published_or_final_version / Medicine / Master / Doctor of Medicine

Diss. (sammanfattning) Stockholm : Karol. inst., 2006. / Härtill 5 uppsatser.

Thesis (M.D.)--University of Hong Kong, 1993.

Thesis (M.D.)--University of Hong Kong, 1993. / Also available in print.

Proefschrift Rijksuniversiteit Groningen. / Datum laatste controle: 15-05-1997. Met bibliogr., lit. opg. - Met een samenvatting in het Nederlands.

Summary: Ventricular cells from adult rats were isolated enzymatically and used as a model system for determining what factors affect the release of adenosine triphosphate (ATP) from myocardial cells. The enzyme systems used to isolate cells were trypsin:collagenase; hya1uronidase:collagenase and dispase:collagenase. Adenosine triphosphate was released in greater amounts in response to hypoxia from cells freed by each of the enzymatic procedures. This occurred while the intracellular concentration of ATP remained constant. Experiments were then performed to determine whether the conditions that occur during myocardial ischaemia or hypoxia altered the release of ATP. Cells suspended in either oxygenated or anoxic buffer at a pH of 6.8 released a significantly lower amount of ATP than cells suspended in either condition at pH 7.4. To test the possibility that ATP was released from nucleotide-protein-Ca2+ complexes located in the sarcolemma, artificial disruption of these structures was carried out. Incubation of oxygenated cells with the chelating agent, ethyleneglycol-bis (B-aminoethyl ether)-N, N-tetraacetic acid (EGTA), stimulated the release of ATP in a hyperbolic relationship while incubation of anoxic cells with ethylenediamine tetraacetate (EDTA) stimulated the release of ATP in such a way that the pattern of release followed a sigmoid response with maximal amounts of ATP, 995±55 pmol·mg-2 protein, occurring in the presence of 0.1 to 2.0 mmol·litre-2 EDTA. By incubating cells with radioactive EDTA, there was no indication that EDTA entered the cells. No release of ATP above control levels occurred when EDTA was chelated with Ca2+ before being applied to isolated cells. These data suggest that the source of ATP found extracellularly may have been nucleotide-protein-Ca2+ complexes located in the sarcolemma, and further support the role of ATP as a coronary vasodilator during hypoxic conditions.

Thesis (Geaggregeerde voor het Hoger Onderwijs)--Catholic University of Leuven, 1988. / Includes bibliographical references (p. 118-129).

Thesis (Geaggregeerde voor het Hoger Onderwijs)--Catholic University of Leuven, 1988. / Includes bibliographical references (p. 118-129).

The metabolism of 5'-AMP by 5'-nucleotidase, adenylate deaminase and adenylate kinase was examined in heart homogenates of rat, rabbit, dog, pigeon and turtle. The study was conducted in consideration of the possibility that adenosine, a catabolic product of 5'-AMP, may control vasotone for the autoregulation of coronary blood flow. The relative activities of homogenates of hearts from various species to form adenosine by the action of 5'-nucleotidase generally supported such a role for this nucleoside. Those species anticipated to have the largest potential requirements for coronary vasodilation, i.e. those whose oxygen consumption is known to increase significantly during physical exertion, had the highest levels of cardiac 5'-nucleotidase. An exception to this was the pigeon which had no detectable cardiac 5'-nucleotidase; the order of levels of this enzyme in hearts of the other species tested was: rat > dog > rabbit > turtle. The turtle ventricle, by virtue of its high content of adenylate deaminase and low content of 5'-nucleotidase appeared to catabolize 5'-AMP largely by deamination to IMP. Homogenates of pigeon ventricle contained the greatest activity of adenylate kinase, indicating that the heart of this species is equipped for preservation of ATP by resynthesis from ADP. Enzyme histochemistry revealed that most 5'-nucleotidase of mammalian hearts was localized in the endothelial cells of capillaries. Therefore, if adenosine is involved in regulation of coronary perfusion, its source may be capillary endothelial cells rather than cardiac muscle cells. 5'-Nucleotidase was partially purified from an acetone powder of rat heart. It was active over a broad range of pH with an optimum at pH 8.5. The enzyme was stimulated up to 5-fold by Mg(++) [formula omitted]; Mn(++) and Ni(++) also stimulated activity. The K for 5'-AMP was 2.1 x 10(-5)M in the absence of 16Mg and 2.3 x 10 M in the presence of 16 mM MgCl(2). Certain of its properties indicated that the production of adenosine might be favoured under conditions in which coronary vasodilation would be required and vice-versa. For example, the enzyme was inhibited by ATP, whose levels are greatest in well oxygenated hearts in which energy charge is high. Not all properties of 5'-nucleotidase were consistent with enhanced adenosine formation at reduced energy charge. Both ADP and orthophosphate, the levels of which increase when energy charge decreases, inhibited the enzyme; in fact ADP was a more powerful inhibitor than ATP. In addition, the enzyme was not specific for 5'-AMP but hydrolyzed a variety of nucleoside 5'-monophosphates; and the hydrolysis of 5'-AMP was competitively inhibited by UMP. In the absence of Mg(++) , inhibition by ADP was of the mixed (competitive- non-competitive) type. In the presence of 16 mM MgCl(2), inhibition was non-competitive. On the basis of these data and Dixon plots of inhibition as a function of ADP concentration, it is suggested that two conformations of the enzyme are possible; one which is competitively inhibited by ADP. The simple non-competitive inhibition by ADP, observed in the presence of 16 mM MgCl(2), is attributed to Mg(++) -induced preference for the latter conformation. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

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