Determination of myoglobin concentration and oxidative capacity in cryostat sections of human and rat skeletal muscle fibres and rat cardiomyocytes

Abstract

Myoglobin plays various roles in oxygen supply to muscle mitochondria. It is difficult, and in some cases impossible, to study the relationship between the myoglobin concentration and the oxidative capacity of individual muscle cells because myoglobin has to be fixed in situ whereas determination of oxidative capacity, for example, succinate dehydrogenase activity, requires unfixed cryostat sections. We have investigated whether a vapour-fixation technique allows the use of serial sections to study the relationship between myoglobin and succinate dehydrogenase activity. The technique is used to study a rat soleus muscle, two human skeletal muscle biopsies and biopsies of two patients with chronic heart failure, and in a control and hypertrophied rat heart. Staining intensities were quantified by microdensitometry. The absorbance values were calibrated using sections cut from gelatine blocks containing known amounts of myoglobin. The results show that it is possible to use serial sections for the determination of the myoglobin concentration and succinate dehydrogenase activity, and indicate that myoglobin can lead to a substantial reduction (18-60%) of the extracellular oxygen tension required to prevent an anoxic core in muscle cells.

Critical determinants of combined sprint and endurance performance: an integrative analysis from muscle fiber to the human body

“…The [Mb] was determined by calibrated histochemistry (52,56). Absorbance was measured at 436 nm (52).…”

Estimated contribution of hemoglobin and myoglobin to near infrared spectroscopy

“…In addition, the concentrations and LAPs of Mb were based on an average value of 0.5 mM (van Beek-Harmsen, 2004), which is known to vary according to gender, muscle, fiber type and species.…”

Immunohistochemical Determination of Cytosolic Cytochrome c Concentration in Cardiomyocytes

“…For other small proteins, parvalbumin, and green fluo-van Beek-Harmsen, van der Laarse rescent protein, this can be prevented through the use of vapor fixation (Füchtbauer et al 1991;Jockusch et al 2003). We have recently developed a vapor fixation technique to determine the myoglobin concentration in individual muscle cells in cryostat sections (van Beek-Harmsen et al 2004). In this report, we show that vapor fixation in combination with immunohistochemistry can be used to determine the cytosolic cytochrome c concentration in individual cardiomyocytes.…”