Myosin heavy chain isoforms in single fibres from m. vastus lateralis of soccer players: effects of strength-training

The myosin heavy chain (MHC) composition of single fibres (n= 2171) was analysed with an electrophoretic technique in biopsy material from m. vastus lateralis of two groups of soccer players before and after a 3-month period of either strength- (n= 8) or non-training (control) (n= 6). Traditional myofibrillar ATPase histochemistry demonstrated a decrease in type IIA fibres with strength-training (35.4 ± 2.1 vs. 26.7 ± 2.4% (P < 0.05)). This was not observed in the non-training group (25.7 ± 4.6 vs. 23.8 ± 1.7%). One-dimensional electrophoresis on muscle homogenates showed no significant change in the amount of MHC isoforms in either of the two groups. The MHC isoform I IB was undetectable in all but three samples. No changes in the proportions of fibres containing any of the MHC isoforms were observed. Fibres containing only MHC isoform I IB were found in very small numbers (only 11 out of 2171). Before the experimental period, between 6 and 10% histochemical type IIB fibres were found in both groups. This was identical with the proportion of fibres showing co-existence of MHC isoforms IIA and IIB, but in contrast to the very few fibres containing only MHC isoform IIB. This suggests that nearly all histochemical type IIB fibres of the soccer players display co-existence of both MHC isoform IIA and IIB. No major change in the muscle fibre area of the two groups was observed. The strength-training group increased mean power output in a short-term dynamic knee-extensor test after strength-training (119 ± 6.6 vs. 136.5 ± 4.7 W (P < 0.01)), whereas no change was observed in the non-training group (151.2 ± 4.5 vs. 148.4 ± 6.5 W).     

There is no slowing of motility speed with increased body size in rat, human, horse and rhinoceros independent on temperature and skeletal muscle myosin isoform

Aim: The predictions of scaling of skeletal muscle shortening velocity made by A.V. Hill 60‐years ago have proven to be remarkably accurate at the cellular level. The current investigation looks to extend the study of scaling of contractile speed to the level of the molecular motor protein myosin at both physiological and unphysiological low temperatures. Methods: A single muscle cell in vitro motility assay to test myosin function, i.e. myosin extracted from short single muscle fibre segments, was used in four species representing a 5 500‐fold difference in body mass (rat, man, horse and rhinoceros) at temperatures ranging from 15 to 35 °C. Results: The in vitro motility speed increased as the temperature of the assay increased, but a more profound effect was observed on the slower isoforms, narrowing the relative differences between fast and slow myosin heavy chain (MyHC) isoforms at physiological temperature in all species. The in vitro motility speed varied according to MyHC isoform within each species: I < IIa < IIx < IIb, but the expected scaling relationship within orthologous myosin isoforms was not observed at any temperature. Conclusion: The scaling effect of body size and limb length on shortening velocity at the muscle fibre level, i.e. the decreasing shortening velocity associated with increasing body weight and limb length, was not confirmed at the motor protein level when including mammals of very large size. Thus, other factors than myosin structure and function appear to cause this scaling effect and thin filament isoform expression or myofilament lattice spacing are forwarded as alternative underlying factors.     

Myosin heavy chain isoforms in single fibres from m. vastus lateralis of sprinters: influence of training

The myosin heavy chain (MHC) composition of single fibres from m. vastus lateralis of a group of male sprint athletes (n= 6) was analysed, before and after a three months period of intensive strength- and interval-training, using a sensitive gel electrophoretic technique. Significant improvements were observed after training in almost all of a series of performance tests. After training the sprinters revealed a decrease in fibres containing only MHC isoform I (52.0 ±3.0% vs. 41.2 ±4.7% (mean±SE) (P < 0.05)) and an increase in the amount of fibres containing only MHC isoform IIA (34.7 ±6.1% vs. 52.3 ±3.6% (P < 0.05)). Fibres showing co-existence of MHC isoforms IIA and IIB decreased with training (12.9 ± 5.0% vs. 5.1 ±3.1%, (P < 0.05)). Only one out of 1000 fibres analysed contained only MHC isoform IIB. In contrast, a higher amount of type IIB fibres (18.8 ± 3.6% vs. 10.5 ± 3.9%, (P < 0.05)) was observed with myofibrillar ATPase histochemistry. The majority of histochemically determined type IIB fibres of sprinters seems therefore to contain both MHC isoforms IIA and IIB. Sprint-training appears to induce an increased expression of MHC isoform IIA in skeletal muscles. This seems related to a bi-directional transformation from both MHC isoforms I and IIB towards MHC isoform IIA.     

Favourable associations between the myosin heavy-chain and light-chain isoforms in human skeletal muscle

Histochemical and biochemical analyses were performed in order to examine the relationship between myosin light-chain (LC) isoforms and fibre-type distributions in whole human skeletal muscle. Muscle biopsies were obtained from the vastus lateralis muscle in six healthy men, and analysed for the relative area occupied by each fibre type (percentage of fibre type area) and the molar ratio of each LC isoform. The percentage of type I fibre area was positively correlated with the molar ratio of slow LC (LC1s and LC2s) to total LC. The regression line was located below the line of unity. Also, the ratio of percentage of type II fibre area to that of type II fibre area was positively correlated with the molar ratio of the fast alkali LC LC1f to fast alkali LCs LC1f and LC3f. These results support previous study, having shown that in human skeletal muscle some type I fibres express various amounts of fast LC in addition to slow LC and suggest that fast myosin heavy-chain HCII a is favourably associated with LC1f, whereas HCIIb is favourably associated with LC3f.     

Human skeletal muscle myosin function at physiological and non-physiological temperatures

Aim: The aim of the study was to assess the function of human skeletal muscle myosin across a wide range of temperatures, including physiological. Methods: We used a single fibre in vitro motility assay. The in vitro motility speed of actin filaments propelled by myosin extracted from fibres expressing type I myosin heavy chain (MyHC; n = 9), IIa MyHC (n = 6), IIax MyHC (n = 4) and I/IIa MyHC (n = 1) was measured at 15, 20, 25, 30 and 35 °C. Results: The motility speed between groups of fibres expressing different MyHC differed significantly (P ? 0.001). The increase in motility speed with an increase in temperature was statistically significant (P ? 0.001) between all temperatures. The relative difference in motility speed between the slow type I and the fast IIax MyHC fibres decreased with increasing temperature, i.e. a 7.5‐fold difference at 15 °C was reduced to twofold at 35 °C. Furthermore, the twofold difference in motility speed between type IIa and IIax MyHC at 15 °C disappeared completely at 35 °C. The activation energy, E_A, and temperature coefficient, Q₁₀, over the 15–35 °C temperature range was higher for type I MyHC, 54.47 ± 4.37 kJ mol^?1 and 2.09 ± 0.12, respectively, than for type IIa MyHC, 45.41 ± 3.12 kJ mol^?1 (P < 0.001) and 1.85 ± 0.08 (P < 0.001), or IIax MyHC, 34.71 ± 1.75 kJ mol^?1 (P ? 0.001) and 1.60 ± 0.04 (P ? 0.001). Conclusion: The present results suggest a significantly reduced difference in shortening velocity between different human muscle fibre types at physiological temperature than previously reported at lower temperatures (12 or 15 °C) with implications for human in vivo muscle function.     

Rate of active tension development from rigor in skinned atrial and ventricular cardiac fibres from swine following photolytic release of ATP from caged ATP

We investigated the rate of tension development (k_td) after photolytical release of ATP from, P³-l-(2-nitrophenyl)-ethyladenosine-5′-triphosphate (‘caged ATP’) of atrial and ventricular fibre bundles from pig. Contraction was initiated from high-tension (HT) and low-tension (LT) rigor at maximal Ca²⁺ activation (pCa 4.5). The k_td of atrial fibre bundles was 6.8 s^_1 from LT and 6.9 s^_1 from HT rigor. Rate of tension development of ventricular fibre bundles was significantly lower (P < 0.001) being 1.06 s“¹ and 0.94 s”¹ from LT and HT rigor, respectively. The k_td of skinned ventricular fibre bundles incubated in a high [K⁺], low [Ca²⁺] (cardioplegic) solution prior to the skinning procedure decreased significantly (P < 0.05) to 0.73 s^-1 and 0.63 s_¹ from LT and HT rigor, respectively, whereas that of skinned atrial fibre bundles remained at 7.1 s^-1 and 6.9 s^-1 from LT and HT rigor, respectively. Phosphorylation levels of the myosin light chain 2 isoform in the atrial fibre bundles (ALC-2) was 15.6±2.7%. The corresponding values for the two ventricular isoforms, VLC-2 and VLC-2*, were 31.2 ± 0.4% and 25.1 ±2.1%, respectively. Phosphorylation levels of fibre bundles incubated in cardioplegic solution prior to skinning were 11.6%, 18.9%, and 15.4% of the ALC-2, VLC-2 and VLC-2*, respectively. The results show that the rate of tension development is more than seven-fold higher in the atrial compared with ventricular fibre bundles. These results correlate with the differences in ATPase activity of the contractile proteins in solution and, most likely, reflect differences in the myosin isoform composition. In ventricular fibre bundles the increased levels of light chain phosphorylation were associated with increased rate of contraction.     

Sarcoplasmic reticulum of human skeletal muscle: age-related changes and effect of trainirig

The effect of ageing on human skeletal muscle was investigated using needle biopsies from young and aged subjects and from aged subjects trained with different activity patterns. Histochemical staining for myofibrillar ATPase of ageing m. vastus lateralis demonstrated an unchanged fibre type distribution but a selective atrophy of type IIa and type IIb fibres. Analysis of myosin heavy chain (MHC) composition showed that type I MHC increased with ageing (P< 0.05). The relative content of the MHC isoforms correlated with the relative area of the respective fibre types. Sarcoplasmic reticulum (SR) proteins were investigated in muscle extracts by electrophoretic and immunoblotting techniques. When compared to a young control group (28 0.1 years old, n = 7) blots of post-myofibrillar supernatant proteins probed with polyclonal antibodies to the rabbit fast SR Ca-ATPase, a marker of extrajunctional SR, showed that the content of Ca-ATPase was significantly lower (P < 0.05) in the old control group (68 ± 0.5 years old, n= 8). On the other hand the content of calsequestrin (CS), the major intraluminal protein of SR terminal cisternae (TC), and of the 350-kDa ryanodine-binding protein, which is localized in the junctional regions of TC, did not show a concomitant decrease. These results suggest that ageing differentially affects extrajunctional and junctional SR of human skeletal muscle. These age-related changes were not observed within a group of old strength-trained subjects.     

Myosin heavy chain isoform distribution in single fibres of bodybuilders

The purpose of this study was to investigate the long-term effects of high intensity resistance training on myosin heavy chain (MHC) isoform composition of single fibres. Muscle biopsies were obtained from the right vastus lateralis of eight bodybuilders (BB) and seven physical education students (PES). Histochemical analyses were used to determine the fibre type distribution and the fibre cross-sectional area. MHC isoform composition of single fibres was determined with protein electrophoresis. The percentage of fibres expressing MHC IIA and MHC I/IIA was larger in BB (P < 0.05), while MHC IIX was completely absent (P < 0.05). In contrast, myofibrilar ATPase histochemistry only revealed a significantly lower percentage of type IIX fibres in BB (P < 0.05). The muscle fibre profile in the vastus lateralis muscle of BB may represent an adaptation based on the mechanical and biochemical demands of the long-term resistance training.     

Force-velocity properties and myosin light chain isoform composition of an identified type of skinned fibres from rat skeletal muscle

Force-velocity relations, myosin heavy chain (MHC) and myosin light chain (MLC) isoform composition of single skinned fibres from rat plantaris muscle were determined. In fibres containing the same (2X) isoform of myosin heavy chain, several parameters derived from the force-velocity relation and isometric force (Po) were tested for relation with the fibre content in alkali myosin light chain (MLC) isoforms. Whereas maximum shortening velocity was found to be proportional to the relative content in the 3f isoform of alkali MLC, velocity of shortening at 5% relative load, maximum power output, and Po were not. These results strengthen the idea that, in mammalian skeletal fibres, alkali MLC isoforms modulate shortening velocity at zero load, but suggest that they do not control the contractile behaviour at loads higher than zero.     

Regional differences in fibre type composition in the human temporalis muscle

Anatomical and electromyographic studies point to regional differences in function in the human temporalis muscle. During chewing and biting the anterior portions of the muscle are in general more intensively activated and they are capable of producing larger forces than the posterior portions. It was hypothetised that this heterogeneity in function is reflected in the fibre type composition of the muscle. The composition and surface area of different fibre types in various anteroposterior portions of the temporalis muscle were investigated in 7 cadavers employing immunohistochemistry with a panel of monoclonal antibodies against different isoforms of myosin heavy chain. Pure slow muscle fibres, type I, differed strongly in number across the muscle. In the most posterior portion of the muscle there were 24% type I fibres, in the intermediate portion 57%, and in the most anterior portion 46%. The mean fibre cross-sectional area (m-fcsa) of type I fibres was 1849 μm², which did not differ significantly across the muscle. The proportion of pure fast muscle fibres, type IIA and IIX, remained more or less constant throughout the muscle at 13% and 11% respectively; their m-fcsa was 1309 μm² and 1206 μm², respectively, which did not differ significantly throughout the muscle. Pure type IIB fibres were not found. The relative proportion of hybrid fibres was 31% and did not differ significantly among the muscle portions. Fibre types I+IIA and cardiac α+I+IIA were the most abundant hybrid fibre types. In addition, 5% of the type I fibres had an additional myosin isoform which has only recently been described by means of electrophoresis and was named Ia. In the present study they were denoted as hybrid type I+Ia muscle fibres. It is concluded that intramuscular differences in type I fibre distribution are in accordance with regional differences in muscle function.     

Buffer capacity and lactate accumulation in skeletal muscle of trained and untrained men

Buffer capacity (β) of skeletal muscle has been determined in trained (n=7) and in sedentary subjects (n=8). The trained subjects were active in ball games where a high degree of anaerobic energy utilization is required. Percentage fibre type occurrence in the thigh muscle was not significantly different in the two groups. However, there was a tendency towards a higher proportion of type I (slow-twitch) fibres (61.5±11.6% vs. 50.2±12.5%) and a lower proportion of type IIB fibres (2.1±3.5% vs 14.1±16.3%) in the trained subjects. The proportion of the cross-sectional area of the muscle biopsies that was made up of type I or type II fibres was not different in the two groups. All subjects performed an isometric contraction of the knee extensors to fatigue at 61% of their maximal voluntary contraction force. Muscle biopsies were taken from the quadriceps femoris muscle at rest and immediately after contraction. The buffer capacity of muscle was calculated from: β= (Muscle lactate (work)-Muscle lactate (rest))/(Muscle pH (rest) -Muscle pH (work)). A higher buffer capacity (p<0.05) was observed in the trained subjects (β=194±30 mmolxpH^-1xkg^-1 dry wt.) compared to the sedentary group (β=164±20) (mean±SD). An unexpected finding was that muscle lactate after contraction to fatigue was lower (30%, p<0.01) and muscle pH was higher (6.80±0.06 vs. 6.61±0.12, p<0.01) in the trained subjects than in the sedentary controls. Creatine phosphate stores were almost completely depleted in both groups. Post-exercise lactate values were related to the proportion of type II fibres in the muscle (p<0.01). There was, however, no statistical correlation betwe β and fibre type occurrence (p>0.05). In summary, the present results indicate that skeletal muscle buffer capacity can be changed by training in man. Furthermore, it is concluded that the lower lactate accumulation and pH decline after an isometric contraction to fatigue that was observed in the trained compared to the sedentary subjects is related to the training per se. However, the tendency towards a lower type I (slowtwitch) fibre percentage in the trained subjects is likely to have contributed to the observed differences.     

Relationship between myosin heavy chain IId isoform and fibre types in soleus muscle of the rat after hindlimb suspension

Summary The relationship between the myosin heavy chain (HC) IId isoform and histochemically defined fibre types was investigated in the rat soleus muscle after hindlimb suspension. After 4 weeks of suspension, right and left muscles were removed and fibre type composition and total fibre number were examined by histochemical myosin adenosine triphosphatase staining sections. Myosin HC isoforms were analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis. After the suspension, there was a significant decrease in the percentage of type I fibres and a concomitant increase in that of type IIa fibres. However, the total number of fibres was not affected by suspension. The synthesis of HC IId isoform, which was not found in the control, and the decrease in the ratio of slow type myosin heavy chain isoform (HC I) were observed after suspension. These results would may suggest that the change of fibre type composition was caused by a shift from type I to IIa fibres after suspension. Furthermore, it could be suggested that the synthesis of HC IId isoform occurred during the stage of type shift from type I to IIa fibres.     

High‐Intensity Resistance Training with Insufficient Recovery Time Between Bouts Induce Atrophy and Alterations in Myosin Heavy Chain Content in Rat Skeletal Muscle

The aim of this study was to test whether high‐intensity resistance training with insufficient recovery time between bouts, could result in a decrease of muscle fiber cross‐sectional area (CSA), alter fiber‐type frequencies and myosin heavy chain (MHC) isoform content in rat skeletal muscle. Wistar rats were divided into two groups: trained (Tr) and control (Co). Tr group were subjected to a high‐intensity resistance training program (5 days/week) for 12 weeks, involving jump bouts into water, carrying progressive overloads based on percentage body weight. At the end of experiment, animals were sacrificed, superficial white (SW) and deep red (DR) portions of the plantaris muscle were removed and submitted to mATPase histochemical reaction and SDS‐PAGE analysis. Throughout the experiment, both groups increased body weight, but Tr was lower than Co. There was a significant reduction in IIA and IID muscle fiber CSA in the DR portion of Tr compared to Co. Muscle fiber‐type frequencies showed a reduction in Types I and IIA in the DR portion and IID in the SW portion of Tr compared to Co; there was an increase in Types IIBD frequency in the DR portion. Change in muscle fiber‐type frequency was supported by a significant decrease in MHCI and MHCIIa isoforms accompanied by a significant increase in MHCIIb isoform content. MHCIId showed no significant differences between groups. These data show that high‐intensity resistance training with insufficient recovery time between bouts promoted muscle atrophy and a transition from slow‐to‐fast contractile activity in rat plantaris muscle. Anat Rec, 2011. © 2011 Wiley‐Liss, Inc.     

Myosin heavy chain isoform composition and stretch activation kinetics in single fibres of Xenopus laevis iliofibularis muscle

Skeletal muscle is composed of specialized fibre types that enable it to fulfil complex and variable functional needs. Muscle fibres of Xenopus laevis , a frog formerly classified as a toad, were the first to be typed based on a combination of physiological, morphological, histochemical and biochemical characteristics. Currently the most widely accepted criterion for muscle fibre typing is the myosin heavy chain (MHC) isoform composition because it is assumed that variations of this protein are the most important contributors to functional diversity. Yet this criterion has not been used for classification of Xenopus fibres due to the lack of an effective protocol for MHC isoform analysis. In the present study we aimed to resolve and visualize electrophoretically the MHC isoforms expressed in the iliofibularis muscle of Xenopus laevis, to define their functional identity and to classify the fibres based on their MHC isoform composition. Using a SDS-PAGE protocol that proved successful with mammalian muscle MHC isoforms, we were able to detect five MHC isoforms in Xenopus iliofibularis muscle. The kinetics of stretch-induced force transients (stretch activation) produced by a fibre was strongly correlated with its MHC isoform content indicating that the five MHC isoforms confer different kinetics characteristics. Hybrid fibre types containing two MHC isoforms exhibited stretch activation kinetics parameters that were intermediate between those of the corresponding pure fibre types. These results clearly show that the MHC isoforms expressed in Xenopus muscle are functionally different thereby validating the idea that MHC isoform composition is the most reliable criterion for vertebrate skeletal muscle fibre type classification. Thus, our results lay the foundation for the unequivocal classification of the muscle fibres in the Xenopus iliofibularis muscle and for gaining further insights into skeletal muscle fibre diversity.     

Gender-related differences in the regulatory influence of thyroid hormone on the expression of myosin isoforms in young and old rats

The effects of 4 weeks of thyroid hormone (3,5,3′-triiodothyronine, T3) treatment on the expression of myosin heavy chain (MyHC) isoforms were examined in young (3–6 months) and old (20–24 months) female rats, and compared with those in age-matched male rats (Larsson et al. 1995). In control rats, ageing was associated with a type IIA to I MyHC isoform switching in the slow-twitch soleus and a type IIB to IIX MyHC isoform switching in the fast-twitch extensor digitorum longus muscle (EDL). Gender- and muscle-specific differences were observed in the regulation of MyHC isoforms by T3. In the soleus, dramatic downregulation of the type I and upregulation of the type IIA MyHC isoform were observed in both females and males, but upregulation of the IIX MyHC isoform was observed only in male rats. In EDL, T3 treatment had no significant influence on the MyHC isoform composition in the males irrespective of the age of the animal. In the females, on the other hand, T3 treatment resulted in a significant MyHC transformation from IIA to IIB, probably via IIX myosin, in spite of the fact that type IIA mRNA has been reported to be downregulated in both females and males. It is concluded that the regulation of MyHC isoforms by thyroid hormone differs between females and males, presumably as a result of a gender-related difference in the translational or post-translational regulation of MyHC synthesis.     

Influence of fast and slow alkali myosin light chain isoforms on the kinetics of stretch-induced force transients of fast-twitch type IIA fibres of rat

This study contributes to understand the physiological role of slow myosin light chain isoforms in fast-twitch type IIA fibres of skeletal muscle. These isoforms are often attached to the myosin necks of rat type IIA fibres, whereby the slow alkali myosin light chain isoform MLC1s is much more frequent and abundant than the slow regulatory myosin light chain isoform MLC2s. In the present study, single-skinned rat type IIA fibres were maximally Ca²⁺ activated and subjected to stepwise stretches for causing a perturbation of myosin head pulling cycles. From the time course of the resulting force transients, myosin head kinetics was deduced. Fibres containing MLC1s exhibited slower kinetics independently of the presence or absence of MLC2s. At the maximal MLC1s concentration of about 75%, the slowing was about 40%. The slowing effect of MLC1s is possibly due to differences in the myosin heavy chain binding sites of the fast and slow alkali MLC isoforms, which changes the rigidity of the myosin neck. Compared with the impact of myosin heavy chain isoforms in various fast-twitch fibre types, the influence of MLC1s on myosin head kinetics of type IIA fibres is much smaller. In conclusion, the physiological role of fast and slow MLC isoforms in type IIA fibres is a fine-tuning of the myosin head kinetics.     

Calcium sensitivity and myofibrillar protein isoforms of rat skinned skeletal muscle fibres

We investigated the calcium sensitivity for tension generation of different fibre types and the possible correlation between calcium sensitivity and the presence of distinct regulatory protein and myosin light chain (MLC) isoforms in rat skinned skeletal muscle fibres. Fibre types 1, 2A and 2B were identified by electrophoretic analysis of myosin heavy chain (MHC) isoforms. Fibres showing more than one MHC isoform were discarded. Type 1 fibres from the soleus showed a higher pCa (–log₁₀ [Ca], where [ ] denotes concentration) threshold and a lower slope of pCa/tension curve than type 2 extensor digitorum longus (EDL) fibres; between type 2 fibres, type 2B showed the higher slope of pCa/tension curve. Type 1 fibres from different muscles showed similar calcium sensitivities when containing only the slow set of regulatory proteins and MLC; when both slow and fast isoforms were present, calcium sensitivity shifted toward fast type fibre values. Type 2A fibres from different muscles showed a similar calcium sensitivity, independently of the set (purely fast or mixed) of regulatory proteins and MLC. It is suggested that when both fast and slow isoforms of regulatory proteins and of MLC are present in a muscle fibre, calcium sensitivity is dictated mainly by the fast isoforms.     

Dependence of cross-bridge kinetics on myosin light chain isoforms in rabbit and rat skeletal muscle fibres

Cross-bridge kinetics underlying stretch-induced force transients was studied in fibres with different myosin light chain (MLC) isoforms from skeletal muscles of rabbit and rat. The force transients were induced by stepwise stretches (< 0.3% of fibre length) applied on maximally Ca²⁺-activated skinned fibres. Fast fibre types IIB, IID (or IIX) and IIA and the slow fibre type I containing the myosin heavy chain isoforms MHC-IIb, MHC-IId (or MHC-IIx), MHC-IIa and MHC-I, respectively, were investigated. The MLC isoform content varied within fibre types. Fast fibre types contained the fast regulatory MLC isoform MLC2f and different proportions of the fast alkali MLC isoforms MLC1f and MLC3f. Type I fibres contained the slow regulatory MLC isoform MLC2s and the slow alkali MLC isoform MLC1s. Slow MLC isoforms were also present in several type IIA fibres. The kinetics of force transients differed by a factor of about 30 between fibre types (order from fastest to slowest kinetics: IIB > IID > IIA ≫ I). The kinetics of the force transients was not dependent on the relative content of MLC1f and MLC3f. Type IIA fibres containing fast and slow MLC isoforms were about 1.2 times slower than type IIA fibres containing only fast MLC isoforms. We conclude that while the cross-bridge kinetics is mainly determined by the MHC isoforms present, it is affected by fast and slow MLC isoforms but not by the relative content of MLC1f and MLC3f. Thus, the physiological role of fast and slow MLC isoforms in type IIA fibres is a fine-tuning of the cross-bridge kinetics.