Coexpression of several types of metabotropic nucleotide receptors in single cerebellar astrocytes.
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, P2Y(1) and P2Y(2) or P2Y(4). In a subpopulation of type 1 astrocytes, ATP prestimulation did not block UTP responses, and UDP elicited clear intracellular Ca(2+) 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 P2Y(1)-specific antagonist N:(6)-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: P2Y(1) and either P2Y(2) or P2Y(4) receptors. Moreover, 30-40% of astrocytes also coexpressed specific pyrimidine receptors of the P2Y(6) subtype, highly selective for UDP coupled to pertussis-toxin insensitive G protein.
Jiménez
A I
AI
Departamento de Bioquímica, Facultad de Veterinaria, Universidad Complutense, Madrid, Spain.
Castro
E
E
Communi
D
D
Boeynaems
J M
JM
Delicado
E G
EG
Miras-Portugal
M T
MT
D000244:Adenosine Diphosphate / Q000378:metabolism / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D007424:Intracellular Fluid / Q000378:metabolism; D037342:Pertussis Toxin; D011493:Protein Kinase C / Q000378:metabolism; D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D051381:Rats; D017208:Rats, Wistar; D018048:Receptors, Purinergic P2 / Q000096:biosynthesis*; D013268:Stimulation, Chemical; D013498:Suramin / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology; D014544:Uridine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D013873:Thionucleotides; D010566:Virulence Factors, Bordetella; D013498:Suramin; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D037342:Pertussis Toxin; D011493:Protein Kinase C; D002118:Calcium; D014544:Uridine Triphosphate; C008501:2-methylthio-ATP