Accelerated degradation of glycogen phosphorylase in denervated and dystrophic mouse skeletal muscle

Pyridoxal phosphate, the cofactor of glycogen phosphorylase, fulfils the criteria needed of a turnover label for this enzyme. The decay of protein-bound label following administration of [³H]pyridoxine is a good index of the rate of degradation of the enzyme in vivo. This method has been applied to the study of catabolism of the enzyme in normal, denervated and dystrophic mouse skeletal muscle. In both of the pathological conditions the enzyme is degraded more rapidly than normal.     

Atypical expression of circadian clock genes in denervated mouse skeletal muscle

Muscle force production and power output in active males, regardless of the site of measurement (hand, leg, or back), are higher in the evening than in the morning. This diurnal variation is attributed to motivational, peripheral and central factors, and higher core and, possibly, muscle temperatures in the evening. This study investigated whether increasing morning rectal temperatures to evening resting values, by active or passive warm-ups, leads to muscle force production and power output becoming equal to evening values in motivated subjects. Ten healthy active males (mean ± SD: age, 21.2 ± 1.9 yrs; body mass, 75.4 ± 8 kg; height, 1.76 ± .06 m) completed the study, which was approved by the University Ethics Committee. The subjects were familiarized with the techniques and protocol and then completed four sessions (separated by at least 48 h): control morning (07:30 h) and evening (17:30 h) sessions (with an active 5-min warm-up) and then two further sessions at 07:30 h but proceeded by an extended active or passive warm-up to raise rectal temperature to evening values. These last two sessions were counterbalanced in order of administration. During each trial, three measures of handgrip strength, isokinetic leg strength measurements (of knee flexion and extension at 1.05 and 4.19 rad.s^?1 through a 90° range of motion), and four measures of maximal voluntary contraction (MVC) on an isometric ergometer (utilizing the twitch-interpolation technique) were performed. Rectal and intra-aural temperatures, ratings of perceived exertion (RPE) and thermal comfort (TC) were measured. Measurements were made after the subjects had reclined for 30 min and after the warm-ups and prior to the measurement of handgrip and isokinetic and isometric ergometry. Muscle temperature was taken after the warm-up and immediately before the isokinetic and MVC measurements. Warm-ups were either active (cycle ergometer at 150 W) or passive (resting in a room at 35°C, relative humidity 45%). Data were analyzed using analysis of variance models with repeated measures. Rectal and intra-aural temperatures were higher at rest in the evening (.56°C and .74°C; p < .05) than in the morning, but there were no differences after the active or passive warm-ups, the subjects' ratings of thermal comfort reflecting this. Muscle temperatures also displayed significant diurnal variation, with higher values in the evening (～.31°C; p < .05). Grip strength, isokinetic knee flexion for peak torque and peak power at 1.05 rad.s^?1, and knee extension for peak torque at 4.19 rad.s^?1 all showed higher values in the evening. All other measures of strength or power showed a trend to be higher in the evening ( .10 > p > .05). There was no significant effect of active or passive warm-ups on any strength or power variable, and subjects reported maximal values for effort for each strength measure. In summary, effects of time of day were seen in some measures of muscle performance but, in this population of motivated subjects, there was no evidence that increasing morning rectal temperature to evening values by active or passive warm-up increased muscle strength to evening values. (Author correspondence: B.J.Edwards@ljmu.ac.uk)     

Relationship of membrane systems in muscle to isomyosin content

The structures and functions of the various subdivisions of the membrane systems of muscle are reviewed. Morphometric data have been recalculated using functional definitions of the membranes as identified by their proteins. Thus, the junctional coupling between the sarcoplasmic reticulum and T system is separated from the remaining longitudinal sarcoplasmic reticulum that bears the calcium ATPase protein. In addition, the morphometry of the membrane systems is related to the various muscle fiber types as defined histochemically and by protein isoforms. The relation of isomyosin type and membrane quantities are compared for guinea pig, chicken, frog, and lobster skeletal muscles and rat and rabbit cardiac muscles. Fiber plasticity is considered in terms of the mixing and matching of amounts and kinds of membranes and proteins.     

Species- and muscle type-dependence of perinatal isomyosin transitions.

The progressive transition from developmental to adult myosin isoforms during perinatal development was quantified in four muscles (diaphragm, gastrocnemius medialis, masseter and tongue) of four mammals (guinea-pig, hamster, rabbit and rat). It was observed that the timing of transition varied for each muscle, and differed according to the mammal as well. This suggests that the synthesis of adult myosin isoforms may be partly related to the specialized contractile function of a given muscle in a given species.     

Protein secretion from mouse skeletal muscle: coupling of increased exocytotic and endocytotic activities in denervated muscle

The secretion of proteins labelled by incorporation of radioactive amino acids was studied in innervated and 10 to 13-day-denervated mouse skeletal muscle. The secretion of ³H-leucine-labelled proteins, expressed per mg muscle wet weight, increased after denervation, and the kinetics of the secretory process was also altered in denervated muscle. Separation of secreted ³⁵S-methionine-labelled proteins by sodium dodecyl sulphate polyacrylamide gel electrophoresis followed by autoradiography revealed some denervation-induced alterations in the pattern of secreted proteins. The secretion from both innervated and denervated muscle was highly temperature sensitive and was reversibly inhibited by brefeldin A, a drug that blocks forward membrane transport from the endoplasmic reticulum/Golgi apparatus. This drug was also found to inhibit the uptake of fluorescein isothiocyanate-labelled dextran in denervated muscle but had no effect on the endocytotic activity of innervated muscle. This lends support to the hypothesis that the increased endocytotic activity in denervated muscle is coupled to a high secretory activity.Abbreviations BF A Brefeldin A - dpm Disintegrations per minute - EDL extensor digitorum longus - FITC fluorescein isothiocyanate - LDH lactate dehydrogenase - SDS-P AGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - TCA trichloroacetic acid     

Expression of adenylyl cyclase mRNAs in the denervated and in the developing mouse skeletal muscle

Changes in theactivity and in the expression of adenylyl cyclase (AC) were examinedin mouse skeletal muscle after denervation and during development. Fourisoforms of AC (AC2, AC6, AC7, and AC9) were detected by Northern blotanalysis in gastrocnemius muscle, AC9 being the most abundant. Afterdenervation, the levels of AC2 and AC9 mRNA decreased, whereas those ofAC6 and AC7 increased. AC activity in response to severalneurotransmitters was increased after denervation. During development,AC activity was high in fetus and neonate and declined in the adult;the sensitivity of AC activity to various neurotransmitters was thehighest on the third postnatal day. The levels of AC6 and AC7 mRNAswere high on the third postnatal day and then decreased in adult,paralleling the decline in AC activity. All the characteristics of ACexpression and activity in fetus and neonate resembled those observedin denervated adult muscle. These results indicate that changes in ACactivity and AC mRNAs play an important role in the various physiopathological states of skeletal muscle, especially during muscleatrophy.

     

The nifk gene is widely expressed in mouse tissues and is up-regulated in denervated hind limb muscle

Denervation of skeletal muscle alters the expression of many genes, which may be important for establishing optimal conditions for reinnervation. Using the differential display technique we have attempted to discover neurally regulated genes in skeletal muscle. An mRNA that is up-regulated in denervated hind limb muscle was identified and cloned. The cDNA encodes an RNA-binding protein, which was discovered during the course of this work to be a nucleolar protein interacting with the fork-head associated domain of the proliferation marker protein Ki-67, and named NIFK. We show that the nifk gene is widely expressed in adult mouse tissues and that the expression is up-regulated in denervated hind limb muscle. No difference between expression in perisynaptic and extrasynaptic portions of muscle was observed. The widespread expression in adult tissues suggests that the NIFK protein has other functions in addition to its interaction with Ki-67, which is only expressed in proliferating cells.     

Effects of zero gravity on myofibril content and isomyosin distribution in rodent skeletal muscle

The purpose of this experiment was to investigate the effects of 12.5 days of zero gravity (0 g) exposure (Cosmos 1887 Biosputnik) on the enzymatic properties, protein content, and isomyosin distribution of the myofibril fraction of the slow-twitch vastus intermedius (VI) and the fast-twitch vastus lateralis (VL) muscles of adult male rats. Measurements were obtained on three experimental groups (n = 5 each group) designated as flight group (FG), vivarium control (VC), and synchronous control (SC). Body weight of the FG was significantly lower than that of the two control groups (P less than 0.05). Compared with the two control groups, VI weight was lower by 23% (P less than 0.10), whereas no such pattern was apparent for the VL muscle. Myofibril yields (mg protein/g muscle) in the VI were 35% lower in the FG than in controls (P less than 0.05), whereas no such pattern was apparent for the VL muscle. When myofibril yields were expressed on a muscle basis (mg/g x muscle weight), the loss of myofibril protein was more exaggerated and suggests that myofibril protein degradation is an early event in the muscle atrophy response to 0 g. Analysis of myosin isoforms indicated that slow myosin (Sm) was the primary isoform lost in the calculated degradation of total myosin. No evidence of loss of the fast isomyosins was apparent for either muscle following spaceflight. Myofibril ATPase activity of the VI was increased in the FG compared with controls, which is consistent with the observation of preferential Sm degradation. These data suggest that muscles containing a high percentage of slow-twitch fibers undergo greater degrees of myofibril protein degradation than muscles containing predominantly fast-twitch fibers in response to a relatively short period of 0 g exposure, and the primary target appears to be the Sm molecule.