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Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, CT 06510.

1. We have measured intracellular pH (pHi) in freshly isolated pyramidal neurones from the CA1 region of the rat hippocampus using the fluorescent indicator 2',7'-bis(carboxy-ethyl)-5-(and-6)-carboxyfluorescein (BCECF). 2. The neurones selected by our isolation procedure, when studied in the nominal absence of CO2-HCO3-, had a mean steady-state pHi of 6.81 +/- 0.02 (n = 163). The recovery of pHi from acid loads was very slow. The rate of recovery from acid loads was reduced by Na+ removal, but only very slightly inhibited by 1 mM amiloride. 3. The addition of 5% CO2-25 mM HCO3- caused steady-state pHi to increase from 6.74 +/- 0.05 to 7.03 +/- 0.03 (n = 28). In the presence of 5% CO2-25 mM HCO3-, the rate of pHi recovery from acid loads was much faster than in its absence. 4. The HCO(3-)-induced alkalinization was reversible, and did not occur in the absence of extracellular Na+ or in the presence of DIDS (4,4'-diisothiocyanatostilbene- 2,2'-disulphonic acid). 5. In the absence of external Cl-, successive exposures to CO2-HCO3- elicited alkalinizations that were progressively reduced in rate and amplitude. This effect, presumably due to gradual depletion of internal Cl-, was rapidly reversed by returning Cl- to the external medium. 6. We conclude that the major acid-extrusion mechanism in pyramidal CA1 neurones is the Na(+)-dependent Cl(-)-HCO3- exchanger. The Na(+)-dependent mechanism that operates in the nominal absence of HCO3- is far less active.

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