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1.
CYTOCHEMICAL LOCALIZATION OF LACTIC DEHYDROGENASE IN WHITE SKELETAL MUSCLE 总被引:3,自引:0,他引:3
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The limitations of the conventional histochemical methods for localization of lactic dehydrogenase (LDH) in white skeletal muscle have been analyzed quantitatively. It is demonstrated that more than 80 per cent of LDH diffuses into the incubation medium within the first 10 minutes of incubation. Furthermore, it is confirmed that the addition of phenazine methosulfate (PMS) to the ingredients of the histochemical reaction for LDH increases substantially the capacity of the white muscle extract to reduce Nitro-BT. Based on these observations, a modified method of cytochemical localization of LDH has been developed. This method prevents the leakage of LDH from tissue sections by the application of all the ingredients of the histochemical reaction to tissue sections in a thin gelatin film. The incubation mixture contains PMS so that the staining system is independent of tissue diaphorase. The application of this method to the adductor magnus muscle of the rabbit revealed a fine reticulum in the sarcoplasm of all muscle fibers, in addition to the staining of mitochondria. The distribution of the staining suggests that LDH is localized in the sarcoplasmic reticulum. 相似文献
2.
Comparative studies were carried out on the Ca2+-transport systems of mitochondria and sarcoplasmic reticulum from longissimus dorsi muscle of genetically selected malignant hyperthermia-prone and normal pigs in order to identify the biochemical lesion responsible for the enhanced release of Ca2+ in the sarcoplasm occurring in porcine malignant hyperthermia. Mitochondria isolated from longissimus dorsi muscle of malignant hyperthermia-prone pigs contained a significantly (P < 0.001) higher amount of endogenous long-chain fatty acids. Similar amounts of endogenous mitochondrial phospholipase A2 were observed in both types of pigs, but the total activity in malignant hyperthermia-prone pigs was at least twice that of normal. Spermine, a phospholipase A2 inhibitor, lowered the activity in both types of mitochondria to a similar final level. Mitochondria of malignant hyperthermia-prone pigs showed a significantly (P < 0.001) higher oligomycin-insensitive (Ca2+ + Mg2+)-ATPase activity, but the Mg2+-ATPase and the (Ca2+ + Mg2+)-ATPase activities were similar in both types of pigs. Sarcoplasmic reticulum isolated from longissimus dorsi muscle of malignant hyperthermia-prone pigs showed a significantly higher (Ca2+ + Mg2+)-ATPase activity and a lower rate of Ca2+ uptake; the maximal amount and the rate of Ca2+ uptake by sarcoplasmic reticulum of malignant hyperthermia-prone pigs were half that of normal. Mitochondria from longissimus dorsi muscle of malignant hyperthermia-prone pigs inhibited the Ca2+-transport system of the sarcoplasmic reticulum of longissimus dorsi from both normal and malignant hyperthermia-prone pigs, but mitochondria from normal pigs had no influence on the sarcoplasmic reticulum from either type. Experimental evidence favours the concept that long-chain fatty acids released from skeletal muscle mitochondria by endogenous mitochondrial phospholipase A2 are responsible for the enhanced release of Ca2+ from mitochondria (Cheah, K.S. and Cheah, A.M. (1981) Biochim. Biophys. Acta 634, 70–84), and also additional release of Ca2+ from sarcoplasmic reticulum into the sarcoplasm during porcine malignant hyperthermia syndrome. 相似文献
3.
David Seiden 《Cell and tissue research》1973,144(4):467-473
Summary Colchicine was intraperitoneally administered chronically to adult male Sprague-Dawley rats. The ultrastructural study of hind-limb muscles revealed that myofilament desorientation resulted. Bundles of myofilaments were found coursing perpendicular or oblique to the longitudinal axis of the muscle fiber. It is concluded that a colchicine-sensitive factor is involved in maintaining normal orientation of myofibrils in mature muscle. Also found in the sarcoplasm of the colchicine treated animals were complex spheromembranous bodies. These bodies enveloped mitochondria or other organelles and appeared to be derived from the sarcoplasmic reticulum. The lysosomal nature of these bodies is indicated by the localization of acid phosphatase activity in them. Acid phosphatase activity was also displayed in the sarcoplasmic reticulum. The spheromembranous bodies seem to be part of a sarcotubulo-lysosomal system in skeletal muscle.This study was supported in part by N.I.H. Grant RR-5576.The author gratefully acknowledges the technical assistance of Mrs. Patricia Driscoll. 相似文献
4.
In adult mammalian muscle cells, energy consuming processes are mainly localized to the sarcolemma, sarcoplasmic reticulum (SR) and myofibrillar compartments, while energy production occurs within mitochondria or glycolytic complexes. Due to the restricted diffusion of adenine nucleotides near the active sites of ATPases involved in contractile activity and calcium homeostasis, there are multiple local systems that can locally rephosphorylate ADP and provide ATP. The creatine kinase (CK) system, with specific isoenzymes localized within each compartment, efficiently controls local adenylate pools and links energy production and utilization. However, mice lacking one or both of the MM-CK and mi-CK isoforms (CK-/-) are viable and develop almost normal cardiac and skeletal muscle function under the conditions of moderate workload, suggesting adaptations or other mechanisms that may ensure efficient energy transfer. While fixed CK is essentially important, other systems could also be involved as well, such as bound glycolytic enzymes or adenylate kinase. We have shown that, additionally, a direct functional interplay exists between mitochondria and sarcoplasmic reticulum, or between mitochondria and myofilaments in muscle cells, that catalyzes direct energy and signal transfer between organelles. In cardiac cells of CK-/- mice, marked cytoarchitectural modifications were observed, and direct adenine nucleotide channeling between mitochondria and organelles was very effective to rescue SR and myofilament functions. In fast skeletal muscles, increased oxidative capacity also indicates compensatory mechanisms. In mutant mice, mitochondrial capacity increases and a direct energy channeling occurs between mitochondria on one hand and ATP consuming sites on the other. However, these systems appear to be insufficient to fully compensate for the lack of CK at high workload. It can be concluded that local rephosphorylation of ADP is a crucial regulatory point in highly differentiated and organized muscle cells to ensure contractile diversity and efficiency and that the CK system is important to control energy fluxes and energy homeostasis. 相似文献
5.
Previous investigations of the T-system in skeletal muscle fibres described the inter-myofibrillar relationships between T-tubules and the sarcoplasmic reticulum. They disregarded the arrangement of the T-system in the myofibril-free sarcoplasm in the area of muscle fibre nuclei. In the present investigation, the T-system was filled by means of lanthanum incubation and the myofibril-free sarcoplasm was ultrastructural examined by means of thin (< or = 100 nm) as well as thick sections (> 300 nm-1 microm) with the electron microscope. The investigation of thick sections revealed that T-tubules meander through this myofibril-free sarcoplasm and tangle up at the poles of muscle fibre nuclei and in the area of fundamental nuclei of the motor end plate. They are, far from myofibrils, in proximity to these nuclei, the Golgi apparatus and mitochondria. On basis of this proximity and their openings at the muscle fibre surface, a contribution at the drainage of metabolic products and at the local calcium control is discussed. 相似文献
6.
Using the isotope exchange technique including 45Ca, the Ca2+-binding and Ca2+-accumulating capacity of mitochondria, sarcolemma and sarcoplasmic reticulum of rat heart was studied. The ATP-independent binding of Ca2+ to isolated membrane fractions is by 1--2 orders of magnitude less than the ATP-dependent Ca2+-accumulating capacity of the fractions. The Ca2+-accumulating capacity of mitochondria is increased 6--8 fold after addition of physiological concentrations of succinate and Pi to the incubation medium. Under these conditions the ratio of Ca2+-accumulating capacity of mitochondria, sarcolemma and sarcoplasmic reticulum of the heart is 100:3,12:2,9. The initial rate of Ca2+-uptake by the sarcoplasmic reticulum is much higher in comparison with sarcolemma and mitochondria. A high Ca2+-accumulating capacity of heart mitochondria probably determines a long-term regulation of the concentration of "troponin-accessible" Ca2+ in the sarcoplasm, whereas the high initial rate of Ca2+ accumulation by the sarcoplasmic reticulum provides for a rapid decrease of Ca2+ concentration during rhythmic contractions of the heart. 相似文献
7.
Average lactate dehydrogenase (LDH) isoenzyme patterns the content of H subunits, total LDH activity, total malate dehydrogenase (MDH) activity and the m- MDH/s-MDH ratio were determined in twelve muscles and the male genital tract of the rabbit. LDH(1) was the predominant form in the heart, soleus and masseter muscles, LDH(3) in the lingual muscles and LDH(5) in the other muscles analysed. In the muscles, an increase in the percentual proportion of M subunits was accompanied, by a proportional increase in total LDH activity and a decrease in total MDH activity, especially m-MDH. LDH isoenzyme patterns and LDH and MDH activities are useful for obtaining some idea about the proportion of individual muscle fibres. Activity accounted for by H subunits was roughly the same in all the muscles analysed, indicating that the synthesis of H subunits is independent of the type of muscle fibre and of the oxygen supply of the muscular tissue, and also that isoenzymes composed chiefly of H subunits are not localized preferentially in the mitochondria. Similar relationships between LDH isoenzymes and LDH and MDH activities were found in the testicular and epididymal tissues. The tests and the head of the epididymis mainly contain LDH isoenzymes composed of H subunits. The total LDH activity in these tissues is relatively low and their MDH activity is relatively high compared with the body and tail of the epididymis. The proportion of H subunits in the ampulla, the seminal vesicles, the coagulating glands and the prostate is also high. Cowper's glands have a high LDH(5) and LDH(4) concentration. One of two LDHx isoenzymes were found in the testes and spermatozoa. 相似文献
8.
S100a0 (αα) Protein, a Calcium-Binding Protein, Is Localized in the Slow-Twitch Muscle Fiber 总被引:2,自引:0,他引:2
We previously showed that, in contrast to the distribution of S100b (beta beta), S100a0 (alpha alpha) is mainly present in human skeletal and heart muscles at the level of 1-2 micrograms/mg of soluble protein and is universally distributed at high levels in skeletal and heart muscles of various mammals. To elucidate cellular and ultrastructural localizations of the alpha subunit of S100 protein (S100-alpha) in skeletal muscle, we used immunohistochemical and enzyme immunoassay methods. The immunohistochemical study revealed that S100-alpha is mainly localized in slow-twitch muscle fibers, whereas the beta subunit of S100 protein (S100-beta) was not detected in both types of muscle fibers, an observation indicating that the predominant form of S100 protein in the slow-twitch muscle fiber is not S100a or S100b, but S100a0. The quantitative analysis using enzyme immunoassay corroborates the immunohistochemical finding: The S100-alpha concentration of mouse soleus muscle (mainly composed of slow-twitch muscle fibers) is about threefold higher than that of mouse rectus femoris muscle (mainly composed of fast-twitch muscle fibers). At the ultrastructural level, S100-alpha is associated with polysomes, sarcoplasmic reticulum, the plasma membrane, the pellicle around lipid droplets, the outer membrane of mitochondria, and thin and thick filaments, by immunoelectron microscopy. 相似文献
9.
B T Storey L C Lin B Tompkins R E Forster 《Archives of biochemistry and biophysics》1989,270(1):144-152
The presence of carbonic anhydrase activity was demonstrated in guinea pig skeletal muscle mitochondria purified by Percoll gradient centrifugation such that contamination by sarcoplasmic reticulum vesicles was less than 5%. Assay of purified heavy sarcoplasmic reticulum vesicles for carbonic anhydrase activity showed these to have somewhat less activity than the mitochondria, so that any contribution by sarcoplasmic reticulum vesicles to mitochondrial activity would be negligible. In agreement with this observation, rabbit skeletal muscle mitochondria prepared by the Percoll method had no detectable activity. Assay of the guinea pig muscle mitochondrial enzyme activity in the presence of Triton X-100 showed a sixfold greater activity than in its absence, indicating a matrix location for the carbonic anhydrase. The enzyme is highly sensitive to the sulfonamide inhibitor ethoxzolamide, with Ki = 8.7 nM. The activation energy obtained from the rate constant for CO2 hydration, kenz with units (mg/ml)-1 s-1, over the range 4 to 37 degrees C was 12.8 kcal/mol. These properties are those expected for a carbonic anhydrase of the CA II class of isozymes, rather than for CA I, CA III, and the liver mitochondrial enzyme CA V. 相似文献
10.
N.Bojji Reddy Katharine L. Oliver Barry W. Festoff W.King Engel 《Biochimica et Biophysica Acta (BBA)/General Subjects》1978,540(3):371-388
The subcellular localization of adenylate cyclase was examined in human skeletal muscle. Three major subcellular membrane fractions, plasmalemma, sarcoplasmic reticulum and mitochondria, were characterized by membrane-marker biochemical studies, by dodecyl sulfate polycrylamide gel electrophoresis and by electron microscopy. About 60% of the adenylate cyclase of the homogenate was found in the plasmalemmal fraction and 10–14% in the sarcoplasmic reticulum and mitochondria. When the plasmalemmal preparation was subjected to discontinuous sucrose gradients, the distribution of adenylate cyclase in different subfractions closely paralleled that of (Na+ + K+)-ATPase. The highest specific activity was found in a fraction which setteled at the 0.6–0.8 M sucrose interface. The electron microscopic study of this fraction revealed the presence of flattened sacs of variable sizes and was devoid of mitochondrial and myofibrillar material. The electron microscopy of each fraction supported the biochemical studies with enzyme markers. The three major membrane fractions also contained a low Km phosphodiesterase activity, the highest specific activity being associated with sarcoplasmic reticulum.The plasmalemmal adenylate cyclase was more sensitive to catecholamine stimulation than that associated with sarcoplasmic reticulum or mitochondria. The catecholamine-sensitive, but not the basal, enzyme was further stimulated by GTP. The plasmalemmal adenylate cyclase had typical Michaelis-Menten kinetics with respect to ATP and the apparent Km for ATP was approx. 0.3. mM. The pH optimum for that enzyme was 7.5. The enzyme required Mg2+, and the concentration to achieve half-maximal stimulation was approx. 3 mM. Higher concentrations of Mg2+ (about 10 mM) were inhibitory. Solubilization of the plasmalemmal membrane fraction with Lubrol-PX resulted in preferential extraction of 106 000- and 40 000-dalton protein components. The solubilized adenylate cyclase lost its sensitivity for catecholamine stimulation, and the extent of fluoride stimulation was reduced to one-sixth of that of the intact membranes. It is concluded that the catalytically active and hormone-sensitive adenylate cyclase is predominantly localized in the surface membranes of the cells within skeletal muscle. (That “plasmalemmal” fraction is considered likely to contain, in addition to plasmalemma of muscle cells, plasmalemma of bloodvessel cells (endothelium, and perhaps smooth muscle) which may be responsible for a certain amount of the adenylate cyclase activity and other propertiesobserved in that fraction.)The method of preparation used in this study provides a convenient material for evaluating the catecholamine-adenylate cyclase interactions in human skeletal muscle. 相似文献
11.
Cytosol from rabbit heart and slow and fast skeletal muscles was fractionated using (NH4)2SO4 to yield three cytosolic protein fractions, viz., CPF-I (protein precipitated at 30% saturation), CPF-II (protein precipitated between 30 and 60% saturation), and cytosol supernatant (protein soluble at 60% saturation). The protein fractions were dialysed and tested for their effects on ATP-dependent, oxalate-supported Ca2+ uptake by sarcoplasmic reticulum from heart and slow and fast skeletal muscles. CPF-I from heart and slow muscle, but not from fast muscle, caused marked inhibition (up to 95%) of Ca2+ uptake by sarcoplasmic reticulum from heart and from slow and fast muscles. Neither unfractionated cytosol nor CPF-II or cytosol supernatant from any of the muscles altered the Ca2+ uptake activity of sarcoplasmic reticulum. Studies on the characteristics of inhibition of sarcoplasmic reticulum Ca2+ uptake by CPF-I (from heart and slow muscle) revealed the following: (a) Inhibition was concentration- and temperature-dependent (50% inhibition with approx. 80 to 100 μg CPF-I; seen only at temperatures above 20°C). (b) The inhibitor reduced the velocity of Ca2+ uptake without appreciably influencing the apparent affinity of the transport system for Ca2+. (c) Inhibition was uncompetitive with respect to ATP. (d) Sarcoplasmic reticulum washed following exposure to CPF-I showed reduced rates of Ca2+ uptake, indicating that inhibition results from an interaction of the inhibitor with the sarcoplasmic reticulum membrane. (e) Concomitant with the inhibition of Ca2+ uptake, CPF-I also inhibited the Ca2+-ATPase activity of sarcoplasmic reticulum. (f) Heat-treatment of CPF-I led to loss of inhibitor activity, whereas exposure to trypsin appeared to enhance its inhibitory effect. (g) Addition of CPF-I to Ca2+-preloaded sarcoplasmic reticulum vesicles did not promote Ca2+ release from the vesicles. These results demonstrate the presence of a soluble protein inhibitor of sarcoplasmic reticulum Ca2+ pump in heart and slow skeletal muscle but not in fast skeletal muscle. The characteristics of the inhibitor and its apparently selective distribution suggest a potentially important role for it in the in vivo regulation of sarcoplasmic reticulum Ca2+ pump, and therefore in determining the duration of Ca2+ signal in slow-contracting muscle fibers. 相似文献
12.
Yi J Ma C Li Y Weisleder N Ríos E Ma J Zhou J 《The Journal of biological chemistry》2011,286(37):32436-32443
Defective coupling between sarcoplasmic reticulum and mitochondria during control of intracellular Ca(2+) signaling has been implicated in the progression of neuromuscular diseases. Our previous study showed that skeletal muscles derived from an amyotrophic lateral sclerosis (ALS) mouse model displayed segmental loss of mitochondrial function that was coupled with elevated and uncontrolled sarcoplasmic reticulum Ca(2+) release activity. The localized mitochondrial defect in the ALS muscle allows for examination of the mitochondrial contribution to Ca(2+) removal during excitation-contraction coupling by comparing Ca(2+) transients in regions with normal and defective mitochondria in the same muscle fiber. Here we show that Ca(2+) transients elicited by membrane depolarization in fiber segments with defective mitochondria display an ~10% increased amplitude. These regional differences in Ca(2+) transients were abolished by the application of 1,2-bis(O-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, a fast Ca(2+) chelator that reduces mitochondrial Ca(2+) uptake. Using a mitochondria-targeted Ca(2+) biosensor (mt11-YC3.6) expressed in ALS muscle fibers, we monitored the dynamic change of mitochondrial Ca(2+) levels during voltage-induced Ca(2+) release and detected a reduced Ca(2+) uptake by mitochondria in the fiber segment with defective mitochondria, which mirrored the elevated Ca(2+) transients in the cytosol. Our study constitutes a direct demonstration of the importance of mitochondria in shaping the cytosolic Ca(2+) signaling in skeletal muscle during excitation-contraction coupling and establishes that malfunction of this mechanism may contribute to neuromuscular degeneration in ALS. 相似文献
13.
The sarcoplasmic reticulum and glycogen pellet derived from rabbit skeletal muscle and the sarcolemma and sarcoplasmic reticulum from pig skeletal muscle contains NAD:dependent mono ADP-ribosyltransferase activity toward the guanidine analog, P- nitrobenzylidine aminoguanidine. No or little activity could be found in the sarcolemma or sarcoplasmic reticulum derived from canine cardiac muscle. Seventy percent of activity extracted from rabbit skeletal muscle is localized in the sarcoplasmic reticulum. The enzyme has a pH optimum of 7.4, and KM of 0.5 mM and 0.35 mM for NAD and p-nitro benzylidine aminoguanidine, respectively. Inorganic phosphate, KCl, and guanidine derivatives inhibit the reaction. Incubation of the sarcoplasmic reticulum or glycogen pellet with (adenylate-32P) NAD or [adenosine-14C(U)]-labeled NAD results in the incorporation of radioactivity into proteins. A large number of proteins are labeled in the sarcoplasmic reticulum fraction. The major labeled band in the glycogen pellet corresponds to a protein of molecular weight of 83 K. 相似文献
14.
THE SARCOTUBULAR SYSTEM OF FROG SKELETAL MUSCLE : A Morphological and Biochemical Study 总被引:6,自引:20,他引:6
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U. Muscatello Ebba Andersson-Cedergren G. F. Azzone Alexandra von der Decken 《The Journal of cell biology》1961,10(4):201-218
In the frog skeletal muscle cell a well defined and highly organized system of tubular elements is located in the sarcoplasm between the myofibrils. The sarcoplasmic component is called the sarcotubular system. By means of differential centrifugation it has been possible to isolate from the frog muscle homogenate a fraction composed of small vesicles, tubules, and particles. This fraction is without cytochrome oxidase activity, which is localized in the mitochondrial membranes. This indicates that the structural components of this fraction do not derive from the mitochondrial fragmentation, but probably from the sarcotubular system. This fraction, called sarcotubular fraction, has a Mg++-stimulated ATPase activity which differs from that of muscle mitochondria in that it is 3 to 4 times higher on the protein basis as compared with the mitochondrial ATPase, and is inhibited by Ca++ and by deoxycholate like the Kielley and Meyerhof ATPase. We therefore conclude that the "granules" of the Kielley and Meyerhof ATPase, which were shown to have a relaxing effect, are fragments of the sarcotubular system. The isolated sarcotubular fraction has a high RNA content and demonstrable activity in incorporating labeled amino acids, even in the absence of added supernatant. 相似文献
15.
Victor H. Engelhard David A. Plut Dan R. Storm 《Biochimica et Biophysica Acta (BBA)/General Subjects》1976,451(1):48-61
Crude homogenates of rat cardiac muscle were fractionated in order to examine the subcellular location of adenylate cyclase in this tissue. The fractionation procedure employed differential centrifugation of homonized material, followed by collagenase treatment, centrifugation on a discontinuous sucrose density gradient and extraction with 1 M KCl. The particulate fraction obtained by this procedure contained a high specific activity and yield of adenylate cyclase, moderate levels of mitochondria and low levels of sarcoplasmic reticulum and contractile protein as judged by marker enzyme activities. Adenylate cyclase was purified 20-fold with a 33% yield from the crude homogenate, while mitochondrial, sarcoplasmic reticulum and contractile protein yields were 5, 0.4 and 0.7% respectively. The membrane fractions prepared in this manner were examined by sodium dodecyl sulfate · gel electrophoresis.Adenylate cyclase copurified with ouabain-sensitive (Na+ + K+)-ATPase, a plasma membrane marker enzyme, and not with Ca2+-accumulating activity, which is associated with the sarcoplasmic reticulum. The distribution of marker enzyme activities indicates that heart adenylate cyclase is not located in the sarcoplasmic reticulum but is localized predominantly, if not exclusively, in the plasma membrane. 相似文献
16.
N.Bojji Reddy Katharine L. Oliver Barry W. Festoff W.King Engel 《Biochimica et Biophysica Acta (BBA)/General Subjects》1978,540(3):389-401
The subcellular localization of adenylate cyclase was examined in human skeletal muscle. Three major subcellular membrane fractions, plasmalemma, sarcoplasmic reticulum and mitochondria, were characterized by membrane-marker biochemical studies, by dodecyl sulfate polycrylamide gel electrophoresis and by electron microscopy. About 60% of the adenylate cyclase of the homogenate was found in the plasmalemmal fraction and 10–14% in the sarcoplasmic reticulum and mitochondria. When the plasmalemmal preparation was subjected to discontinuous sucrose gradients, the distribution of adenylate cyclase in different subfractions closely paralleled that of (Na+ + K+)-ATPase. The highest specific activity was found in a fraction which setteled at the 0.6–0.8 M sucrose interface. The electron microscopic study of this fraction revealed the presence of flattened sacs of variable sizes and was devoid of mitochondrial and myofibrillar material. The electron microscopy of each fraction supported the biochemical studies with enzyme markers. The three major membrane fractions also contained a low Km phosphodiesterase activity, the highest specific activity being associated with sarcoplasmic reticulum.The plasmalemmal adenylate cyclase was more sensitive to catecholamine stimulation than that associated with sarcoplasmic reticulum or mitochondria. The catecholamine-sensitive, but not the basal, enzyme was further stimulated by GTP. The plasmalemmal adenylate cyclase had typical Michaelis-Menten kinetics with respect to ATP and the apparent Km for ATP was approx. 0.3. mM. The pH optimum for that enzyme was 7.5. The enzyme required Mg2+, and the concentration to achieve half-maximal stimulation was approx. 3 mM. Higher concentrations of Mg2+ (about 10 mM) were inhibitory. Solubilization of the plasmalemmal membrane fraction with Lubrol-PX resulted in preferential extraction of 106 000- and 40 000-dalton protein components. The solubilized adenylate cyclase lost its sensitivity for catecholamine stimulation, and the extent of fluoride stimulation was reduced to one-sixth of that of the intact membranes. It is concluded that the catalytically active and hormone-sensitive adenylate cyclase is predominantly localized in the surface membranes of the cells within skeletal muscle. (That “plasmalemmal” fraction is considered likely to contain, in addition to plasmalemma of muscle cells, plasmalemma of bloodvessel cells (endothelium, and perhaps smooth muscle) which may be responsible for a certain amount of the adenylate cyclase activity and other propertiesobserved in that fraction.)The method of preparation used in this study provides a convenient material for evaluating the catecholamine-adenylate cyclase interactions in human skeletal muscle. 相似文献
17.
A O Jorgensen W Arnold D R Pepper S D Kahl F Mandel K P Campbell 《Cell motility and the cytoskeleton》1988,9(2):164-174
Ca2+-ATPase of the sarcoplasmic reticulum was localized in cryostat sections from three different adult canine skeletal muscles (gracilis, extensor carpi radialis, and superficial digitalis flexor) by immunofluorescence labeling with monoclonal antibodies to the Ca2+-ATPase. Type I (slow) myofibers were strongly labeled for the Ca2+-ATPase with a monoclonal antibody (II D8) to the Ca2+-ATPase of canine cardiac sarcoplasmic reticulum; the type II (fast) myofibers were labeled at the level of the background with monoclonal antibody II D8. By contrast, type II (fast) myofibers were strongly labeled for Ca2+-ATPase of rabbit skeletal sarcoplasmic reticulum. The subcellular distribution of the immunolabeling in type I (slow) myofibers with monoclonal antibody II D8 corresponded to that of the sarcoplasmic reticulum as previously determined by electron microscopy. The structural similarity between the canine cardiac Ca2+-ATPase present in the sarcoplasmic reticulum of the canine slow skeletal muscle fibers was demonstrated by immunoblotting. Monoclonal antibody (II D8) to the cardiac Ca2+-ATPase binds to only one protein band present in the extract from either cardiac or type I (slow) skeletal muscle tissue. By contrast, monoclonal antibody (II H11) to the skeletal type II (fast) Ca2+-ATPase binds only one protein band in the extract from type II (fast) skeletal muscle tissue. These immunopositive proteins coelectrophoresed with the Ca2+-ATPase of the canine cardiac sarcoplasmic reticulum and showed an apparent Mr of 115,000. It is concluded that the Ca2+-ATPase of cardiac and type I (slow) skeletal sarcoplasmic reticulum have at least one epitope in common, which is not present on the Ca2+-ATPase of sarcoplasmic reticulum in type II (fast) skeletal myofibers. It is possible that this site is related to the assumed necessity of the Ca2+-ATPase of the sarcoplasmic reticulum in cardiac and type I (slow) skeletal myofibers to interact with phosphorylated phospholamban and thereby enhance the accumulation of Ca2+ in the lumen of the sarcoplasmic reticulum following beta-adrenergic stimulation. 相似文献
18.
D O Levitski? M K Aliev T S Levchenko I Ia Lipitskaia V N Smirnov 《Biokhimii?a (Moscow, Russia)》1976,41(5):854-863
A microsomal preparation with a high ability for Ca2+ uptake has been isolated from pigeon heart. A method of further purification of Ca2+-accumulating system of heart, based on the ability of sarcoplasmic reticulum for the energy-dependent Ca2+ accumulation in the presence of oxalate, has been developed. Upon centrifugation in the gradient of sucrose and KCl concentration the fragments of sarcoplasmic reticulum, rendered "heavy" by calcium oxalate, can be separated from foreign cell membranes. The main component of heart "calcium pump" is Ca2+-dependent ATPase (making up to about 50% of all proteins of the purified reticulum), having a molecular weight of 100.000--105.000. Specific activity of heart Ca2+-ATPase as well as the ability of purified heart sarcoplasmic reticulum for Ca2+ uptake are only slightly less than those of the skeletal muscle reticulum. The data obtained suggest that heart sarcoplasmic reticulum may be efficient for providing heart muscle relaxation. 相似文献
19.
T Mayahara T Kamimura M Tanaka S Shibanoki K Tokuyoshi I Maruyama S Yamada 《Physiologia Bohemoslovaca》1982,31(4):297-303
In order to investigate the mechanism of skeletal muscle relaxation induced by dimethyl sulfoxide, 2-butoxyethanol and dimethyl sulfoxide were examined for their effects on 1) Ca2+ uptake into and efflux from sarcoplasmic reticulum vesicles prepared from rabbit fast skeletal muscle and crayfish tail muscle by the murexide method, 2) ATPase activities of rabbit reticulum vesicles, 3) the isolated phrenic nerve-diaphragm preparation of the rat and 4) crayfish opener muscle preparation. Ca2+ efflux rate from rabbit reticulum vesicles was markedly decreased with increasing concentrations (5-20% v/v) of dimethyl sulfoxide without affecting the maximum Ca2+ uptake by the reticulum. 2-Butoxyethanol showed quite contrary effects. Dimethyl sulfoxide strongly inhibited the activity of basal ATPase rather than of Ca2+-dependent ATPase. 2-Butoxyethanol did not significantly inhibit the activity of basal ATPase, but markedly increased Ca2+-dependent ATPase activity. Antagonisms between dimethyl sulfoxide and caffeine were demonstrated either in contractions of crayfish opener muscles or in the Ca2+ release from crayfish sarcoplasmic reticulum vesicles. These results indicate a possibility that dimethyl sulfoxide reversibly induces skeletal muscle relaxation mainly in the sarcoplasmic reticulum by means of decreasing the rate and the amount of Ca2+ release from the reticulum. 相似文献
20.
THE ULTRASTRUCTURE OF THE NEUROMUSCULAR JUNCTIONS OF MAMMALIAN RED, WHITE, AND INTERMEDIATE SKELETAL MUSCLE FIBERS 总被引:8,自引:3,他引:5
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Distinct ultrastructural differences exist at the neuromuscular junctions of red, white, and intermediate fibers of a mammalian twitch skeletal muscle (albino rat diaphragm). The primary criteria for recognizing the three fiber types are differences in fiber diameter, mitochondrial content, and width of the Z line. In the red fiber the neuromuscular relationship presents the least sarcoplasmic and axoplasmic surface at each contact. Points of contact are relatively discrete and separate, and axonal terminals are small and elliptical. The junctional folds are relatively shallow, sparse, and irregular in arrangement. Axoplasmic vesicles are moderate in number, and sarcoplasmic vesicles are sparse. In the white fiber long, flat axonal terminals present considerable axoplasmic surface. Vast sarcoplasmic surface area is created by long, branching, closely spaced junctional folds that may merge with folds at adjacent contacts to occupy a more continuous and widespread area. Axoplasmic and sarcoplasmic vesicles are numerous. Both axoplasmic and sarcoplasmic mitochondria of the white fiber usually contain intramitochondrial granules. The intermediate fiber has large axonal terminals that are associated with the most widely spaced and deepest junctional folds. In all three fiber types, the junctional sarcoplasm is rich in free ribosomes, cisternae of granular endoplasmic reticulum, and randomly distributed microtubules. 相似文献