Activation and desensitization of N-methyl-D-aspartate receptors in nucleated outside-out patches from mouse neurones.
1. Activation and desensitization of N-methyl-D-aspartate (NMDA) receptors were studied in large outside-out patches excised from cultured embryonic neurones dissociated from mouse forebrain. The patches were exposed to rapid changes of NMDA or L-glutamate concentrations in the presence of glycine at concentrations (10-20 microM) saturating the modulatory site of the NMDA receptor. 2. Immediately after formation of the patch the responses to NMDA and L-glutamate showed a slow and small desensitization, even with high concentrations of agonist. During the following hour, the peak response either decreased or remained relatively stable, but in all cases the desensitization increased and accelerated until it stabilized. In this 'stabilized' state, the desensitization produced by high concentrations of NMDA (1 mM) or L-glutamate (300 microM) had an exponential time course, with a time constant of about 30 ms. The ratio of the peak over the steady-state current was in the order of 40 for NMDA and about 30 for L-glutamate. 3. Concentration-response curves were built for the peak and the plateau responses, for NMDA and for L-glutamate. The comparison of these curves indicated that (i) the EC50 of the peak (K(app) was always higher than the EC50 of the plateau (Kss); (ii) the two EC50 values for NMDA (K(app) and Kss) were higher than those for L-glutamate; (iii) the Hill coefficient was close to 1.4 for each of the four curves. 4. The application of NMDA or L-glutamate at a low concentration for 3 s periods reduced the response to a subsequent application of the same agonist at a saturating concentration. The IC50 of this 'predesensitization', termed Kpre, was lower than the EC50 of the steady-state response, Kss. 5. The onset rates of desensitization increased with the concentration of agonist. The EC50 of this relation was close to the value of K(app). 6. The decay of the currents at the end of a 3 s application of agonist was usually well described by the sum of two exponentials both of which were faster for NMDA than for L-glutamate. 7. The recovery from desensitization after a long (3 s) pulse of agonist was approximately exponential, with a time constant of about 0.5 s for NMDA and about 3.5 s for L-glutamate.(ABSTRACT TRUNCATED AT 400 WORDS)
Laboratoire de Neurobiologie, Ecole Normale Supérieure, Paris, France.
W
W
Sather
S
S
Dieudonné
J F
JF
MacDonald
P
P
Ascher
D000818:Animals; D002478:Cells, Cultured; D004594:Electrophysiology; D005971:Glutamates / Q000378:metabolism; D018698:Glutamic Acid; D007700:Kinetics; D051379:Mice; D008956:Models, Chemical; D016202:N-Methylaspartate / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism* / Q000502:physiology
D005971:Glutamates; D016194:Receptors, N-Methyl-D-Aspartate; D018698:Glutamic Acid; D016202:N-Methylaspartate