title: P2X4 activation modulates volume-sensitive outwardly rectifying chloride channels in rat hepatoma cells.
abstract: Volume-sensitive outwardly rectifying (VSOR) Cl(-) channels are critical for the regulatory volume decrease (RVD) response triggered upon cell swelling. Recent evidence indicates that H(2)O(2) plays an essential role in the activation of these channels and that H(2)O(2) per se activates the channels under isotonic isovolumic conditions. However, a significant difference in the time course for current onset between H(2)O(2)-induced and hypotonicity-mediated VSOR Cl(-) activation is observed. In several cell types, cell swelling induced by hypotonic challenges triggers the release of ATP to the extracellular medium, which in turn, activates purinergic receptors and modulates cell volume regulation. In this study, we have addressed the effect of purinergic receptor activation on H(2)O(2)-induced and hypotonicity-mediated VSOR Cl(-) current activation. Here we show that rat hepatoma cells (HTC) exposed to a 33% hypotonic solution responded by rapidly activating VSOR Cl(-) current and releasing ATP to the extracellular medium. In contrast, cells exposed to 200 microm H(2)O(2) VSOR Cl(-) current onset was significantly slower, and ATP release was not detected. In cells exposed to either 11% hypotonicity or 200 microm H(2)O(2), exogenous addition of ATP in the presence of extracellular Ca(2+) resulted in a decrease in the half-time for VSOR Cl(-) current onset. Conversely, in cells that overexpress a dominant-negative mutant of the ionotropic receptor P2X4 challenged with a 33% hypotonic solution, the half-time for VSOR Cl(-) current onset was significantly slowed down. Our results indicate that, at high hypotonic imbalances, swelling-induced ATP release activates the purinergic receptor P2X4, which in turn modulates the time course of VSOR Cl(-) current onset in a extracellular Ca(2+)-dependent manner.
authors.affiliation: Centro de Estudios Moleculares de la Célula & Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, 838-0453 Santiago, Chile. dvarela@bitmed.med.uchile.cl
authors.forename: Diego
authors.initials: D
authors.lastname: Varela
authors.forename: Antonello
authors.initials: A
authors.lastname: Penna
authors.forename: Felipe
authors.initials: F
authors.lastname: Simon
authors.forename: Ana Luisa
authors.initials: AL
authors.lastname: Eguiguren
authors.forename: Elías
authors.initials: E
authors.lastname: Leiva-Salcedo
authors.forename: Oscar
authors.initials: O
authors.lastname: Cerda
authors.forename: Francisco
authors.initials: F
authors.lastname: Sala
authors.forename: Andrés
authors.initials: A
authors.lastname: Stutzin
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D045744:Cell Line, Tumor; D048429:Cell Size; D018118:Chloride Channels / Q000235:genetics / Q000378:metabolism*; D007038:Hypotonic Solutions; D008113:Liver Neoplasms; D018408:Patch-Clamp Techniques; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism*; D058848:Receptors, Purinergic P2X4
chemical_list: D018118:Chloride Channels; D007038:Hypotonic Solutions; C548228:P2rx4 protein, rat; D018048:Receptors, Purinergic P2; D058848:Receptors, Purinergic P2X4; D000255:Adenosine Triphosphate; D002118:Calcium