The release of purine and pyrimidine nucleotide catabolites from working blood vessels

This study examined the release of purine and pyrimidine nucleotide catabolites from perfused rat hindlimb and mesenteric arcade, and incubated aortas in response to vasoconstriction, stretch-induced tone and skeletal muscle contraction. The release of uric acid and uracil from non-recirculating, perfused rat hindlimb was stimulated from 0.95 and 0.4nmol/min per g, respectively, by a factor of 2-5fold by noradrenaline, vasopressin or angiotensin II, in a dose-dependent manner, coinciding with increases in V02 and perfusion pressure. Angiotensin-mediated increases were blocked by the nitrovasodilator nitroprusside. Skeletal muscle contraction increased the release of inosine and hypoxanthine but did not alter uracil or uric acid release. Perfused mesenteric arcade and incubated aortas released nucleotide catabolites in response to serotonin and tension respectively. The main products released from aortas were uridine, cytidine, urate and hypoxanthine. Following vasoconstriction pyrimidine compounds represented 23-38 % of the nucleotide catabolites found in venous effluent or incubation medium from hindlimb, mesentery or aorta. Pyrimidine nucleotides represented 25 % of the intracellular nucleotides in aorta. In contrast, only 2.9% of the hindlimb nucleotide pool consisted of pyrimidine nucleotides. It was concluded that nucleotide catabolite release during vasoconstriction derived from vascular tissue. The possibility that uracil derived from the breakdown of UTP released with a vasomodulatory function was examined. UTP was a potent vasodilator in the perfused rat hindlimb, opposing noradrenaline- and angiotensin-mediated vasoconstriction. Exogenous UTP was rapidly catabolized by perfused hindlimb to non-vasoactive products, UMP, uridine and uracil. ˜í¬±, ˜í‚â§-MethyleneADP, an ecto-5'nucleotidase inhibitor, blocked extracellular UMP degradation during infusion of UTP, but failed to reduce uracil or uridine release and did not lead to the accumulation of UMP during angiotensin infusion. Hence, extracellular UTP degradation is unlikely to be the source of hindlimb uracil release. Lactate release from hindlimb correlated well with uracil release during vasoconstriction, vasodilation or cyanide poisoning (r=0.831; P<0.001) but not during B-adrenergic stimulation. High [Ca2+], (‚Äöv¢vñ10mM) but not K+-depolarization, stimulated nucleotide catabolite release from aortas incubated under tension. This data provides evidence of a link between glycolysis, raised cytoplasmic [Na+] and pyrimidine release. Pyrimidine nucleotide (but not RNA) levels were reduced in hindlimb and aorta after intensive pyrimidine release. However, the relationship between catabolite release and tissue nucleotide depletion was not stoichiometric, suggesting that pyrimidine de novo synthesis rates may be high in vascular tissue. Radiolabel from [3H]cytidine entered aortic uracil nucleotides, suggesting that uridine may derive from cytosine nucleotides. Aorta homogenate UTP hydrolysis was stimulated by 10-150mM Na+ in the absence of other cations was separate from Na+/K+-ATPase and not ouabain inhibited. Highest activity was observed with UTP and the activity was probably intracellular but not soluble. Overall the study shows: (i) that high workloads result in the release of nucleotide catabolites from VSM, of which a high proportion are pyrimidine compounds; (ii) that nucleotide catabolites released in response to vascular work are likely to be generated intracellularly; (iii) and that pyrimidine nucleotide catabolite generation and release from VSM is associated with raised glycolysis and high intracellular [Na+].