ATP-DIPHOSPHOHYDROLASE ACTIVITY IN RAT RENAL MICROVILLAR MEMBRANES AND VASCULAR TISSUE

Ecto-nucleotidases may have a role in the regulation of purinoceptor-m ediated responses. ATP-diphosphohydrolase or apyrase has been describe d as an ecto-nucleotidase, which is characterized by a low specificity for its substrates and bivalent cations, The aim of this work was to demonstrate the presence of apyrase as an ecto-enzyme in the rat kidne y, ATPase-ADPase activities of the renal microvillar membrane preparat ion, which correspond to 'right side out' membranes, were characterize d, The detection of ATP-diphosphohydrolase in the renal vasculature wa s done through perfusion of isolated rat kidney, ATPase-ADPase activit ies of the microvillar membrane preparation and apyrase share similar kinetic properties, These include: low substrate and bivalent metal sp ecificities and insensitivity towards inhibitors like: oligomycin, oua bain, verapamil, levamisole and Ap(5)A. The M(r) of native ATPase and ADPase activities,vas determined by the Co-60 irradiation-inactivation technique being around 65 kDa for both hydrolytic activities, Immunow estern blot analysis also supports the presence of apyrase in microvil li, Perfusion of isolated rat kidney with ATP and ADP, in the presence or absence of different inhibitors or apyrase antibodies indicated th e existence of this enzyme in the vascular endothelium. The identifica tion of ATP-diphosphohydrolase as an ecto-enzyme both in microvilli an d vasculature support the proposal that the enzyme may have an importa nt role in the extracellular metabolism of nucleotides. Copyright (C) 1996 Elsevier Science Ltd.