Richard Grace

Studies of mitochondrial development during embryogenesis in the rat

Abstract Marked and progressive increases in the activities of DPNH oxidase, cytochrome c oxidase, succinic dehydrogenase, and mitochondrial ATP-ase were shown to occur between days 10 and 14 of gestation in the rat embryo-fetus. The rise in enzyme activity was paralleled by striking changes in the number and structure of the cristae of the heart mitochondria of the embryo-fetus as demonstrated by electron microscopy.

Studies of mitochondrial energy systems during embryogenesis in the rat

Abstract Mitochondria isolated from rat embryos of Day 11 to Day 14 of gestation showed an equal capacity to carry out oxidative phosphorylation with P:O ratios of approximately 3 for pyruvate-malate and 2 for succinate as substrates. Activities of succinic dehydrogenase, cytochrome oxidase, and mitochondrial ATPase remained at nearly constant levels in the preparations during this period of gestation, but DPNH oxidase activity increased over 2-fold. The increase in DPNH oxidase activity could be prematurely induced at Day 11 of gestation of the embryos by prior treatment of the pregnant rats with an atmosphere containing 85% oxygen and 0.4% CO2, but no changes in the growth and development of the treated embryos were found by gross examination.

Iron deficiency in the rat: effects on oxidative metabolism in distinct types of skeletal muscle.

Summary: Studies performed on iron-deficient and control rats demonstrated that oxidative energy production (phosphorylation) by mitochondria from iron-deficient red and intermediate skeletal muscles was greatly reduced with pyruvate-malate, succinate, and a-glycerophosphate as substrates. Although phosphorylation was also decreased in iron-deficient white skeletal muscle with succinate and pyruvate-malate as substrates, no change was found with a-glycerophosphate as substrate.

Maternal barbiturate administration and offspring response to shock.

Gravid Sprague-Dawley-derived rats were injected SC twice daily with either 20 or 40 mg/kg pentobarbital sodium (PT), sodium phenobarbital (PH), or the same volume of the saline vehicle on days 9–21 of pregnancy. Pair-feeding was employed. Vital, developmental, and activity measures were obtained on the neonates and locomotor activity was measured from 3–10 months of age. Avoidance was measured sequentially in a shuttle box, and in an operant chamber beginning at 3 months of age. The PH-80 dams gained less weight over the gestational period, and PH-80 and PH-40 offspring had more neonatal deaths. These male offspring were hyperactive at maturity, and PH-80 rats were initially slower to escape experimenter-initiated shock. PT exposure caused transient neonatal and juvenile hyperactivity. PT rats performed more poorly on both the conditioned avoidance and Sidman shock schedules, and had significantly lower brain: body weight ratios at 1 year of age. All four drug groups outperformed the saline offspring on subject-initiated shock schedules (punishment). Sex of offspring was determined on postnatal day 4 and the sex ratio was shifted towards male births with both drugs relative to controls.

Pyridine nucleotide transhydrogenations in yeast

Though previously described as very low or absent in yeast, we find significant pyridine nucleotide transhydrogenation (NADPH + acetyl pyridine-NAD+----NADP+ + acetyl pyridine-NADH) activity in yeast extracts when assayed at pH 8-9, and describe here the subcellular distribution and separation of the various molecular forms contributing to the total activity in two yeast species. Gentle subcellular fractionation reveals transhydrogenase activity only in the cytosolic fraction of both Saccharomyces cerevisiae and Candida utilis while intact mitochondria and microsomes are without activity. On sucrose gradient centrifugation, this soluble cytosolic activity proves to be primarily in a high-molecular-weight (greater than 10(6)) band which has salmon-colored fluorescence on uv illumination. Sonication of the particulate subcellular fractions solubilizes substantial transhydrogenase activity from mitochondria of C. utilis (but not from S. cerevisiae) which on sucrose gradients consists of both high (greater than 10(6))- and low-molecular-weight active fractions, each with yellow-green fluorescence. Ammonium sulfate fractionation and sucrose gradient centrifugation of protein solubilized from whole yeast of both species by vigorous homogenization with glass beads confirms the presence and fluorescence of these various molecular weight forms. The relationship of these activities to other enzymatic activities (especially the mitochondrial external NADH dehydrogenase) is discussed.

Iron deficiency in the rat: effects on energy metabolism in brown adipose tissue.

ABSTRACT.: Mitochondria were prepared from the brown adipose tissue of control rats and animals made iron deficient by means of a low iron diet. The specific activities of the mitochondrial electron transport system (NADH, succinate and α-glycerophosphate oxidase activities) were markedly and significantly reduced in preparations of brown adipose tissue from the iron-deficient rats as compared with preparations from the control animals. In contrast, concentrations of the cytochrome pigments a + a3, and c + c1 were normal and cytochrome b was slightly reduced (18%) in the mitochondrial preparations from the iron-deficient animals. Treatment of the iron-deficient animals with triiodothyronine significantly increased the amount of brown fat present per kilogram of body weight in both control and iron-deficient rats, but did not significantly affect the specific activities of the mitochondrial electron transport system.

Cytochrome a3 deficiency in human achondroplasia

Mitochondria prepared from tissue culture cells (skin fibroblasts) from normal subjects and subjects with homozygous achondroplasia were studied to determine the concentrations of cytochromes a and a3 in the preparations. Cytochrome a3 was markedly decreased (80%) in the achondroplastic preparations with cytochrome a present in normal amounts. Determination of total heme a (as the pyridine hemochromogen) in the normal and achondroplastic preparations demonstrated that the observed decrease in concentration of cytochrome a3 in the achondroplastic preparations was due to an absence of cytochrome a3 and not to a change in its absorbancy (extinction coefficient). The decreased concentrations of cytochrome a3 in the achondroplastic cells may decrease the reactivity or affinity of the mitochondrial oxidative systems for oxygen and result in the phenotypic expression of the disease.