Abstract:
Objective: It was designed to execute in vitro expression and functional dection of one subunit (NR
1) of NMDA receptor and to investigate NR
1′s electrophysiology characteristics.
Methods: Primitive NR
1 plasmid DNA from Nakanishi (Japan) was transformed into DH
5α and identified by enzyme Not I cut. Then large scale plasmid DNA was prepared and transcripted into RNA. The obtained RNA was microinjected into matured Xenopus oocytes (50 nl, 1 ng/nl) for receptor expression over 24 h incubation in modified Bath′s solution at a temperature of -19 ℃. The functions of these expressed NR
1 subunit were detected via two electrode voltage clamp technique.
Results: An inward current was induced by 10
-4 mol/L NMDA in Xenopus oocytes microinjected with the obtained RNA transcripted from NR
1 plasmid DNA. The equibrilium potential of the current was -22 mV, which is close to those of chloride ions. This current could be completely inhibited by the specific antagoinst DAP 5 of NMDA receptor and was dependent on glycine (10
-4 mol/L). This current was also blocked by the presence of Mg
2+ , which was inhibited by the replacement of CaCl
2 with MgCl
2 in Ringer′s solution.
Conclusion: The NR
1 subunit of NMDA receptor is functionally in vitro expressed in Xenopus oocytes. Its physiological and pharmacological characteristic is similar to those of whole NMDA receptor.