Pharmacological characterization of extracellular pH transients evoked by selective synaptic and exogenous activation of AMPA, NMDA and GABA A receptors in the rat hippocampal slice. Voipio, J., P. Paalasmaa, T. Taira, and K. Kaila. Department of Biosciences, Division of Animal Physiology, P.O. Box 17, and the Institute of Biomedicine, Department of Physiology, P.O. Box 9, FIN-00014 University of Helsinki, Finland, Fax: +358-0-1917301.
APStracts 2:0099N, 1995.
SUMMARY AND CONCLUSIONS
1. Inhibitors of extracellular carbonic anhydrase (CA o ) offer much promise as diagnostic tools in the study of the synaptic basis of activity-induced alkaline transients in the brain. However, most of the present information related to the effects of CA o blockers in nervous tissue comes from experiments which involve simultaneous synaptic activation of various types of postsynaptic receptor channels. In the present work, double-barreled H + - selective microelectrodes were used to study alkaline shifts in extracellular pH (pH o ) evoked by selective synaptic and pharmacologic activation of glutamate and _-aminobutyric acid (GABA) receptors in the CA1 cell body layer in rat hippocampal slices. Inhibition of CA o was achieved using the poorly- permeant carbonic anhydrase inhibitor, benzolamide (10 [mu]M; applied in the bath solution) or the impermeant macromolecular inhibitor, prontosil-dextran 5000 (PD 5000; applied in microdrops). 2. Alkaline transients that were exclusively attributable to synaptic activation of glutamate receptors were induced by stimulation of Schaffer collaterals in the presence of picrotoxin (PiTX, 100 [mu]M). An enhancement by the CA o inhibitors of these alkaline transients took place at all stimulus frequencies (5-200 Hz) and stimulus train durations (0.5-20 s) examined. 3. Inhibition of CA o enhanced the alkaline transients evoked by selective synaptic activation of [alpha]-amino- 3-hydroxy-5-methylioxazolate-4-propionic acid (AMPA)/kainate receptors in experiments involving stimulation of Schaffer collaterals in the simultaneous presence of PiTX and D-2-amino-5-phosphonopentoate (AP5, 40-80 _M). 4. Alkaline shifts evoked by selective synaptic activation of ¯N -methyl-D- aspartate (NMDA) receptors were enhanced following inhibition of CA o as seen in experiments where Schaffer collaterals were stimulated in the simultaneous presence of PiTX and 6-cyano-2,3-dihydroxy-7-nitroquinoxaline (CNQX, 20-40 _M) in an Mg 2+ -free solution. 5. Benzolamide and PD 5000 also enhanced the alkaline shifts seen upon pressure injection of glutamate, AMPA or NMDA. The glutamate-induced alkaline shifts were inhibited by AP5 + CNQX suggesting that uptake of glutamate did not significantly contribute to their generation. 6. Stimuli applied at 5-10 Hz in stratum radiatum close (within 0.5 mm) to the recording site evoked alkaline shifts which were blocked by CNQX plus AP5. In the continuous presence of the two glutamate antagonists, PiTX-sensitive alkaline transients were observed in response to brief high-frequency (20-100 Hz) trains consisting of 100 stimuli. Upon application of pentobarbital (100 [mu]M), these apparently monosynaptically-evoked GABA A receptor-mediated alkaline transients were evident also at low stimulation frequencies (5-10 Hz). 7. The amplitude of the synaptic GABA A receptor-mediated alkaline shift evoked by a train of 100 stimuli showed a monotonic dependence on frequency within the range examined, 20-100 Hz. At 100 Hz, its amplitude was comparable to that seen in the absence of the glutamate antagonists. That this was not due to a saturation of the postsynaptic acid sink was evident from the pronounced enhancement of the GABAergic alkalosis by pentobarbital which took place at all frequencies of stimulation. 8. The monosynaptically-evoked GABAergic alkaline shifts were enhanced in the presence of GABA B receptor antagonists (2-hydroxy-saclofen or CGP 35348), and blocked by the two CA inhibitors. 9. Pressure injection of GABA and of the selective GABA A agonist, muscimol, produced a fast alkalosis which was blocked by benzolamide and by PD 5000. The GABA B receptor agonist, baclofen, had no effect on pH o . 10. We conclude that activity-induced alkaline transients evoked in the CA1 cell body region result from the activation of three kinds of ligand-gated receptor channels: AMPA, NMDA and GABA A . Each of these mechanisms is likely to provide a variable contribution to alkaline transients associated with various patterns of activity of the Schaffer collaterals. The present observations also demonstrate that the glutamatergic and GABAergic components of an alkaline transient do not sum up in a simple algebraic manner when evoked simultaneously under control conditions.The clearcut profile of action of CA o inhibitors will provide a useful diagnostic criterion in the study of the relative contributions of glutamatergic vs. GABAergic neuronal activity to pH o transients evoked under physiological conditions in brain tissue. 

Received 29 September 1994; accepted in final form 23 March 1995.
APS Manuscript Number J612-4.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 25 April 1995.