多巴胺在吗啡成瘾记忆诱导的vSUB-NAc通路突触易化中的作用及其分子机制研究

Drug addiction is considered as a pathological form of learning and memory which partly shares synaptic and molecular mechanisms that serve normal learning and memory. Dopamine (DA) and Glutamate have been considered to play key roles in drug addictive memory. D1 receptor activation is believed to selectively enhance surface expression and function of NR2B-NMDAR. Our previous studies found that the increased DA in NAc was paralleled by increased NR2B-NMDAR and facilitated LTP in vSUB-NAc pathway after the consolidation of morphine addictive memory. But whether the DA is involved in the synaptic facilitation remains unclear. Using the Conditioned Place Preference (CPP) as an animal model that mimics associative learning and memory between drug effect and drug-related cues, our study is intended to determine if (1) D1 receptor activation promotes the dissociation of RACK1 from the NR2B-RACK1-Fyn trimolecular complex, thereby facilitating Fyn-mediated phosphorylation of NR2B-Tyr 1472, which enhance synaptic plasticity (2) DA is required for the facilitation of LTP induction in vSUB-NAc pathway.

多巴胺（DA）和谷氨酸是药物成瘾记忆形成的关键递质。研究发现D1受体的激活可通过上调NR2B-NMDAR的数量和功能来增强突触效能。我们的研究发现吗啡成瘾记忆形成后NAc内DA升高，此时腹侧海马下托(vSUB)至伏核(NAc)兴奋性投射的LTP发生易化，但是DA是否参与该LTP易化的发生尚不清楚。本课题欲采用吗啡CPP大鼠模型确认吗啡CPP形成后（1）NAc内升高的DA是否通过激活D1受体促进RACK1与NR2B和Fyn的解离，使Fyn对NR2B亚基Tyr1472位点的磷酸化水平上调，从而稳定NR2B亚基在突触膜表面的表达，使兴奋性突触效能增强（2）D1受体激活是否是vSUB-NAc通路LTP诱导易化的关键。旨在基本阐明NAc内DA对吗啡成瘾记忆诱导的vSUB-NAc通路LTP易化的调节作用及其分子机制。

多巴胺（DA）和谷氨酸是药物成瘾记忆形成的关键递质。研究发现D1受体的激活可通过上调GluN2B-NMDAR的数量和功能来增强突触效能。本课题欲采用吗啡条件性位置偏爱（CPP）大鼠模型来确认吗啡成瘾记忆形成后，（1）腹侧海马下托（vSUB）至伏核（NAc）通路是否发生可塑性变化；（2）伏核的DA是否通过激活D1受体促进RACK1与GluN2B和Fyn的解离，使Fyn对GluN2B亚基Tyr1472位点的磷酸化水平上调，从而稳定GluN2B-NMDAR在神经元膜表面的表达并增强vSUB-NAc通路的功能。结果显示：（1）吗啡CPP测试后取材，在vSUB可以观察到很多FG/NeuN/c-fos阳性的神经元，说明吗啡CPP表达后vSUB-NAc通路被激活。（2）利用化学遗传学特异性失活vSUB-NAc通路，吗啡CPP的表达可被抑制，说明vSUB-NAc通路在吗啡CPP表达中发挥了重要作用。（3）200 Hz电刺激后，吗啡CPP组的LTP幅度明显高于对照组，10 Hz在对照组不能诱导出LTP，仅在吗啡CPP组可以诱导出LTP，提示吗啡CPP表达后vSUB-NAc通路的LTP诱导发生易化。（4）给予NMDAR拮抗剂AP-5或GluN2B-NMDAR的拮抗剂RO可以抑制vSUB-NAc通路LTP诱导的易化，提示LTP易化的产生可能由GluN2B-NMDAR介导。（5）吗啡CPP表达后NAc壳部神经元突触体内GluN2B亚基的蛋白水平升高。（6）吗啡CPP表达后NAc壳部神经元的pY1472 GluN2B、pY416 Src蛋白水平升高而D1受体拮抗剂SCH23390可以翻转这一变化，Fyn和RACK1的蛋白水平没有明显变化。（7）药理学失活一侧vSUB，同时抑制对侧NAc壳部的GluN2B-NMDAR或Fyn可以抑制吗啡CPP的表达。这些结果提示再暴露于吗啡相关环境可易化vSUB-NAc通路的LTP，其机制与NAc壳部GluN2B-NMDAR的激活有关。吗啡CPP表达后DA可能会激活D1受体，使Fyn激酶激活，从而将GluN2B亚基C末端Tyr1472位点磷酸化，稳定GluN2B亚基在神经元膜表面的表达并增强vSUB-NAc突触的功能，介导成瘾记忆的形成。这些结果为成瘾记忆的机制研究提供了新的线索同时为将GluN2B-NMDAR作为开发治疗成瘾药物新靶点的可能性提供依据。