Intracellular pH,intracellular free Ca,and junctional cell-cell coupling |
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Authors: | Birgit Rose Roger Rick |
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Institution: | (1) Department of Physiology and Biophysics, University of Miami School of Medicine, P.O. Box 016430, 33101 Miami, Florida;(2) Present address: Physiologisches Institut d. Univ. München, 8 München 2, F.R. Germany |
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Abstract: | Summary Intracellular pH (pH
i
) and intracellular Ca2+ (Ca2+]
i
) were determined inChironomus salivary gland cells under various conditions of induced uncoupling. pH
i
was measured with aThomas-type microelectrode, changes in Ca2+]
i
and their spatial distribution inside the cell were determined with the aid of intracellularly injected aequorin and an image intensifier-TV system, and cell-to-cell coupling was measured electrically. Treatments with NaCN (5mm), DNP (1.2mm), or ionophore A23187 (2 m) caused fall in junctional conductance (uncoupling) that was correlated with Ca2+]
i
elevation, as was shown before (Rose & Loewenstein, 1976,J. Membrane Biol.
28:87) but not with changes in pH
i
: during the uncoupling induced by CN, the pH
i
(normally 7.5) decreased at most by 0.2 units; during the uncoupling induced by the ionophore, pH
i
fell by 0.13 or rose by 0.3; and in any one of these three agents' uncouplings, the onset of uncoupling and recovery of coupling were out of phase with the changes in pH
i
. Intracellular injection of Ca-citrate or Ca-EGTA solutions buffered to pH 7.2 or 7.5 produced uncoupling with little or no pH
i
change when their free Ca2+]
i
was >10–5
m. On the other hand, such a solution at pH 4, buffered to Ca2+]<10–6
m, lowered pH
i
to 6.8 but produced no uncoupling. Thus, a decrease in pH
i
is not necessary for uncoupling in any of these conditions. In fact, uncoupling ensued also during increase in pH
i
: exposure to NH4HCO3 or withdrawal of propionate following exposure to a propionate-containing medium caused pH
i
to rise to 8.74, accompanied by Ca2+]
i
elevation and uncoupling at pH
i
>7.8.Cell acidification itself can cause elevation of Ca2+]
i
: injection (iontophoresis) of H+ invariably caused Ca2+]
i
elevation and uncoupling. These effects were produced also by an application of H+-transporting ionophore Nigericin at extracellular pH 6.5 which caused pH
i
to fall to 6.8. Exposure to 100% CO2 produced a fall in pH
i
, associated in 10 out of 25 cases with Ca2+]
i
elevation and, invariably, with uncoupling. The absence of a demonstrable Ca2+]
i
elevation in a proportion of these trials is attributable to depression in Ca2+-measuring sensitivity; inin vivo tests, detection sensitivity for Ca2+]
i
by aequorin was found to be depressed by the CO2 treatment. Upon CO2 washout, pH
i
and coupling recovered, but onset of recoupling set in at pH
i
as low as 6.32–6.88, generally lower than at the pH
i
at which uncoupling had set in. Exposure to 5% CO2 lowered pH
i
on the average by 0.3 and depressed coupling (in initially poorly coupled cells). After CO2-washout, pH
i
and coupling recovered. During the recovery phase Ca2+]
i
was elevated, an elevation associated with renewed uncoupling or decrease in rate of recoupling. The results are discussed in connection with possible regulatory mechanisms of junctional permeability. |
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Keywords: | |
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