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Hypotonic stimulation induced Ca2+ release from IP3‐sensitive internal stores in a green monkey kidney cell line.
Author(s) -
Ishii T,
Hashimoto T,
Ohmori H
Publication year - 1996
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.1996.sp021389
Subject(s) - thapsigargin , egta , ryanodine receptor , stimulation , chemistry , biophysics , tonicity , medicine , endocrinology , calcium , caffeine , membrane potential , fura 2 , intracellular , biology , biochemistry , enzyme , cytosol
1. Hypotonic stimulation (180 +/‐ 5 mosmol l‐1) increased [Ca2+]i in fura‐2‐loaded Green monkey kidney cells (COS‐7 cells) and depolarized the membrane. 2. COS‐7 cells were depolarized up to ‐3.5 +/‐ 4.4 mV from a resting membrane potential of ‐35.2 +/‐ 2.3 mV in response to hypotonic stimulation, when the patch electrode was filled with a 160 mM KCl‐0.5 mM EGTA‐based intracellular medium. 3. The increase in [Ca2+]i induced by hypotonic stimulation was divided into two phases. One was transient and oscillatory, and observed in Ca(2+)‐free medium; the other was persistent, blocked by 100 microM La3+, and observed only in Ca(2+)‐containing medium. 4. The increase in [Ca2+]i in Ca(2+)‐free medium was blocked by pretreatment with 10 microM thapsigargin. The increase in [Ca2+]i induced by 10 microM thapsigargin was reduced after hypotonic stimulation which induced an increase in [Ca2+]i in Ca(2+)‐free medium. 5. The increase in [Ca2+]i in Ca(2+)‐free medium was not affected by treatment with 5 mM caffeine or 1‐10 microM ryanodine. Neither caffeine nor ryanodine induced an increase in [Ca2+]i. 6. Adenosine 5'‐O‐2‐thiodiphosphate (ADP‐beta‐S; a P2Y receptor agonist) induced an increase in [Ca2+]i in Ca(2+)‐free medium and caused phosphoinositide breakdown in COS‐7 cells. Exposure to 10 microM ADP‐beta‐S blocked the increase in [Ca2+]i induced in the Ca(2+)‐free medium by hypotonic stimulation. The results of summary points 4, 5, and 6 suggest that the increase in [Ca2+]i induced by hypotonic stimulation is due to Ca2+ release from inositol 1,4,5‐trisphosphate (IP3)‐sensitive internal stores. 7. The hypotonic stimulation‐activated hydrolysis of phosphoinositides was decreased by pertussis toxin (PTX) in a dose‐dependent manner. 8. These observations strongly suggest that hypotonic stimulation induced an increase in [Ca2+]i in Ca(2+)‐free medium through activation of cascades using PTX‐sensitive guanine nucleotide binding protein (G protein) and IP3.