A combined myosin ATPase and acetylcholinesterase histochemical method for the demonstration of fibre types and their innervation pattern in skeletal muscle

An improved, combined staining method for myofibrillar ATPase (m-ATPase) and for acetylcholinesterase activity is described. This method allows the observations, on the same slide, of the classical histochemical m-ATPase profile following the Brooke and Kaiser technique and the neuromuscular junction morphology. Thus the pattern of innervation, nerve ending structure and number of nerve endings along the fibres is shown simultaneously for the basic differentiation between slow and fast fibres. The use of acidic and alkaline preincubation allows better visualization of endplate morphology and avoids the masking effect of a positive m-ATPase reaction. The technique has been validated on skeletal muscles from avian and mammalian species.     

The reliability of histochemical fibre typing of human necropsy muscles

The reliability of muscle fibre typing of post mortem specimens was investigated with special reference to the influence of time and temperature. In specimens stored at +4 degrees C, muscle fibre typing could be reliably performed up to at least ten and fifteen days post mortem for the masseter and biceps brachii muscles respectively. The corresponding figures for storage at room temperature were three and six days. The difference in the preservation of enzyme activity between masticatory and limb muscles might be related to the demonstrated difference in the fibre type composition and thus the enzyme content and energy sources.     

The reliability of histochemical fibre typing of human necropsy muscles

Summary The reliability of muscle fibre typing of post mortem specimens was investigated with special reference to the influence of time and temperature. In specimens stored at +4° C, muscle fibre typing could be reliably performed up to at least ten and fifteen days post mortem for the masseter and biceps brachii muscles respectively. The corresponding figures for storage at room temperature were three and six days. The difference in the preservation of enzyme activity between masticatory and limb muscles might be related to the demonstrated difference in the fibre type composition and thus the enzyme content and energy sources.     

Differences in two species-at-risk classification schemes for North American mammals

Several classification systems are used to rank species’ extinction risk. Assessments from two of these, IUCN and NatureServe, are often used to inform prioritisation of conservation resources and management strategies. However, despite their widespread use, they have rarely been compared. No research has assessed rank concordance specifically for mammals, while factors increasing the chance of mismatches between systems have not been investigated. In this study, consistency of IUCN/NatureServe extinction risk categorisation is compared for 409 classified extant American and Canadian mammals. Taxonomic bias in between-system mismatches is then analysed, and common ecological factors associated with mismatches are also identified. There was a significant positive correlation between IUCN and NatureServe ranks, although this was not strong (r_s = 0.504). Agreement was good for non-threatened categories: 97% of species classified as non-threatened by one system were classified likewise by the other. However, there was considerable discord in threatened categories, with 40% of species classified as threatened by one system and non-threatened by the other. In 89% of such cases, this was due to higher ranking by NatureServe, suggesting that this system is more conservative. Mismatches were identified for 102 of the 373 species with exact rankings on both systems (27%), and these were biased taxonomically with significantly more mismatches for Cetacea and fewer for Rodentia. Mismatches were more common for species with longer gestation periods, fewer offspring per year, and longer life expectancies (all traits associated with K-strategist species), as well as for species in higher trophic levels. Many mismatched species also had fragmented ranges and/or uncertain data. Recognition that IUCN and NatureServe ranks are not synonymous is essential. Assessments should be viewed as complementary and dual results should be used to inform species management. The need for more detailed population demographic data to improve extinction risk calculations should also be addressed.     

The histochemical demonstration of myofibrillar ATPase in elasmobranch muscle

Synopsis A modification of the histochemical method of Guth & Samaha (1970) is presented, which gives good results with dogfish material. The routine involves pre-incubation in diethanolamine buffers, and the addition of urea to the preincubation and incubating media. The results obtained accord with the view that contraction speed is related to myofibrillar ATPase activity.     

Myosin-ATPase fibre typing of chemically skinned muscle fibres

Summary The efficacy of myosin (M)-ATPase fibre typing to differentiate fibre types in chemically (EGTA) skinned muscle fibres was investigated. Cryosections or single fibres from isolated bundles of chemically skinned rat extensor digitorum longus (EDL) and soleus (SOL) muscles were stained for M-ATPase activity. The results indicate that two major fibre types (type I and II, Brooke & Kaiser, 1970) can be indentified, as well as subgrouping of the type II fibres into types IIa and IIb. Thus, chemically skinning muscle fibres appears to have no detrimental effects on subsequent M-ATPase fibre typing.     

Comparative lectin histochemical analysis of the duodenal glands in various mammals

Composition and histotopography of lectin receptors have been studied in 12 species of mammals with various nutritional specialization: carnivorous, phytophagous and omnivorous. In cells of the duodenal glands of the carnivorous and omnivorous receptors to concanavalin A and lentil lectin (D-mannosoglycans ) are absent and they are present in the glands of the phytophagous animals. In cells of some parts of the glands presence of receptors to soya bean lectin (N-acetyl-D-galactosamine++) is the most characteristic sign of the duodenal glands in the carnivorous and phytophagous animals. Together with certain differences, depending on the nutritional way of the animals, specific peculiarities of lectins binding with glandulocytes of the duodenal glands are demonstrated. The data on rearrangement of the lectin receptors are obtained during the process of cellular differentiation. Presence of N-acetyl-D-galactosamine++ remnants-biding soya bean lectin in composition of oligosaccharide++ chains of glycoconjugates is a sign of low differential degree of the glandular cells. In more differentiated cells concealment in oligosaccharide chains of D-galactose remnants (peanut and castor-oil lectins receptors) by L-fucose, N-acetil-D-glucosamin remnants and sialic acid can have place; this is demonstrated as accumulation of receptors to wheat germ and Laburnum anagyroides lectins in the glandular cells.     

The histochemical profiles of fibre types in porcine skeletal muscle

Using a variety of histochemical methods -mATPase staining after alkaline and acid preincubations, NADH-TR and alpha-MGPDH- we have investigated the fibre types in porcine skeletal muscle. The results reveal that four major fibre types -I, IIA, IIB and II*- can be separated histochemically in Longissimus lumborum muscle of Landrace pigs. The histochemical properties of the muscle fibre type II* are very similar to that of type IIX described in other mammals. The existence of IIX fibres in pig muscle has been recently demonstrated by molecular biology techniques and our results validate the use of histochemistry (mATPase) as an easy methodology to differentiate the three fast myosins (type II fibres) in pig muscle.     

Simultaneous histochemical demonstration of capillaries and muscle fibre types

Immunohistochemical staining of fibronectin is proposed as a good method for demonstrating capillaries in skeletal muscle tissue. The reaction superimposed on the histochemical reaction for myofibrillar ATPase enables simultaneous demonstration of fibre typing and capillary supply on the same tissue section. The method is quick and makes possible automated image analysis.     

A modified histochemical technique for sarcoplasmic reticular ATPase

Summary A calcium precipitation technique is described for the histochemical demonstration of the activity of sarcoplasmic reticular ATPase (SR ATPase) in skeletal muscle at pH 7.2–7.4. The specificity of this technique is discussed with reference to the problems involved in previous methods.     

Ultrastructural and histochemical comparison in two haplotaxids

Pelodrilus leruthi Hrabe, 1958 and Haplotaxis gordioides Hartman, 1821 have been investigated from a histochemical point of view. A different distribution and number of fast and slow fibres has been detected in the two species. Pelodrilus leruthi specimens living in different caves, show some differences in the body wall muscle thickness and in the structure of the ventral nerve cord. A correlation with their different life styles is proposed.     

A comparison of the molar efficiency of two insect-eating mammals

The extent to which the different molar morphologies of two similar-sized mammals, Petaurus breviceps (sugar glider) and Dasyuroides byrnei (kowari), could affect the digestion of larval insect nitrogen was investigated. Nitrogen digestibility of Tenebrio larvae treated with a proteolytic enzyme (trypsin) increased with decreasing larval piece size. However, it was found that the molars of P. breviceps , with their minimal shearing capacity compared with D. byrnei , can only compress rather than finely comminute Tenebrio larvae. As a result, P. breviceps have less efficient nitrogen digestibility than D. byrnei whose shearing molars can finely comminute larvae. Petaurus breviceps molars are inefficient at breaking down insects possibly because of functional constraints, where the molars must perform several different tasks with competing morphological demands, or because of phylogenetic constraints.     

A new method for myofibrillar Ca++-ATPase reaction based on the use of metachromatic dyes: its advantages in muscle fibre typing

A new method for Ca++-ATPase reaction in human muscle fibres is presented as an alternative to previous ATPase stains. The method is based on the use of metachromatic dyes, namely Azure A and Toluidine Blue, and has the advantages of speed, ease of performance and production of an elegant and clearcut fibre typing. The method distinguishes fibre types because of their metachromatic or orthochromatic staining, due to their different content of phosphate after incubation in the reaction medium. The comparison of serial sections stained by cationic dyes and by ammonium sulphide revealed close correspondence of fibre typing. Fibre type differentiation was also obtained with Acridine Orange; however this method was less reproducible.     

The value of enzyme histochemical techniques in classifying fibre types of human skeletal muscle

Summary Fibre-type classification of human skeletal muscle into type I and type II fibres is mostly based on their slight or strong staining with the myosin adenosine triphosphatase reaction. In order to evaluate the reliability of this screening technique a combined histochemical and biochemical study was performed on normal and diseased skeletal muscle of human subjects. In the present investigation activities of enzymes which play a role in the aerobic and anaerobic pathways and which can characterize fibre type, were examined in muscle specimens, with no apparent disease of the neuromuscular system. Special attention is given to the maximal activities of phosphofructokinase and fructose-1,6-diphosphatase, the rate limiting enzymes for the regulation of the glycolysis and glyconeogenesis, respectively. A most important feature of the biochemical findings is the constancy of the activity ratios of the examined enzymes. From these results and from the histochemical results it can be concluded that in apparently normal adult human skeletal muscle the ATP-ase technique for type I and type II typing is reliable. For fibres with an intermediate intensity of staining with the myosin ATPase technique of typing it is also necessary to apply other enzyme histochemical techniques.