Effects of repeated biopsying on muscle tissue in horses.

The collection of muscle samples in horses to estimate their performance potential, adaptations to training and diagnosis of exertional myopathies is of increasing interest. This study was designed to examine whether repeated biopsying of the gluteus medius muscle at 7 week intervals alters morphology and muscle fibre variables in the sampling area. The gluteus had been biopsied 3 times previously, each biopsy separated by a 7 week interval. Seven weeks after the last biopsy, this study was initiated. Percutaneous needle biopsies were taken from left and right muscles at 2 depths (2 and 6 cm) and at 2 sites for each depth (22 and 25 cm from the tuber coxae) of 7 mature Thoroughbreds. The 22 cm site was located close (2 cm caudal) to an area of the muscle that had been biopsied previously. The 25 cm site was 5 cm apart from this area. A total of 52 samples were available for histology, electron microscopy and immunohistochemistry. The latter was used to study percentages, sizes and capillaries of muscle fibre types. Most muscle samples collected (75%) had normal morphology, but the remaining 25% showed signs of muscle repair. In some circumstances, they showed regenerative signs of complete muscle repair, while in others ineffective muscle repair (scar formation) was evident. Nevertheless, sufficient normal tissue was always available for measuring routine muscle variables. Samples collected 3 cm laterally apart showed large differences with regard to muscle fibre type variables examined. These results show that repeated muscle samples in intervals of 7 weeks do not have effects that would impair evaluation of muscle fibre variables for diagnosis of effectivity of conditioning programmes. Furthermore, they emphasise that the only way of keeping variations through sample collection technique small is to standardise all biopsying techniques accurately.

Intramuscular distribution of fibre types in the gluteus medius of the horse: a histochemical analysis.

The results from a recent study by Bruce and Schurg (1990) indicate that the gluteus medius (Gm) of the horse is a complex muscle subdivided by an internal tendinous sheet into two regions (dorsal and ventral) which may subserve different functions. In the present study, both regions were found to contain a similar proportion of histochemically-determined muscle fibre types. Fibre type composition did not generally vary over the length of the muscle at the same relative depth. In contrast, there was a significant tendency for the percentage of type I fibres to increase gradually in accordance with depth. These histochemical results do not suggest that the two regions of the Gm assume distinct functional roles; they do, however, indicate that at a given relative depth the muscle fibre type distribution is relatively uniform over the length of the muscle. Nevertheless, care should be exercised in sampling and interpreting data obtained from limited biopsy of equine Gm, due to the substantial histochemical variation of fibre type composition as a function of sampling depth.

Variability of muscle fibre composition and fibre size in the horse gluteus medius: an enzyme-histochemical and morphometric study.

To determine the variability in fibre types and fibre sizes in the equine gluteus medius muscle, biopsy specimens were removed from 5 sites, at 4 different depths, within the right and left muscles of 3 Andalusian stallions. The percentage, lesser fibre diameter and cross-sectional area of the various fibre types were measured systematically in myosin ATPase and NADH-tetrazolium reductase-stained, serial cryostat sections of these multiple samples. Significant differences in muscle fibre type composition were recorded, with a lower percentage of type I fibres (high myosin ATPase activity at pH 4.5) being observed towards superficial regions of the muscle and a greater percentage towards the deep areas. Type II B fibres (moderate myosin ATPase activity at pH 4.5), including both II B nonoxidative (low NADH-TR activity) and II B oxidative (moderate NADH-TR activity), displayed the opposite tendency, and the percentage of type II A fibres (low myosin ATPase activity at pH 4.5) did not change with depth. Types I and II A fibres in the deep regions were larger than superficially, whereas the II B fibres in the deep regions were smaller than in the superficial parts of the muscle. The results also imply that type I fibres tend to be larger than type II fibres in the deep regions. The size of type I fibres is more homogeneous in the deep parts than in the superficial regions of the muscle, while II B fibres vary more in size in the peripheral portions than in deep regions. A single biopsy taken from the gluteus medius muscle of the horse is therefore a poor representative of the whole muscle and care should be exercised in sampling and interpreting data obtained from limited biopsy of this muscle. The pattern of variation in fibre types and fibre sizes between the different depths of the muscle probably reflect different functional demands on the gluteus medius muscle.

Skeletal muscle fiber size in untrained and endurance-trained horses.

The mean area and minimal diameter of 3 histochemically determined myofiber types (1, 2A, and 2B; myosin ATPase in acid buffer) were calculated in middle gluteal muscle biopsy specimens from 62 stallions, 47 Andalusians and 15 Arabians, ranging in age from 6 to 12 years. Fourteen Andalusians and 7 Arabians were untrained, and the remainder were actively endurance-trained. The 6-month training schedules involved walking, slow trotting, and cantering. Fourteen Andalusians were moderately endurance-trained, whereas the other 19 Andalusians and 8 Arabians were strongly endurance-trained. Significant differences were not recorded between untrained and endurance-trained Arabians with respect to the area (type 1, 3,194 +/- 869 microns 2 and 3,150 +/- 370 microns 2; type 2A, 3,819 +/- 890 microns 2 and 3,380 +/- 356 microns 2; and type 2B, 4,872 +/- 962 microns 2 and 4,417 +/- 646 microns 2) or minimal diameter (type 1, 52.2 +/- 7.4 microns and 52.8 +/- 3.1 microns; type 2A, 58.1 +/- 6.7 microns and 55.0 +/- 2.8 microns; and type 2B, 65.3 +/- 6.4 microns and 63.4 +/- 4.3 microns) of the 3 fiber types, nor between untrained and endurance-trained Andalusians with respect to the area (untrained, 3,990 +/- 690 microns 2; moderately endurance-trained, 3,882 +/- 347 microns 2; and strongly endurance-trained, 3,758 +/- 510 microns 2) and minimal diameter (untrained, 58.1 +/- 4.7 microns; moderately endurance-trained, 59.7 +/- 2.7 microns; and strongly endurance-trained, 58.7 +/- 4.5 microns) of 2A fibers.(ABSTRACT TRUNCATED AT 250 WORDS)

Muscle fibre type composition in untrained and endurance-trained Andalusian and Arab horses.

Muscle biopsies were taken from the middle gluteal muscle of 68 stallions (52 Andalusian [AN] and 16 Arab [AR]) ranging from six to 12 years of age. Seventeen AN horses and eight AR horses were untrained, while the remainder underwent active endurance training for 6 months. Fifteen AN horses were moderately endurance-trained while the other 20 AN horses and eight AR horses were strongly endurance-trained. Percentages of type I and type II fibres were similar in all groups (P greater than 0.05). The group of horses with the hardest training had a higher percentage of type IIA fibres (AN P less than 0.01; AR P less than 0.02) and a lower percentage of type IIB fibres (P less than 0.05 for AN and AR) than the untrained horses. All trained horses had a higher percentage of type IIB oxidative fibres and a lower percentage of type IIB non-oxidative fibres than the untrained horses (P less than 0.01 for both). These results suggest that the proportions of type I and type II fibres are highly stable within a given breed, but the stimulus of training facilitates changes both in contractile properties and, particularly, in the oxidative capacity of type II subgroups.

Type IIC fibres in certain muscles of the adult rat (sedentary and exercised).

Samples were taken, at fixed levels, of the vastus lateralis, the caput lateralis of the gastrocnemius muscle and the longissimus lumbaris of 72 Wistar rats which were either sedentary or subjected to various exercise schedules. The samples were analyzed using the histochemical technique of myosin ATPase (m-ATPase) after preincubation at pH 4.2, and the fibre-types I, II (IIA and IIB) and IIC were identified, calculating the percentage of type IIC fibres as well as their minimum diameter. The percentages of these IIC fibres found in the red and mixed parts of the gastrocnemius (caput lateralis) and the longissimus lumbaris were between 0.7% and 2.6%. However, their presence was not detected in the vastus lateralis or in the white part of the gastrocnemius (caput lateralis). The lack of differences in this fibre type between the males and females of the population was shown statistically. Likewise, no significant modification of the IIC fibres between sedentary and exercised animals was seen. With regard to fibrillar size, females showed a smaller minimum diameter than males, the results showing a small increase in the size of these fibres in both sexes after exercise, although in most cases this was not statistically significant.

Degree of correspondence between contractile and oxidative capacities in horse muscle fibres: a histochemical study.

Samples taken from the middle gluteal muscle of 95 untrained adult horses of different ages and sex were subjected to histochemical analysis using the myosin adenosine triphosphatase (m-ATPase) and nicotinamide adenine dinucleotide tetrazolium reductase (NADH-TR) staining techniques. Fibres were classified into types I, IIA and IIB according to m-ATPase activity after preincubation at pH 4.4. The percentage of FT (Fast-Twitch Glycolytic) fibres and the proportion of IIB fibres with "high" and "low" oxidative capacity were determined in serial sections stained for NADH-TR. Statistical analysis revealed a significantly higher proportion of IIB fibres than FT fibres (P less than 0.001), though both percentages were correlated. Thus, 72.2 +/- 17.6% of type IIB fibres showed low oxidative capacity, but the remaining 27.8 +/- 17.6% showed high aerobic potential, and thus did not correspond to FT fibres. These results confirm that the contractile capacity of a muscle fibre does not determine its oxidative profile. The different types of muscle fibre should thus be classified solely according to m-ATPase activity, since this characteristic is related to the molecular structure of contractile proteins. Oxidative capacity should be assessed separately, and not be used as a criterion for fibre classification in horses.