Single-Channel Currents from Recombinant NMDA NR1a/NR2D Receptors Expressed in Xenopus Oocytes

David J. A. Wyllie, Philippe BEhe, Mohammed Nassar, Ralf Schoepfer, David Colquhoun

Abstract

We have investigated the single-channel and whole-cell behaviour of recombinant N-methyl-D-aspartate (NMDA) receptors formed from NR1a and NR2D receptor subunits expressed in Xenopus oocytes. The EC$_{50}$ for apparent steady-state activation of NR1a/NR2D receptors by glutamate was 450 nM, while extracellular Mg$^{2+}$ produced a voltage-dependent block of glutamate responses with an IC$_{50}$ of 440 $\mu $M at -70 mV. At negative holding potentials glutamate-activated NR1a/NR2D single-channel currents, in 0.85 mM external Ca$^{2+}$, had slope conductances of 35 pS for the main level, and 17 pS for the sublevel; direct transitions occurred between these two conductance levels. On average 35 pS events had mean open times of 1.01 $\pm $ 0.04 ms, whereas the mean open times of 17 pS events were consistently longer (1.28 $\pm $ 0.06 ms). In 5 mM external Ca$^{2+}$ the larger conductance level was reduced to 20 pS whereas in Ca$^{2+}$-free solutions it was increased to 50 pS. The frequency of transitions between the main and subconductance levels showed temporal asymmetry: 35-17 pS transitions were more frequent (61%) than 17-35 pS transitions. This asymmetry was not affected by alterations in the external Ca$^{2+}$ concentration (up to 5 mM). In conclusion, the NR1a/NR2D channel is, like NR1a/NR2C, a `low conductance' NMDA channel, but it can be distinguished from NR1a/NR2C channels on the basis of transition asymmetry and differences in the open times of its main and sub-conductance levels.