The actions of polyamines on NMDA receptors: Electrophysiological and molecular studies

Abstract

Polyamines such as spermine regulate the activity of many neuronal channels, including both non-NMDA and NMDA glutamate receptors. Because of the fundamental role of NMDA receptors in brain function, it is important to understand the mechanism of action of polyamines. The overall objective of this research was to elucidate the molecular mechanisms underling spermine's action on NMDA receptors using a combination of patch clamp recordings from hippocampal neuronal cultures together with recordings from recombinant NMDA receptors expressed in Xenopus oocytes.;In whole-cell recordings from hippocampal neurons, spermine produced a dose-dependent potentiation of NMDA responses at all potentials {dollar}\rm (EC\sb{lcub}50{rcub}=200\ \mu M){dollar}. In outside-out patches, NMDA channel currents had a conductance of {dollar}\sim{dollar}45 pS. Spermine had two different effects on NMDA channels, potentiation of channel activity and decrease in channel conductance. Potentiation by spermine was due to an increase in open probability of the receptors by two mechanisms; increase in the frequency of channel openings and increase in the open burst duration.;The decrease in channel conductance produced by spermine resembled fast channel block. Spermine reduced the NMDA channel current with an {dollar}\rm EC\sb{lcub}50{rcub} = 278{dollar} at {dollar}-{dollar}60 mV. A Woodhull analysis of voltage-dependence of block indicated deviation from that expected for an open channel blocker, as the inhibitory effect was reduced at potentials negative to {dollar}-{dollar}60 mV, suggesting permeation of spermine. In agreement with this possibility, a simple one-binding site two-barriers permeation model described the effects of spermine on NMDA currents.;Further evidence that spermine interacts with the permeation pore of NMDA receptors was obtained in studies with recombinant NMDA receptors. As with receptors from hippocampal neurons, spermine reduced the conductance of recombinant NR1{dollar}\sb{lcub}100{rcub}{dollar}/NR2A channels. In the NMDA receptor (TM2) channel mutant NR{dollar}\sb{lcub}100{rcub}{dollar}(N598R)/NR2A, which has reduced Mg{dollar}\sp{lcub}2+{rcub}{dollar} block and Ca{dollar}\sp{lcub}2+{rcub}{dollar} permeability, spermine inhibition was reduced.;In summary, spermine interacts with at least two sites on the NMDA receptor. Interaction at one site increases the channel open probability (potentiation of the responses). At a different site, at negative potentials, spermine binds and blocks the channel but can also permeate.