Department of Bioengineering, University of Colorado - Denver, Anschutz Medical Campus, Aurora, CO 80045, USA.
Department of Bioengineering, University of Colorado - Denver, Anschutz Medical Campus, Aurora, CO 80045, USA; Rocky Mountain Regional VA Medical Center, Aurora, CO 80045, USA.
J Neurosci Methods. 2024 Nov;411:110268. doi: 10.1016/j.jneumeth.2024.110268. Epub 2024 Aug 25.
Delivering optogenetic genes to the peripheral sensory nervous system provides an efficient approach to study and treat neurological disorders and offers the potential to reintroduce sensory feedback to prostheses users and those who have incurred other neuropathies. Adeno-associated viral (AAV) vectors are a common method of gene delivery due to efficiency of gene transfer and minimal toxicity. AAVs are capable of being designed to target specific tissues, with transduction efficacy determined through the combination of serotype and genetic promoter selection, as well as location of vector administration. The dorsal root ganglia (DRGs) are collections of cell bodies of sensory neurons which project from the periphery to the central nervous system (CNS). The anatomical make-up of DRGs make them an ideal injection location to target the somatosensory neurons in the peripheral nervous system (PNS).
Previous studies have detailed methods of direct DRG injection in rats and dorsal horn injection in mice, however, due to the size and anatomical differences between rats and strains of mice, there is only one other published method for AAV injection into murine DRGs for transduction of peripheral sensory neurons using a different methodology.
NEW METHOD/RESULTS: Here, we detail the necessary materials and methods required to inject AAVs into the L3 and L4 DRGs of mice, as well as how to harvest the sciatic nerve and L3/L4 DRGs for analysis. This methodology results in optogenetic expression in both the L3/L4 DRGs and sciatic nerve and can be adapted to inject any DRG.
将光遗传学基因递送到周围感觉神经系统为研究和治疗神经紊乱提供了一种有效的方法，并为假肢使用者和那些遭受其他神经病变的人重新引入感觉反馈提供了可能。腺相关病毒（AAV）载体是一种常见的基因传递方法，因为其具有高效的基因转移和最小的毒性。AAV 可以被设计成靶向特定的组织，通过血清型和遗传启动子选择的结合以及载体给药的位置来确定转导效率。背根神经节（DRG）是感觉神经元细胞体的集合，从周围投射到中枢神经系统（CNS）。DRG 的解剖结构使它们成为靶向周围神经系统（PNS）中的感觉神经元的理想注射位置。
以前的研究已经详细描述了在大鼠中直接 DRG 注射和在小鼠中背角注射的方法，然而，由于大鼠和小鼠品系之间的大小和解剖差异，只有另一种已发表的方法可用于使用不同方法学将 AAV 注射到小鼠的 DRG 中，以转导周围感觉神经元。
新方法/结果：在这里，我们详细介绍了将 AAV 注射到小鼠的 L3 和 L4 DRG 中所需的必要材料和方法，以及如何收获坐骨神经和 L3/L4 DRG 进行分析。这种方法学导致 L3/L4 DRG 和坐骨神经中的光遗传学表达，可以适应于注射任何 DRG。
