Stimulus-induced extracellular pH transients in the in vitro turtle cerebellum

Abstract

In a number of CNS preparations, neuronal activation has been shown to result in a rapid extracellular alkaline transient, followed by a prolonged acid shift. The isolated turtle cerebellum was used to investigate the early alkaline transient. Double-barreled ion-sensitive microelectrodes for H +, K + and tetramethylammonium were used to measure field potentials and extracellular ion and volume shifts in response to bipolar electrical stimulation of the parallel fibers. Transition from 15 mM HEPES to 35 mM HCO 3 − -buffered Ringer decreased the amplitude of the alkaline shift, presumably due to a marked increase in extracellular buffering power. In HEPES-buffered Ringer, repetitive stimulation produced alkaline shifts as large as 0.3–0.4 pH. Single shocks produced an alkaline shift of 0.006 ± 0.0002 pH with a latency as short as 70ms. Kynurenic acid (an excitatory amino acid antagonist), or Mn 2+, blocked the alkaline shift and the postsynaptic component of the field potential. The alkaline shift was not blocked by the Na-H exchange inhibitor amiloride. The relationship between pH o and extracellular volume transients was studied using tetramethylammonium as an extracellular volume indicator. In nominally HCO 3 −free Ringer, stimulation at 5 Hz for 10 s caused a decrease in extracellular volume of 3.0 ± 0.2 per cent. The volume transient was unaffected by 3 mM Mn 2+, while the alkaline shift was completely abolished. The data for the alkaline shift are consistent with a channel-mediated transmembrane flux of proton equivalents. The size of the pH change and the underlying perturbation it represents, indicate that acid-base shifts may be a functionally important consequence of neuronal activity.