The effect of glibenclamide on the production of interstitial adenosine by inhibiting ecto-5'-nucleotidase in rat hearts.

Abstract

1. Adenosine exerts cardioprotective effects on the ischaemic myocardium. The production of adenosine in the ischaemic myocardium is attributed primarily to the enzymatic dephosphorylation of adenosine 5'-monophosphate (AMP) by 5'-nucleotidase. We determined the activity of 5'-nucleotidase in rat hearts. The objective of the study was to determine the effects of ATP-sensitive K+ (K[ATP]) channel antagonists (glibenclamide and 5-hydroxydecanoate) on the production of adenosine, by use of a flexibly mounted microdialysis technique. 2. Rats were anaesthetized and the microdialysis probe was implanted in the left ventricular myocardium, followed by perfusion with Tyrode solution. The baseline level of dialysate adenosine was 0.51 +/- 0.09 microM (n = 16). Introduction of AMP (100 microM) through the probe increased the dialysate adenosine markedly to 9.79 +/- 0.43 microM (n = 12, P < 0.001 vs baseline), and this increase was inhibited by the ecto-5'-nucleotidase inhibitor, alpha,beta-methyleneadenosine 5'-diphosphate (100 microM), to 0.76 +/- 0.12 microM (n = 8). Thus, the dialysate adenosine noted during the perfusion of AMP originated from dephosphorylation of AMP by ecto-5'-nucleotidase, and the dialysate level of adenosine attained reflects the ecto-5'-nucleotidase activity in the tissue in situ. 3. Glibenclamide (0.1-100 microM) decreased the adenosine concentration measured during the perfusion of AMP (100 microM) in a concentration-dependent manner (IC50 = 10.5 microM). In contrast, 5-hydroxydecanoate (10-100 microM) did not affect the concentrations of dialysate adenosine, measured in the presence of AMP (100 microM). These results suggest that glibenclamide inhibits the activity of endogenous ecto-5'-nucleotidase and decreases the concentration of adenosine in the interstitial space of rat ventricular muscles in situ.