Histochemical identification of fiber types in canine skeletal muscle.

Proximal and distal skeletal muscles from pectoral and pelvic limbs were histochemically examined in 18 neuromuscular disease-free dogs. On the basis of the human system of classification and nomenclature and results of standard adenosine triphosphatase (ATPase) and glycine-formaldehyde preincubation procedures, the fiber types identified in immature and mature canine skeletal muscles were I, IIA, and IIC.     

A comparative histochemical study of plant polyphenols in southern African grasses

Recent anatomical studies have shown that tannin‐like substances (TLS) occur in the epidermal cells of a number of southern African tropical grasses, and the presence of condensed tannins in grasses has been confirmed by chemical analyses. A number of species from four of the five subfamilies of the Poaceae were compared for their responses to a range of histochemical tests which differ in their specificity for phenolic compounds. These included: ferrous sulphate, acidified vanillin, diazotized sulphanilic acid, Fast Blue‐BB, dimethoxybenzaldehyde and nitrous acid, Safranin and Fast Green. In addition, the radial diffusion test for protein precipitation was used. Comparative histochemical tests indicated that most taxa known to contain TLS showed comparable responses to the tests used here, with variations in intensity and hue of the coloured products formed. These qualitative differences suggest the presence of a number of different compounds including oligomeric procyanidins, oligomeric prodelphinidins, monomeric and/or dimeric flavan‐3‐ols and flavan‐3,4‐diols. The presence flavan‐4‐ols has been confirmed in the andropogonoid grasses by previous workers. Histochemical tests are adequate to identify the presence of condensed tannins and their precursors in plant tissue. However, they do not provide a means to identify those compounds which precipitate protein and function as digestibility‐reducing compounds in plant‐herbivore interactions.     

A morphometric study of the canine colon: comparison of control dogs and cases of colonic disease.

The microstructure of the canine colon was described morphometrically. The artifacts induced by administration of enemas and biopsy technique were studied by comparing biopsy specimens to tissues obtained at necropsy from 15 normal dogs. Biopsies from control dogs and clinical cases of colonic disease were then evaluated quantitatively, and histological abnormalities which might clarify mechanisms underlying large bowel dysfunction in the dog were sought. In control dogs, gland length and diameter, epithelial, goblet cell and mucosal mast cell numbers, and intraepithelial lymphocyte and mitotic indices were remarkably uniform throughout the colon. Minor variations were found in the proximal and distal regions of the colon. An apparent shortening of glands, and a reduction in mucous goblets and intraepithelial lymphocytes in biopsies were attributed to suboptimal orientation and irritation caused by enemas. The only significant difference from controls identified by morphometric analysis of biopsies from clinical cases was fewer epithelial cells lining longitudinal sections of glands. It was concluded that failure to identify morphometric variations in the colonic mucosa of clinical cases might reflect either a biased, homogeneously mild clinical syndrome in this group, or the possibility that in many of the clinical cases, a functional rather than physical abnormality was involved. The proprial inflammatory cell population was not examined quantitatively; further investigation of this component is merited.     

Effect of dietary protein on calpastatin in canine skeletal muscle

The cysteine proteinases, mu- and m-calpain, along with their inhibitor, calpastatin, have been hypothesized to play a role in skeletal muscle protein degradation. Because nutrition has previously been shown to influence the expression of calpastatin, the working hypothesis of this study was that the quantity and source of dietary protein could influence regulation of the calpain system in muscle. The objectives to support this hypothesis were to determine the effects of dietary protein (amount and source) on the expression of calpastatin in canine skeletal muscle. This study comprised eight diets with seven dogs per diet. A biopsy was taken from the biceps femoris of all 56 dogs before and after 10 wk on their respective diets. This experimental design allowed examination of change within individual dogs. Diets 1 to 4 contained 12% total protein derived from chicken and/or corn gluten meal in ratios of 100:0, 67:33, 33:67, and 0:100%, respectively. Diets 5 to 8 contained 28% total protein with protein sources and ratios identical to Diets 1 to 4. Differences in calpastatin were examined qualitatively using SDS-PAGE and immunoblotting, and semiquantitatively with densitometric analyses. The majority of the calpastatin blots showed three distinct calpastatin bands, the uppermost appearing at approximately 110 kDa. Diet 5 (28% CP, 100% chicken) resulted in an increase in the expression of the 110-kDa calpastatin band compared with the other two lower molecular weight bands in the same samples. Muscle from dogs fed Diet 5 showed greater increase in (P < 0.05) calpastatin intensity of the topmost band than those fed Diet 8 (0:100; chicken:corn gluten meal). Diet 5 (100:0; chicken:corn gluten meal) showed greater total calpastatin intensity than Diet 8 (0:100; chicken:corn gluten meal). These data suggest that dogs fed a diet containing a higher total percentage of chicken protein may have a greater potential to regulate calpain-mediated degradation of muscle protein than dogs fed diets containing corn gluten meal.     

Histochemical differentiation of fiber types in neonatal canine skeletal muscle

Differentiation of fiber types in developing canine skeletal muscle was studied, using morphologic, morphometric, and histochemical techniques. Sample collections were made from 6 muscles from the pectoral and pelvic limbs of 16 healthy pups between 1 day and 12 weeks of age. In newborn pups, 90% to 95% of the fibers in the 6 muscles were classified as undifferentiated or type IIC; the remaining fibers were classified either normal or large-size type I. Large-size type I fibers usually accounted for 2% to 4% of the total population and were considered analogous with the B fiber of Wohlfart. These fibers were larger than all other fiber types and disappeared after pups reached 4 to 5 weeks of age. After 2 to 4 weeks, the number of undifferentiated fibers decreased with the appearance of, and the concomitant numerical increases of, normal size type I and type IIA fibers. The percentages of type I and IIA fibers approached proportions of the adult dog by 12 weeks, at which time a type IIA fiber predominance was present in biceps femoris, lateral head of the gastrocnemius, cranial tibial, and long head of the triceps. Type I fibers predominated in medial head of the triceps and superficial digital flexor after 4 to 5 weeks. The mean fiber diameters of type I and IIA fibers were similar to any given muscle throughout the postnatal development. All fiber types stained uniformly with the oxidative stain nicotinamide adeninedinucleotide-tetrazolium reductase during the first 12 weeks of life, whereas a distinction between type I and II fibers was evident after 3 to 4 weeks with the periodic acid-Schiff stain reaction.     

Technique for physiological examination of canine skeletal muscle in vivo

A technique is described for studying the physiological function of canine skeletal muscle in vivo. The contractile properties of the tarsal flexor muscles were examined in three beagle dogs under general anaesthesia. The force responses to electrical stimulation of the common peroneal nerve were measured at various frequencies to determine the frequency:force relationship for this muscle group. Fatigue characteristics were also examined during intermittent stimulated activity delivered in a set pattern of frequencies. The results provide quantitative characterisation of muscle function which is repeatable. The technique described could be applied to other animals and is a potentially powerful tool for evaluating the effects of drugs on muscle performance.     

A comparative pathological study on canine necrotizing meningoencephalitis and granulomatous meningoencephalomyelitis

Canine necrotizing meningoencephalitis (NME) and granulomatous meningoencephalomyelitis (GME) were compared pathologically. Gross observation exhibited lateral ventricular dilation and discoloration, malacia and/or cavitation of the cerebrum in NME. On the contrary, gross changes were milder in GME, except for occasional visible granulomatous mass formation. Histopathologically, the lesions of NME were distributed predominantly in the cerebral cortex and various degrees of inflammatory and necrotic changes were observed according to clinical stages. Besides, microscopic lesions of GME were mainly distributed in the white matter of the cerebrum, cerebellum and brainstem, which are characterized by perivascular cuffing, multiple granulomas and leptomeningeal infiltrates. Although macrophages and lymphocytes were predominant in the inflammatory lesions of both disorders, macrophages in GME transformed into epithelioid cells and exhibited more massive infiltration. Although lectin RCA-1-reactive cells were numerous in both disorders, lysozyme immunoreactive cells in NME were fewer than that in GME. Parenchymal infiltration of MAC387-positive cells was common in GME and limited in NME. The number of CD3-positive lymphocytes in the GME lesions tended to be greater than in NME, though the difference was not statistically significant. Morphological and immunohistochemical differences of the lesions, in particular, the characteristics of infiltrative macrophages may reflect these different pathogeneses of the two disorders.     

A morphometric comparative study of the medial geniculate body of the rabbit and the fox

SUMMARY: Unbiased stereological methods were used to morphometrically examine and compare the medial geniculate body (MGB) of two species from different mammalian orders. The MGB had a similar nuclear pattern, and it was parcelled into three major cytoarchitectural areas: the dorsal nucleus (MGd), the ventral nucleus (MGv) and the medial nucleus (MGm). The MGd was predominant in the fox, where it contributed nearly 50% to the total MGB volume, while in the rabbit, the MGv was insignificantly larger than the MGd. In both species, the percentage contribution of the MGm was the lowest. The MGd in the fox was also characterized by twice as many neurons per mm(3) as in the rabbit, whereas a reverse proportion was observed in the MGm, although the numerical density in the MGv was very similar in both species. The total number of MGB neurons in the fox was over twice higher than that in the rabbit. The variability in the percentage contribution of the MGd, MGv and MGm cells to the total neuronal population of the MGB was different in both mammals. In the rabbit, there was a larger contribution from the MGv and MGm, while in the fox, the MGd was predominant. These data demonstrate that the main areas of the MGB complex differ in terms of the morphometric characteristics in both species. Our results also show that the negative correlation between the volume and numerical density in the sensory centres of the brain might not be as distinct as in the non-sensory brain structures.     

Biochemical evaluation of mitochondrial respiratory chain enzymes in canine skeletal muscle

OBJECTIVE: To perform respiratory chain enzymatic activity assays on canine skeletal muscle biopsy specimens and establish reference range values of skeletal muscle enzyme activities for dogs. SAMPLE POPULATION: Biopsy specimens from the vastus lateralis muscle were obtained from 24 dogs (8 sexually intact males and 14 sexually intact females) ranging from 15 months to 6 years of age. PROCEDURE: Mean values of citrate synthase, cytochrome-c oxidase, succinate dehydrogenase, succinate dehydrogenase-cytochrome-c reductase, nicotinamide adenine dinucleotide (NADH) dehydrogenase, and NADH dehydrogenase-cytochrome-c reductase activities were established by use of 6 standard spectrophotometric assays for respiratory chain enzyme analysis. RESULTS: Compared with published data for skeletal muscle enzyme activities in humans, skeletal muscle enzyme activities in dogs were 2- to 4-fold higher. Additionally, citrate synthase activity, a marker for mitochondrial volume, was positively correlated with age in dogs, suggesting that mitochondrial volume increases with age, although no apparent change in respiratory chain enzymatic activity with an increase in age was found. CONCLUSIONS AND CLINICAL RELEVANCE: Reference range values for skeletal muscle enzyme activities of dogs are needed to accurately interpret results of respiratory chain enzymatic activity assays. During investigation of metabolic myopathies, if skeletal muscle biopsy specimens are evaluated for respiratory chain enzyme kinetics, they should be performed and evaluated in concert with skeletal muscle biopsy specimens from clinically normal animals of the same species.     

Simplified technique for histochemical determination of three fiber types in equine skeletal muscle

For determination of 3 muscle fiber types in equine skeletal muscle, a comparison of 2 preincubation buffers, each followed by myosin adenosine triphosphatase staining, was made. Serial sections of the muscle samples (n = 75) were preincubated in an acid buffer (pH 4.6) or a formaldehyde-glycine buffer (pH 7.25) and then were stained for myosin adenosine triphosphatase. Differentiation of muscle fibers into type I, IIA, and IIB was identical with both techniques; however, in the samples prepared at pH 4.6, type I fibers were black; type IIA, light gray; and type IIB, dark gray. In the samples prepared at pH 7.25, types I, IIA, and IIB fibers were white, light gray, and dark gray respectively. The formaldehyde-glycine preincubation buffer (at pH 7.25) gave more consistent results, was easier to prepare, and retained cytoarchitecture better, compared with the samples prepared at pH 4.6.     

Oxidative capacity of skeletal muscle fibres in racehorses: histochemical versus biochemical analysis

The oxidative capacity of skeletal muscle fibre types was evaluated histochemically using the nicotinamide dinucleotide diaphorase (NADH-D) staining, and biochemically by measuring the activity of citrate synthase (CS) in both whole muscle samples and in pools of fibres of identified type. Duplicate determinations of the NADH-D staining pattern resulted in standard deviations (sd) between duplicates of 6 and 11 per cent for two observers. The NADH-D pattern was found to differ between observers. Duplicate determinations of CS activity in the same fibre pools resulted in an sd value of 2.9 mumol/g/min. Measurements of whole muscle CS activity did not provide information about the distribution of oxidative capacity among fibre types. The NADH-D stain and CS activity in fibre pools both showed that, in general, type I and IIA fibres had a higher oxidative capacity than type IIB fibres. Biochemical techniques also showed, however, that the CS activity in type I and IIA fibres of different horses could vary as much as twofold, whereas the NADH-D rating showed a high intensity staining for most type I and IIA fibres in all horses. Furthermore, type IIB fibres received a lower NADH-D rating than the other fibre types even when the CS activities were quite similar. For purposes of research, biochemical measurement of oxidative capacity in individual muscle fibre types provides valuable quantitative and comparative information. The ease of histochemical NADH-D staining in comparison to fibre dissections makes this technique more practical for routine estimates of the distribution of oxidative capacity among muscle fibres.(ABSTRACT TRUNCATED AT 250 WORDS)     

Masticatory and skeletal muscle myositis in canine leishmaniasis (Leishmania infantum)

Twenty-four dogs with a parasitologically and serologically established diagnosis of leishmaniasis were studied to investigate the atrophy of the masticatory muscles which commonly occurs in this disease, and to compare the lesions in the masticatory muscles with those in the cranial tibial muscles. The 24 animals were divided into three groups of eight, group A dogs with no muscular atrophy, group B dogs with different degrees of atrophy in the masticatory and skeletal muscles, and group C dogs with similar degrees of atrophy in the masticatory and skeletal muscles. Increased activities of creatine phosphokinase and lactate dehydrogenase were recorded in only some of the dogs in groups B and C, but there were no significant differences between the mean activities in the three groups. Electromyographic changes indicating myopathy and involving both the temporalis and cranial tibial muscles, were observed in two of the dogs in group A, seven of those in group B, and in all the dogs in group C. Muscle histopathology revealed a variable degree of muscle fibre necrosis and atrophy, mononuclear infiltrates and neutrophilic vasculitis in all the dogs except two in group A. Leishmanial amastigotes were found within macrophages and myofibres in 16 of the dogs, some in each group. IgG immune complexes were detected in muscle samples, and circulating antibodies against myofibres were detected in serum samples from all the 24 dogs.     

The effect of testosterone on gastrocnemius muscle fibres in growing and adult male and female rats: a histochemical,morphometric and ultrastructural study

In this study, the effect of testosterone on gastrocnemius muscle fibres in growing and adult rats (male and female) was examined using histochemical, morphometric and ultrastructural techniques. After physiological saline (PS), olive oil (OvO) or olive oil + testosterone (OvOT) injections on 72 rats (growing and mature, 36 male and 36 female), the sample tissues of fibre types of the gastrocnemius muscle taken were examined by histochemical [alkaline adenosine triphosphatase (alk-ATPase), acid ATPase (ac-ATPase)], morphometric and ultrastructural techniques. In PS-injected control groups, the gastrocnemius muscle of both sexes contained all the fibre types studied [slow-oxidative muscle fibres (type I), fast-oxidative glycolytic muscle fibres (type IIA) and fast-glycolytic muscle fibres (type IIB)]. The type I fibres had the smallest diameter, type IIA had a medium diameter and type IIB fibres had the largest diameter. In OvO-injected groups, it was observed that the OvO had little effect on the gastrocnemius muscles of either sex, although there was significant enlargement of type IIB fibres. After the injection of OvOT, hypertrophy of muscle fibres was determined by morphometric study. The biggest increase in diameter was on type I fibres. In addition, degenerations on some mitochondria, accumulation of lipid droplets on type I and type II fibres, an increase in glycogen particles, bifurcation of myofibrils, an increase in the number and diameter of units resembling T tubules and an increase in ribosomal content were also observed in the same group by transmission electron microscope. Consequently, it was determined that testosterone can induce protein synthesis in gastrocnemius muscle fibres, and induces changes in shape and size, and also can change the appearance and the number of fibres.     

A comparative enzyme histochemical and histological study of the effect of ischaemia and post mortem change on rat kidneys.

A histochemical study of the effect of ischaemia on rat kidneys showed that changes were demonstrable in adenosine triphosphatase, alkaline phosphatase and succinic dehydrogenase within 2 h. Further changes occurred with increasing time. The activity of acid phosphatase was little affected up to 24 h although at this time there was marked tubular disruption. Paraffin embedded H and E sections also showed marked changes within 2 h. Enzyme histochemical and histological changes in kidneys taken at varying periods after the death of the animal showed very similar changes to those in ischaemic kidneys. Differences were mainly in the rate and extent of the changes.     

Lectin histochemical characteristics of the canine female mammary gland.

Twelve biotinylated lectins and an avidin-biotin-peroxidase method were used to detect and localize specific carbohydrate residues on formalin-fixed, paraffin-embedded female canine mammary gland sections. Histologic sections from 3 lactating and 7 nonlactating mixed-breed dogs (age 5.6 +/- 0.35 years) were incubated with Arachis hypogea agglutinin (peanut agglutinin; PNA), Concanavalia ensiformis agglutinin (conA), Dolichos biflorus agglutinin (DBA), Glycine max agglutinin (SBA), Griffonia simplicifolia agglutinin-I (GS-I), Lens culinaris agglutinin (LCA), Lycopersicon esculentum agglutinin (LEA), Phytolacca americana mitogen (pokeweed mitogen; PWM), Ricinus communis agglutinin-I and -II (RCA-I and -II), Triticum vulgaris (WGA), and Ulex europaeus agglutinin-I (UEA-I). Each lectin had a specific binding pattern, except SBA and DBA. In nonlactating glands, PNA, conA, LEA, and UEA-I stained duct cells in a linear-binding pattern, with a mean percentage of positive ducts per section of 28.7 (+/- 0.6), 65.7 (+/- 0.3), 100 (+/- 0), and 8.4 (+/- 0.2), respectively. Strong apical, lateral, basal, and cytoplasmic positivity on duct cells was seen after incubation of the sections with RCA-I, RCA-II, and WGA in all ducts. In acinar cells, the binding pattern and the staining distribution of all the lectins studied were similar to those in duct cells. However, for PNA, conA, and UEA-I, the mean percentage of positive lobules per section was 33.7 (+/- 0.9), 62 (+/- 0.5), and 10.5 (+/- 0.2), respectively. In glands from lactating dogs, conA and UEA-I did not stain. The cytoplasm of all myoepithelial cells was moderately stained with RCA-I, RCA-II, and WGA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Nutritional aspects of post exercise skeletal muscle glycogen synthesis in horses: A comparative review

Carbohydrate (CHO) stored in the form of skeletal muscle glycogen is the main energy source for glycolytic and oxidative ATP production during vigorous exercise in mammals. In man, horse and dog both short‐term high intensity and prolonged submaximal exercise deplete muscle glycogen. In horses, however, muscle glycogen synthesis is 2–3‐fold slower than in man and rat, even when a diet high in soluble CHO is fed. There appear to be significant differences in CHO and glycogen metabolism between horses and other mammals, and it is becoming increasingly clear that many conclusions drawn from human exercise physiology do not apply to horses. This review aims to provide a comprehensive, comparative summary of the research on muscle glycogen synthesis in horse, man and rodent. Species differences in CHO uptake and utilisation are examined and the issues with feeding high soluble CHO diets to horses are discussed. Alternative feeding strategies, including protein and long and short chain fatty acid supplementation and the importance of rehydration, are explored.     

Morphological, histochemical and morphometric study of the myotomal muscle tissue of the pacu (Piaractus mesopotamicus Holmberg 1887: Serrasalminae, Characidae, Teleostei)

Histochemical, ultrastructural and morphometric methods were used to study growth patterns of red, pink and white muscle fibres and their relation to body weight and total length in the fast-growing freshwater fish Piaractus mesopotamicus Holmberg. The correlations amongst body weight, body length and diameter of red, pink and white fibres were low. From 10-15 to 40-50 cm, body weight increased 102.7 times, while the diameter of each type of fibre increased by factors of 0.94, 0.74 and 0.70, respectively. Muscle fibres revealed different morphological and histochemical stages of maturation. The frequencies of < 20 microns fibres of red, pink and white muscle tissue in the youngest and oldest classes were 64.5 and 11.0, 38.2 and 7.7 and 24.0 and 1.4%, respectively. In 30-40 cm fish, the frequency of < 20 microns fibres in the red and pink tissue was 24.5 and 25.5%, while in the white tissue it was 11.5%. During sexual maturity (40-50 cm), the recruitment of < 20 microns fibres in white muscle was 1.4%. Muscle fibres of this species showed continuous growth by both hyperplastic and hypertrophic mechanisms, and hyperplasia was particularly active in the juvenile phase.     

A cohort study of canine testicular neoplasia.

A prospective epidemiologic study of canine testicular neoplasia was undertaken in the Philadelphia area in 1971, with the cooperation of private veterinary practitioners. By the end of 1975, 938 dogs had been monitored for an average of 2 years. The cohort consisted of 609 cryptorchid and 329 age- and breed-matched controls. The incidence of testicular neoplasia in the cryptorchid subcohort was 12.7/1,000 dog-years at risk. Testicular neoplasms did not develop in controls. A large proportion of the dogs were below the average age at onset for this neoplasm. Among dogs over 6 years of age, the incidence was 68.1/1,000 dog-years at risk. The incidence of Sertoli cell tumors and seminoma was approximately twice as high in dogs with unilaterally retained inguinal testicles as in abdominal cryptorchids. Sertoli cell tumors developed in 10 dogs and seminoma developed in 6. One half of the testicular neoplasms that developed did so within the first year of observation. This study demonstrated the feasibility of conducting prospective epidemiologic studies of canine diseases with the assistance of practicing veterinarians.