Comparative subcellular fractionation of control and cold-adapted rat brown and white adipose tissue with special reference to peroxisomal and mitochondrial distributions.

A new technique for single-step subcellular fractionation of adipose tissue homogenates by analytical sucrose density gradient centrifugation in a vertical pocket reorientating rotor is described. The density gradient distributions of mitochondrial and peroxisomal marker enzymes in brown and white adipose tissue of control and cold exposed rats are compared. The equilibrium density of brown fat mitochondria was found to be significantly increased compared with white fat mitochondria. GDP binding activity was localized solely to the mitochondria in both control and cold-adapted brown adipose tissue. Brown and white fat mitochondria fractions were isolated by differential centrifugation and the specific activities of various enzymes in the homogenate and mitochondrial preparations determined. The specific activity of creatine kinase in brown adipose tissue was found to be ten-fold higher than in white fat and subcellular fractionation studies showed the activity to have an exclusively cytosolic distribution in both tissues. GDP binding activity and some of the mitochondrial enzymes showed, in brown adipose, a striking increase in total activity in cold adapted rats compared to control animals. For some enzyme activities there was a small increase when expressed per mg tissue or per mg mitochondrial protein. When expressed per mg DNA i.e. per cell, there was a reduced specific activity of the mitochondrial and peroxisomal enzymes in both brown and white adipose tissue on cold adaptation.

Comparison of effects of inhibitors on adenosine triphosphatase and adenosine diphosphatase activities in rat-liver mitochondria.

Adenosine diphosphatase (ADPase) activity and ATPase activity were assayed in rat liver mitochondria and outer mitochondrial membrane preparations with [beta-32P]ADP and [gamma-32P]ATP as substrates. Inhibition studies were performed with the mitochondrial ATPase inhibitor oligomycin and the adenine nucleotide transport inhibitor, carboxyatractyloside. Kinetic studies were also performed with the nucleotide thiophosphate analogs adenosine 5'-O-thiophosphate, adenosine 5'-O-(2-thiodiphosphate) and adenosine 5'-O-(3-thiotriphosphate) which can act as inhibitors of phosphohydrolases. It is concluded that part of the apparent ADPase activity of intact mitochondria is mediated via ATPase, presumably in conjunction with adenylate kinase. In addition the outer mitochondrial membrane appears to show a distinct ADPase not attributable to contamination by inner membrane ATPase.

Studies on the nature of adenosine diphosphatase activity from rat liver mitochondria.

Adenosine diphosphatase (ADPase) activity was studied in rat liver with [beta-32P]ADP as a substrate. Mitochondria and outer mitochondrial membrane fractions were isolated and assayed for ADPase and various marker enzymes. ADPase activity was strikingly reduced when the outer membranes were removed from the mitochondria whether by digitonin treatment or osmotic shock. Addition of the inter-membrane space subfraction to the purified outer membranes resulted in enhanced ADPase activity. Addition of the inter-mitochondrial membrane enzyme adenylate kinase to outer membranes also produced a large stimulation of activity. The ADPase activity could also be reconstituted in vitro with adenylate kinase and either mitoplast ATPase or ouabain-sensitive (Na+ + K+ + Mg2+)-ATPase. Chloroform-released ATPase, however, was not capable of producing an ADPase activity when combined with adenylate kinase. Gel permeation chromatography of Triton-solubilised outer mitochondrial membranes was unable to resolve ADPase activity from contaminating ATPase. These results suggest that the majority of ADPase activity in rat liver mitochondria consists of the coupled activity of adenylate kinase and ATPase.

Tissue distribution of adenosine diphosphatase activity in the rat.

The specific activity of adenosine diphosphatase (ADPase) was determined in rat tissue homogenates using a specific radioassay. 14 different tissues were sampled and activity was found in most homogenates examined. The levels ranged from very little activity in the pancreas to high levels in the duodenum and ileum. High levels of ADPase activity found in the gastrointestinal tract could be partially attributed to non-specific alkaline phosphatase. It is concluded that ADPase activity is widely distributed in rat tissues.

Human figure drawing characteristics related to juvenile delinquents.

Human figure drawings were scored on seven characteristics popularly attributed to juvenile delinquents, i.e., head size, shading, and three indicators of emotional conflict, i.e., transparencies, omissions, and erasures. Ss were adolescents incarcerated at training schools. Delinquency was defined in terms of MMPI profiles associated with delinquency rather than by social-legal or medical-diagnostic classification. The results fail to support popular hypotheses concerning human figure drawings of delinquents. Delinquents drew significantly more transparencies; however, no other emotional indicator was significant. It was concluded that little support exists for hypotheses concerning human figure drawings of delinquents and drawings should be used with caution in the clinical assessment of a delinquent's personality.