Electrogenic uptake contributes a major component of the depolarizing action of l‐glutamate in rat hippocampal slices

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Abstract
1 A grease-gap technique has been used to measure d.c. potentials, in response to the application of excitatory amino acids and electrical stimulation of the Schaffer collateral-commissural pathway, in the CA1 region of rat hippocampal slices. The actions of l-glutamate (l-Glu) have been quantified and compared to those of structurally related compounds. 2 Perfusion of l-Glu (90s applications) depolarized the tissue with a threshold of approximately 50 μm and a maximum response in excess of 10 mm.l-Aspartate (l-Asp) produced a similar dose-response relationship. By comparison N-methyl-d-aspartate (NMDA) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) were more potent excitants, producing dose-dependent depolarizations over the range 2–50 μm. 3 Application of the agonists depressed the amplitude of electrically-evoked synaptic responses; an effect that presumably reflects depolarization of neuronal tissue. However, for a given agonist-induced d.c. potential, l-Glu or l-Asp caused smaller depressions of synaptic responses than did either NMDA or AMPA. 4 The combined application of 50 μm d-2-amino-5-phosphonopentanoate (AP5) and 10 μm 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) substantially depressed synaptic responses and antagonized responses to NMDA and AMPA producing mean (±s.e.) dose-ratios of 12.2 ± 1.2 and 7.0 ± 0.8, respectively. However, these compounds produced minimal antagonism of responses to l-Glu and l-Asp (dose-ratios of 1.5 ± 0.1 and 1.5 ± 0.2, respectively). 5 Responses to the stereoisomers of homocysteate (HCA) were compared over the range 50 μm to 10 mm.d-HCA was approximately 3.6 times more potent than l-HCA and was antagonized to a greater extent by the combined application of 50 μm AP5 and 10 μm CNQX; the dose ratios were 8.7 ± 0.8 and 5.1 ± 0.9 for the d- and l- isomers, respectively. 6 The application of high doses of an excitant (e.g., 50 mm l-Glu or 5 mm d-HCA) caused an irreversible loss of sensitivity to NMDA and AMPA and abolished synaptic transmission. Responses to the other excitants were depressed by this excitotoxic lesion in the following order: d-HCA > l-HCA > l-Glu = l-Asp. In slices treated in this manner, l-Glu, l-Asp and l-HCA produced very similar dose-response curves. 7 Some slices were unresponsive to NMDA, AMPA and electrical stimulation from the onset of the experiment but had sensitivity to l-Glu, l-Asp and l-HCA similar to that of slices that had received an excitotoxic lesion. 8 Slices that were experimentally lesioned, such that they became unresponsive to synaptic stimulation, AMPA and NMDA, were depolarized by a variety of compounds when tested at 5 mm. The d- and l- enantiomers of Asp and Glu produced similar responses and were slightly more active than quisqualate and threo-hydroxy-l-aspartate (THLA). Glycine, l-serine, d-serine, l-alanine and 4-aminobutanoate (GABA) elicited responses 15–30% of the size, while l-lysine, α-amino-isobutanoate and l-proline produced depolarizations of less than 10% of the size of those induced by 5 mm l-Glu. NMDA and dihydrokainate were either inactive or induced small negative shifts in the d.c. potential. 9 We conclude that when hippocampal slices are perfused with l-Glu (or l-Asp) the main source of the depolarization is due to an interaction with an electrogenic carrier system.