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Characterization of the binding of DL-[3H]-2-amino-4-phosphonobutyrate to L-glutamate-sensitive sites on rat brain synaptic membranes.

Abstract：

:The binding of DL-[3,4-3H] 2-amino-4-phosphonobutyric acid DL[3H]-APB to rat whole brain synaptic membranes was investigated. Binding was linear with membrane protein concentration, and optimal at physiological pH and temperature. The association rate was rapid, achieving equilibrium within 10 min. Prolonged incubations (greater than 20 min) revealed additional sites, which apparently possessed identical binding characteristics to those detected with 10 min incubations. Binding of DL-[3H]-APB was enriched in synaptic membrane fractions, and assessment of the regional distribution, indicated greatest binding in those areas with a rich glutamatergic innervation. The binding of DL-[3H]-APB in HEPES-KOH buffer exhibited an absolute requirement for Cl-. The addition of Ca2+ resulted in a further enhancement of binding. Saturation analysis revealed the presence of specific glutamate-sensitive DL-[3H]-APB binding sites, with a KD = 1.26 microM and Bmax = 12.08 pmol mg-1 protein. A Hill plot revealed a slope slightly greater than unity, which could possibly be a reflection of a contribution to binding of a further site which is relatively insensitive to glutamate. Analysis of 60 min incubation data indicated an approximately 3 fold increase in the capacity of the system, but a relatively unchanged KD. Examination of the pharmacological specificity of binding, showed that for both agonist and antagonist molecules, the L-enantiomers were invariably more active than the D-forms. For example, the L-(+)-2-amino-4-phosphonobutyrate isomer was 15 times more active than the D-(-)-form in inhibiting the binding of DL-[3H]-APB. This is in close agreement with the ability of these compounds to produce depression of synaptic transmission. The most potent inhibitor of binding was quisqualate. It is suggested that APB may interact with a quisqualate-preferring class of excitatory amino acid receptors, possibly localised predominantly on presynaptic terminals.

journal_name

Br J Pharmacol

journal_title

British journal of pharmacology

authors

Butcher SP,Collins JF,Roberts PJ

doi

10.1111/j.1476-5381.1983.tb10041.x

subject

Has Abstract

pub_date

1983-10-01 00:00:00

pages

355-64

issue

eissn

0007-1188

issn

1476-5381

journal_volume

pub_type

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Characterization of the binding of DL-[3H]-2-amino-4-phosphonobutyrate to L-glutamate-sensitive sites on rat brain synaptic membranes.