Antoniazzi Aldana M, Unda Santiago R, Norman Sofya, Pomeranz Lisa E, Marongiu Roberta, Stanley Sarah A, Friedman Jeffrey M, Kaplitt Michael G
Laboratory of Molecular Neurosurgery, Department of Neurological Surgery, Weill Cornell Medical College, Cornell University; New York, NY, USA.
Laboratory of Molecular Genetics, Rockefeller University; New York, NY, USA.
bioRxiv. 2025 Mar 19:2025.03.18.644041. doi: 10.1101/2025.03.18.644041.
Primary nociceptors in the dorsal root ganglion (DRG) receive sensory information from discrete parts of the body and are responsible for initiating signaling events that in supraspinal regions will be interpreted as physiological or pathological pain. Genetic, pharmacologic and electric neuromodulation of nociceptor activity in freely moving non-transgenic animals has been shown to be challenging due to many factors including the immunogenicity of non-mammalian proteins, procedure invasiveness and poor temporal precision. Here, we introduce a magnetogenetic strategy that enables remote bidirectional regulation of nociceptor activity. Magnetogenetics utilizes a source of direct magnetic field (DMF) to control neuronal activity in cells that express an anti-ferritin nanobody-TRPV1 receptor fusion protein (Nb-Ft-TRPV1). In our study, AAV2retro-mediated delivery of an excitatory Nb-Ft-TRPV1 construct into the sciatic nerve of wild-type mice resulted in stable long-term transgene expression accompanied by significant reduction of mechanical withdrawal thresholds during DMF exposure, place aversion of the DMF zone and activity changes in the anterior cingulate (ACC) nucleus. Conversely, delivery of an inhibitory variant of the Nb-Ft-TRPV1 construct, engineered to gate chloride ions in response to DMF, led to reversed behavioral manifestations of mechanical allodynia and showed place preference for the DMF zone, suggestive of functional pain relief. Changes in DRG activity were confirmed by post-mortem levels, immediately following DMF exposure, of the activity-induced gene , which increased with the excitatory construct in normal mice and decreased with the inhibitory construct in pain models Our study demonstrates that magnetogenetic channels can achieve long-term expression in the periphery without losing functionality, providing a stable gene therapy system for non-invasive, magnetic field regulation of pain-related neurons for research and potential clinical applications.
背根神经节（DRG）中的初级伤害感受器从身体的离散部位接收感觉信息，并负责启动信号传导事件，这些事件在脊髓上区域将被解释为生理性或病理性疼痛。由于包括非哺乳动物蛋白的免疫原性、手术侵入性和时间精度差等多种因素，在自由活动的非转基因动物中对伤害感受器活动进行基因、药理和电神经调节已被证明具有挑战性。在这里，我们引入了一种磁遗传学策略，能够对伤害感受器活动进行远程双向调节。磁遗传学利用直接磁场（DMF）源来控制表达抗铁蛋白纳米抗体-TRPV1受体融合蛋白（Nb-Ft-TRPV1）的细胞中的神经元活动。在我们的研究中，通过AAV2retro介导将兴奋性Nb-Ft-TRPV1构建体递送至野生型小鼠的坐骨神经，导致稳定的长期转基因表达，同时在DMF暴露期间机械退缩阈值显著降低，并出现对DMF区域的位置厌恶以及前扣带回（ACC）核的活动变化。相反，递送经过工程改造以响应DMF来控制氯离子的抑制性Nb-Ft-TRPV1构建体变体，导致机械性异常性疼痛的行为表现逆转，并显示出对DMF区域的位置偏好，表明功能性疼痛缓解。在DMF暴露后立即通过死后水平确认DRG活动的变化，即活动诱导基因的水平，在正常小鼠中该基因随兴奋性构建体增加，在疼痛模型中随抑制性构建体降低。我们的研究表明，磁遗传学通道可以在外周实现长期表达而不丧失功能，为非侵入性磁场调节疼痛相关神经元提供了一个稳定的基因治疗系统，用于研究和潜在的临床应用。
