title: Effects of extracellular pH on receptor-mediated Ca2+ influx in A7r5 rat smooth muscle cells: involvement of two different types of channel.
abstract: 1. The effects of extracellular pH (pHo) on receptor (vasopressin or endothelin-1)-mediated Ca2- entry and Ca(2+)-permeable channels were investigated in aortic smooth muscle cells (A7r5) from rat embryonic thoracic aorta. Intracellular Ca2+ ([Ca2+]i) was measured using fura-2 AM and whole-cell voltage clamp techniques were employed. 2. Vasopressin and endothelin-1 (100 nM) in the presence of nicardipine (10 microM) evoked a sustained rise in [Ca2+]i due to calcium entry. Extracellular acidosis decreased receptor (vasopressin or endothelin-1)-mediated Ca2+ entry, while extracellular alkalosis potentiated it. 3. Depletion of intracellular Ca2+ stores with thapsigargin (1 microM) also evoked Ca2+ entry activated by emptying of intracellular Ca2+ stores (capacitative Ca2+ entry). Extracellular acidosis decreased this capacitative Ca2+ entry, while extracellular alkalosis potentiated it. 4. Under voltage-clamp conditions with Ca+ internal solution, vasopressin and endothelin-1 activated non-selective cation currents (ICAT). Ba2+ or Ca2+ were also charge carriers of ICAT. Reducing the pHo inhibited ICAT, while increasing pHo potentiated it in a reversible manner. 5. Intracellular pH (pHi) changes did not cause the same marked effects as pHo changes, and a high concentration of Hepes (50 mM) in the patch pipette did not inhibit the effects of pHo on ICAT. 6. Similar results were obtained when ICAT was activated by GTP gamma S (1 mM) applied through the patch pipette, even in the absence of agonists, probably because of direct activation of GTP-binding proteins coupled to the receptors. 7. In cells treated with thapsigargin, addition of Ca2+ to the bath solution induced Ca(2+)-dependent K+ currents activated by capacitative Ca2+ entry. However, no measurable ionic currents activated by capacitative Ca2+ entry (ICRAC) were observed under conditions with Cs+ internal solution and EGTA (5 mM), although vasopressin still activated ICAT. 8. These results suggest that the contractile agonists vasopressin and endothelin-1 evoked Ca2+ entry through two different types of Ca(2+)-permeable channel (ICAT and ICRAC) and pHo affects these channels, which may modulate receptor-mediated Ca2+ influx in A7r5 cells. Thus, pH-induced changes of these channels may play a pathophysiological role in the control of receptor-mediated contractions.
authors.affiliation: Second Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Iwasawa
authors.forename: T
authors.initials: T
authors.lastname: Nakajima
authors.forename: H
authors.initials: H
authors.lastname: Hazama
authors.forename: A
authors.initials: A
authors.lastname: Goto
authors.forename: W S
authors.initials: WS
authors.lastname: Shin
authors.forename: T
authors.initials: T
authors.lastname: Toyo-oka
authors.forename: M
authors.initials: M
authors.lastname: Omata
mesh_terms: D000818:Animals; D002021:Buffers; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D004558:Electric Stimulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006531:HEPES / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D051381:Rats; D017466:Receptors, Endothelin / Q000187:drug effects / Q000378:metabolism*; D017483:Receptors, Vasopressin / Q000187:drug effects / Q000378:metabolism*; D019284:Thapsigargin / Q000494:pharmacology
chemical_list: D002021:Buffers; D015220:Calcium Channels; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D017466:Receptors, Endothelin; D017483:Receptors, Vasopressin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D004533:Egtazic Acid; D019284:Thapsigargin; D006531:HEPES; D002118:Calcium