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Hypotonic stress activates BK channels in clonal kidney cells via purinergic receptors, presumably of the P2Y 1 subtype
Author(s) -
Hafting T.,
Haug T. M.,
Ellefsen S.,
Sand O.
Publication year - 2006
Publication title -
acta physiologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.591
H-Index - 116
eISSN - 1748-1716
pISSN - 1748-1708
DOI - 10.1111/j.1748-1716.2006.01601.x
Subject(s) - purinergic receptor , p2y receptor , tonicity , receptor , purinergic signalling , microbiology and biotechnology , chemistry , biology , endocrinology , medicine , adenosine receptor , agonist
Aim: Membrane stretch due to cell swelling may cause a minute leakage of adenosine triphosphate (ATP) that stimulates endogenous purinergic receptors. The following elevation of the cytosolic‐free Ca 2+ concentration ([Ca 2+ ] i ) may then participate in cell volume regulation. The aim of the present study was to test if purinergic receptors and large conductance Ca 2+ activated K + (BK) channels are activated in response to hypotonic stress in clonal kidney cells (Vero cells). Methods: The methods used are fura‐2 microfluorometry, cell‐attached patch clamp and reverse‐transcriptase polymerase chain reaction (RT‐PCR). Results: Subjecting cells to hypotonic stress for 10 s by exposure to a solution with 45% reduced osmolality induced a transient rise in [Ca 2+ ] i . This response persisted in virtually Ca 2+ ‐free extracellular solution, demonstrating that Ca 2+ was mainly released from intracellular stores. The hypotonically induced elevation of [Ca 2+ ] i was completely inhibited by the P2 receptor antagonists suramine (100 μ m ) and pyridoxalphosphate‐6‐azophenyl‐2′4′‐disulphonate (PPADS; 20 μ m ), indicating that extracellular ATP is crucial for the [Ca 2+ ] i increase. RT‐PCR revealed the expression of mRNA for P2Y 1 receptors in Vero cells. The putatively selective P2Y 1 antagonist PPADS did completely block Ca 2+ responses to both ATP and hypotonic stress, suggesting that P2Y 1 receptors are mediating the response. Furthermore, patch clamp recordings in cell‐attached configuration revealed that BK channels are activated in response to hypotonic stress. Conclusion: Vero cells express functional purinergic receptors, presumably of the P2Y 1 subtype. These receptors are responsible for the elevation of [Ca 2+ ] i evoked by hypotonic stress. The concurrent activation of BK channels permits K + efflux that may contribute to regulatory volume decrease.