title: Voltage-dependent interaction of open-channel blocking molecules with gating of NMDA receptors in rat cortical neurons.
abstract: 1. The mechanisms by which four adamantane derivatives (IEM-1857, -1592, -1460 and -1754) block the open NMDA-activated channel were studied at membrane voltages (Vm) from -170 to +30 mV. The rate constants of channel block (k+) and of channel unblock (k-) were measured from the fully resolvable flicker of single-channel currents induced by each compound. 2. The k+ of each compound exhibited a similar exponential dependence on voltage over the Vm range studied. 3. The k- of IEM-1857 and IEM-1592 over the Vm range studied, and of IEM-1754 and IEM-1460 from -30 to -90 mV, exhibited similar exponential dependencies on voltage. However, the k- of IEM-1754 and IEM-1460 at Vm values more hyperpolarized than -90 mV were much more steeply voltage dependent, suggesting that at these Vm values the two drugs can occupy a deeper binding site. 4. Each of the drugs induced a concentration-dependent prolongation of the mean burst length at -90 mV, suggesting that while blocking they can interfere with channel closure. 5. The prolongation of mean burst length induced by the largest drug (IEM-1857) increased with hyperpolarization. The increase was consistent at each Vm with the predictions of the sequential scheme of block, suggesting that channel closure is prevented when IEM-1857 is bound. The prolongation of burst length induced by the smallest drug (IEM-1754) was less than predicted by the sequential scheme and the deviation increased with hyperpolarization. 6. The IEM-1857 concentration-dependence of number of blockages per unit open time had a slope equal to k+ at -150 mV. The IEM-1754 concentration-dependence of number of blockages per unit open time revealed a slope about two times less than k+ for this compound at -150 mV. 7. The mean patch current was not significantly altered by 3 microM IEM-1857 at Vm values from -90 to -150 mV, as expected of a drug that prevents channel closure when blocking. Mean patch current significantly decreased with hyperpolarization beyond -90 mV in the presence of 1 microM IEM-1754. 8. The data suggest that there are two blocking sites at different depths within the NMDA-activated channel. Channel closure is prevented when any of the IEM drugs occupy the shallow blocking site. Channel closure is permitted during occupation of a deeper blocking site that can be reached only by the smaller IEM drugs at hyperpolarized voltages.
authors.affiliation: Department of Neuroscience, University of Pittsburgh, PA 15260, USA. S. M. Antonov: antonov@bns.pitt.edu
authors.forename: S M
authors.initials: SM
authors.lastname: Antonov
authors.forename: J W
authors.initials: JW
authors.lastname: Johnson
mesh_terms: D000218:Adamantane / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001369:Axons / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004594:Electrophysiology; D015640:Ion Channel Gating / Q000502:physiology*; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D009474:Neurons / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D018408:Patch-Clamp Techniques; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors*
chemical_list: C092754:IEM 1754; C118520:IEM 1857; D000644:Quaternary Ammonium Compounds; D016194:Receptors, N-Methyl-D-Aspartate; D008274:Magnesium; D000218:Adamantane