Cre recombinase mediated deletion of the NMDA receptor in dopamine neurons

在可卡因致敏方案中，DAT-NR1KO小鼠在VTA内植入一个双侧套管，以输送特定的NMDAR拮抗剂，我们的结果表明，通过在VTA内同时注射NMDAR拮抗剂和外周可卡因，DAT-NR1KO和WT动物的致敏作用可以同样被阻断。我们的实验现在的目的是排除或包括中皮质DA细胞群作为VTA NMDA依赖敏化的中介。我们目前正在对DAT-NR1KO中投射到前额叶皮质的逆行标记神经元进行膜片钳记录，以确定这些多巴胺神经元是否可能表达NMDAR电流，这是由于DAT水平较低，因此缺乏CRE。此外，TH-NR1 KO动物正在生成，如果可行，将用于确定对可卡因敏化的反应，因为在这些KO动物中，所有多巴胺细胞中的NMDAR电流将被消除。当使用TH-NR1动物时，可能的陷阱是NR1缺失的非特异性，因为NMDAR将在所有中儿茶酚胺能脑区被删除。然而，如果TH-NR1KO动物没有发展出可卡因诱导的敏化，这将表明DA中皮层投射可能在敏化的启动中发挥积极作用。

DAT-NR1 KO mice were implanted with a bilateral cannula in the VTA to deliver specific NMDAR antagonists during a cocaine sensitization regimen, and our results have shown that sensitization can be equally blocked in DAT-NR1 KO and WT animals by simultaneous injection of an NMDAR antagonist in the VTA along with peripheral cocaine. Our experiments are now aimed at ruling out or including the mesocortical DA cell population as the mediator of VTA NMDA dependent sensitization. We are currently performing patch-clamp recordings from retrograde labeled neurons projecting to the prefrontal cortex in DAT-NR1 KO to determine if these dopamine neurons, due to low DAT levels and therefore lack of Cre, may express NMDAR currents. In addition, TH-NR1 KO animals are being generated, and if viable, will be used to determine the response to cocaine sensitization, as in these KO animals NMDAR currents will be eliminated in all dopamine cells. Possible pitfalls when using TH-NR1 animals is the unspecificity of the NR1 deletion, as NMDAR will be deleted in all cathecholaminergic brain areas. However, if TH-NR1 KO animals do not develop cocaine induced sensitization it will suggest the likelihood of an active role of the DA mesocortical projection for the initiation of sensitization.