Effects of divalent cations on slow unblock of native NMDA receptors in mouse neocortical pyramidal neurons

Abstract

The N-methyl-d-aspartate receptor (NMDAR) exhibits strong voltage-dependent block by extracellular Mg 2+, which is relieved by sustained depolarization and glutamate binding, and which is central to the function of the NMDAR in synaptic plasticity. Rapid membrane depolarization during agonist application reveals a slow unblock of NMDARs, which has important functional implications, for example in the generation of NMDAR spikes, and in determining the narrow time window for spike-timing-dependent plasticity. However, its mechanism is still unclear. Here, we study unblock of divalent cations in native NMDARs in nucleated patches isolated from mouse cortical layer 2/3 pyramidal neurons. Comparing unblock kinetics of NMDARs in the presence of extracellular Mg 2+or in nominally zero Mg 2+, and with Mn 2+or Co 2+substituting for Mg 2+, we found that the properties of slow unblock were determined by the identity of the blocking metal ion at the binding site, presumably by affecting the operation of a structural link to channel gating. The time course of slow unblock was not affected by zinc, or the zinc chelator TPEN [N,N,N′,N′-tetrakis-(2-pyridylmethyl)-ethylenediamine], while the slower fraction of unblock was reduced by ifenprodil, an NR2B-selective antagonist. Slow unblock was only weakly temperature dependent, speeding up with rise in temperature with a Q 10 of ≈1.5. Finally, using action potential waveform voltage-clamp, we show that this slow relief from divalent cation block is a prominent feature in physiologically realistic patterns of changing membrane potential. © 2011 The Authors. European Journal of Neuroscience © 2011 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.