thioinosine and Brain-Ischemia

Adenosine is an endogenous nucleoside that signals through G-protein coupled receptors. Extracellular adenosine is required for receptor activation and two pathways have been identified for formation and cellular release of adenosine. The CLASSICAL pathway relies on intracellular formation of adenosine from adenine nucleotides and cellular efflux of adenosine via equilibrative nucleoside transporters (ENTs). The ALTERNATE pathway involves cellular release of adenine nucleotides, hydrolysis via ecto-5'-nucleotidases and extracellular formation of adenosine.. A rat model of cerebral ischemia and primary cultures of rat forebrain astrocytes and neurons were used.. Using a rat model of cerebral ischemia, the ENT1 inhibitor nitrobenzylmercaptopurine ribonucleoside (NBMPR) significantly increased post-ischemic forebrain adenosine levels and significantly decreased hippocampal neuron injury relative to saline-treatment. NBMPR-induced increases in adenosine receptor activation were not detected, suggesting that altering the intracellular:extracellular distribution of adenosine can affect ischemic outcome. Using primary cultures of rat forebrain astrocytes and neurons, adenosine release was evoked by ischemic-like conditions. Dipyridamole, an inhibitor of ENTs, was more effective at inhibiting adenosine release from neurons than from astrocytes. In contrast, alpha , beta-methylene ADP, an inhibitor of ecto-5'-nucleotidase, was effective at inhibiting adenosine release from astrocytes, but not from neurons. Thus, during ischemic-like conditions, neurons released adenosine via the CLASSICAL pathway, while astrocytes released adenosine via the ALTERNATE pathway.. These cell type differences in pathways for adenosine formation during ischemia may allow transport inhibitors to block simultaneously adenosine release from neurons and adenosine uptake into astrocytes. In principle, this could improve neuronal ATP levels without decreasing adenosine receptor activation.

Topics: Adenosine; Affinity Labels; Animals; Astrocytes; Brain Ischemia; Cells, Cultured; Deoxyglucose; Dipyridamole; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Glucose; Hypoxia; Inosine; Models, Biological; Neurons; Oligomycins; Phosphodiesterase Inhibitors; Prosencephalon; Purines; Rats; Thioinosine; Tritium

We have used rats with streptozotocin-induced diabetes to investigate the effects of hyperglycaemia-mediated impaired nucleoside uptake on the actions of endogenous adenosine in hippocampal slices. In control tissue under conditions of anoxia and aglycaemia the rise in the extracellular adenosine concentration resulted in complete inhibition of synaptic activity in about 2 min. In slices from previously hyperglycaemic rats the inhibition of synaptically mediated responses occurred significantly faster, although this change could be prevented by insulin treatment. Application of the selective adenosine A1 receptor antagonist [8-cyclopentyl-1,3-dipropylxanthine (DPCPX)] prevented the anoxia/aglycaemia-mediated inhibition and, furthermore, abolished the differences in the electrophysiological responses between control and diabetic tissue. The effects of impaired nucleoside uptake could be mimicked in control slices by applying the nucleoside uptake blocker hydroxynitrobenzylthioinosine (HNBTI). This had the effect of speeding up the rate of anoxia/aglycaemia-induced synaptic inhibition in control tissue to that seen in diabetic tissue. However, such treatment had no effect on the responses in diabetic tissue as expected if the HNBTI-sensitive uptake process was already inhibited by the chronic hyperglycaemia. The impairment of nucleoside uptake by chronic hyperglycaemia results in the potentiation of the modulatory actions of endogenous adenosine in the central nervous system. Such an alteration in adenosine function may be important in explaining behavioural and pathological changes associated with diabetes mellitus.

Topics: Action Potentials; Adenosine; Affinity Labels; Animals; Brain Ischemia; Diabetes Mellitus, Experimental; Hippocampus; Hypoglycemia; Hypoxia, Brain; Male; Rats; Rats, Wistar; Synaptic Transmission; Thioinosine; Xanthines

Cerebral ischemia is often followed by a period of delayed hypoperfusion that may contribute to tissue injury. We tested the hypothesis that augmentation of interstitial adenosine can improve tissue perfusion under this condition 10 min global ischemia was produced in two groups of isoflurane-anesthetized newborn pigs by occlusion of subclavian and brachiocephalic arteries, and changes in local cortical blood flow and cortical interstitial purine metabolites were measured using the combined hydrogen clearance-microdialysis technique. In one group, the dialysis probe was perfused with artificial cerebrospinal fluid buffer containing nitrobenzyl-thioinosine (NBT1, 100 mumol/l), a competitive inhibitor of adenosine transport. In the untreated group (n = 9), baseline cortical blood flow (39 +/- 3 ml/min/100 g) was depressed by 51 +/- 5% and 42 +/- 6% at 40 and 60 min, respectively, of postischemic reperfusion. NBTI increased baseline interstitial adenosine levels 2.4-fold which increased baseline cortical blood flow 1.5-fold to 60 +/- 4 ml/min/100 g, and increased both absolute adenosine levels as well as adenosine as a percentage of total purine metabolites throughout ischemia and reperfusion. As a result, the extent of postischemic hypoperfusion was significantly lessened in NBTI-treated animals (n = 9), with reductions in cortical blood flow of only 28 +/- 3% and 24 +/- 5% at 40 and 60 min of reperfusion, respectively. These results indicate that inhibition of adenosine transport by NBTI elevates interstitial adenosine concentration during and following cerebral ischemia, and concomitantly improves cortical perfusion in the post-ischemic period. The latter effect may contribute to the documented neuroprotective efficacy of adenosinergic therapy in cerebral ischemia.

Topics: Adenosine; Animals; Biological Transport; Brain; Brain Ischemia; Cerebrovascular Circulation; Purines; Reperfusion; Swine; Thioinosine

The neuroprotective effects of adenosine are well-recognized. Recently, propentofylline, a xanthine derivative, has been shown to increase extracellular concentrations of adenosine in ischemic brain and to limit the extent of neuronal damage in experimental models of cerebral ischemia. Since the concentration of adenosine in brain is controlled, in part, by nucleoside transporter proteins, the action of propentofylline was proposed to be due to inhibition of mediated transfer of adenosine across cell membranes. To determine the likelihood of this mechanism, we examined the inhibitory effects of propentofylline on [3H]adenosine transport by the three best-characterized nucleoside transport processes, es, ei, and cif in cultured cell lines under conditions where only a single transporter type was operative. Propentofylline inhibited [3H]adenosine uptake by each of the three transport processes in a concentration-dependent manner. The greatest inhibitory potency was for es transporters (L1210/B23.1 cells), with an IC50 value of 9 microM, followed by ei transporters, with IC50 values of 170 microM (L1210/C2 cells) and 166 microM (Walker 256 cells). Propentofylline was a weak inhibitor of cif transporter, with an IC50 value of 6 mM. These results demonstrate that propentofylline is an inhibitor of adenosine transport processes and suggest that its neuroprotective effects may be due to an increase in extracellular concentrations of adenosine by virtue of inhibition of es transporter function.

Topics: Adenosine; Animals; Biological Transport; Brain Ischemia; Cell Line; Mice; Rats; Thioinosine; Tritium; Tumor Cells, Cultured; Xanthines