title: Extracellular ATP activates a P2 receptor in necturus erythrocytes during hypotonic swelling.
abstract: We recently reported that ATP is released from Necturus erythrocytes via a conductive pathway during hypotonic swelling and that extracellular ATP potentiates regulatory volume decrease (RVD). This study was designed to determine whether extracellular ATP exerts its effect via a purinoceptor. This was accomplished using three different experimental approaches: 1) hemolysis studies to examine osmotic fragility, 2) a Coulter counter to assess RVD, and 3) the whole-cell patch-clamp technique to measure membrane currents. We found extracellular ATP and ATPgammaS, two P2 agonists, decreased osmotic fragility, enhanced cell volume recovery in response to hypotonic shock, and increased whole-cell currents. In addition, 2-methylthio-ATP potentiated RVD. In contrast, UTP, alpha,beta-methylene-ATP, and 2'-& 3'-O-(4-benzoyl-benzoyl) adenosine 5'-triphosphate and the P1 agonist adenosine had no effect regardless of experimental approach. Furthermore, the P2 antagonist suramin increased osmotic fragility, inhibited RVD, and reduced whole-cell conductance in swollen cells. Consistent with a previous study that indicated cell swelling activates a K+ conductance, suramin had no effect in the presence of gramicidin (a cationophore used to maintain a high K+ permeability). We also found the P2 antagonist pyridoxal-5-phosphate-6-azophenyl-2'4-disulfonic acid (PPADS) increased osmotic fragility; however, reactive blue 2 and the P1 antagonists caffeine and theophylline had no effect. Our results show that extracellular ATP activated a P2 receptor in Necturus erythrocytes during hypotonic swelling, which in turn potentiated RVD by stimulating K+ efflux. Pharmacological evidence suggested the presence of a P2X receptor subtype.
authors.affiliation: Department of Biology, Ripon College, 300 Seward St, Ripon, WI 54971, USA.
authors.forename: D B
authors.initials: DB
authors.lastname: Light
authors.forename: P K
authors.initials: PK
authors.lastname: Dahlstrom
authors.forename: R T
authors.initials: RT
authors.lastname: Gronau
authors.forename: N L
authors.initials: NL
authors.lastname: Baumann
mesh_terms: D000241:Adenosine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D048429:Cell Size / Q000502:physiology*; D004912:Erythrocytes / Q000187:drug effects / Q000378:metabolism*; D006096:Gramicidin / Q000494:pharmacology; D006461:Hemolysis; D009338:Necturus maculosus; D009996:Osmotic Fragility; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism; D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D013498:Suramin / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology
chemical_list: D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D013873:Thionucleotides; D006096:Gramicidin; C022571:adenosine 5'-O-(3-thiotriphosphate); D013498:Suramin; D000255:Adenosine Triphosphate; D000241:Adenosine; C002630:alpha,beta-methyleneadenosine 5'-triphosphate; D011188:Potassium; C008501:2-methylthio-ATP