Mechanisms Regulating Serotonin Clearance In Vivo: Studies Using KO Mice

描述（由申请方提供）：5-羟色胺转运蛋白（5-HTT）通过高亲和力摄取释放的5-羟色胺来调节多巴胺能神经传递。它是许多精神治疗和成瘾药物的关键作用部位。根据个体是否携带5-HTT启动子区多态性（5-HTTLPR）的短变体，5-HTT表达水平在人类中变化。s等位基因携带者表达的5-HTT比长（/）等位基因纯合子少约50%。与/等位基因纯合子相比，s等位基因携带者似乎更容易患上各种精神疾病，包括抑郁症和药物滥用，并且通常对选择性5-羟色胺再摄取抑制剂（SSRIs）治疗有抵抗力。为了研究5-HTT基因型和药物效应之间关系的神经化学基础，我们使用了5-HTT无效突变的小鼠。杂合子5-HTT突变体类似于s等位基因的携带者，因为它们表达的5-HTT比野生型小鼠少50%，并且从细胞外液中清除5-羟色胺的速度比野生型小鼠慢。无效突变体缺乏5-HTT。我们最有趣的发现之一是5-HTT突变小鼠对“摇头丸”（MDMA）和酒精的5-羟色胺清除抑制作用更敏感，其中无效突变体对5-羟色胺清除的抑制作用最大。因此，这些药物抑制血清素清除，但通过5-HTT独立的机制。重要的是，这种机制可能在5-HTT突变体中发生了补偿性上调。这里提出的研究旨在确定这些替代机制的5-羟色胺运输。已知多巴胺和去甲肾上腺素转运蛋白摄取5-羟色胺，但初步数据表明，至少在海马CA 3区，这些转运蛋白在5-HTT突变小鼠的5-羟色胺清除中不起显著作用。相比之下，有机阳离子（OCT）和脯氨酸（PROT）转运体的阻断剂深刻抑制5-HTT突变小鼠的5-羟色胺清除。在这里，我们将确定5-HTT表达的遗传定义缺陷与（1）OCT和PROT表达，（2）OCT和PROT功能和（3）行为之间的关系，以评估SSRI和OCT和PROT阻滞剂在临床前抗抑郁疗效和治疗酒精中毒措施中的有效性变化。我们还将研究OCT和PROT作为抗抑郁药和滥用药物的作用部位，以及它们的疗效如何随5-HTT基因型而变化。除了增加我们对多巴胺能神经传递的基本理解外，这些研究可能会导致更适合个人的情感和成瘾性疾病的新治疗方法。当5-HTT表达受损时，上调5-羟色胺摄取的替代机制的临床意义是深远的。也就是说，针对OCT或PROT的药物可能是对SSRIs治疗反应不佳的个体的一线或连续治疗。

DESCRIPTION (provided by applicant): The serotonin transporter (5-HTT) regulates serotonergic neurotransmission by high affinity uptake of released serotonin. It is a key site of action for many psychotherapeutic and addictive drugs. The level of 5-HTT expression varies in humans according to whether an individual carries the short (s) variant of the 5-HTT promoter region polymorphism (5-HTTLPR). Carriers of the s allele express ~50% fewer 5-HTTs than those homozygous for the long (/) allele. Carriers of the s allele appear to be more prone to a variety of psychiatric disorders, including depression and drug abuse, and are often resistant to treatment with selective serotonin reuptake inhibitors (SSRIs) compared to individuals homozygous for the / allele. To study the neurochemical underpinnings of the relationship between 5-HTT genotype and drug effect, we have made use of mice with a null mutation of the 5-HTT. Heterozygote 5-HTT mutants resemble carriers of the s allele in that they express 50% fewer 5-HTTs and clear serotonin from extracellular fluid more slowly than wild-type mice. The null mutants lack 5-HTTs. One of our most intriguing findings is that 5-HTT mutant mice are more sensitive to the serotonin clearance inhibiting effect of "Ecstasy" (MDMA) and alcohol where greatest inhibition of serotonin clearance occurs in null mutants. Thus, these drugs inhibit serotonin clearance but via a 5-HTT independent mechanism. Importantly, this mechanism has presumably undergone compensatory upregulation in 5-HTT mutants. The studies proposed here seek to identify these alternative mechanisms for serotonin transport. It is known that the dopamine and norepinephrine transporters take up serotonin, but preliminary data indicate that at least in the CA3 region of hippocampus these transporters do not play a significant role in serotonin clearance in 5-HTT mutant mice. By contrast, blockers of the organic cation (OCT) and proline (PROT) transporters profoundly inhibit serotonin clearance in 5-HTT mutant mice. Here we will determine the relationship between genetically defined deficiencies in 5-HTT expression and (1) OCT and PROT expression, (2) OCT and PROT function and (3) behavior to assess changes in effectiveness of SSRIs and blockers of the OCT and PROT in preclinical measures for antidepressant efficacy and treatment of alcoholism. We will also investigate OCT and PROT as sites of action for antidepressant and abused drugs and how their efficacy varies with 5-HTT genotype. In addition to increasing our fundamental understanding of serotonergic neurotransmission, these studies may lead to novel treatments for affective and addictive disorders that are better tailored to the individual. The clinical implications for upregulation of alternative mechanisms for serotonin uptake when 5-HTT expression is compromised are far reaching. That is, drugs targeted at OCTs or PROT may represent first line or adjunctive therapies for individuals who do not respond well to treatment with SSRIs.