Relationship between availability of NMDA receptor subunits and their expression at the synapse.
The effect of increasing the expression of NMDA subunits in cerebellar granule cells (CGCs) by transfection was studied to determine how the availability of various NMDA subunits controls both the total pool of functional receptors and the synaptic pool. Overexpression of either NR2A or NR2B, but not splice variants of NR1, by transfection caused a significant increase in the total number of functional NMDA receptors and in surface NR1 subunit cluster density in CGCs in primary culture. These data solidify the central role of NR2 subunit availability in determining the number of cell surface receptors. Overexpression of either NR2A or NR2B significantly altered the deactivation kinetics of NMDA-mediated miniature EPSCs (NMDA-mEPSCs). However, there was no significant effect of NR2 subunit overexpression on the mEPSC amplitude or single-channel conductance. NR2 subunit overexpression did not change the rate of block by MK-801 of NMDA-mediated currents in excised patches from CGCs, indicating that subunit composition does not regulate peak open probability of the channel in CGCs. With the overexpression of a mutant of NR2B lacking the PDZ binding domain, there was an increase in the total number of NMDA receptors without a change in mEPSC kinetics. Therefore, the entry of NMDA receptors into the synapse requires a PDZ binding domain and is limited by means other than receptor subunit availability.
Prybylowski
Kate
K
Laboratory of Neurochemistry, National Institute on Deafness and Other Communication Disorders, National Institutes of Health, Bethesda, Maryland 20892, USA. prybylow@nidcd.nih.gov
Fu
Zhanyan
Z
Losi
Gabriele
G
Hawkins
Lynda M
LM
Luo
JianHong
J
Chang
Kai
K
Wenthold
Robert J
RJ
Vicini
Stefano
S
D000818:Animals; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology; D016291:Dizocilpine Maleate / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D019706:Excitatory Postsynaptic Potentials / Q000502:physiology; D049452:Green Fluorescent Proteins; D007150:Immunohistochemistry; D008164:Luminescent Proteins / Q000235:genetics; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D018408:Patch-Clamp Techniques; D021122:Protein Subunits; D051381:Rats; D017207:Rats, Sprague-Dawley; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013569:Synapses / Q000378:metabolism*; D009435:Synaptic Transmission / Q000502:physiology; D014162:Transfection
D018691:Excitatory Amino Acid Antagonists; D008164:Luminescent Proteins; C409466:NR1 NMDA receptor; C120997:NR2A NMDA receptor; C121001:NR2B NMDA receptor; D021122:Protein Subunits; D016194:Receptors, N-Methyl-D-Aspartate; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D016291:Dizocilpine Maleate