The chloride cotransporter NKCC1 in the embryonic etiology and treatment of FASD

摘要 虽然怀孕期间禁欲是可以接受的，但母亲的乙醇消费是一个重要的公共健康问题。 这是一个令人担忧的问题，因为相当比例的孕妇饮酒;有些人甚至狂饮到危险水平。 怀孕期间摄入乙醇会导致胎儿酒精谱系障碍（FASD），其特征是生命- 后代长期神经行为和认知异常。事实上，那些有过 在子宫内暴露于甚至中等水平的乙醇中， 神经行为和认知结果。然而，FASD/FAS缺乏有效的靶向治疗策略或存在缺陷。 定义了推动这项研究项目的首要动力是更好地了解细胞和 需要FASD相关异常的分子基础。为此，我们将测试一种新的机械 乙醇破坏胚胎神经元中氯稳态的假设，并提出评估治疗 靶向氯共转运蛋白的潜力，以开发和推进药理学 子宫内FASD的干预。 具体目的1：检验用其拮抗剂布美他尼阻断NKCC 1的功能可预防 和/或挽救乙醇对迁移的胚胎GABA能皮层中间神经元中氯稳态的作用 并减轻它们由于子宫内乙醇暴露而引起的异常切向迁移。 具体目标2：检验布美他尼治疗预防和/或挽救受损mPFC的假设。 在子宫内暴露于乙醇的年轻成年小鼠的依赖性行为灵活性和注意力能力。 具体目标3：检验布美他尼治疗预防和/或挽救（1）改变的数量 和/或GABA能皮质中间神经元的分布和/或（2）异常的GABA能和谷氨酸能 在子宫内暴露于乙醇的年轻成年小鼠的mPFC中的神经传递。 我们先前的工作证实GABA促进GABA能皮层中间神经元的切向迁移， 并且乙醇会影响这一过程，导致“神经元间病变”， 皮质形式、突触功能和神经行为。然而，这种神经间病变的机制是 不知道。这三个具体目标将填补这一知识空白，并在这样做的过程中，促进我们对 FASD的病因学。总体而言，NKCC 1将首次作为一个关键的统一机制进行调查 子宫内乙醇暴露对切线迁移、氯化物稳态和长时程的不利影响， 长期神经行为和认知结果，以及作为FASD潜在治疗的靶向治疗。在这 轻，这一建议解决了紧迫的问题，在FASD是非常重要的，采取多层次的 神经解剖学、电生理学和行为学方法，具有创新性、影响力， 为FASD的治疗管理提供了线索。

ABSTRACT Preventable as it is by abstinence during pregnancy, maternal ethanol consumption is a significant public health concern because a significant percentage of pregnant women drink; some even binge drink to risky levels. Consumption of ethanol during pregnancy can lead to fetal alcohol spectrum disorders (FASD), hallmarked by life- long neurobehavioral and cognitive abnormalities in the offspring. Indeed, children with history of having been exposed in utero to even moderate levels of ethanol frequently present with varying degrees of deleterious neurobehavioral and cognitive outcomes. Yet, effective targeted treatment strategies for FASD/FAS are lacking or ill defined. The overarching impetus driving this research project is that a better understanding of the cellular and molecular underpinnings of FASD-associated abnormalities is needed. To this end, we will test a novel mechanistic hypothesis that ethanol disrupts chloride homeostasis in embryonic neurons, and propose to assess the therapeutic potential of targeting chloride co-transporters in order to develop and advance strategies for the pharmacological intervention of FASD in utero. Specific Aim 1: Test the hypothesis that blocking the function of NKCC1 with its antagonist bumetanide prevents and/or rescues ethanol’s effect on chloride homoeostasis in migrating embryonic GABAergic cortical interneurons and mitigates their abnormal tangential migration due to ethanol exposure in utero. Specific Aim 2: Test the hypothesis that bumetanide treatment prevents and/or rescues the impaired mPFC- dependent behavioral flexibility and attentional capacity in young adult mice exposed in utero to ethanol. Specific Aim 3: Test the hypothesis that bumetanide treatment prevents and/or rescues the (1) altered number and/or distribution of GABAergic cortical interneurons and/or (2) abnormal GABAergic and glutamatergic neurotransmission in the mPFC of young adult mice exposed in utero to ethanol. Our previous work established that GABA promotes the tangential migration of GABAergic cortical interneurons, and that ethanol affects this process to result in an “interneuronopathy” with enduring adverse consequences on cortical form, synaptic function and neurobehavior. However, the mechanism underlying this interneuronopathy is not known. The three specific aims will fill this knowledge gap and, in so doing, advance our understanding of the etiology underlying FASD. Overall, for the first time, NKCC1 will be investigated as a key unifying mechanism underlying the untoward effects of ethanol exposure in utero on tangential migration, chloride homeostasis and long- term neurobehavioral and cognitive outcomes, and pharmacologically targeted as potential therapy for FASD. In this light, this proposal addresses pressing questions in FASD that are highly significant, taking a multi-level neuroanatomical, electrophysiological and behavioral approach that is innovative, impactful, with great potential for shedding light on the therapeutic management of FASD.