Differential modulation of ATP-induced calcium signalling by A1 and A2 adenosine receptors in cultured cortical astrocytes

1 Despite the accumulating evidence that under various pathological conditions the extracellular elevation of adenine-based nucleotides and nucleosides plays a key role in the control of astroglial reactivity, how these signalling molecules interact in the regulation of astrocyte function is still largely elusive. 2 The action of the nucleoside adenosine in the modulation of the intracellular calcium signalling ([Ca2+](i)) elicited by adenosine 5'-triphosphate (ATP)-induced activation of P2 purinoceptors was investigated on neocortical type-1 astrocytes in primary culture by using single-cell microfluorimetry. 3 Astrocyte challenge with ATP (1-10 muM) elicited biphasic [Ca2+](i) responses consisting of an initial peak followed by a sustained elevation. The stable adenosine analogue 2-chloroadenosine (2-ClA) potentiated the transient [Ca2+](i) rise induced by activation of metabotropic P2Y receptors. Among the various P1 receptor agonists tested, the nonselective agonist 5'-N-ethylcarboxamidoadenosine (NECA) mimicked the 2-ClA action, whereas the selective A1 R(-) N6-(2-phenylisopropyl)-adenosine (R-PIA), the A2A 2-[4-(2-carboxyethyl)phenethylamino]-5'-N-ethylcarboxamidoadenosine (CGS-21680) and A3 1-deoxy-1-(6-[([3-lodophenyl]methyl)-amino]-9H-purin-9-yl)-N-methyl-beta-D-ribofuranuronamide (IB-MECA) agonists were ineffective. 4 Application of R-PIA > NECA greater than or equal to 2-ClA depressed the [Ca2+](i) plateau reversibly. Moreover, in the presence of R-PIA or 2-ClA, the prolonged [Ca2+](i) signal was maintained by application of the A1 antagonist 1,3-diethyl-8-phenylxanthine(DPX). Finally, preincubation of the astrocytes with pertussis toxin abrogated the 2-ClA inhibition of the ATP-elicited sustained [Ca2+](i) rise without affecting the transient [Ca2+](i) potentiation. 5 Taken together, these findings indicate that stimulation of A1 and A2 adenosine receptors mediates a differential modulation of [Ca2+](i) signalling elicited by P2 purinoceptors. Since variations in [Ca2+](i) dynamics also affect cell proliferation and differentiation, our data suggest that tuning of the extracellular levels of adenosine may be relevant for the control of astrogliosis mediated by adenine nucleotides.