A voltage-clamp study of isolated stingray horizontal cell non-NMDA excitatory amino acid receptors

Abstract

1. Horizontal cells enzymatically isolated from retinas of the Atlantic stingray (Dasyatis sabina) were voltage-clamped using the patch electrode in the whole-cell mode. A rapid microsuperfusion system was used to apply excitatory amino acid agonists and antagonists. 2. The isolated cells responded to glutamate (GLU), kainate (KA), quisqualate (QA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA). Responses elicited by GLU, QA, and AMPA but not KA exhibited a concentration-dependent and concanavalin A- (Con-A) sensitive desensitization. No responses were elicited by aspartate, N-methyl-D-aspartate, or quinolinate at concentrations as high as 1.0 mM. 3. Judging from the concentration producing one-half of the maximal current response (EC50), the rank order affinities of the agonists was QA greater than or equal to GLU greater than AMPA greater than KA. Whereas KA had the lowest affinity of the agonists tested it was the most efficacious, producing the largest currents. Hill coefficients of the concentration-response data were near two for KA and GLU and near one for QA and AMPA. 4. The agonists differed in their sensitivity to various excitatory amino acid receptor antagonists. Kynurenate (KYN) produced a nearly complete block of horizontal cell responses to GLU and KA at concentrations that had little effect on QA and AMPA. Piperidine-2,3-dicarboxylic acid (cis-PDA), 1-(4-chlorobenzoyl)-piperazine-2,3-dicarboxylic acid (pCB-PzDA), and folic acid were less potent antagonists than KYN but were also better blockers of KA and GLU responses than of QA- and AMPA-elicited responses. 5. When QA, AMPA, or GLU were applied in combination with 55.0 microM KA the current was less than that produced by KA alone. The rank order potency for the inhibition of KA-elicited responses was QA greater than AMPA greater than GLU. In the presence of low concentrations of KA (1.0-20.0 microM), QA- and AMPA-elicited responses were potentiated. This potentiation was prevented by KYN. 6. Single-channel conductance and mean open time were estimated from the current noise fluctuations in the presence of agonist. The mean single-channel conductance for QA was 9 pS that was almost twice as large as the conductance for KA (5.9 pS) and GLU (5.7 pS). The mean open time in the presence of QA or GLU was approximately 1 ms, which was about one-half of that for KA (2.0 ms). 7. These results are best explained by the existence of a single receptor protein with multiple but not identical ligand-binding sites.(ABSTRACT TRUNCATED AT 400 WORDS)