Slow deactivation kinetics of NMDA receptors containing NR1 and NR2D subunits in rat cerebellar Purkinje cells

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Abstract
We have examined the deactivation kinetics of native N-methyl-D-aspartate receptors (NMDARs) containing NR1 and NR2D subunits by patch-clamp recording from Purkinje cells in cerebellar slices from young rats. NMDAR-mediated whole-cell currents were elicited in response to bath application of 20 microM NMDA and 50 microM glycine. The NMDAR-mediated currents were small, with an average whole-cell conductance of approximately 750 pS. Following the rapid application of brief pulses (1-10 ms) of 1 mM glutamate to outside-out membrane patches, we observed a low-conductance type of single-channel activity which lasted up to 30 s after the removal of agonist. Analysis of individual channel openings revealed asymmetry of transitions between the main- and subconductance states - a characteristic of NR1/NR2D-containing NMDARs. The averaged macroscopic current exhibited a decay time course which was well described by a single exponential function with a time constant of approximately 3 s. We conclude that native NR1/NR2D-containing NMDARs, like their recombinant counterparts, display very slow deactivation kinetics. This feature should provide a means for identification of these receptors at synapses, and indicates that they do not contribute to the synaptic NMDAR currents so far described.