P2Y receptor expression on astrocytes in the nucleus accumbens of rats

Abstract

The expression of purinoceptor (P2)Y-subtypes on astrocytes in vivo under physiological conditions and after stab wound injury was investigated. Reverse transcriptase-polymerase chain reaction with specific primers for the receptor-subtypes P2Y 1,2,4,6,12 in tissue extracts of the nucleus accumbens of untreated rats revealed the presence of all P2Y receptor mRNAs investigated. Double immunofluorescence visualized with laser scanning microscopy indicated the expression of the P2Y 1,4 receptors on glial fibrillary acidic protein (GFAP)-labeled astrocytes under physiological conditions. After stab wound injury the additional expression of the P2Y 2 and P2Y 6 receptors, and an up-regulation of the P2Y 1,4 receptor-labeling on astrocytic cell bodies and/or processes was observed. Astrocytes of cortical, in contrast to accumbal areas exhibited P2Y 1,2,4,6 receptor-immunoreactivity (IR) under control conditions, which was up-regulated after stab would injury. Labeling for the P2Y 12 receptor was not observed on GFAP-positive cortical and accumbal astrocytes under any of the conditions used. For the first time, the co-localization of different P2 receptor-subtypes (e.g. P2Y 1 and P2X 3) on the same astrocyte was shown immunocytochemically. The up-regulation of P2Y 1 receptor-IR on astrocytes and non-glial cells after mechanical injury could be facilitated by microinfusion of the P2Y 1,12,13 receptor agonist adenosine 5′- O-(2-thiodiphosphate) (ADPβS). Proliferative changes after ADPβS-microinjection were characterized by means of double-staining with antibodies against GFAP and 5-bromo-2′-deoxyuridine. The non-selective P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2′,4′-disulphonic acid, the P2Y 1 receptor antagonist N 6-methyl-2′-deoxyadenosine 3′,5′-bisphosphate and the P2Y 1 receptor-antibody itself inhibited the agonist-induced effects. The data indicate the region-specific presence of P2Y receptors on astrocytes in vivo and their up-regulation after injury as well as the co-localization of P2X and P2Y receptor-subtypes on the same astrocyte. The dominant role of P2Y 1 receptors in proliferation and the additional stimulation of non-P2Y 1 receptors has been demonstrated in vivo suggesting the involvement of this receptor-type in the gliotic response under physiological and pathological conditions.