Determination of extracellular bicarbonate and carbon dioxide concentrations in brain slices using carbonate and pH-selective microelectrodes

摘要

The extracellular pH of the brain is subject to shifts during neural activity. To understand these pH changes, it is necessary to measure [H⁺], [HCO3−],[CO32−] and [CO2]. In principle, this can be accomplished using CO32− and pH-sensitive microelectrodes; however, interference from HCO3− and Cl⁻, and physiological changes in [ HCO3−], complicate measurements with CO32− electrodes. Calibration requires knowledge of slope response, interference constants and corrections for [ HCO3−] shifts. We show that when [ HCO3−] is altered at constant [CO2] in the absence of Cl⁻, the HCO3− interference cancels and the Nikolsky equation reduces to the Nernst equation for CO32−. Measurement of CO32− slope response by this method yielded a value of 28.5 ± 0.72 mV per decade change in [ CO32−]. In Cl⁻-containing solutions, interference coefficients for HCO3− and Cl⁻ were determined by altering [ HCO3−] at constant [CO2], changing [CO2] at constant [ HCO3−], then solving the simultaneous Nikolsky equations for each transition. The mean interference constants corresponded to selectivity ratios of 245:1 and 1150:1 for CO32− over HCO3− and Cl⁻ respectively. To correct for possible changes in [HCO3−], the equilibrium relation between CO32− and HCO3− was substituted into the Nikolsky equation to yield an equation in [CO32−] and [H⁺]. By simultaneously measuring shifts in [H⁺] with a pH microelectrode, this equation is readily solved for [CO32−]. These methods were tested by measuring [HCO3−] and [CO2] in experimental solutions, and in the extracellular fluid of rat hippocampal slices.