Genetic Approaches To Characterizing Drug Responses And Vulnerabilities: Mice

人与动物在对滥用药物的行为、生理和毒理反应方面存在很大的个体差异。许多个体对药物的行为反应差异显示出很大的遗传成分。转基因动物提供了实现几个相互关联的目标的手段：1）确定特定基因的引入或破坏的生物化学和行为后果; 2）确定在人类研究中鉴定的候选基因的过表达或低表达的后果; 3）阐明产生细胞类型特异性表达和跨突触基因调节的基因元件; 4）研究不同基因座上变异体之间相互作用的影响; 5）阐明体内表达差异的单倍型特异性水平，6）允许我们监测突触连接性及其通过药物给药和细胞粘附和其他重要脑分子中单倍型的改变进行的调节，以及7）允许“去风险”或评估可能伴随靶向其表达被修饰的基因的潜在药物治疗的风险水平。 奖励，强化和学习的兴趣机制导致了今年对这些系统的持续关注，因为我们已经为识别大脑中的布线差异奠定了基础。 在对候选基因的敲除的持续研究中，这些候选基因是有趣的，因为它们编码药物靶点和/或在感兴趣的回路中表达，我们已经报道了关于这些基因敲除对a）可卡因适应性反应B）社会缺陷引起的压力的不同影响的新数据（mu/OPRM 1敲除）c）抗抑郁药和镇痛药（OPRM 1敲除）d）物质诱导的多巴胺释放e）脑回路（DAT/SLC 6A 3敲除）和f）疼痛反应（DAT/SLC 6A 4、SERT/SLC 6A 4和mu/OPRM 1敲除）。 我们已经通过对四种基因表达改变的小鼠的研究，进一步研究了细胞粘附基因改变的影响，我们的人类全基因组关联基因组提供了最一致的复制。我们已经提交了数据，表明我们在人类中有最强证据的两个基因CDH 13、PTPRD和CSMD 1纯合或杂合缺失的小鼠减少了对与10 mg/kg可卡因剂量配对的环境的偏好。这些结果显示特异性，因为可卡因保留运动刺激，因为这些敲除小鼠可以在Morris水迷宫和其他比较行为中以正常（或超常）水平进行。初步数据支持这样的可能性，即在CDH 13敲除中灭绝和恢复的程度也降低。因此，这些动物提供了来自复杂的多基因人类基因座的全基因组关联结果的最佳小鼠确认之一。有趣的是，这些小鼠在5 mg/kg可卡因剂量下也表现出更大的奖励。这些数据与可卡因反应的个体差异和对初始酒精剂量反应的个体差异的人类实验室研究结果一致。 我们已经开始寻找这些行为变化的神经和神经化学相关性。在CDH 13基因敲除小鼠的初步研究中，我们已经确定了大脑皮质多巴胺水平和前额叶皮质多巴胺能纤维密度的变化。如果得到证实，这些结果将为小鼠和人类中伴随CDH 13表达改变的选择性行为结果提供基础。

There are large individual differences among humans and animals in behavioral, physiological and toxicological responses to drugs of abuse. Many of these individual differences in behavioral responses to drugs display substantial genetic components. Transgenic animals provide means for approaching several interrelated goals: 1)Ascertainment of biochemical and behavioral consequences of the introduction of or disruption of specific genes; 2)Ascertainment of the consequences of over- or under-expressing candidate genes identified in human studies; 3) Elucidation of gene elements yielding cell-type specific expression and trans-synaptic gene regulation; 4) Studying influences of interactions between variants at different genomic loci; 5)Elucidating haplotype-specific levels of expression differences in vivo, 6) allowing us to monitor synaptic connectivities and their modulation by drug administration and alterations in haplotypes in cell adhesion and other important brain molecules and 7) allowing "derisking" or assessment of the levels of risk likely to accompany potential pharmacotherapeutics that target the gene whose expression is modified. Interest mechanisms of reward, reinforcement and learning have led to continuing focus on these systems during this year, as we have established much of the groundwork for identification of wiring differences in brain. In continuing studies of knockouts of candidate genes that are interesting because they encode drug targets and/or are expressed in circuits of interest, we have reported novel data concerning the differential influences of these knockouts on a) adaptive responses to cocaine b) stress engendered by social defect (mu/OPRM1 knockouts)c) antidepressants and analgesics (OPRM1 knockouts) d) substance-induced dopamine release (mu/OPRM1 knockouts) e) brain circuitry (DAT/SLC6A3 knockouts) and f) pain responses (DAT/SLC6A4, SERT/SLC6A4 and mu/OPRM1 knockouts). We have moved forward with studies of the influences of cell adhesion gene alterations by studies of mice with altered expression of the four genes for which our human genome wide association datasests provide the most consistent replication. We have submitted data that mice with homozygous or heterozygous deletion of the two genes for which we have the strongest evidence in humans, CDH13,PTPRD and CSMD1, reduce preference for environments paired with 10 mg/kg cocaine doses. These results display specificity, since cocaine retains locomotor stimulation and since these knockout mice can perform at normal (or supranormal) levels in Morris water maze and other comparison behaviors. Initial data supports the possibility that the magnitude of extinction and reinstatement are also reduced in CDH13 knockouts. These animals thus provide one of the best mouse confirmations of a genome wide association result from a complex, polygenic human locus. Interestingly, these mice also display greater reward from 5 mg/kg cocaine doses. These data are convergent with results from human laboratory studies of individual differences in cocaine responses, and individual differences in responses to initial alcohol doses. We have begun to seek neural and neurochemical correlates of these behavioral changes. In initial studies of CDH13 knockout mice, we have identified alterations in levels of cerebral cortical dopamine and in densities of prefrontal cortical dopaminergic fibers. If confirmed, these results would provide a basis for the selective behavioral results that accompany altered expression of CDH13 in mice and in humans.