Brains fixed in paraformaldehyde or in Clarke's solution were blocked coronaily. Blocks from brains fixed in paraformaldehyde were either frozen in liquid nitrogen or embedded in paraffin. Tissue fixed in Clarke's solution was embedded in paraffin. Sections from each block were stained by the peroxidase-antiperoxidase method for adenosine deaminase or complexing protein using affinity-purified goat antibodies. Adenosine deaminase and complexing protein did not co-localize. Adenosine deaminase was detected in oligodendroglia and in endotheial cells lining blood vessels, whereas complexing protein was concentrated in neurons. The subcellular location and appearance of the peroxidase reaction product associated with individual cells was also quite distinctive. The cell bodies ofadenosine deaminase-positive oligodendroglia were filled with intense deposits ofperoxidase reaction product. In contrast to oligodendroglia, the reaction product as-If the regulatory actions of adenosine depend on binding to cell surface receptors, it seems reasonable that the concentration ofextracellular adenosine should also be subject to regulation. Wu and Phillis (1984) have suggested that uptake and metabolism of

The distribution of adenosine deaminase and adenosine deaminase complexing protein in rabbit heart has been compared using immunohistochemical staining procedures. Sections (4-5 ,um) of tissue fixed in Clarke's solution or paraformaldehyde and embedded in paraffin were stained by the peroxidase anti-peroxidase method for adenosine deaminase or complexing protein, using affinity purified antibodies. Staining for adenosine deaminase and complexing protein was observed in the central myocardium of all heart chambers. Adenosine deaminase was detected in endothelial cells of blood vessels and adjacent pericytes. The nuclei of arteries stained heavily for adenosine deaminase, whereas those of venules and small veins, although positive, stained much more lightly. The cytoplasm of blood vessel endothelial cells and smooth muscle cells of the tunica media were also weakly positive for adenosine deaminase. Endothelial cells of the endocardium and epicardium did not stain. Randomly distributed mononuclear inflammatory cells and interstitial connective tissue fibroblasts were also negative for adeno-sine deaminase. These results raise the possibility that endothelial cells containing adenosine deaminase could serve as a metabolic barrier preventing the free exchange of plasma and interstitial adenosine. Positive staining for complexing protein was restricted to blood vessel endothelial cells, especially cytoplasmic processes. Colocalization experiments carried out with biotinylated primary antibodies indicate that some vessels are positive forboth adenosine deaminase and complexing protein. This is the first experimental evidence of possible in situ association of adenosine deaminase and 1990;66:754-762) nhe vasoactive properties of adenosine have been recognized for over 50 years.1 It was the proposal by Berne in 1963,2 that adenosine may help to regulate coronary circulation, that seems to have most stimulated interest in the role played by this nucleoside in cardiac muscle. Since then, numerous studies have demonstrated that there is a correlation between adenosine production and the oxygen supply/demand ratio in cardiac muscle.34 According to the hypothesis proposed by Berne, adenosine is generated when the supply of oxygen to the myocar-dium does not meet the demand. The supply/demand ratio is brought back into balance by an increase in complexing protein. (Circulation Research coronary flow caused by adenosine. Although numerous studies provide strong support for the adenosine hypothesis, it is acknowledged that a cause and effect relation has not been established. Proving that adenosine participates in the regulation of coronary flow is made difficult by the complexity of its metabolism. The modes of production and degradation of adenosine both require additional clarification. A better understanding of the enzymes involved in these processes would also be helpful in determining if the adenosine hypothesis is correct. Advances in the characterization of adeno-sine deaminase serve as a good illustration. Most models o...

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