Abstract
Metabolic acidosis (MAc)-an extracellular pH (pH(o)) decrease caused by a [HCO(3) (-)](o) decrease at constant [CO(2)](o)-usually causes intracellular pH (pH(i)) to fall. Here we determine the extent to which the pH(i) decrease depends on the pH(o) decrease vs the concomitant [HCO(3) (-)](o) decrease. We use rapid-mixing to generate out-of-equilibrium CO(2)/HCO(3) (-) solutions in which we stabilize [CO(2)](o) and [HCO(3) (-)](o) while decreasing pH(o) (pure acidosis, pAc), or stabilize [CO(2)](o) and pH(o) while decreasing [HCO(3) (-)](o) (pure metabolic/down, pMet↓). Using the fluorescent dye 2',7'-bis-2-carboxyethyl)-5(and-6)carboxyfluorescein (BCECF) to monitor pH(i) in rat hippocampal neurons in primary culture, we find that-in naïve neurons-the pH(i) decrease caused by MAc is virtually the sum of those caused by pAc (∼70%) + pMet↓ (∼30%). However, if we impose a first challenge (MAc(1), pAc(1), or pMet↓(1)), allow the neurons to recover, and then impose a second challenge (MAc(2), pAc(2), or pMet↓(2)), we find that pAc/pMet↓ additivity breaks down. In a twin-challenge protocol in which challenge #2 is MAc, the pH(o) and [HCO(3) (-)](o) decreases during challenge #1 must be coincident in order to mimic the effects of MAc(1) on MAc(2). Conversely, if challenge #1 is MAc, then the pH(o) and [HCO(3) (-)](o) decreases during challenge #2 must be coincident in order for MAc(1) to produce its physiological effects during the challenge #2 period. We conclude that the history of challenge #1 (MAc(1), pAc(1), or pMet↓(1))-presumably as detected by one or more acid-base sensors-has a major impact on the pH(i) response during challenge #2 (MAc(2), pAc(2), or pMet↓(2)).