Fast excitatory synaptic transmission in the mammalian central nervous system is mediated by glutamate-activated α-amino-5-methyl-3-hydroxy-4-isoxazole propionate (AMPA) receptors. In neurons, AMPA receptors coassemble with transmembrane AMPA receptor regulatory proteins (TARPs). Assembly with TARP γ8 alters the biophysical properties of the receptor, producing resensitization currents in the continued presence of glutamate. Using single-channel recordings, we show that under resensitizing conditions, GluA2 AMPA receptors primarily transition to higher conductance levels, similar to activation of the receptors in the presence of cyclothiazide, which stabilizes the open state. To study the conformation associated with these states, we have used single-molecule FRET and show that this high-conductance state exhibits tighter coupling between subunits in the extracellular parts of the receptor. Furthermore, the dwell times for the transition from the tightly coupled state to the decoupled states correlate to longer open durations of the channels, thus correlating conformation and function at the single-molecule level.

The synaptic proteins cerebellin-1 and neurexin-1β permit ion channel activity in delta subtype of ionotropic glutamate receptors.

Significance: NMDA receptors are the main excitatory receptors in the mammalian central nervous system. They require the binding of both glutamate and glycine for the activation of the protein. However, the binding of one agonist lowers the affinity of the second. The mechanism and physiological significance of this negative cooperativity between the two agonists is currently unknown. Using single molecule FRET and MD simulations we show that binding of one agonist destabilizes the second agonist-binding domain, making it less favorable to bind the second agonist, thus leading to lower affinity. Additionally, there is minimal effect on the transmembrane segments in the presence of a single agonist thus ensuring that the protein is activated only when bound to both agonists.