Proton (H+) conductive pathways are suggested to play roles in the regulation of intracellular pH. We characterized temperature-sensitive whole cell currents in mouse bone marrow–derived mast cells (BMMC), immature proliferating mast cells generated by in vitro culture. Heating from 24 to 36°C reversibly and repeatedly activated a voltage-dependent outward conductance with Q10 of 9.9 ± 3.1 (mean ± SD) (n = 6). Either a decrease in intracellular pH or an increase in extracellular pH enhanced the amplitude and shifted the activation voltage to more negative potentials. With acidic intracellular solutions (pH 5.5), the outward current was detected in some cells at 24°C and Q10 was 6.0 ± 2.6 (n = 9). The reversal potential was unaffected by changes in concentrations of major ionic constituents (K+, Cl−, and Na+), but depended on the pH gradient, suggesting that H+ (equivalents) is a major ion species carrying the current. The H+ current was featured by slow activation kinetics upon membrane depolarization, and the activation time course was accelerated by increases in depolarization, elevating temperature and extracellular alkalization. The current was recorded even when ATP was removed from the intracellular solution, but the mean amplitude was smaller than that in the presence of ATP. The H+ current was reversibly inhibited by Zn2+ but not by bafilomycin A1, an inhibitor for a vacuolar type H+-ATPase. Macroscopic measurements of pH using a fluorescent dye (BCECF) revealed that a rapid recovery of intracellular pH from acid-load was attenuated by lowering temperature, addition of Zn2+, and depletion of extracellular K+, but not by bafilomycin A1. These results suggest that the H+ conductive pathway contributes to intracellular pH homeostasis of BMMC and that the high activation energy may be involved in enhancement of the H+ conductance.
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1 June 1997
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June 01 1997
A Highly Temperature-sensitive Proton Current in Mouse Bone Marrow–derived Mast Cells
Miyuki Kuno,
Miyuki Kuno
From the Department of Physiology, Osaka City University Medical School, Abeno-ku, Osaka 545, Japan
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Junko Kawawaki,
Junko Kawawaki
From the Department of Physiology, Osaka City University Medical School, Abeno-ku, Osaka 545, Japan
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Fusao Nakamura
Fusao Nakamura
From the Department of Physiology, Osaka City University Medical School, Abeno-ku, Osaka 545, Japan
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Miyuki Kuno
From the Department of Physiology, Osaka City University Medical School, Abeno-ku, Osaka 545, Japan
Junko Kawawaki
From the Department of Physiology, Osaka City University Medical School, Abeno-ku, Osaka 545, Japan
Fusao Nakamura
From the Department of Physiology, Osaka City University Medical School, Abeno-ku, Osaka 545, Japan
Address correspondence to Miyuki Kuno, MD, Ph.D., Department of Physiology, Osaka City University Medical School, Abeno-ku, Osaka 545, Japan. Fax: 06-645-2015; E-mail: [email protected]
1
Abbreviations used in this paper: BCECF, 2′,7′-bis-(2-carboxyethyl)- 5(and 6)carboxy-fluorescein; BMMC, bone marrow-derived mast cells; EH, equilibrium potential for H+; pHi, intracellular pH; pHo, extracellular pH; pHp, the pH of pipette solution; Vrev, reversal potential.
Received:
October 31 1996
Accepted:
March 18 1997
Online ISSN: 1540-7748
Print ISSN: 0022-1295
1997
J Gen Physiol (1997) 109 (6): 731–740.
Article history
Received:
October 31 1996
Accepted:
March 18 1997
Citation
Miyuki Kuno, Junko Kawawaki, Fusao Nakamura; A Highly Temperature-sensitive Proton Current in Mouse Bone Marrow–derived Mast Cells . J Gen Physiol 1 June 1997; 109 (6): 731–740. doi: https://doi.org/10.1085/jgp.109.6.731
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