To answer some of the as yet unresolved questions about the formation, metabolism, and release of purine catabolites in hypoxic myocardium, we compared their release from isolated rabbit hearts during hypoperfusion, anoxia, and after ischemia, with and without nucleoside-transport inhibition. Results provide evidence to suggest the following. Besides the supply-to-demand ratio for O2, other factors may affect the formation of adenosine. The myocyte is the major source of purine catabolites. Adenosine is not produced within the myocyte. Once in the interstitium, adenosine is (largely) taken up in the endothelial cells, where it is catabolized to inosine and hypoxanthine and released that way into the lumen. Some of the adenosine can reach the lumen unchanged through clefts. Nucleoside transport inhibition prevents the escape through the endothelial cells and thus the formation and release of inosine and hypoxanthine. As a consequence, more adenosine will accumulate in the interstitium and more will reach the lumen through the clefts. The pathway, as proposed, explains the long known paradox of increased extracellular levels of adenosine after inhibition of nucleoside transport.

Yeast cells exposed to different doses of the antimycotic agent miconazole revealed important cytochemical changes in the topographic distribution of the phosphatases. A strong effect was observed on the behavior of oxidative and peroxidative enzymes. Decreased cytochrome c oxidase and peroxidase activity and increased catalase activity were seen after treatment with a fungistatic dose ofmiconazole, whereas a complete disappearance ofthese enzymes was observed after treatment with a minimal fungicidal dose of miconazole. This was in complete agreement with the quantitative biochemical data. A hypothesis is advanced concerning the possible involvement of peroxidase and catalase in the mechanism of action of this drug., is an imidazole nitrate that has demonstrated broad-spectrum activity against most pathogenic fungi and gram-positive bacteria (7,10,18,19,36). The studies on the uptake and utilization of substances in Candida albicans by Van den Bossche (37) indicated that miconazole induced changes in the permeability of the plasmalemma and the cell wall. At the ultrastructural level, C. albicans cells show changes in a dosedependent manner when grown in the presence of 10`8 to 10-M concentrations of miconazole. These changes range from slight alterations in the cell periphery and increase in the number of peroxisomes and lipid globules, to progressive cytoplasmic deterioration, prominent shape changes, and complete cellular necrosis (11, 12; S. De Nollin, Ph.D thesis, University of Brussels, Brussels, Belgium, 1976). Recent investigations with scanning and transmission electron microscopy (12) on C. albicans cultures treated with a fungicidal dose of miconazole (10-4 M) have shown that the sequence of alterations differs totally from that obtained with lower-dose treatments.The cytochemistry of untreated yeast cells has already been described by several authors (2, 3, 13, 22, 40; S. De Nollin, Ph.D. thesis, 1976). In earlier experiments (16, 13), we tried to optimize the preparatory conditions for the cytochemical examination of yeasts cells by using a modification of the conventional procedure for preservation. This yields, besides a good micromorphology of the cells, sufficient enzyme activities and allows an adequate penetration of substrates and captation ions during the incubation. For the biochemical studies, Van Belle, Goossens, and Van Roy (in preparation) have recently developed a method for obtaining adequate homogenization of the very rigid cells of yeast, thereby yielding reliable enzyme activities that are directly proportional to the number of cells (more than 99.9% of the cells are thoroughly homogenized). Using these modifications of the cytochemical and biochemical preparative techniques, we have tried in this study to evaluate the effects of miconazole on the activities and the distribution pattem of phosphohydrolases and of oxidative and peroxidative enzymes in C. albicans and Saccharomyces cerevisiae. Our particular aim was to see whether there was any correlation between the observed morph...

In this study evidence is provided to suggest that nucleoside formation with hypoxia in myocardial tissue from the guinea-pig follows a different course from that in the rat, rabbit or dog. 1) After ischemia, tissue levels of adenosine remain barely detectable in the guinea-pig but rise considerably in the rat and the dog. 2) IMP, remaining almost absent in the dog, does not change in the rat but strongly increases (X 6) in the guinea-pig heart with ischemia. 3) Mioflazine, a nucleoside transport inhibitor, completely reverses the ratio adenosine/inosine in dog myocardium after 8 min of ischemia, making adenosine by far the major nucleoside. No effect could be detected in the guinea-pig. 4) In contrast with the rat and rabbit, ischemia in the guinea-pig does not lead to any considerable release of adenosine upon reperfusion. 5) In the rabbit, the presence of a nucleoside transport inhibitor completely reverses the adenosine/inosine ratio in reperfusates after ischemia. Although the release is strongly inhibited under these conditions in the guinea-pig, adenosine release remains negligible when compared with inosine. 6) Even in the presence of high concentrations of an adenosine deaminase inhibitor, inosine remains the major metabolite released upon reperfusion after ischemia, in the guinea-pig heart.

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Van Belle, H., J. Wynants and F. Goossens: Nucleoside transport inhibition and the release of purine metabolites from hypoperfused isolated guinea pig hearts. Drug Dev. Res. 8:425-431, 1986. Hypoperfusion elicited a marked but transient release from isolated guinea pig hearts of inorganic phosphate, inosine, hypoxanthine, xanthine, and uridine, but had no effect on the release of uric acid, uracil, and adenosine. Lack of increased formation of adenosine, as well as a clearcut effect of hypoperfusion on uridine release, suggest that the purine catabolites did not originate from either cyclic AMP or from ATP released with catecholamines during ischemia. They merely point to a particular sensitivity of some cells to ischemia whereby the nucleotides are rapidly degraded and released into the interstitium.Inclusion at low concentrations (5 x lo-* M) of a nucleoside transport inhibitor, such as the water soluble analogue of mioflazine, drastically reduced the basal release of most of the metabolites and completely suppressed the effect of hypoperfusion. This effect is specifically ascribed to nucleoside-transport inhibition since no marked change was observed on the release of inorganic phosphate.