AJP - Gastrointestinal and Liver Physiology, Vol 250, Issue 5 625-G632, Copyright © 1986 by American Physiological Society
Measurements of intracellular pH in Necturus antral mucosa by microelectrode technique
S. W. Ashley, D. I. Soybel and L. Y. Cheung
Intracellular pH (pHi) was measured in the surface epithelial cells of
Necturus antrum using pH-sensitive intracellular microelectrodes.
Electrodes were prepared by filling 10- to 20-M omega resistance glass
microelectrodes with a H+ ion-selective exchange resin and calibrated
before use in solutions of known pH 4.0-8.0. The electrode response (n =
15) was linear (r = 0.93; P less than 0.001) with a slope of 52.1 +/- 2.3
mV/pH unit. Antral mucosa was mounted in a modified Ussing chamber and pHi
was determined from the difference between the potentials recorded by
intracellular H+-selective and conventional microelectrodes. These
measurements of pHi were validated by examining the response of the
intracellular microelectrodes to 1) depolarization of the cell membrane
produced by K+ substitution for Na+, and 2) alkalinization and
acidification of pHi produced by NH+4 substitution for Na+ in the bathing
solutions. In tissues bathed with HCO-3-Ringer solution (pH 7.0), the mean
pHi was 7.34 +/- 0.02 with a range from 7.24 to 7.43. In
N-2-hydroxymethylpiperazine-N' -ethanesulfonic acid (HEPES)-Ringer solution
(pH 7.0), pHi was reduced to 7.02 +/- 0.05 (P less than 0.01).
Acidification of the luminal solution to pH 6.8 with CO2 produced a 0.22
+/- 0.04-pH unit fall in pHi (P less than 0.001). In contrast,
acidification to pH 4.0 with HCl had no significant effects on pHi. These
findings indicate that HCO-3 may play an important role in pHi regulation
in this tissue. In addition, they suggest that, in contrast to CO2, the
surface epithelial cells of Necturus antrum are relatively resistant to
acidification by luminal HCl.
