Allosteric modulation of [3H]dizocilpine binding to N-methyl-D-aspartate receptor by an endogenous Na+, K+-ATPase inhibitor: dependence on receptor activation.

An endogenous Na(+), K(+)-ATPase inhibitor, termed endobain E, has been isolated from rat brain and proved to decrease [3H]dizocilpine binding to cerebral cortex N-methyl-D-aspartate (NMDA) receptor, an effect independent of sodium pump activity. The purpose of this study was to disclose the mechanism of [3H]dizocilpine binding reduction by endobain E by performing saturation, kinetic and competitive assays. In saturation binding assays, endobain E increased K(d) without modifying B(max) value. To determine whether competitive or allosteric interaction was involved, kinetics of [3H]dizocilpine binding to cerebral cortex membranes was studied. Endobain E increased [3H]dizocilpine dissociation rate constant and induced an initial fast phase, without modifying association rate constant, indicating an allosteric interaction. In competitive [3H]dizocilpine binding assays, no additive effect was observed with endobain E plus competitive antagonists for glutamate or glycine sites (2-amino-5-phosphonopentanoic acid (AP-5) and 7-chlorokynurenic acid, respectively), indicating that coagonist site blockade interferes with endobain E effect. However, the higher glutamate and glycine concentration, the greater its effect. Endobain E binding reduction was partially additive with that induced by ketamine or Mg(2+) (receptor-associated channel blockers). Results suggest that the greater the channel activation by glutamate and glycine, the greater endobain E allosteric effect. Furthermore, as ketamine and Mg(2+) interfere with endobain E effect, this factor most likely binds to the inner surface of the NMDA associated channel.