The Oxidation of β-Hydroxybutyric Acid by Small Quantities of Type-pure Red and White Skeletal Muscle

R.A Pieter Kark,John P Blass,Joel Avigan,W King Engel

doi:10.1016/s0021-9258(18)62048-4

Abstract

Abstract Small samples of fiber type-pure red (type I) and pure white (typeII) skeletal muscle were obtained from guinea pigs. The oxidation of 14C-labeled d,l-β-hydroxybutyrate, pyruvate, succinate, and palmitate was measured in type-pure muscle slices of 2 to 8 mg fresh weight, and in a mitochondrial fraction prepared from 100- to 600-mg samples of such muscles. Slices from red muscle oxidized all four substrates more rapidly than did slices from white muscle, the differences being most striking for β-hydroxybutyrate (19-fold) and pyruvate (9-fold). Equal amounts of mitochondria from red muscle oxidized β-hydroxybutyrate 8 times and pyruvate twice as rapidly as mitochondrial fractions from white muscle, but there was no difference with respect to the oxidation of succinate nor of palmitate. d-(-)-β-hydroxybutyrate dehydrogenase activity could readily be detected in sonicated mitochondria from red muscle, but was usually not detectable in sonicated mitochondria from white muscle. Mitochondrial preparations from white muscle did not inhibit those from red. The differences between the rates of oxidation of β-hydroxybutyrate in slices of the two types of muscle is largely due to the differences in dehydrogenase activity in the respective mitochondria from the two sources.

Highlights

The oxidation of 14C-labeled D, L-P-hydroxybutyrate, pyruvate, succinate, and palmitate was measured in type-pure muscle slices of 2 to 8 mg fresh weight, and in a mitochondrial fraction prepared from lOO- to 600-mg samples of such muscles
Histochemical Methods-Samples of the fresh red muscle were mounted beside samples of fresh white muscle on a chuck, rapidly frozen in isopentane cooled to -160” by liquid nitrogen, and further prepared as a unit by methods previously described [18]
The white muscle was darkly and uniformly stained by the reaction for menadione-mediated a-glycerophosphate dehydrogenase [4]. It was virtually unstained by the reaction for P-hydroxybutyrate dehydrogenase

Summary

Methods

Preparationof Type-pureA1uscle-Male guinea pigs from t,he National Institutes of Health general purpose strain were decapitated. The light peach anterolateral layers of the vastus lateralis, dissected with gross color as the sole guide, provided histochemically type-pure white muscle. Sodium 2-i4C-pyruvate (3.44 mCi per mmole) was obtained from New England Nuclear. L-14C-Palmitic acid (55.1 mCi per mmole) (of greater than 99% radiopurity by gasliquid chromatography) was obtained from Amersham Searle. Radioactive and nonradioactive substrates were dissolved in Krebs-Ringer phosphate medium to buive solutions containing lo cpm per ml and final concentrations as indicated in the legend to Table I. Oxidation of TXaboled substrates by mitochondrial fraction from red and from white muscle of guinea pigs. Values are means f standard rrrors of four (P-hydroxybutyrate), six (pyruvate), four (succinate), and five (palmitate) experiments. Final concentrations of substrate were 1.5 MM fi-hydroxybutyrate, 2.2 PM pyruvate, 0.7 PM succinate, and 0.14 MM palmitate.

Results

Discussion

Conclusion