Synaptic Vesicle Distribution and Release at Rat Diaphragm Neuromuscular Junctions

2007 ◽  
Vol 98 (1) ◽  
pp. 478-487 ◽  
Author(s):  
Katharine L. Rowley ◽  
Carlos B. Mantilla ◽  
Leonid G. Ermilov ◽  
Gary C. Sieck
Keyword(s):  
Synaptic Vesicle ◽  
Phrenic Nerve ◽  
Active Zone ◽  
Synaptic Vesicles ◽  
Fiber Type ◽  
Repetitive Stimulation ◽  
Diaphragm Muscle ◽  
Type I ◽  
Fiber Types ◽  
Rat Diaphragm

Synaptic vesicle release at the neuromuscular junction (NMJ) is highly reliable and is vital to the success of synaptic transmission. We examined synaptic vesicle number, distribution, and release at individual type-identified rat diaphragm NMJ. Three-dimensional reconstructions of electron microscopy images were used to obtain novel measurements of active zone distribution and the number of docked synaptic vesicles. Diaphragm muscle-phrenic nerve preparations were used to perform electrophysiological measurements of the decline in quantal content (QC) during repetitive phrenic nerve stimulation. The number of synaptic vesicles available for release vastly exceeds those released with a single stimulus, thus reflecting a relatively low probability of release for individual docked vesicles and at each active zone. There are two components that describe the decline in QC resulting from repetitive stimulation: a rapid phase (<0.5 s) and a delayed phase (<2.5 s). Differences in the initial rapid decline in QC were evident across type-identified presynaptic terminals (fiber type classification based on myosin heavy chain composition). At terminals innervating type IIx and/or IIb fibers, the initial decline in QC during repetitive stimulation matched the predicted depletion of docked synaptic vesicles. In contrast, at terminals innervating type I or IIa fibers, a faster than predicted decline in QC with repetitive stimulation suggests that a decrease in the probability of release at these terminals plays a role in addition to depletion of docked vesicles. Differences in QC decline likely reflect fiber-type specific differences in activation history and correspond with well-described differences in neuromuscular transmission across muscle fiber types.

scholarly journals Denervation effects on myonuclear domain size of rat diaphragm fibers

2006 ◽  
Vol 100 (5) ◽  
pp. 1617-1622 ◽  
Author(s):  
Bharathi Aravamudan ◽  
Carlos B. Mantilla ◽  
Wen-Zhi Zhan ◽  
Gary C. Sieck
Keyword(s):  
Fiber Type ◽  
Cross Sectional Area ◽  
Diaphragm Muscle ◽  
Type I ◽  
Fiber Types ◽  
Sectional Area ◽  
Rat Diaphragm ◽  
Cross Sectional ◽  
Fiber Size ◽  
Myonuclear Domain

Denervation (DNV) of rat diaphragm muscle (DIAm) leads to selective atrophy of type IIx and IIb fibers, whereas the cross-sectional area of type I and IIa fibers remains unchanged or slightly hypertrophied. DIAm DNV also increases satellite cell mitotic activity and myonuclear apoptosis. Similar to other skeletal muscles, DIAm fibers are multinucleated, and each myonucleus regulates the gene products in a finite fiber volume, i.e., myonuclear domain (MND). MND size varies across DIAm fiber types in rank order, I < IIa < IIx < IIb [fiber type based on myosin heavy chain isoform expression]. We hypothesized that, after DNV, the total number of myonuclei per fiber does not change and, accordingly, that MND changes proportionately to the change in fiber size regardless of fiber type. Adult rats underwent unilateral (right side) DIAm DNV, and after 2 wk single fibers were dissected. Fiber cross-sectional area, myonuclear number, and MND were measured by confocal microscopy, and these values in DNV DIAm were compared with those obtained in controls. After DNV, type I fibers hypertrophied, type IIa fiber size was unchanged, and type IIx and IIb fibers atrophied compared with control. The total number of myonuclei per fiber was not affected by DNV. Accordingly, after DNV, type I fiber MND increased by 25%, whereas it decreased in type IIx and IIb fibers by 50 and 70%, respectively. These results suggest that MND is not maintained after DNV-induced DIAm fiber hypertrophy or atrophy. These results are interpreted with respect to consequent effects of DNV on myonuclear transcriptional activity and protein turnover.

Extreme endurance training and fiber type adaptation in rat diaphragm

1989 ◽  
Vol 66 (4) ◽  
pp. 1914-1920 ◽  
Author(s):  
H. J. Green ◽  
M. J. Plyley ◽  
D. M. Smith ◽  
J. G. Kile
Keyword(s):  
Endurance Training ◽  
Fiber Type ◽  
Succinic Dehydrogenase ◽  
Diaphragm Muscle ◽  
Type I ◽  
Fiber Types ◽  
Rat Diaphragm ◽  
Oxidative Potential ◽  
Glycerophosphate Dehydrogenase ◽  
Type I Fibers

Extreme endurance training was used to investigate the adaptability of the rat diaphragm muscle fibers. During the final phase of the 14-wk training program, the animals were running for 240 min/day at an estimated requirement of 80% of pretraining maximal O2 consumption. Analysis of a sample of the costal diaphragm indicated that training resulted in a 34% reduction (P less than 0.05) in the percent distribution of type IIa fibers [27.7 +/- 1.1 vs. 18.3 +/- 2.6 (SE)] and a 15% increase (P less than 0.05) in the percent of type IIb fibers (40.0 +/- 1.2 vs. 46.1 +/- 2.4). No change (P greater than 0.05) was found in the distribution of the type I fibers (32.3 +/- 1.2 vs. 35.7 +/- 1.3). Oxidative potential as assessed with NADH-tetrazolium reductase and measured microphotometrically increased (P less than 0.05) by 19% in type I fibers but did not change in either the type IIa or type IIb fibers. No effect of training was found when a different oxidative marker, succinic dehydrogenase, was employed. Similarly glycolytic potential based on the activity of alpha-glycerophosphate dehydrogenase was not affected by training. Glycogen concentration was elevated by 60% (P less than 0.01) in type I fibers and 77% (P less than 0.01) in type IIb fibers with training but was not altered (P greater than 0.05) in type IIa fibers. Reductions (P less than 0.05) in fiber area ranging from 11 to 20% were observed in all fiber types as a result of training, whereas the number of capillaries per fiber remained static.(ABSTRACT TRUNCATED AT 250 WORDS)

SDH and actomyosin ATPase activities of different fiber types in rat diaphragm muscle

1995 ◽  
Vol 79 (5) ◽  
pp. 1629-1639 ◽  
Author(s):  
G. C. Sieck ◽  
W. Z. Zhan ◽  
Y. S. Prakash ◽  
M. J. Daood ◽  
J. F. Watchko
Keyword(s):  
Atpase Activity ◽  
Rank Order ◽  
Cross Sectional Area ◽  
Diaphragm Muscle ◽  
Type I ◽  
Fiber Types ◽  
Sectional Area ◽  
Rat Diaphragm ◽  
Cross Sectional ◽  
Actomyosin Atpase

In the rat diaphragm muscle, the histochemical classification of type I, IIa, IIb, or IIx fibers was correlated with myosin heavy chain (MHC) immunoreactivity. Expression of MHC isoforms in single dissected fibers was also assessed electrophoretically. Most fibers (approximately 86%) expressed a single MHC isoform, and when present, coexpression of MHC-2X and MHC-2B isoforms was most prevalent. Type I and IIa fibers were the smallest, type IIb fibers were the largest, and type IIx fibers were intermediate. Succinate dehydrogenase (SDH) and calcium-activated myosin adenosinetriphosphatase (actomyosin ATPase) activities were measured with quantitative histochemical procedures. Type I and IIa fibers had the highest SDH activities, followed in rank order by type IIx and IIb fibers. Type I fibers had the lowest actomyosin ATPase activity, followed in rank order by type IIa, IIx, and IIb fibers. Across all fibers, there was an inverse relationship between fiber SDH activity and cross-sectional area and a positive correlation between fiber actomyosin ATPase activity and cross-sectional area. The SDH and actomyosin ATPase activities of muscle fibers were also inversely correlated. These phenotypic differences in SDH and ATPase activities may be important in determining the contractile and fatigue properties of different fiber types in the rat diaphragm muscle.

Morphological Adaptations of Neuromuscular Junctions Depend on Fiber Type

1997 ◽  
Vol 22 (3) ◽  
pp. 197-230 ◽  
Author(s):  
Gary C. Sieck ◽  
Y. S. Prakash
Keyword(s):  
Neuromuscular Junctions ◽  
Fiber Type ◽  
Muscle Fibers ◽  
Motor Units ◽  
Diaphragm Muscle ◽  
Dimensional Structure ◽  
Type I ◽  
Fiber Types ◽  
Structural And Functional Properties ◽  
Fast Twitch

The neuromuscular junction (NMJ) forms the communicative types of motor units, which are recruited selectively to accomplish various link between motoneurons and muscle fibers. The properties of motoneurons and muscle fibers are matched in different motor behaviors. Motor units and muscle fibers can be classified based upon structural and functional properties, reflecting the essential match between motoneuron and muscle fiber. Using a three-color immunofluorescence technique combined with confocal microscopy, we examined the three-dimensional structure of pre- and postsynaptic elements of NMJs on different fiber types in the rat diaphragm muscle. On type I and IIa fibers, comprising slow-twitch and fast-twitch fatigue-resistant motor units, the structure of NMJs is far less complex than on type IIx and IIb fibers comprising fast-twitch fatigue-intermediate and fast-twitch fatigable motor units. We also found a greater extent of overlap between pre- and postsynaptic elements of NMJs on type I and IIa fibers. This review focuses on these normal phenotypic differences in NMJ properties and on the adaptations that occur under various conditions of altered use. Key words: nerve terminal, motor endplate, morphology, neuromuscular transmission, fatigue, plasticity

Rostrocaudal pattern of fiber-type changes in an overloaded rat ankle extensor

1991 ◽  
Vol 71 (2) ◽  
pp. 558-564 ◽  
Author(s):  
P. F. Gardiner ◽  
B. J. Jasmin ◽  
P. Corriveau
Keyword(s):  
Fiber Type ◽  
Muscle Length ◽  
Similar Degree ◽  
Complex Nature ◽  
Type I ◽  
Fiber Types ◽  
Type Ii ◽  
Cross Sectional ◽  
Hypertrophic Response ◽  
Type I Fibers

Our aim was to quantify the overload-induced hypertrophy and conversion of fiber types (type II to I) occurring in the medial head of the gastrocnemius muscle (MG). Overload of MG was induced by a bilateral tenotomy/retraction of synergists, followed by 12–18 wk of regular treadmill locomotion (2 h of walking/running per day on 3 of 4 days). We counted all type I fibers and determined type I and II mean fiber areas in eight equidistant sections taken along the length of control and overloaded MG. Increase in muscle weights (31%), as well as in total muscle cross-sectional areas (37%) and fiber areas (type I, 57%; type II, 34%), attested to a significant hypertrophic response in overloaded MG. An increase in type I fiber composition of MG from 7.0 to 11.5% occurred as a result of overload, with the greatest and only statistically significant changes (approximately 70–100%) being found in sections taken from the most rostral 45% of the muscle length. Results of analysis of sections taken from the largest muscle girth showed that it significantly underestimated the extent of fiber conversion that occurred throughout the muscle as a whole. These data obtained on the MG, which possesses a compartmentalization of fiber types, support the notion that all fiber types respond to this model with a similar degree of hypertrophy. Also, they emphasize the complex nature of the adaptive changes that occur in these types of muscles as a result of overload.

Glucose transporter protein content and glucose transport capacity in rat skeletal muscles

1990 ◽  
Vol 259 (4) ◽  
pp. E593-E598 ◽  
Author(s):  
E. J. Henriksen ◽  
R. E. Bourey ◽  
K. J. Rodnick ◽  
L. Koranyi ◽  
M. A. Permutt ◽  
...  
Keyword(s):  
Protein Content ◽  
Glucose Transport ◽  
Glucose Transporter ◽  
Contractile Activity ◽  
Fiber Type ◽  
Linear Regression Analysis ◽  
Type I ◽  
Fiber Types ◽  
Transport Activity ◽  
Glut 4

The relationships among fiber type, glucose transporter (GLUT-4) protein content, and glucose transport activity stimulated maximally with insulin and/or contractile activity were studied by use of the rat epitrochlearis (15% type I-20% type II2a-65% type IIb), soleus (84-16-0%), extensor digitorum longus (EDL, 3-57-40%), and flexor digitorum brevis (FDB, 7-92-1%) muscles. Insulin-stimulated 2-deoxy-D-glucose (2-DG) uptake was greatest in the soleus, followed (in order) by the FDB, EDL, and epitrochlearis. On the other hand, contractile activity induced the greatest increase in 2-DG uptake in the FDB, followed by the EDL, soleus, and epitrochlearis. The effects of insulin and contractile activity on 2-DG uptake were additive in all the muscle preparations, with the relative rates being FDB greater than soleus greater than EDL greater than epitrochlearis. Quantitation of the GLUT-4 protein content with the antiserum R820 showed the following pattern: FDB greater than soleus greater than EDL greater than epitrochlearis. Linear regression analysis showed that whereas a relatively low and nonsignificant correlation existed between GLUT-4 protein content and 2-DG uptake stimulated by insulin alone, significant correlations existed between GLUT-4 protein content and 2-DG uptake stimulated either by contractions alone (r = 0.950) or by insulin and contractions in combination (r = 0.992). These results suggest that the differences in maximally stimulated glucose transport activity among the three fiber types may be related to differences in their content of GLUT-4 protein.

scholarly journals Lifetime of autofluorescence: A novel method to determine murine skeletal muscle fiber types

2021 ◽  
Vol 154 (9) ◽  
Author(s):  
C. Manno ◽  
E. Tammineni ◽  
Y. Oropeza ◽  
L. Figueroa ◽  
E. Rios
Keyword(s):  
Fiber Type ◽  
Myosin Heavy Chain Isoforms ◽  
Muscle Fiber Types ◽  
Fluorescence Lifetime Imaging ◽  
Type I ◽  
Fiber Types ◽  
Mean Values ◽  
Lifetime Imaging ◽  
Oxidation Reduction ◽  
Murine Skeletal Muscle

This work describes a simple way to identify fiber types in living muscles by fluorescence lifetime imaging microscopy (FLIM). We quantified the mean values of lifetimes derived from a two-exponential fit (τ1 and τ2) in freshly dissected mouse FDB and soleus muscles. While τ1 values did not change between muscles, the distribution of τ2 shifted to higher values in FDB. To understand the origin of this difference, we obtained maps of autofluorescence lifetimes in cryosections of both muscles and paired them with immunofluorescence images of myosin heavy chain isoforms (MHC), which allow identification of fiber types. In soleus, τ2 was 3.1 ns for type I (SEM = 0.009, n = 49), 3.4 ns for type IIA (SEM = 0.01, n = 30), and 3.3 ns for type IIX (SEM = 0.01, n = 21). In FDB muscle, τ2 was 3.17 ns for type I (SEM = 0.04, n = 18), 3.5 ns for type IIA (SEM = 0.03, n = 27), and 3.62 ns for type IIX (SEM = 0.03, n = 22). From the distribution of measures, it follows that an FDB fiber with τ2 &gt;3.3 ns is expected to be of type II, and of type I otherwise. This simple classification method has first- and second-class errors estimated at 0.06 and 0.27, respectively. Studies in progress aim at further elucidating the reasons for the different lifetimes, not just among fiber types but between fibers of the same type in the two muscles. Preliminary results point at differences in both the oxidation-reduction and protein-bound versus free states of flavins as causes for the observed divergence of fluorescence lifetimes. Lifetime maps of autofluorescence therefore constitute a tool to identify fiber type that, being practical, fast, and noninvasive, can be applied in living tissue without compromising other experimental interventions.

scholarly journals Dual pools of actin at presynaptic terminals

2012 ◽  
Vol 107 (12) ◽  
pp. 3479-3492 ◽  
Author(s):  
Adam Bleckert ◽  
Huzefa Photowala ◽  
Simon Alford
Keyword(s):  
Synaptic Transmission ◽  
Synaptic Vesicle ◽  
Synaptic Vesicles ◽  
Repetitive Stimulation ◽  
Filamentous Actin ◽  
Cortical Actin ◽  
Vesicle Recycling ◽  
Latrunculin A ◽  
Kinetics Of

We investigated actin's function in vesicle recycling and exocytosis at lamprey synapses and show that FM1-43 puncta and phalloidin-labeled filamentous actin (F-actin) structures are colocalized, yet recycling vesicles are not contained within F-actin clusters. Additionally, phalloidin also labels a plasma membrane-associated cortical actin. Injection of fluorescent G-actin revealed activity-independent dynamic actin incorporation into presynaptic synaptic vesicle clusters but not into cortical actin. Latrunculin-A, which sequesters G-actin, dispersed vesicle-associated actin structures and prevented subsequent labeled G-actin and phalloidin accumulation at presynaptic puncta, yet cortical phalloidin labeling persisted. Dispersal of presynaptic F-actin structures by latrunculin-A did not disrupt vesicle clustering or recycling or alter the amplitude or kinetics of excitatory postsynaptic currents (EPSCs). However, it slightly enhanced release during repetitive stimulation. While dispersal of presynaptic actin puncta with latrunculin-A failed to disperse synaptic vesicles or inhibit synaptic transmission, presynaptic phalloidin injection blocked exocytosis and reduced endocytosis measured by action potential-evoked FM1-43 staining. Furthermore, phalloidin stabilization of only cortical actin following pretreatment with latrunculin-A was sufficient to inhibit synaptic transmission. Conversely, treatment of axons with jasplakinolide, which induces F-actin accumulation but disrupts F-actin structures in vivo, resulted in increased synaptic transmission accompanied by a loss of phalloidin labeling of cortical actin but no loss of actin labeling within vesicle clusters. Marked synaptic deficits seen with phalloidin stabilization of cortical F-actin, in contrast to the minimal effects of disruption of a synaptic vesicle-associated F-actin, led us to conclude that two structurally and functionally distinct pools of actin exist at presynaptic sites.

Gene Polymorphisms and Fiber-Type Composition of Human Skeletal Muscle

2012 ◽  
Vol 22 (4) ◽  
pp. 292-303 ◽  
Author(s):  
Ildus I. Ahmetov ◽  
Olga L. Vinogradova ◽  
Alun G. Williams
Keyword(s):  
Skeletal Muscle ◽  
Muscle Fiber ◽  
Human Skeletal Muscle ◽  
Fiber Type ◽  
Vastus Lateralis Muscle ◽  
Type I ◽  
Fiber Types ◽  
Fiber Type Composition ◽  
Type Composition ◽  
Type I Fibers

The ability to perform aerobic or anaerobic exercise varies widely among individuals, partially depending on their muscle-fiber composition. Variability in the proportion of skeletal-muscle fiber types may also explain marked differences in aspects of certain chronic disease states including obesity, insulin resistance, and hypertension. In untrained individuals, the proportion of slow-twitch (Type I) fibers in the vastus lateralis muscle is typically around 50% (range 5–90%), and it is unusual for them to undergo conversion to fast-twitch fibers. It has been suggested that the genetic component for the observed variability in the proportion of Type I fibers in human muscles is on the order of 40–50%, indicating that muscle fiber-type composition is determined by both genotype and environment. This article briefly reviews current progress in the understanding of genetic determinism of fiber-type proportion in human skeletal muscle. Several polymorphisms of genes involved in the calcineurin–NFAT pathway, mitochondrial biogenesis, glucose and lipid metabolism, cytoskeletal function, hypoxia and angiogenesis, and circulatory homeostasis have been associated with fiber-type composition. As muscle is a major contributor to metabolism and physical strength and can readily adapt, it is not surprising that many of these gene variants have been associated with physical performance and athlete status, as well as metabolic and cardiovascular diseases. Genetic variants associated with fiber-type proportions have important implications for our understanding of muscle function in both health and disease.

Carnitine metabolism in human muscle fiber types during submaximal dynamic exercise

1996 ◽  
Vol 80 (3) ◽  
pp. 1061-1064 ◽  
Author(s):  
D. Constantin-Teodosiu ◽  
S. Howell ◽  
P. L. Greenhaff
Keyword(s):  
Muscle Fiber ◽  
Fiber Type ◽  
Vastus Lateralis ◽  
Group Formation ◽  
Free Carnitine ◽  
Muscle Fiber Types ◽  
Type I ◽  
Fiber Types ◽  
Carnitine Metabolism ◽  
Type I Fibers

The effect of prolonged exhaustive exercise on free carnitine and acetylcarnitine concentrations in mixed-fiber skeletal muscle and in type I and II muscle fibers was investigated in humans. Needle biopsy samples were obtained from the vastus lateralis of six subjects immediately after exhaustive one-legged cycling at approximately 75% of maximal O2 uptake from both the exercised and nonexercised (control) legs. In the resting (control) leg, there was no difference in the free carnitine concentration between type I and II fibers (20.36 +/- 1.25 and 20.51 +/- 1.16 mmol/kg dry muscle, respectively) despite the greater potential for fat oxidation in type I fibers. However, the acetylcarnitine concentration was slightly greater in type I fibers (P < 0.01). During exercise, acetylcarnitine accumulation occurred in both muscle fiber types, but accumulation was greatest in type I fibers (P < 0.005). Correspondingly, the concentration of free carnitine was significantly lower in type I fibers at the end of exercise (P < 0.001). The sum of free carnitine and acetylcarnitine concentrations in type I and II fibers at rest was similar and was unchanged by exercise. In conclusion, the findings of the present study support the suggestion that carnitine buffers excess acetyl group formation during exercise and that this occurs in both type I and II fibers. However, the greater accumulation of acetylcarnitine in type I fibers during prolonged exercise probably reflects the greater mitochondrial content of this fiber type.

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