Evaluation of agonist selectivity for the NMDA receptor ion channel in bilayer lipid membranes based on integrated single-channel currents.
A new method for evaluating chemical selectivity of agonists to activate the N-methyl-D-aspartate (NMDA) receptor was presented by using typical agonists NMDA, L-glutamate and (2S, 3R, 4S)-2-(carboxycyclopropyl)glycine (L-CCG-IV) and the mouse epsilon1/zeta1 NMDA receptor incorporated in bilayer lipid membranes (BLMs) as an illustrative example. The method was based on the magnitude of an agonist-induced integrated single-channel current corresponding to the number of total ions passed through the open channel. The very magnitudes of the integrated single-channel currents were compared with the different BLMs as a new measure of agonist selectivity. The epsilon1/zeta1 NMDA receptor was partially purified from Chinese hamster ovary (CHO) cells expressing the epsilon1/zeta1 NMDA receptor and incorporated in BLMs formed by the tip-dip method. The agonist-induced integrated single-channel currents were obtained at 50 microM agonist concentration, where the integrated current for NMDA was shown to reach its saturated value. The obtained integrated currents were found to be (4.5 +/- 0.55) x 10(-13) C/s for NMDA, (5.8 +/- 0.72) x 10(-13) C/s for L-glutamate and (6.6 +/- 0.61) x 10(-13) C/s for L-CCG-IV, respectively. These results suggest that the agonist selectivity in terms of the total ion flux through the single epsilon1/zeta1 NMDA receptor is in the order of L-CCG-IV approximately = L-glutamate > NMDA.
Hirano
A
A
Department of Chemistry, School of Science, The University of Tokyo, Japan.
Sugawara
M
M
Umezawa
Y
Y
Uchino
S
S
Nakajima-Iijima
S
S
D000600:Amino Acids, Dicarboxylic / Q000494:pharmacology; D000818:Animals; D015374:Biosensing Techniques / Q000379:methods*; D016466:CHO Cells; D006224:Cricetinae; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D018698:Glutamic Acid / Q000494:pharmacology; D066298:In Vitro Techniques; D007473:Ion Channels / Q000819:agonists*; D008051:Lipid Bilayers; D051379:Mice; D016202:N-Methylaspartate / Q000494:pharmacology; D016194:Receptors, N-Methyl-D-Aspartate / Q000819:agonists*; D011994:Recombinant Proteins / Q000819:agonists
D000600:Amino Acids, Dicarboxylic; D018690:Excitatory Amino Acid Agonists; D007473:Ion Channels; D008051:Lipid Bilayers; D016194:Receptors, N-Methyl-D-Aspartate; D011994:Recombinant Proteins; C035261:(alpha-carboxycyclopropyl)glycine; D018698:Glutamic Acid; D016202:N-Methylaspartate