L-(³H) glutamate binding to a membrane preparation from the optic lobe of the giant freshwater prawn Macrobrachium rosenbergii de Man

Abstract

Membrane preparation from the optic lobe of the giant freshwater prawn, Macrobrachium rosenbergii de Man, was examined for the presence of specific L-(3H) glutamate binding. The optic lobes were isolated from live animals. The tissue was homogenized and the membrane fraction isolated by differential centrifugation. The membrane suspension was incubated with 10-1,000 nM of L-(3H) glutamate at 37 ° C for 60 min. Nonspecific binding was determined by incubating the mixture with 100 μM L-glutamate. L-(3H) glutamate specifically bound to the membrane fraction with a dissociation equilibrium constant (Kd) of 205 nM and maximum number of binding sites (Bmax) of 2.04 n mol/mg protein. By using LIGAND computerized program, the saturation isotherm binding pattern indicates a single type of binding. To determine the type of glutamate receptors, competitive inhibition and IC50of several glutamate agonists and antagonists were determined. The study reveals a metabotropic type of binding site. © 1991.