THE REGULATION OF INTRACELLULAR PH STUDIED BY P-31-NMR AND H-1-NMR SPECTROSCOPY IN SUPERFUSED GUINEA-PIG CEREBRAL-CORTEX SLICES

(1) The intracellular pH (pH(i)) of superfused slices of guinea-pig cerebral cortex was measured in P-31-NMR spectra using the chemical shifts of intracellular inorganic phosphate (Pi) and of 2-deoxyglucose 6-phosphate (DOG6P). The pH(i) was found to be 7.30 +/- 0.04 (SD, n = 15) in bicarbonate-buffered medium and 7.20 +/- 0.05 (n = 10, P < 0.001) in bicarbonate-free HEPES buffer of the same pH (7.4). (2) Decreases in pH(e) below 7.05 resulted in pH(i) falling to similar values, with a decrease in the energy state. There was no change in intracellular lactate as assessed by H-1-NMR. (3) The tissues showed an, ability to buffer higher pH: increasing pH(e) to 8.0 had no effect on pH(i), PCr or lactate. (4) In order to characterize possible mechanisms of pH regulation in the tissue, the recovery from acid insult was investigated under various conditions. Initially pH(i) was decreased to 6.44 +/- 0.15 (n = 15) by exposure to media containing 6 mM bicarbonate gassed with O2/CO2, 80: 20 (pH(e) 6.4). When this medium was replaced by normal bicarbonate buffer (pH 7.4) there was full recovery of pH(i) to 7.31 +/- 0.05 (n = 15), whereas replacing the buffer with HEPES resulted in incomplete recovery of pH(i) to 6.88 +/- 0.15 (n = 15, P < 0.001). (5) In the presence of the carbonic anhydrase inhibitor, acetazolamide (1 mM), or the sodium/proton exchange inhibitor, amiloride (1 mM), there was an incomplete return of pH(i) to the control value (pH(i) 6.90 +/- 0.20, n = 5, P < 0.001). (6) There was partial recovery of pH(i) in the presence of an inhibitor of the sodium/bicarbonate co-transporter, 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS), to 7.11 +/- 0.15 (n = 5, P < 0.02). (7) These results indicate that both exchangers, bicarbonate/chloride and sodium/proton, are involved in pH(i) regulation in the intact metabolizing cerebral cortex. The role of the sodium/bicarbonate co-transporter is less clear.