Centre for Integrative Physiology, University of Edinburgh School of Biomedical Sciences, Hugh Robson Building, George Square, Edinburgh EH8 9XD, UK.
Neuropharmacology. 2013 Nov;74:119-25. doi: 10.1016/j.neuropharm.2013.01.024. Epub 2013 Feb 10.
MK-801 is a use-dependent NMDA receptor open channel blocker with a very slow off-rate. These properties can be exploited to 'pre-block' a population of NMDARs, such as synaptic ones, enabling the selective activation of a different population, such as extrasynaptic NMDARs. However, the usefulness of this approach is dependent on the stability of MK-801 blockade after washout. We have revisited this issue, and confirm that recovery of NMDAR currents from MK-801 blockade is enhanced by channel opening by NMDA, and find that it is further increased when Mg(2+) is also present. In the presence of Mg(2+), 50% recovery from MK-801 blockade is achieved after 10' of 100 μM NMDA, or 30' of 15 μM NMDA exposure. In Mg(2+)-free medium, NMDA-induced MK-801 dissociation was found to be much slower. Memantine, another PCP-site antagonist, could substitute for Mg(2+) in accelerating the unblock of MK-801 in the presence of NMDA. This suggests a model whereby, upon dissociation from its binding site in the pore, MK-801 is able to re-bind in a process antagonized by Mg(2+) or another PCP-site antagonist. Finally we show that even when all NMDARs are pre-blocked by MK-801, incubation of neurons with 100 μM NMDA in the presence of Mg(2+) for 2.5 h triggers sufficient unblocking to kill >80% of neurons. We conclude that while synaptic MK-801 'pre-block' protocols are useful for pharmacologically assessing synaptic vs. extrasynaptic contributions to NMDAR currents, or studying short-term effects, it is problematic to use this technique to attempt to study the effects of long-term selective extrasynaptic NMDAR activation. This article is part of the Special Issue entitled 'Glutamate Receptor-Dependent Synaptic Plasticity'.
MK-801 是一种具有非常缓慢失活率的 NMDA 受体开放通道阻滞剂。这些特性可用于“预先阻断” NMDA 受体群体，如突触 NMDA 受体，从而选择性地激活另一群体，如突触外 NMDA 受体。然而，这种方法的有效性取决于 MK-801 阻断作用在洗脱后的稳定性。我们重新研究了这个问题，并确认 NMDA 通道的开放可增强 MK-801 阻断后 NMDA 受体电流的恢复，并且发现当存在 Mg2+时，恢复进一步增强。在存在 Mg2+的情况下，100 μM NMDA 作用 10'或 15 μM NMDA 作用 30'后，MK-801 阻断的 50%恢复得以实现。在无 Mg2+的培养基中，发现 NMDA 诱导的 MK-801 解离要慢得多。另一种 PCP 位点拮抗剂美金刚可替代 Mg2+，在 NMDA 存在时加速 MK-801 的去阻断。这表明，在从其在孔中的结合位点解离后，MK-801 能够重新结合，这一过程受到 Mg2+或另一种 PCP 位点拮抗剂的拮抗。最后，我们发现即使所有的 NMDA 受体都被 MK-801 预先阻断，在有 Mg2+存在的情况下，用 100 μM NMDA 孵育神经元 2.5 小时，足以引起足够的去阻断，杀死超过 80%的神经元。我们得出结论，虽然突触 MK-801“预先阻断”方案可用于药理学评估 NMDA 电流的突触与突触外贡献，或研究短期效应，但使用该技术试图研究长期选择性突触外 NMDA 激活的效应存在问题。本文是题为“谷氨酸受体依赖性突触可塑性”的特刊的一部分。
