L-methionine counteracts cocaine-conditioned place preference in mice through Glutamatergic plasticity pathways

Drug-induced alterations in gene expression play an important role in the development of addictive behavior. Methionine has been proven to inhibit addictive behaviors of cocaine dependence. However, the mechanisms underlying how methionine use corresponds to drug-induced behaviors still remain unclear. We performed mRNA and miRNA high-throughput sequencing of the prefrontal cortex in a mouse model of cocaine CPP combined with L-methionine in order to identify L-methionine target miRNAs and genes that participate in the cocaine conditioned place preference (CPP). We found that the L-methionine inhibits cocaine CPP. Sequencing data analysis showed that L-methionine down-regulates genes enriched in the Glutamatergic Synapse pathways and significantly reversed the cocaine-induced expression changes of the substance dependence pathways (Morphine addiction and Nicotine addiction) and the neurotransmitter synapse pathways (Glutamatergic Synapse, Cholinergic Synapse and GABAergic Synapse). Furthermore, the Glutamatergic synapse was either overlapped between DEGs with DEGs-miRNA induced by cocaine CPP, or with the MET effects on cocaine CPP. Nineteen targeted genes were investigated and five were identified (Gria4, Grid1, Grik4, Grik5 and Grin3a) to belong to iGluR family. Interestingly, there were several miRNAs that had the same sequence which targets the iGluR family: Mmu-miR-30e-50p and mmu-miR-380-5p share UUGAC motif and targets Grik4; mmu-miR-6940-3p and mmu-miR-212-5p both share UGGCU motif which targets Gria4 and Grid1 respectively. Thus, we demonstrated the efficacy of L-methionine in counteracting the effects of cocaine CPP and identified specific genes of synaptic plasticity pathways, especially the Glutamatergic synapse pathway, which is modulated by L-methionine in response to cocaine dependence. Overall design: PFC region of cocaine-conditioned place preference mice and methionine treated cocaine-conditioned place preference mice were performed mRNA and miRNA sequenceing

药物诱导的基因表达改变在成瘾行为的发生发展中发挥关键作用。蛋氨酸已被证实可抑制可卡因依赖相关的成瘾行为。然而，蛋氨酸干预对应药物诱导成瘾行为的潜在分子机制仍未明确。本研究针对可卡因条件性位置偏好（conditioned place preference, CPP）小鼠模型联合L-蛋氨酸干预，对其前额叶皮层（prefrontal cortex, PFC）开展信使RNA（messenger RNA, mRNA）与微小RNA（microRNA, miRNA）高通量测序，以期筛选参与可卡因CPP的L-蛋氨酸靶向miRNA及靶基因。
研究结果显示，L-蛋氨酸可抑制可卡因CPP。测序数据分析表明，L-蛋氨酸下调了富集于谷氨酸能突触通路的基因，并显著逆转了可卡因诱导的物质依赖通路（吗啡成瘾、尼古丁成瘾）以及神经递质突触通路（谷氨酸能突触、胆碱能突触、γ-氨基丁酸能突触）的表达变化。此外，差异表达基因（differentially expressed genes, DEGs）所富集的谷氨酸能突触通路，既与可卡因CPP诱导的差异表达miRNA靶基因存在重叠，也与L-蛋氨酸对可卡因CPP的调控效应相关。本研究共筛选出19个候选靶基因，其中5个属于离子型谷氨酸受体（ionotropic glutamate receptor, iGluR）家族，分别为Gria4、Grid1、Grik4、Grik5及Grin3a。值得关注的是，部分miRNA拥有靶向iGluR家族的保守基序：Mmu-miR-30e-50p与mmu-miR-380-5p共享UUGAC基序，共同靶向Grik4；mmu-miR-6940-3p与mmu-miR-212-5p共享UGGCU基序，分别靶向Gria4与Grid1。
综上，本研究证实了L-蛋氨酸可拮抗可卡因CPP的生物学效应，并筛选出受L-蛋氨酸调控的突触可塑性通路相关特异性基因，尤其是谷氨酸能突触通路——该通路在可卡因依赖进程中受L-蛋氨酸的调控。整体实验设计：本研究对可卡因条件性位置偏好小鼠、L-蛋氨酸干预的可卡因条件性位置偏好小鼠的前额叶皮层组织进行了mRNA与miRNA测序。