title: Electrophysiological characterization of CGP68730A a N-methyl-D-aspartate antagonist acting at the strychnine-insensitive glycine site.
abstract: 1. Electrophysiological experiments were performed in vitro and in vivo. Voltage clamp recordings were done in Xenopus oocytes. Extracellular recordings were done in vitro in the neocortical slice and in the CA1 region of the hippocampal slice and in vivo in the CA1 region of the hippocampus of the anaesthetized rat. 2. In oocytes expressing either the human NMDAR1A/2A or 1A/2B subunit combinations, CGP68730A [sodium (-)-9-bromo-2,3,6,7-tetrahydro-5,6-dioxo-5H-pyrazino[1,2,3-de]-1,4-benzo thiazine-3-acetic acid] antagonized L-glutamate / glycine induced currents with calculated IC50s of 20.5 and 81.6 nM, respectively. 3. In vitro, CGP68730A was tested on NMDA induced depolarizations in the neocortical slice preparation and on epileptiform activity in hippocampal slices bathed in Mg2+-free-medium, which is known to be NMDA mediated. In both in vitro models CGP68730A exhibited antagonistic effects on the NMDA receptor mediated responses. 4. In vivo CGP68730A was tested on NMDA induced excitations in the CA1 region. CGP68730A abolished NMDA induced excitations when applied microiontophoretically. However, only weak effects on NMDA induced excitation were observed after systemic administration at 100 mg/kg i.v.. These results indicate that CGP68730A has poor central nervous system bioavailability. 5. In oocytes, an increase in the glycine concentration from the EC80 to the EC95.99 shifted the inhibition curves for CGP68730A to the right. Furthermore, in neocortical slices and in anaesthetized rats CGP68730A inhibited NMDA mediated depolarizations, and this effect could be reversed by the addition of the glycine mimetic D-serine. This indicates that these effects of CGP68730A are mediated by an action on the strychnine-insensitive glycine site. 6. Selectivity tests in oocytes and in the neocortical slice preparation, using NMDA, kainate and AMPA showed that CGP68730A was selective in antagonizing NMDA receptor mediated responses. In oocytes, the compound was about 1000 times less potent on the rat GluR3 and the human GluR6 receptors than on the human NMDAR1A/2A subunit combination. In the neocortical slice preparationCGP68730A had no effects on AMPA or kainate induced depolarizations at concentrations of 3 and 10 microM. At 30 microM CGP68730A reduced the effects of each of the three agonists tested. 7. Thus, CGP68730A seems to be a selective antagonist at the strychnine-insensitive glycine coagonist site of the NMDA receptor. However, the compound showed no obvious central NMDA antagonistic effects following intravenous application.
authors.affiliation: Novartis Pharma AG, NS Research, Neuromodulation, Basel, Switzerland. mario_f.pozza@pharma.novartis.com
authors.forename: M F
authors.initials: MF
authors.lastname: Pozza
authors.forename: K
authors.initials: K
authors.lastname: Zimmerman
authors.forename: S
authors.initials: S
authors.lastname: Bischoff
authors.forename: K
authors.initials: K
authors.lastname: Lingenhöhl
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000502:physiology; D004594:Electrophysiology; D006624:Hippocampus / Q000187:drug effects / Q000502:physiology*; D016202:N-Methylaspartate / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D019579:Neocortex / Q000187:drug effects / Q000502:physiology; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D018408:Patch-Clamp Techniques; D016194:Receptors, N-Methyl-D-Aspartate / Q000187:drug effects / Q000502:physiology*; D013331:Strychnine / Q000494:pharmacology; D014981:Xenopus
chemical_list: D016194:Receptors, N-Methyl-D-Aspartate; D016202:N-Methylaspartate; D013331:Strychnine