Young Adult
8.
J Muscle Res Cell Motil ; 28(2-3): 141-51, 2007.
Article in English | MEDLINE | ID: mdl-17610135

ABSTRACT

The transverse (t)-tubule is responsible for the rapid inward spread of excitation from the sarcolemma to the inside of the muscle fiber, and the compartments of the t-tubule become highly and regularly organized during development. Although it is known that skeletal muscle fibers lengthen by adding sarcomeres at the myotendinous junction (MTJ) during development, no specific model exists for the assembly of new t-tubule architecture at the MTJ. We performed an electron-microscopic examination of the assembly of t-tubule architecture at the MTJ in developing rat skeletal muscle fibers. Although the longitudinally oriented t-tubule elements represent only a small fraction of the total t-tubule system in adult muscle fibers, they were observed at both A-band and I-band regions of middle and MTJ regions in early developmental stages, and gradually disappeared in the middle regions of muscle fibers during development; however, they remained in the MTJ even in adult muscle fibers. The frequency of pentads and heptads (two or three t-tubule elements with three or four elements of terminal cisternae, closely aligned with terminal cisternae of the sarcoplasmic reticulum) decreased during development, with sudden decrease between 7 and 10 weeks of age in the middle regions. Interestingly, although the frequency of decrease appeared to be higher in the middle region than in the MTJ regions in early (3- to 7-week) development, this pattern reversed, and the frequency of decrease was higher in the MTJ in later development (after 10 weeks of age). The MTJ maintained the features of immature membrane systems involved in e-c coupling much longer than the middle region of the fiber during development. The assembly of t-tubule architecture during postnatal development thus follows different processes in the middle and MTJ regions of skeletal muscle fibers.


Subject(s)
Muscle Development/physiology , Muscle Fibers, Skeletal/ultrastructure , Muscle, Skeletal/growth & development , Muscle, Skeletal/ultrastructure , Sarcolemma/ultrastructure , Tendons/ultrastructure , Action Potentials/physiology , Aging/physiology , Animals , Body Weight/physiology , Calcium Signaling/physiology , Cell Differentiation/physiology , Female , Microscopy, Electron, Transmission , Microtubules/physiology , Microtubules/ultrastructure , Muscle Contraction/physiology , Muscle Fibers, Skeletal/physiology , Muscle Proteins/metabolism , Muscle Proteins/ultrastructure , Myofibrils/physiology , Myofibrils/ultrastructure , Organ Size/physiology , Rats , Rats, Inbred F344 , Sarcolemma/physiology , Sarcomeres/physiology , Sarcomeres/ultrastructure , Tendons/physiology
9.
Proc Natl Acad Sci U S A ; 103(52): 19760-5, 2006 Dec 26.
Article in English | MEDLINE | ID: mdl-17172444

ABSTRACT

We have defined regions of the skeletal muscle ryanodine receptor (RyR1) essential for bidirectional signaling with dihydropyridine receptors (DHPRs) and for the organization of DHPR into tetrad arrays by expressing RyR1-RyR3 chimerae in dyspedic myotubes. RyR1-RyR3 constructs bearing RyR1 residues 1-1681 restored wild-type DHPR tetrad arrays and, in part, skeletal-type excitation-contraction (EC) coupling (orthograde signaling) but failed to enhance DHPR Ca(2+) currents (retrograde signaling) to WT RyR1 levels. Within this region, the D2 domain (amino acids 1272-1455), although ineffective on its own, dramatically enhanced the formation of tetrads and EC coupling rescue by constructs that otherwise are only partially effective. These findings suggest that the orthograde signal and DHPR tetrad formation require the contributions of numerous RyR regions. Surprisingly, we found that RyR3, although incapable of supporting EC coupling or tetrad formation, restored a significant level of Ca(2+) current, revealing a functional interaction with the skeletal muscle DHPR. Thus, our data support the hypotheses that (i) the structural/functional link between RyR1 and the skeletal muscle DHPR requires multiple interacting regions, (ii) the D2 domain of RyR1 plays a key role in stabilizing this interaction, and (iii) a form of retrograde signaling from RyR3 to the DHPR occurs in the absence of direct protein-protein interactions.


Subject(s)
Muscle, Skeletal/metabolism , Ryanodine Receptor Calcium Release Channel/metabolism , Signal Transduction , Calcium/metabolism , DNA, Complementary/genetics , Muscle Contraction , Potassium/metabolism , Protein Binding , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Ryanodine Receptor Calcium Release Channel/genetics
10.
J Muscle Res Cell Motil ; 27(8): 607-15, 2006.
Article in English | MEDLINE | ID: mdl-17051345

ABSTRACT

We evaluated the effects of brief, temporary denervation caused by ischiadic nerve-freezing on the processes of degeneration and regeneration of ultrastructural features in neuromuscular junction (NMJ) architecture in different types of rat skeletal muscle fibers. Nerve terminal (NT) area was decreased significantly 12 h after nerve freezing in both fast-twitch (FT) and slow-twitch (ST) fibers. One day after nerve freezing, some terminal axons were absent; decrease in NT area was remarkable in ST fibers, and there was retraction of Schwann cells and perineural epithelial cells. Fiber type-specific differences were observed in pattern of decrease in NT area between 24 h and 7 days after nerve freezing (there was significantly more decrease in FT fibers). The primary synaptic cleft became shallow, and the secondary junctional folds shorter and wider, but the basement lamina filling the subneural apparatus was unaltered. The number of secondary junctional folds decreased gradually between 6 h and 14 days after nerve freezing in both types of fiber. In control muscle fibers, synaptic vesicle density (SVD) per terminal area was significantly higher in FT fibers. The SVD densities decreased following nerve freezing-induced destruction of NMJs, and were minimal 3 days in FT fibers or 7 days ST fibers after nerve freezing. At 3 weeks, regeneration of both FT and ST fibers was well advanced, and all parameters had recovered to control values in FT fibers 28 days after nerve freezing. Severe degradation of the ultrastructural features in NMJs occurred due to temporary denervation during muscle fiber degeneration processes, and these structural changes were all reversible and fiber type-specific.


Subject(s)
Muscle Fibers, Fast-Twitch/ultrastructure , Muscle Fibers, Slow-Twitch/ultrastructure , Neuromuscular Junction/ultrastructure , Animals , Female , Freezing , Muscle Denervation , Neuromuscular Junction/injuries , Rats , Rats, Inbred F344 , Time Factors
11.
Eur J Appl Physiol ; 98(3): 323-7, 2006 Oct.
Article in English | MEDLINE | ID: mdl-16896720

ABSTRACT

While intramuscular wire electrodes (IWE) for the measurement of neuromuscular function offer high spatial resolution for examining single motor unit activity, the resulting damage to muscle tissue and mechanical instability should be considered. We examined the influence of IWE type and component parts on muscle damage using light microscopy in rats and confirmed that intramuscular pressure influences the mechanical stability of IWE. Three types of electrode, coiled electrodes with or without suture material inside and a straight electrode, were inserted into the soleus, gastrocnemius and tibialis anterior muscles. Transverse serial sections (5 microm) of these muscles in the vicinity of the electrodes were stained with haematoxylin and eosin. Less structural damage was observed in the vicinity of the recording points (leading-off surface; 50 microm diameter) for all electrode types compared to the electrode body. No differences in the extent of tissue damage were observed around the recording points for all electrodes. However, compared to straight electrodes, the extent of damaged tissue around the bodies of coiled electrodes was significantly (P < 0.0001) greater. The average distance between the recording points and the electrode body was <1 mm for all electrodes. Intramuscular pressure at rest and maximal twitch contraction were 1.1 +/- 0.5 and 49.4 +/- 4.0 mmHg, respectively. Coiled IWEs became well integrated with muscle fibres, stabilizing electrode localization and facilitating electromyographic recordings without causing significant muscle damage.


Subject(s)
Electrodes , Electromyography/instrumentation , Electromyography/methods , Muscle Fibers, Skeletal/pathology , Muscle, Skeletal/anatomy & histology , Animals , Male , Rats , Rats, Wistar , Signal Processing, Computer-Assisted
12.
Free Radic Res ; 39(9): 905-11, 2005 Sep.
Article in English | MEDLINE | ID: mdl-16087471

ABSTRACT

We investigated whether habitual exercise (HE) modulates levels of oxidative DNA damage and responsiveness to oxidative stress induced by renal carcinogen Fe-nitrilotriacetic acid (Fe-NTA). During a ten week protocol, two groups of rats either remained sedentary or underwent swimming for 15--60 min per day, 5 days per week, with or without a weight equivalent to 5% of their body weight. Then we injected Fe-NTA and sacrificed the rats 1 h after the injection. We determined the activity of superoxide dismutase (SOD) in diaphragm and kidney, evaluated levels of 8-hydroxydeoxyguanosine (8OHdG), catalase, and glutathione peroxidase, and assayed OGG1 protein levels in kidney. SOD activity in the diaphragm and kidney was increased in HE rats. By itself, HE had no effect on the level of 8OHdG, but it did significantly suppress induction of 8OHdG by Fe-NTA, and the amount of suppression correlated with intensity of exercise. These results suggest that HE induces resistance to oxidative stress and, at least at the initiation stage, inhibits carcinogenesis.


Subject(s)
DNA Damage/physiology , Kidney/metabolism , Oxidative Stress/physiology , Physical Exertion , 8-Hydroxy-2'-Deoxyguanosine , Animals , Body Weight , Carcinogens , DNA Glycosylases/metabolism , Deoxyguanosine/analogs & derivatives , Deoxyguanosine/analysis , Diaphragm/enzymology , Diaphragm/metabolism , Ferric Compounds , Kidney/chemistry , Kidney/drug effects , Male , Nitrilotriacetic Acid/analogs & derivatives , Oxidative Stress/drug effects , Rats , Swimming
13.
Mol Biol Cell ; 15(12): 5408-19, 2004 Dec.
Article in English | MEDLINE | ID: mdl-15385628

ABSTRACT

The plasmalemmal dihydropyridine receptor (DHPR) is the voltage sensor in skeletal muscle excitation-contraction (e-c) coupling. It activates calcium release from the sarcoplasmic reticulum via protein-protein interactions with the ryanodine receptor (RyR). To enable this interaction, DHPRs are arranged in arrays of tetrads opposite RyRs. In the DHPR alpha(1S) subunit, the cytoplasmic loop connecting repeats II and III is a major determinant of skeletal-type e-c coupling. Whether the essential II-III loop sequence (L720-L764) also determines the skeletal-specific arrangement of DHPRs was examined in dysgenic (alpha(1S)-null) myotubes reconstituted with distinct alpha(1) subunit isoforms and II-III loop chimeras. Parallel immunofluorescence and freeze-fracture analysis showed that alpha(1S) and chimeras containing L720-L764, all of which restored skeletal-type e-c coupling, displayed the skeletal arrangement of DHPRs in arrays of tetrads. Conversely, alpha(1C) and those chimeras with a cardiac II-III loop and cardiac e-c coupling properties were targeted into junctional membranes but failed to form tetrads. However, an alpha(1S)-based chimera with the heterologous Musca II-III loop produced tetrads but did not reconstitute skeletal muscle e-c coupling. These findings suggest an inhibitory role in tetrad formation of the cardiac II-III loop and that the organization of DHPRs in tetrads vis-a-vis the RyR is necessary but not sufficient for skeletal-type e-c coupling.


Subject(s)
Calcium Channels, L-Type/chemistry , Calcium Channels, L-Type/metabolism , Muscle, Skeletal/metabolism , Myocardium/metabolism , Animals , Calcium Channels, L-Type/genetics , Cell Membrane/metabolism , Freeze Fracturing , Houseflies , Particle Size , Protein Structure, Quaternary , Protein Subunits/genetics , Protein Subunits/metabolism , Rabbits , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism
14.
J Muscle Res Cell Motil ; 24(7): 439-51, 2003.
Article in English | MEDLINE | ID: mdl-14677647

ABSTRACT

We have studied the effects of short term denervation followed by reinnervation on the ultrastructure of the membrane systems and on the content of and distribution of key proteins involved in calcium regulation of fast-twitch (FT) extensor digitorum longus (EDL) and slow-twitch (ST) soleus (SOL) muscle fibres. Ischiadic nerve freezing resulted in total lack of neuromuscular transmission for 3 days followed by a slow recovery, but no decline in twitch force elicited by direct stimulation. The latter measurements indicate no significant atrophy within this time frame. The membrane systems of skeletal muscle fibres were visualized using Ca92+)-K3Fe(CN)6-OsO4 techniques and observed using a high voltage electron microscope. [3H]nitrendipine binding was used to detect levels of dihydropyridine receptor (DHPR) expression. The Ca2+ pumping free sarcoplasmic reticulum domains were not affected by the denervation, but the Ca2+ release domains were dramatically increased, particularly in the FT-EDL muscle fibres. The increase is evidenced by a doubling up of the areas of contacts between SR and transverse (t-) tubules, so that in place of the normal triadic arrangement, pentadic and heptadic junctions, formed by multiple interacting layers of ST and t-tubules are seen. Frequency of pentads and heptads increases and declines in parallel to the denervation and reinnervation but with a delay. Immunofluorecence and electron microscopy observations show presence of DHPR and ryanodine receptor clusters at pentads and heptads junctions. A significant (P < 0.01) positive correlation between the level of [3H]nitrendipine binding component and the frequency pentads and heptads was observed in both the FT-EDL and ST-SOL muscle fibres indicating that overexpression of DHPRs accompanies the build up extra junctional contacts. The results indicate that denervation reversibly affects the domains of the membrane systems involved in excitation-contraction coupling.


Subject(s)
Intracellular Membranes/ultrastructure , Muscle Fibers, Fast-Twitch/ultrastructure , Muscle Fibers, Slow-Twitch/ultrastructure , Muscle, Skeletal/ultrastructure , Sarcoplasmic Reticulum/ultrastructure , Animals , Calcium/metabolism , Calcium Channels, L-Type/metabolism , Calcium Signaling , Female , Microscopy, Electron , Muscle Contraction/physiology , Muscle Denervation , Muscle, Skeletal/innervation , Nerve Crush , Rats
16.
J Muscle Res Cell Motil ; 24(8): 527-37, 2003.
Article in English | MEDLINE | ID: mdl-14870968

ABSTRACT

We evaluated the degeneration and regeneration of neuromuscular junctions (NMJs) on the extensor digitorum longus muscle of Fischer 344 rats between 4 h and 3 weeks after bupivacaine hydrochloride (BPVC) injection, which induces muscle fiber necrosis, using histochemical staining by acetylcholine esterase (AchE)-silver and electron microscopy. Degeneration of muscle fibers and NMJs was observed 4 h after BPVC injection. One week after BPVC injection, some terminal axons were almost completely retracted, and the level of basal lamina-associated AchE in some NMJ regions had gradually disappeared. At that time, the depression contained a few, mostly pit-like or elongated oval invaginations: the incipient junctional folds and some NMJs did not have any secondary junctional fold. By 2 weeks after the BPVC injection, secondary junctional folds began to develop: however, the number of secondary junctional folds was clearly less than that in normal NMJs. At 3 weeks when regeneration of muscle fibers was well advanced, the staining for AchE at the end-plates became stronger and better-defined. The volume density of mitochondria in the terminal area of the terminal significantly decreased upon BPVC-induced destruction of the NMJ, and the density reached the lowest value 24 h after BPVC injection. Significant changes in the ultrastructural features of the architecture of NMJs occurred in skeletal muscle fibers damaged by BPVC during both the degeneration and regeneration processes. The changes in the ultrastructural and morphological features of the NMJ architecture during the regeneration of degenerated muscle fibers resembled those that occur during the differentiation of normal muscle fibers.


Subject(s)
Esterases/metabolism , Myofibrils/metabolism , Neuromuscular Junction/metabolism , Regeneration/physiology , Animals , Bupivacaine/toxicity , Microscopy, Electron, Scanning Transmission , Myofibrils/drug effects , Neuromuscular Junction/drug effects , Neuromuscular Junction/injuries , Rats
17.
Biophys J ; 83(5): 2742-53, 2002 Nov.
Article in English | MEDLINE | ID: mdl-12414707

ABSTRACT

The relative disposition of ryanodine receptors (RyRs) and L-type Ca(2+) channels was examined in body muscles from three arthropods. In all muscles the disposition of ryanodine receptors in the junctional gap between apposed SR and T tubule elements is highly ordered. By contrast, the junctional membrane of the T tubule is occupied by distinctive large particles that are clustered within the small junctional domain, but show no order in their arrangement. We propose that the large particles of the junctional T tubules represent L-type Ca(2+) channels involved in excitation-contraction (e-c) coupling, based on their similarity in size and location with the L-type Ca(2+) channels or dihydropyridine receptors (DHPRs) of skeletal and cardiac muscle. The random arrangement of DHPRs in arthropod body muscles indicates that there is no close link between them and RyRs. This matches the architecture of vertebrate cardiac muscle and is in keeping with the similarity in e-c coupling mechanisms in cardiac and invertebrate striated muscles.


Subject(s)
Calcium/metabolism , Muscles/metabolism , Animals , Biophysical Phenomena , Biophysics , Calcium Channels, L-Type/chemistry , Cell Membrane/metabolism , Diptera , Freeze Fracturing , Insecta , Muscle, Skeletal/metabolism , Myocardium/metabolism , Scorpions
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