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  • Title: Hypotonic stress activates BK channels in clonal kidney cells via purinergic receptors, presumably of the P2Y subtype.
    Author: Hafting T, Haug TM, Ellefsen S, Sand O.
    Journal: Acta Physiol (Oxf); 2006 Sep; 188(1):21-31. PubMed ID: 16911250.
    Abstract:
    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). METHODS: 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 microM) and pyridoxalphosphate-6-azophenyl-2'4'-disulphonate (PPADS; 20 microM), 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.
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