The regulation of intracellular pH studied by 31P- and 1H-NMR spectroscopy in superfused guinea-pig cerebral cortex slices.
(1) The intracellular pH (pHi) of superfused slices of guinea-pig cerebral cortex was measured in 31P-NMR spectra using the chemical shifts of intracellular inorganic phosphate (Pi) and of 2-deoxyglucose 6-phosphate (DOG6P). The pHi 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 pHe below 7.05 resulted in pHi falling to similar values, with a decrease in the energy state. There was no change in intracellular lactate as assessed by 1H-NMR. (3) The tissues showed an ability to buffer higher pH: increasing pHe to 8.0 had no effect on pHi, 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 pHi was decreased to 6.44 +/- 0.15 (n = 15) by exposure to media containing 6 mM bicarbonate gassed with O2/CO2, 80:20 (pHe 6.4). When this medium was replaced by normal bicarbonate buffer (pH 7.4) there was full recovery of pHi to 7.31 +/- 0.05 (n = 15), whereas replacing the buffer with HEPES resulted in incomplete recovery of pHi 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 pHi to the control value (pHi 6.90 +/- 0.20, n = 5, P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)
Brooks
K J
KJ
Division of Biochemistry, U.M.D.S., St. Thomas's Campus, London, U.K.
Bachelard
H S
HS
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000086:Acetazolamide / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism; D002021:Buffers; D002352:Carrier Proteins / Q000378:metabolism; D002540:Cerebral Cortex / Q000378:metabolism*; D019298:Glucose-6-Phosphate / Q000031:analogs & derivatives*; D005958:Glucosephosphates / Q000378:metabolism; D006168:Guinea Pigs; D006859:Hydrogen; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism; D010758:Phosphorus; D027982:Sodium-Bicarbonate Symporters
D001639:Bicarbonates; D002021:Buffers; D002352:Carrier Proteins; D005958:Glucosephosphates; D010710:Phosphates; D027982:Sodium-Bicarbonate Symporters; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D010758:Phosphorus; C015785:2-deoxyglucose-6-phosphate; D019298:Glucose-6-Phosphate; D000584:Amiloride; D006859:Hydrogen; D000086:Acetazolamide; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid