The intrinsic intracellular H+ buffering power of snail neurones

Abstract

1. We measured intracellular pH (pHi) in snail neurones using pH-sensitive glass microelectrodes. We then calculated the intracellular buffering power (beta i) from the pHi changes associated with the influx or efflux of a variety of weak acids or bases. 2. The weak acid anions butyrate and propionate (20 mM) gave similar values for beta i but those measured using 20 mM-acetate were on average twice as great. 3. Although solutions were nominally CO2-free, blockage of pHi regulation with SITS (4-acetamido-4'-isothiocyanatostilbene-2,2'-disulphonic acid) increased the sizes of the pHi changes upon weak acid addition and removal. The corresponding measured values of beta i were on average 26% lower with SITS than without. 4. With pHi regulation blocked, the use of 2.7% CO2 to measure beta i gave beta i values similar to those measured with butyrate or propionate. These values were about 50% less than those previously measured in snail neurones using CO2. 5. beta i values calculated from the pHi changes due to the removal of 5 mM of the weak bases trimethylamine, procaine and NH4Cl were all similar and comparable to those measured using butyrate or propionate. Removing the influence of pHi regulation on the undershoots after NH4Cl removal was found to decrease the apparent measured values of beta i by 10%. 6. Combining all the data (except the values obtained using CO2 and acetate), and adjusting for the errors due to pHi regulation reducing the sizes of the pHi changes, we found that the mean value for beta i was 10.4 +/- 0.6 mM (+/- S.E.M.) at a mean pHi of 7.36 +/- 0.05. 7. We also investigated the relationship between beta i and pHi using ionophoretic acid injection. By means of step-wise injections, with pHi regulation blocked, we found that at normal pHi levels beta i remained relatively constant. However, at a pHi of less than about 6.8 beta i increased with decreasing pHi.