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。我们目前正在进行分子研究，以确定这些多巴胺神经元是否可能表达NMDA受体，因为DAT水平较低，因此缺乏CRE。 使用NR1(DATCre)小鼠转基因模型，我们证明，虽然在DA神经元中NMDAR的选择性失活消除了导致突触加强的分子变化的诱导，但在NR1(DATCre)小鼠中，诸如可卡因诱导的运动敏化和条件性位置偏爱等行为措施保持不变。由于投射到前额叶皮质和杏仁核的VTA DA神经元几乎没有表达可检测到的多巴胺转运体，因此推测在NR1(DATCre)小鼠中，投射到这些脑区的DA神经元中的NMDA受体可能是免疫的。我们已经证明，在大多数VTA DA神经元中，NMDA受体基因被消融，包括那些在我们的NR1(DATCre)转基因模型中表现出不可检测的DAT表达水平的神经元，并且在NR1(DATCre)动物的VTA内应用NMDAR拮抗剂仍然可以阻断对可卡因的敏化。这些结果排除了我们的NR1(DATCre)小鼠模型中不同DA神经元亚群中NMDAR介导的神经可塑性的可能性，因此提示VTA内非DA神经元上的NMDAR在可卡因相关成瘾行为中可能起主要作用。

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 molecular studies to determine if these dopamine neurons, due to low DAT levels and therefore lack of Cre, may express NMDA receptors. Using an NR1(DATCre) mouse transgenic model, we demonstrate that while the selective inactivation of NMDARs in DA neurons eliminates the induction of molecular changes leading to synaptic strengthening, behavioral measures such as cocaine induced locomotor sensitization and conditioned place preference remain intact in NR1(DATCre) mice. Since VTA DA neurons projecting to the prefrontal cortex and amygdala express little or no detectable levels of the dopamine transporter, it has been speculated that NMDA receptors in DA neurons projecting to these brain areas may have been spared in NR1(DATCre) mice. We have demonstrated that the NMDA receptor gene is ablated in the majority of VTA DA neurons, including those exhibiting undetectable DAT expression levels in our NR1(DATCre) transgenic model, and that application of an NMDAR antagonist within the VTA of NR1(DATCre) animals still blocks sensitization to cocaine. These results eliminate the possibility of NMDAR mediated neuroplasticity in the different DA neuronal subpopulations in our NR1(DATCre) mouse model and therefore suggest that NMDARs on non-DA neurons within the VTA must play a major role in cocaine-related addictive behavior.