Physiological effects of micropauses in isometric handgrip exercise

The physiological response to continuous and intermittent handgrip exercise was evaluated. Three experiments were performed until exhaustion at 25% of maximal voluntary contraction (MVC): experiment 1, continuous handgrip (CH) (n = 8); experiment 2, intermittent handgrip with 10-s rest pause every 3 min (IH) (n = 8); and experiment 3, as IH but with electrical stimulation (ES) of the forearm extensors in the pauses (IHES) (n = 4). Before, during, and after exercise, recordings were made of heart rate (HR), arterial blood pressure (BP), exercising forearm blood flow, and concentrations of potassium [K+] and lactate [La-] in venous blood from both arms. The electromyogram (EMG) of the exercising forearm extensors and perceived exertion were monitored during exercise. Before and up to 24 h after exercise, observations were made of MVC, of force response to electrical stimulation and of the EMG response to a 10-s test contraction (handgrip) at 25% of the initial MVC. Maximal endurance time (tlim) was significantly longer in IH (23.1 min) than in CH (16.2 min). The ES had no significant effect on tlim. During exercise, no significant differences were seen between CH and IH in blood flow, venous [K+] and [La-], or EMG response. The HR and BP increased at the same rate in CH and IH but, because of the longer duration of IH, the levels at exhaustion were higher in this protocol. The subjects reported less subjective fatigue in IH. During recovery, return to normal MVC was slower after CH (24 h) than after IH (4 h).(ABSTRACT TRUNCATED AT 250 WORDS)     

Biotin carboxylases in mitochondria and the cytosol from skeletal and cardiac muscle as detected by avidin binding

Biotin carboxylases in mammalian cells are regulatory enzymes in lipogenesis and gluconeogenesis. In this study, endogenous biotin in skeletal and cardiac muscle was detected using avidin conjugated with alkaline phosphatase and applied in high concentrations to muscle sections. The avidin binding was subsequently visualized by histochemical demonstration of the alkaline phosphatase activity. All cardiac muscle cells showed high affinity for avidin with only the nuclei and the intercalated discs remaining unstained. In skeletal muscle a diffuse reaction could be detected in the sarcoplasm of the muscle fibres. A granular reaction was noted in the same fibres that showed activity for succinic dehydrogenase. The specificity of the coloured reaction product in the muscle sections was investigated and is suggested to be caused by avidin binding to biotin moieties in mitochondria and the cytosol. Mitochondrial and cytosolic preparations of skeletal muscle were electrophoresed in sodium dodecyl sulphate gels. After blotting and incubation with conjugated avidin, two bands with molecular weights of 75 kDa and 130 kDa respectively were evident in the mitochondrial preparation. It is suggested that the 75-kDa band represents comigration of the biotin-containing subunits of propionyl-CoA carboxylase and methylcrotonyl-CoA carboxylase. The 130-kDa band may represent the biotin-containing pyruvate carboxylase. In the cytosolic preparation a 270-kDa band was stained in blots that had been incubated with conjugated avidin; this band is suggested to represent acetyl-CoA carboxylase. A 190-kDa cytosolic band might be a cleavage product of acetyl-CoA carboxylase. We propose that using alkaline phosphatase-conjugated avidin it is possible to detect the mitochondrial and cytosolic biotin-dependent carboxylases in striated muscle.     

The effect of sugars on (pro)insulin biosynthesis

Rates of incorporation of [4,5-(3)H]leucine into insulin plus proinsulin, designated ;(pro)insulin', and total protein in rat pancreatic islets were measured. Glucose stimulates rates of total protein and (pro)insulin biosynthesis, but (pro)insulin biosynthesis is stimulated preferentially. Mannose and N-acetylglucosamine also stimulate (pro)insulin and total protein biosynthesis; inosine and dihydroxyacetone stimulate (pro)insulin biosynthesis specifically. Fructose does not stimulate (pro)insulin biosynthesis when tested alone, but does so in the presence of low concentrations of glucose, mannose or N-acetylglucosamine. Many glucose analogues do not stimulate (pro)insulin biosynthesis. Mannoheptulose inhibits synthesis of (pro)insulin and total protein stimulated by glucose or mannose but not by dihydroxyacetone, inosine or N-acetylglucosamine; phloretin (9mum) inhibits N-acetylglucosamine-stimulated (pro)insulin biosynthesis preferentially. The data are in agreement with the view that the same glucose-sensor mechanism may control both insulin release and biosynthesis, and ;substrate-site' model is suggested. The threshold for stimulation of biosynthesis of (pro)insulin and total protein is lower than that found for glucose-stimulated insulin release; moreover the biosynthetic response to an elevation of glucose concentration is slower than that found for insulin release. The physiological implication of these findings is discussed. Caffeine and isobutylmethylxanthine, at concentrations known to increase islet 3':5'-cyclic AMP and potentiate glucose-induced insulin release, were without effect on rates of glucose-stimulated (pro)insulin biosynthesis.     

Vibratome Sections Of Difficult Tissues

After 1 hour of aldehyde fixation, 4 to 20 hours of soaking in 2% BSA (bovine serum albumin) solution and another 17 hours in the same fixative, the vibratome will produce smooth, even sections from agar embedded guinea pig thyroid, skeletal muscle or larynx with the same ease as it will from soft, homogeneous tissues such as liver or spleen.     

Methods for demonstration of enzyme activity in muscle fibres at the muscle/bone interface in demineralized tissue

Summary A method for demonstration of activity for ATPase and various oxidative enzymes (succinic dehydrogenase, -glycerophosphate dehydrogenase, and lactic dehydrogenase) in muscle/bone sections of fixed and demineralized tissue has been developed. It was found that it is possible to preserve considerable amounts of the above mentioned enzymes in the muscle fibres at the muscle/bone interfaces. The best results were obtained after 20 min fixation, and 2–3 weeks of storage in MgNa₂EDTA containing media. As the same technique previously has been used to describe patterns of resorption and deposition with the aid of a mapping of presence of phosphomonoesterases on bone surfaces, the method may be used to study possible biochemical interactions between bone and muscle tissue at the muscle/bone interface.     

A newborn child with karyotype 47,XX,+der(12) (12pter→12q12::8q24→8qter),t(8;12) (q24;q12) pat

Summary A case of Meckel or Gruber syndrome is reported, together with a survey of the relevant literature of recent years (1971–1977), in reference to a probably autosomal recessive inheritance of this malformation.     

Coumarin glycosides of seseli montanum

Three new coumarin glycosides isolated from roots of Seseli montanum were shown to be the 3′-O-,2′-O-, and the 7-O-β-D-glucopyranosides respectively of 2′(R)-6-(2′,3′-dihydroxy-3′-methylbutyl)-7-hydroxy-coumarin. The structures were elucidated by spectroscopic and chemical methods. (R)-Configuration was assigned to the aglycone, also known as ( + )-peucedanol, and to its 7-methyl ether, ( + )-ulopterol, by chemical correlation. Additionally, apterin was obtained and characterized.     

Lectin binding in skeletal muscle. Evaluation of alkaline phosphatase conjugated avidin staining procedures

Summary Cryostat sections from rat gracilis muscles were incubated with different biotinylated lectins: Con A (Concanavilin A), WGA (Wheat germ agglutinin), SBA (soybean agglutinin), GS I and GS II (Griffonia simplicifolia agglutinin), LCA (Lens culinaris agglutinin), PNA (peanut agglutinin) and PSA (Pisum sativum agglutinin). The sections were subsequently treated with alkaline phosphatase conjugated avidin. The lectin binding sites were visualized after incubation in substrate media containing: (1) 5-bromo-4-chloro indoxyl phosphate and Nitro Blue tetrazolium or copper sulphate; (2) naphthol AS-MX phosphate or naphthol AS-BI phosphate and various types of diazonium salts; (3) -naphthylphosphate and Fast Blue BB; (4) -glycerophosphate according to the method of Gomori. The results obtained with the alkaline phosphatase methods were compared with those seen with a streptavidin-horseradish peroxidase procedure. Several chromogen protocols for visualizing alkaline phosphatase activity showed differences in the ability to detect lectin binding sites. A sarcoplasmic reaction was evident for Con A, GS II, WGA, LCA, and PSA after incubation in the indoxyl phosphate medium. Sarcoplasmic reaction for GS II was also noticed after incubation with naphthol AS-MX Fast Blue BB and -glycerophosphate. The latter substrate also gave rise to a sarcoplasmic Con A reaction. With the indoxylphosphate tetrazolium salt method some muscle fibres showed a very strong intracellular reaction after incubation with Con A and GS II while the staining intensity was weak in other fibres. The same muscle fibres were stained with PAS. No sarcoplasmic reactions were observed with either naphthol phosphate media or with the diaminobenzidine peroxidase methods. Further, the staining of the muscle fibre periphery, connective tissue, and capillaries was intensified using the indoxyl method. The indoxylphosphate-tetrazolium salt method seems to be suitable for future investigations of lectin binding sites in muscle sections.     

SH groups in the α-naphthyl acetate esterase in the thyroid of the guinea-pig

Summary The -naphthyl acetate esterase in both group I and group II thyroid cells is shown to contain SH groups since there is a decline in activity in both cell groups when certain sulfhydryl reagents [DTNB; 5,5-Dithiobis-(2-nitrobenzoic acid)-AgNO₃-Mersalyl-PCMB (parachloro mercuribenzoate)+urea] are added to the incubation media. Thus the inhibition is by far the greatest in group I cells, which also show the greatest activity after incubation in conventional media, when long fixation and storage times are used. In all cases the inhibiting effect was complete or almost completely reversed if cysteine was added to the incubation media in equivalent concentrations to the SH blocker. There were great differences among the sulfhydryl reagents used in their ability to bring about enzyme inhibition. The alkylating agents NEM (N-ethylmaleimide) and iodoacetamide had no or little effect while PCMB could only inhibit the activity of the -naphthylacetate esterase if the enzyme was denaturated with 5 m urea. The maximal inhibitory effect of PCMB was only obtained when NaCl was added to the incubation media. The most effective inhibitor was AgNO₃.