Coexpression of Several Types of Metabotropic Nucleotide Receptors in Single Cerebellar Astrocytes

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Abstract
We have examined the expression of mRNA for several P2Y nucleotide receptors by northern blot analysis in purified type 1 cerebellar astrocyte cultures. These results suggest that different P2Y subtypes could be responsible for ATP metabotropic calcium responses in single type 1 astrocytes. To identify these subtypes we have studied the pharmacological profile of ATP calcium responses using fura‐2 microfluorimetry. All tested astrocytes responded to ATP and UTP stimulations evoking similar calcium transients. Most astrocytes also responded to 2‐methylthioATP and ADP challenges. The agonist potency order was 2‐methylthioATP > ADP > ATP = UTP. Cross‐desensitization experiments carried out with ATP, UTP, and 2‐methylthioATP showed that 2‐methylthioATP and UTP interact with different receptors, P2Y1 and P2Y2 or P2Y4. In a subpopulation of type 1 astrocytes, ATP prestimulation did not block UTP responses, and UDP elicited clear intracellular Ca2+ concentration responses at very low concentrations. 2‐MethylthioATP and UTP calcium responses exhibited different sensitivity to pertussis toxin and different inhibition patterns in response to P2 antagonists. The P2Y1‐specific antagonist N6‐methyl‐2′‐deoxyadenosine 3′,5′‐bisphosphate (MRS 2179) specifically blocked the 2‐methylthio‐ATP responses. We can conclude that all single astrocytes coexpressed at least two types of P2Y metabotropic receptors: P2Y1 and either P2Y2 or P2Y4 receptors. Moreover, 30‐40% of astrocytes also coexpressed specific pyrimidine receptors of the P2Y6 subtype, highly selective for UDP coupled to pertussis‐toxin insensitive G protein.