RESUMEN
Denervation of mouse soleus muscle followed by self-reinnervation causes a significant increase in slow twitch (type I) muscle fiber content, suggesting preferential reinnervation by slow alpha-motoneurons. Since intracellular Ca2+ influences both axonal elongation rate and branching, we examined the process of self-reinnervation in mouse soleus muscles in the presence of the L-type Ca2+ channel blocker nimodipine. Soleus muscles in both control and experimental animals were denervated by crushing the soleus nerve where it enters the muscle. Experimental animals received a daily i.p. injection of a 0.1% nimodipine solution beginning 4 days prior to denervation and ending 2 weeks postdenervation. At 2 months postdenervation reinnervated and contralateral muscles from both control and experimental animals were sectioned and histochemically stained for myosin ATPase to determine the percentage of slow twitch fibers in the muscles. It was found that, in agreement with previous experiments, untreated reinnervated muscles had a significantly higher percentage of slow twitch fibers than did their contralateral controls (91.3 versus 74. 6%). However, in nimodipine-treated animals only a small, but not statistically significant, difference between reinnervated and contralateral control muscles was observed (76.5 versus 72.8%). These results suggest that Ca2+ influx through L-type calcium channels in growing neurites may play a role in the outcome of the reinnervation process.
Asunto(s)
Bloqueadores de los Canales de Calcio/farmacología , Neuronas Motoras/fisiología , Músculo Esquelético/inervación , Regeneración Nerviosa/efectos de los fármacos , Nimodipina/farmacología , Animales , Canales de Calcio/fisiología , Tamaño de la Célula , Dihidropiridinas/farmacología , Femenino , Ratones , Neuronas Motoras/efectos de los fármacos , Neuronas Motoras/ultraestructura , Desnervación Muscular , Fibras Musculares de Contracción Rápida/citología , Fibras Musculares de Contracción Rápida/fisiología , Fibras Musculares de Contracción Lenta/citología , Fibras Musculares de Contracción Lenta/fisiología , Músculo Esquelético/citología , Fibras Nerviosas/química , Fibras Nerviosas/efectos de los fármacos , Fibras Nerviosas/fisiologíaRESUMEN
Mouse soleus muscles were denervated by crushing the soleus nerve where it enters the muscle to determine if denervation followed by self-reinnervation can permanently alter the mix of fiber types in a muscle. Reinnervated and contralateral control muscles were sectioned at 2 and 7 months postdenervation and histochemically stained for myosin ATPase to determine the percentages of fast and slow twitch fibers in the muscles. It was found that, at both 2 and 7 months postdenervation, reinnervated muscles had a significantly higher percentage of slow twitch fibers than did contralateral control muscles (86.7 versus 67.8% at 2 months and 90.0 versus 69.3% at 7 months). Soleus muscles were also denervated by crushing the soleus nerve where it exists the gastrocnemius muscle (approximately 4 mm proximal to where the nerve enters the soleus muscle) to ascertain if the location of the nerve lesions plays a role in the ultimate outcome of the process of self-reinnervation. Reinnervated muscles and their contralateral muscles were sectioned at 2 months postdenervation and histochemically stained for myosin ATPase as before. It was found that, in contrast to muscles denervated at the point of nerve entry, muscles denervated 4 mm more proximal exhibited only a small increase in their percentage of slow twitch fibers which was not statistically significant (71.4 versus 68.4%). These results suggest that denervation followed by self-reinnervation can cause a permanent change in a muscle's fibers type mix and that the location of the nerve lesion strongly influences the final outcome of the reinnervation process.
Asunto(s)
Desnervación Muscular , Músculo Esquelético/inervación , Regeneración Nerviosa/fisiología , Animales , Femenino , Miembro Posterior , Estudios Longitudinales , Ratones , Fibras Musculares de Contracción Lenta/ultraestructura , Músculo Esquelético/enzimología , Músculo Esquelético/ultraestructura , Miosinas/metabolismoRESUMEN
Na channel densities were measured in fast and slow twitch mouse skeletal muscle fibres using the loose patch voltage clamp technique. It was found that Na channel density was approximately four times greater in fast twitch fibres than in slow. Computer simulations of action potential propagation in these fibres strongly suggest that the higher channel densities in fast twitch fibres are necessary to maintain action potential amplitude and fidelity of transmission across the neuromuscular junction, especially during the periods of rapid stimulation that these fibres are subjected to by their motoneurons. Transplantation of a foreign nerve containing axons which had previously innervated fast twitch fibres on to a slow twitch muscle resulted in an approximate doubling of the Na channel density in fibres innervated by the foreign nerve. These results suggest that motoneurons may exert considerable control over Na channel density in the muscle fibres they innervate.
Asunto(s)
Neuronas Motoras/trasplante , Fibras Musculares de Contracción Rápida/química , Fibras Musculares de Contracción Lenta/química , Plasticidad Neuronal/fisiología , Sarcolema/química , Canales de Sodio/análisis , Trasplante Heterotópico , Animales , Simulación por Computador , Ratones , Fibras Musculares de Contracción Lenta/fisiología , Músculo Esquelético/inervación , Músculo Esquelético/ultraestructura , Unión Neuromuscular/química , Unión Neuromuscular/fisiología , Técnicas de Placa-Clamp , Canales de Sodio/fisiologíaRESUMEN
An observational study of 429 Holstein cows in 13 herds (mean 305-FCM yield of 7225 kg) was conducted to determine the relationship between body condition score and its changes with milk yield, reproductive performance, and disease incidence. Cows were scored once for body condition during the dry period, near calving, and then every 14 d until termination of lactation. Condition score at calving had no effect on either peak of 305-d milk yields. Condition loss averaged .73 and .83 points for primiparous and multiparous cows, respectively. The duration and magnitude of condition loss depended primarily on score at calving and was greater for cows that calved with higher condition scores. After reaching minimum score, cows gained an average of .53 condition points in the remainder of lactation. Total amount of gain was not affected by body condition score at calving or by milk yield. No significant differences were found for days to first observed estrus, days to first breeding, days to conception, or number of times bred for cows grouped by body condition score at calving or by condition loss between calving and first breeding. Body condition score at calving was not significantly different for diseased cows. Cows that were diagnosed with a disease lost more condition than undiseased cows, but the difference generally was < .25 points.
Asunto(s)
Fenómenos Fisiológicos Nutricionales de los Animales , Enfermedades de los Bovinos , Bovinos/fisiología , Lactancia , Reproducción , Animales , Femenino , Paridad , Isla del Principe EduardoRESUMEN
Sodium channel distributions were measured in fast and slow twitch rodent skeletal muscle fibres using the loose patch voltage clamp technique. Large differences were found between these fibre types with respect to Na channel density in the perijunctional region. Fast twitch fibres exhibited a large increase in Na channel density near the endplate, while slow twitch fibres did not.
Asunto(s)
Placa Motora/fisiología , Músculos/fisiología , Canales de Sodio/fisiología , Animales , Potenciales de la Membrana , Ratones , Placa Motora/ultraestructura , Músculos/ultraestructura , Ratas , Canales de Sodio/ultraestructuraRESUMEN
Individual ion channels are electrically isolated and studied in living cells with the tight patch voltage clamp method. Channels are identified, categorized, and sometimes named on the basis of the biophysical properties obtained with this method. Although it is usually presumed that these recordings are from native, undisturbed membrane, the physical basis of this technique is not well established. Observations that lipid blebs readily form when suction is applied to patch clamp electrodes suggest that many single channel recordings are from ion channels in these blebs.
Asunto(s)
Canales Iónicos/fisiología , Animales , Membrana Celular/fisiología , Membrana Celular/ultraestructura , Conductividad Eléctrica , Electrofisiología , Canales Iónicos/ultraestructura , Lípidos de la Membrana/fisiología , Proteínas de la Membrana/fisiología , Métodos , Ratones , Músculos/citología , Músculos/fisiología , Músculos/ultraestructuraRESUMEN
When suction was applied to loose patch clamp pipettes while recording from enzymatically dissociated muscle fibers, large membrane blebs formed within the pipettes. We initiated a study of these suction-induced blebs because ion channels in the blebs would complicate or possibly invalidate loose patch voltage clamp measurements of membrane current density. The low lateral mobility (Stühmer and Almers, 1982) and steep gradients of Na channels at the end-plate and tendon (Caldwell et al., 1986) imply tight binding of Na channels to cytoskeletal elements and led us to expect few, if any, Na channels in the blebs. Bleb formation produced an increase in membrane capacitance, as expected from the increase in membrane area. Bleb formation also increased the Na current, indicating that the blebs contained Na channels. Assuming that the increased capacitance and Na current were due to lipid and Na channels moving from membrane outside the pipette, ejection of the bleb from the pipette was expected to bring the capacitance and Na current back to their original values. Capacitance did return to its original value, but Na current was lower than expected. The decrease in Na current is explained by Na channels moving from the patch membrane into the bleb. Normalization of bleb and patch Na current to their respective capacitances revealed that bleb membrane had a Na channel density almost 50% that of normal surface membrane. Thus, bleb membrane is neither devoid of proteins nor truly representative of the normal surface membrane from which it arose. It is enriched in membrane lipids and is relatively protein poor. Two conclusions can be drawn.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Músculos/metabolismo , Canales de Sodio/fisiología , Sodio/fisiología , Animales , Transporte Biológico , Membranas/anatomía & histología , Membranas/fisiología , Ratones , Músculos/anatomía & histología , Canales de Sodio/metabolismo , Succión , Distribución TisularRESUMEN
1. Sodium channel current density was measured using the loose-patch voltage clamp technique. Innervated rat, mouse and snake muscle had the highest density of Na+ channels in the end-plate region. These high Na+ channel densities were maintained in denervated muscle. 2. Perijunctional membrane had a Na+ current density 5- to 10-fold greater than the density several hundred micrometres from the end-plate. In all muscles this concentration of channels near the end-plate persisted following denervation. 3. At the tendon Na+ current density fell to low values (approximately 1 mA/cm2). The decrease in density began about 300-500 microns from the tendon. This pattern was found in all snake twitch fibres and fast-twitch (EDL) rat and mouse muscle fibres. This reduction in channel density near the tendon was not affected by denervation. 4. Sodium channels in all regions of innervated rat and snake muscle fibres were highly sensitive to tetrodotoxin (TTX). Sodium channels in snake muscle remained sensitive to TTX after denervation. Sodium channels that are relatively resistant to TTX appeared in rat muscle after denervation. TTX-resistant channels were even more concentrated near the end-plate than were TTX-sensitive channels in innervated muscle. At the tendon TTX-resistant Na+ channel density decreased. 5. We conclude that although the nerve presumably directs the localization of Na+ channels during development, the ability to maintain this distribution and to control the distribution of newly appearing channels persists long after the nerve has been removed.
Asunto(s)
Canales Iónicos/fisiología , Músculos/fisiología , Sodio/fisiología , Potenciales de Acción/efectos de los fármacos , Animales , Técnicas In Vitro , Canales Iónicos/efectos de los fármacos , Ratones , Placa Motora/fisiología , Desnervación Muscular , Fibras Nerviosas/fisiología , Ratas , Ratas Endogámicas , Serpientes , Tetrodotoxina/farmacologíaRESUMEN
Charge movements in intact frog twitch fibres were studied using a three-microelectrode voltage-clamp technique. When high potassium solution was applied transiently to the muscle fibres at low temperature in the presence of D600, the fibres became paralysed and, concomitantly, charge movement disappeared. The amount of charge suppressed by the paralysis treatment was about 70-100% of that in control experiments. This paralysing action of D600 is not shared by its derivative D890. The requirement of conditioning potassium contracture is, most likely, related to prolonged membrane depolarization, as voltage-clamped depolarization to 0 mV lasting tens of seconds also suppressed charge movement. When paralysed fibres were warmed, the main charge component (Q beta) was reprimed. By contrast, the hump charge component (Q gamma) was only reprimed in some of the fibres. Other than by warming, as paralysed fibre could be revived by stimulating it with large suprathreshold pulses but not by voltage-clamped hyperpolarization to -160 mV for tens of seconds. The paralysing action of D600 described here appears to be unrelated to its ability in blocking Ca2+ channels.
Asunto(s)
Galopamilo/farmacología , Músculos/efectos de los fármacos , Animales , Calcio/metabolismo , Galopamilo/análogos & derivados , Técnicas In Vitro , Canales Iónicos/efectos de los fármacos , Canales Iónicos/metabolismo , Potenciales de la Membrana/efectos de los fármacos , Contracción Muscular/efectos de los fármacos , Músculos/fisiología , Potasio/farmacología , Rana temporariaRESUMEN
The electrical properties of the end of a muscle fiber were determined using three microelectrodes, one passing sinusoidal current, the other two recording the resulting voltages. An electrical model was constructed from the morphology of the fiber, including the resistance of the extracellular space between cells; the parameters of this model were determined by fitting the model to the observed voltage responses. Our results, analyzed directly or by curve fits, show that the end of muscle fibers contains a large capacitance resulting from the extensive membrane folds at the myotendon junction. Analysis and simulations show that the extra capacitance at the myotendon junction has substantial effects on measurements of linear properties, in particular on estimates of the capacitance of the membranes. There is little qualitative effect on classical measurements of nonlinear charge movement (provided they were made with one set of electrode locations) if the linear components have been subtracted. Quantitative estimates of nonlinear charge movement and ionic currents are significantly affected, however, because these estimates are customarily normalized with respect to the linear capacitance.
Asunto(s)
Contracción Muscular , Músculos/fisiología , Animales , Conductividad Eléctrica , Matemática , Microelectrodos , Modelos Biológicos , Rana temporaria , Retículo Sarcoplasmático/fisiologíaRESUMEN
The force produced within skeletal muscle fibers is transmitted to the bone via a myotendinous junction. This junctional region was examined by light and electron microscopy in the sartorius muscles of three Rana temporaria. The muscle fibers tapered and inserted at an angle of about 25 degrees with the connective tissue fascia near the bone. The composition of the structures within the last 100 microns of the fiber was analyzed morphometrically. The T-system, terminal cisternae, and caveolae were the same as in the central region of the muscle fiber. However, the mitochondrial content was higher and the volume of longitudinal sarcoplasmic reticulum was lower than elsewhere in the fiber. The membrane at the end of the fiber had extensive villiform processes interdigitating with the tendon. The surface area of the membrane around the villiform processes was estimated with point-counting techniques and calculated from the stereological equations appropriate for partially anisotropic structures. The extra membrane involved in the myotendinous junction was about 32 times that of the cross-sectional area of the fiber. Part of this additional membrane contained specialized adherens junctions through which the contractile proteins of the muscle are anchored to collagen. The increased area at the myotendinous junction presumably provides greater mechanical strength than a flat termination. The high values of membrane capacitance and specific resistance measured electrophysiologically at the end of the fiber also can be attributed to the characteristics of the terminal membrane structure.
Asunto(s)
Músculos/anatomía & histología , Rana temporaria/anatomía & histología , Tendones/anatomía & histología , Animales , Microscopía Electrónica , Músculos/ultraestructura , Miofibrillas/ultraestructura , Tendones/ultraestructuraRESUMEN
We report measurements of nonlinear charge movement in frog skeletal muscle fibers paralyzed by the calcium-entry blocker [Schwartz, A. & Taira, N., eds. (1983) Circ. Res. 52, Part II, Number 2, 1-181.] D600 (methoxyverapamil, recently renamed gallopamil). Nonlinear charge movement was not seen in such fibers, suggesting that the drug severs the link between membrane depolarization and the main components of charge movement. This is the only pharmacological agent that blocks the main components of charge movement.
Asunto(s)
Galopamilo/toxicidad , Músculos/fisiopatología , Parálisis/fisiopatología , Verapamilo/toxicidad , Animales , Conductividad Eléctrica/efectos de los fármacos , Potenciales de la Membrana/efectos de los fármacos , Músculos/efectos de los fármacos , Potasio/farmacología , Rana temporariaRESUMEN
The Ca2+ channel blocker D-600 (methoxyverapamil) paralyses single muscle fibres of the frog: fibres exposed to the drug at 7 degrees C give a single K+ contracture after which they are paralysed, unable to contract in response to electrical stimulation or further applications of K+. Paralysed fibres contract in response to caffeine and have normal resting potentials and action potentials. Fibres treated with D-600 at 22 degrees C are not paralysed. Paralysed fibres warmed to 22 degrees C recover contractile properties: they twitch and give K+ contractures. Other workers have shown that D-600 blocks a Ca2+ channel at room temperature; thus, the paralytic action of D-600 is probably mediated by a different membrane protein, perhaps a different Ca2+ channel from that blocked at room temperature. These results suggest that the binding of D-600 can disrupt the mechanism coupling electrical potential changes across the T membrane to Ca2+ release from the sarcoplasmic reticulum.