ABSTRACT
Mixed neuronal and glial cell spinal cord cultures from neonates express ADP sensitive P2Y(1,12&13) receptors. ADP (10microM) evoked increases in intracellular calcium that were essentially abolished by the P2Y(1) receptor antagonist MRS2179 (10microM), responses were also absent in preparations from P2Y(1) receptor deficient mice however UTP (100microM) evoked calcium rises were unaffected. ADP also evoked a robust increase in extracellular signal-regulated protein kinase (ERK) phosphorylation that was of similar magnitude in the cultures from wild type and P2Y(1) receptor deficient mice. These results suggest that ADP acts through P2Y(1) receptors to mediate an increase in intracellular calcium but not to stimulate ERK phosphorylation in the spinal cord.
Subject(s)
Adenosine Diphosphate/metabolism , Receptors, Purinergic P2/physiology , Signal Transduction/physiology , Spinal Cord/physiology , Adenosine Diphosphate/analogs & derivatives , Adenosine Diphosphate/pharmacology , Aniline Compounds/metabolism , Animals , Calcium/metabolism , Dose-Response Relationship, Drug , Drug Interactions , Female , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Purinergic P2 Receptor Antagonists , Receptors, Purinergic P2/deficiency , Receptors, Purinergic P2Y1 , Signal Transduction/drug effects , Spinal Cord/drug effects , Time Factors , Tissue Culture Techniques , Uridine Triphosphate/pharmacology , Xanthenes/metabolismABSTRACT
P2X receptors for adenosine tri-phosphate (ATP) are a distinct family of ligand-gated cation channels with two transmembrane domains, intracellular amino and carboxy termini and a large extracellular ligand binding loop. Seven genes (P2X(1-7)) have been cloned and the channels form as either homo or heterotrimeric channels giving rise to a wide range of phenotypes. This review aims to give an account of recent work on the molecular basis of the properties of P2X receptors. In particular, to consider emerging information on the assembly of P2X receptor subunits, channel regulation and desensitisation, targeting, the molecular basis of drug action and the functional contribution of P2X receptors to physiological processes.
Subject(s)
Adenosine Triphosphate/chemistry , Adenosine Triphosphate/metabolism , Ion Channel Gating/physiology , Receptors, Purinergic P2/chemistry , Receptors, Purinergic P2/metabolism , Amino Acid Sequence , Animals , Binding Sites , Humans , Mice , Models, Chemical , Models, Molecular , Molecular Sequence Data , Protein Binding , Receptors, Purinergic P2X2 , Structure-Activity RelationshipABSTRACT
1 P2Y receptors are expressed in the nervous system and are involved in calcium signalling in neurons and glia. In the superior cervical ganglion (SCG), RT-PCR analysis indicated the presence of P2Y(1,2&6) receptors. Rises in intracellular calcium in response to P2Y receptor stimulation were determined from adult mouse cultured SCG neurons and glia. 2 ADP evoked suramin (100 microM)- and pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS, 30 microM)-sensitive rises in intracellular calcium in approximately 80% of SCG neurons (EC50 approximately 20 microM). ADP-evoked responses were abolished in neurons from P2Y1 receptor-deficient mice (responses to UTP were unaffected). 3 The pyrimidines UTP (EC50 approximately 85 microM) and UDP (EC50>90 microM) evoked PPADS- and suramin-sensitive responses in approximately 70 and approximately 20% of SCG neurons, respectively. 4 In SCG glial cells, ADP (EC50 approximately 30 microM) evoked calcium responses in approximately 50% of glia. These were suramin and PPADS sensitive and essentially abolished in SCG glial cells cultured from adult P2Y1 receptor-deficient mice. 5 UTP (EC50 approximately 25 microM) and UDP (EC50>200 microM) evoked suramin- and pyridoxalphosphate-6-azophenyl-2',5'-disulphonate-sensitive rises in calcium in approximately 60 and 20% SCG glial cells, respectively. 6 These results indicate the presence of several P2Y receptors coupled to an increase in intracellular calcium in the SCG: ADP-sensitive P2Y1 receptors and UDP-sensitive P2Y6 receptors in SCG neurons and glial cells, a novel UTP-sensitive P2Y receptor in SCG neurons and UTP- and ATP-sensitive P2Y2 receptors in SCG glia.
