Method for isolating single cardiac cells in the guinea pig using pronase. The role of calcium ions

A method for isolation of guinea-pig cardiomyocytes with pronase has been developed. The method has been assessed in hearts perfused with solutions containing pronase (1 U/ml) and 200 microM Ca2+. Eighty per cent of the cells released were rod-shaped and 1.2 mM Ca2+ tolerant. Enriched medium 199 was used for all solutions. Sodium and slow inward currents recorded from cells dispersed with pronase were similar to those recorded from cells isolated after prolonged exposure to collagenase. Two principal factors are to be marked: (a) presence of high enough amounts of Ca2+ in enzyme solution (up to 200 microM); (b) use of the enriched medium in all the stages of the procedure.     

Characteristics of adenosine activity on adenine nucleotide metabolism of rat thymocytes

The effects of adenosine on adenine nucleotide metabolism in [14C]adenine-labeled rat thymocytes were studied. It was shown that adenosine increases the intracellular pool of adenine nucleotides, predominantly ATP, which is accompanied by marked acceleration of their catabolism and a release of labeled products (especially inosine, hypoxanthine and adenosine) from the thymocytes. The effect of adenosine depends on its concentration and manifests itself already at 10(-6) M. 2-Deoxycoformycin partly relieves the effect of adenosine on adenine nucleotide metabolism. Exogenous deoxyadenosine, inosine, hypoxanthine and adenine, unlike adenosine, do not significantly affect the adenine nucleotide catabolism and the label release from the cells. All the effectors under study strongly increase inosine transport from the thymocytes, and inhibit, with the exception of adenosine, the hypoxanthine release from the cells.     

Different secretory pathways of renin from mouse cells transfected with the human renin gene

Mammalian cells in culture, transfected with human renin gene, can provide a useful tool for studying renin biosynthesis and secretion. We transfected fibroblast cells (mouse L929 and Chinese hamster ovary cells) and pituitary tumor cells (mouse AtT-20) with the human renin gene and a selectable plasmid (pSV2Neo). Transfected fibroblasts synthesize prorenin only. Prorenin is secreted by fibroblasts constitutively and the secretion is not influenced by 8-bromo-cAMP. On the other hand, transfected AtT-20 cells synthesized both prorenin and mature active renin. Transfected AtT-20 cells release prorenin by constitutive secretion but mature renin is secreted by a regulated mechanism since the secretion of the former is not influenced by 8-bromo-cAMP but the release of the latter is significantly stimulated. Our studies demonstrate that human renin may be secreted by at least two cellular pathways: prorenin by a constitutive pathway and mature renin by a regulated pathway. These transfected cells may provide useful models for studies of human renin synthesis, processing, and secretion.     

Measurement of cerebral blood flow by transcranial Doppler ultrasonography in children presenting with autistic behavior. Preliminary results

Transcranial Doppler ultrasonographic recordings of the middle cerebral arteries were performed on eight children with autistic behavior compared to eight controls. Blood flow measurements were assessed at rest and during auditory and visual stimulations. The main result was obtained during the auditory stimulations and concerned the left artery blood flow which is lower in autistics than in controls in these conditions (p less than .02). This result confirms the possibility of a left hemisphere dysfunctioning in autistics and may be related to clinical features as language disabilities and paradoxical reactivity to auditory stimuli.     

Characterization of the specific binding of rat apolipoprotein E-deficient HDL to rat hepatic plasma membranes

We have used a preparation of rat liver plasma membranes to study the binding of rat apolipoprotein E-deficient HDL to rat liver. The membranes were found to bind HDL by a saturable process that was competed for by excess unlabeled HDL. The binding was temperature-dependent and was 85% receptor-mediated when incubated at 4, 22 and 37 degrees C. The affinity of the binding site for the HDL was consistent at all temperatures, while the maximum binding capacity increased at higher temperatures. The specific binding of HDL to the membranes did not require calcium and was independent of the concentration of NaCl in the media. The effect of varying the pH of the media on HDL binding was small, being 30% higher at pH 6.5 than at pH 9.0. Both rat HDL and human HDL3 were found to compete for the binding of rat HDL to the membranes, whereas rat VLDL remnants and human LDL did not compete. At 4 degrees C, complexes of dimyristoylphosphatidylcholine (DMPC) and apolipoproteins A-I, A-IV and the C apolipoproteins, but not apolipoprotein E, competed for HDL binding to the membranes. At 22 and 37 degrees C, all DMPC-apolipoprotein complexes competed to a similar extent, DMPC vesicles that contained no protein did not compete for the binding of HDL. These results suggest that the rat liver possesses a specific receptor for apolipoprotein E-deficient HDL that recognizes apolipoproteins A-I, A-IV and the C apolipoproteins as ligands.