Existence of high and low affinity dinucleotides pentaphosphate-induced calcium responses in individual synaptic terminals and lack of correlation with the distribution of P2X 1–7 subunits

Abstract

Individual analysis of synaptic terminals calcium responses, induced by dinucleotides pentaphosphate, Ap 5A or Gp 5G, demonstrates the presence of two main groups considering the concentration required for stimulation. The first group corresponds to those responding to Ap 5A or Gp 5G at nanomolar concentration, representing 16% and 12%, respectively, and the second one responds to micromolar concentration and represents, respectively, 17% and 14%, of the total functional synaptosomal population in rat midbrain. Dose–response curves in single terminals showed an Ap 5A EC 50 values of 0.9 ± 0.2 nM and 11.8 ± 0.9 μM, being the maximal intrasynaptosomal calcium increase of 200 ± 0.3 and 125 ± 0.2 nM for the high and low affinity responding terminals, respectively. Combination of microfluorimetric and immunocytochemical studies showed lack of correlation between dinucleotides pentaphosphate responses and P2X receptor subunits expression, in spite of the abundance of P2X 2, P2X 3 and P2X 7 at the presynaptic level in rat midbrain synaptosomes. Pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS), a P2X receptors antagonist, showed no effect on low affinity dinucleotides receptors population, and partial inhibition on the high affinity one. On the other hand, diinosine pentaphosphate (Ip 5I) completely abolished the low affinity dinucleotides responses, and 60% inhibition of the high affinity ones.