Title

Binding Sites for l‐[3H]Glutamate on Hippocampal Synaptic Membranes: Three Populations Differentially Affected by Chloride and Calcium Ions

Document Type

Journal Article

Publication Date

1-1-1985

Journal

Journal of Neurochemistry

Volume

44

Issue

6

DOI

10.1111/j.1471-4159.1985.tb07170.x

Keywords

Calcium; Chloride; Glutamate; Hippocampus; Neurotransmitter receptors

Abstract

The effects of Cl− and Ca2+ were studied on the specific binding of l‐[3H]glutamate to multiple sites on rat hippocampal synaptic membranes. Quisqualate (5 μM) or dl‐2‐amino‐4‐phosphonobutyrate (2‐APB) (300 μM) was used to discriminate two previously identified classes of binding sites. Saturation isotherms and displacement curves constructed under different ionic conditions suggested that the effects of Cl− and Ca2+ could best be explained by postulating the existence of three major binding site populations in this preparation rather than two. The binding of l‐glutamate to Glu A sites exhibits an absolute dependence on Cl−, and Ca2+ markedly increases the maximum density of these sites. Glu A sites bind quisqualate and 2‐APB with relatively high affinity. Cl− (47 μM) more than doubles the maximum density of Glu B sites, but Ca2+ appears to have no effect. Glu B sites can be discriminated from the other classes by their relatively low affinity for quisqualate and 2‐APB. There is reason to think that the Glu B population is heterogeneous. The novel Glu C population can be virtually selectively labeled by exposing 2‐APB‐sensitive binding sites to radioligand in Tris‐HOAc buffer with Ca2+. Binding of l‐[3H]glutamate to these sites is enhanced by both Cl− and Ca2+, but requires neither ion. Ca2+ appears to increase both the affinity of Glu C sites for l‐glutamate and their maximum binding site density. In the presence of Ca2+ and Cl−, Glu C sites bind the radioligand with micromolar affinity (KD > 2 μM) and high capacity (Bmax > 160 pmol/mg protein). The structural specificity of these binding sites closely resembles that of Glu A sites, but these sites bind excitants with about an order of magnitude lower affinity. The Glu C binding sites exhibit characteristics compatible with a synaptic receptor function in the hippocampal formation. Copyright © 1985, Wiley Blackwell. All rights reserved

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