Characterizing antidepressant-like effects of a novel peptide HCN channel inhibitor

严重抑郁障碍(MDD)影响着全球数百万人。大多数可用于治疗MDD的药物 靶向涉及单胺类神经递质的大脑通路。因为多达50%的患者 没有针对现有疗法的改善，针对新机制的新疗法是 需要的。超极化激活的环核苷酸门控(HCN)通道介导Ih，一个重要的 用于控制神经元兴奋性的电流。大脑HCN通道由一个辅助分子严格调控 TRIP8b亚基(含四肽重复序列的Rab8b相互作用蛋白)。缺乏TRIP8b的动物 和其他HCN亚基表现出抗抑郁剂样的表型，这表明抑制HCN通道 可以有效地治疗抑郁症。因为这些通道是直接控制心率的关键 靶向全身的HCN通道不是一种可行的治疗方法。我们最近 证实TRIP8b在海马区的病毒过表达导致HCN的变化 通道功能伴随着与抗抑郁药使用相关的行为变化。在……里面 特别是，恢复TRIP8b基因敲除小鼠海马区TRIP8b的表达可促进抑郁- 类似的行为，而TRIP8b的一个版本与HCN通道成孔亚基结合受损 抗抑郁药样行为增加。这些结果表明，TRIP8b与HCN的相互作用可能是 治疗MDD的新药理靶点。 尽管我们最近对病毒基因疗法的研究加强了抑制脑内HCN可能 对抑郁症的治疗有用，这种方法需要手术，技术不是很容易 可翻译给患有MDD的人类患者。另一方面，TRIP8b-HCN相互作用的抑制剂提供 提高了在探测HCN通道在行为中的作用方面的易用性，以及更好的可译性 潜在的治疗方法。沿着这些思路，我们已经合成了一种小肽(11个氨基酸)，能够 体外结合TRIP8b并阻断TRIP8b-HCN相互作用。这种多肽的细胞渗透型 合成了SNL-CP，使其能够穿透血脑屏障和细胞膜。在……里面 初步研究表明，SNL-CP能穿过CA1神经元的质膜。我们 假设多肽介导抑制TRIP8b与HCN的相互作用 亚基将促进抗抑郁药样行为。在目标1中，我们将评估SNL-CP的能力 阻断小鼠慢性海马区注射后TRIP8b-HCN的相互作用。这些实验将 确定治疗干预所需的剂量策略。在目标2中，我们将评估 SNL-CP可促进小鼠的抗抑郁药样行为。本提案中描述的工作将评估 一种在小鼠身上产生抗抑郁药样行为的新方法，并可能为新的 治疗对现有疗法无效的人类MDD患者的方法。

Major depressive disorder (MDD) affects millions of people worldwide. Most drugs available to treat MDD target brain pathways involving monoaminergic neurotransmitters. Because up to fifty percent of patients do not improve in response to existing therapies, new treatments that target novel mechanisms are needed. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels mediate Ih, an important current for controlling neuronal excitability. Brain HCN channels are tightly regulated by an auxiliary subunit, TRIP8b (tetratricopeptide repeat-containing Rab8b-interacting protein). Animals lacking TRIP8b and other HCN subunits exhibit an antidepressant-like phenotype, suggesting that inhibiting HCN channels could effectively treat depression. Because the channels are critical in controlling heart rate, directly targeting HCN channels throughout the body is not a viable therapeutic approach. We recently demonstrated that viral overexpression of TRIP8b in the hippocampus leads to changes in HCN channel function with concomitant changes in behaviors associated with antidepressant use. In particular, restoring TRIP8b expression in the hippocampi of TRIP8b knockout mice promoted depression- like behaviors, while a version of TRIP8b with impaired binding to HCN channel pore-forming subunits increased antidepressant-like behavior. These results indicate that the TRIP8b-HCN interaction might be a novel pharmacological target for treating MDD. Although our recent studies with viral gene therapy strengthen the evidence that inhibiting brain HCN might be useful for depression treatment, the approach requires surgery and the technology is not readily translatable to human patients with MDD. On the other hand, inhibitors of the TRIP8b-HCN interaction offer increased ease of use in probing the role of HCN channels in behavior as well as greater translatability as potential therapies. Along theses lines, we have synthesized a small (11 amino acids) peptide capable of binding TRIP8b and blocking the TRIP8b-HCN interaction in vitro. A cell permeable version of this peptide (SNL-CP) was synthesized to allow the peptide to penetrate the blood-brain barrier and cell membranes. In preliminary studies, we show that SNL-CP crosses the plasma membrane of CA1 neurons. We hypothesize that peptide mediated inhibition of TRIP8b's interaction with HCN pore-forming subunits will promote antidepressant-like behavior. In aim 1, we will assess the ability of SNL-CP to disrupt the TRIP8b-HCN interaction after chronic intrahippocampal delivery in mice. These experiments will determine the dosing strategy necessary for therapeutic intervention. In aim 2, we will evaluate the ability of SNL-CP to promote antidepressant-like behavior in mice. The work described in this proposal will evaluate a novel approach to producing antidepressant-like behaviors in mice and could pave the way for a new approach to treating MDD in human patients refractory to existing therapies.