Format of Original
Alcoholism, Clinical & Experimental Research
Original Item ID
The N-methyl-D-aspartate (NMDA) glutamate receptor is a major target of ethanol in the brain. Previous studies have identified positions in the third and fourth membrane-associated (M) domains of the NMDA receptor GluN1 and GluN2A subunits that influence alcohol sensitivity. The structural model of the NMDA receptor, predicted from the structure of the related GluA2 subunit, indicates a close apposition of the alcohol-sensitive positions in M3 and M4 between the two subunit types. We investigated possible interactions between the M3 and M4 domain positions of the two subunit types affecting the ethanol sensitivity of the receptor by using dual substitution mutants. In an initial screen of single-substitution mutants, we found that a position in both subunits adjacent to one previously identified, GluN1(G638) and GluN2A(F636), can strongly regulate ethanol sensitivity. Significant interactions affecting ethanol inhibition were observed at four pairs of positions in GluN1/GluN2A: G638/M823, F639/L824, M818/F636, and L819/F637. Two of these interactions involve a position in M4 of both subunits, GluN1(M818) and GluN2A(L824), that does not by itself alter ethanol sensitivity, and one of the previously identified positions affecting ethanol sensitivity, GluN2A(A825), did not appear to interact with any other position tested. These results also indicate a shift by one position of the predicted alignment of the GluN1 M4 domain. These findings have allowed for the refinement of the NMDA receptor M domain structure, and support the existence of four sites of alcohol action on the NMDA receptor at the M3-M4 domain intersubunit interfaces. These studies were supported by grants R01 AA015203-01A1 and AA015203-06A1 from the NIAAA to R.W.P.