Selective Targeting of a Defined Subpopulation of Corticospinal Neurons using a Novel Klhl14-Cre Mouse Line Enables Molecular and Anatomical Investigations through Development into Maturity.

Jake Lustig, Alexander Lammers, Julia Kaiser, Payal Patel, Aidan Raghu, James M Conner, Phong Nguyen, Eiman Azim, Vibhu Sahni
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Abstract

The corticospinal tract (CST) facilitates skilled, precise movements, which necessitates that subcerebral projection neurons (SCPN) establish segmentally specific connectivity with brainstem and spinal circuits. Developmental molecular delineation enables prospective identification of corticospinal neurons (CSN) projecting to thoraco-lumbar spinal segments; however, it remains unclear whether other SCPN subpopulations in developing sensorimotor cortex can be prospectively identified in this manner. Such molecular tools could enable investigations of SCPN circuitry with precision and specificity. During development, Kelch-like 14 (Klhl14) is specifically expressed by a specific SCPN subpopulation, CSNBC-lat, that reside in lateral sensorimotor cortex with axonal projections exclusively to bulbar-cervical targets. In this study, we generated Klhl14-T2A-Cre knock-in mice to investigate SCPN that are Klhl14+ during development into maturity. Using conditional anterograde and retrograde labeling, we find that Klhl14-Cre is specifically expressed by CSNBC-lat only at specific developmental time points. We establish conditional viral labeling in Klhl14-T2A-Cre mice as a new approach to reliably investigate CSNBC-lat axon targeting and confirm that this identifies known molecular regulators of CSN axon targeting. Therefore, Klhl14-T2A-Cre mice can be used as a novel tool for identifying molecular regulators of CST axon guidance in a relatively high-throughput manner in vivo. Finally, we demonstrate that intersectional viral labeling enables precise targeting of only Klhl14-Cre+ CSNBC-lat in the adult central nervous system. Together, our results establish that developmental molecular delineation of SCPN subpopulations can be used to selectively and specifically investigate their development, as well as anatomical and functional organization into maturity.

使用一种新的Klhl14-Cre小鼠系选择性靶向皮质脊髓神经元亚群,可以通过发育到成熟进行分子和解剖研究。
皮质脊髓束(CST)促进熟练、精确的运动,这需要脑下投射神经元(SCPN)与脑干和脊髓回路建立特定的节段性连接。发育分子描述使皮质脊髓神经元(CSN)投射到胸腰椎节段的前瞻性鉴定;然而,目前尚不清楚在感觉运动皮层发育中的其他SCPN亚群是否可以用这种方式前瞻性地识别。这种分子工具可以精确和特异性地研究SCPN电路。在发育过程中,kelch样14 (Klhl14)由特定的SCPN亚群CSN BC-lat特异性表达,该亚群位于外侧感觉运动皮层,轴突投射仅针对球颈目标。在这项研究中,我们生成了Klhl14- t2a - cre敲入小鼠,以研究发育成熟过程中Klhl14+的SCPN。通过条件顺行和逆行标记,我们发现Klhl14-Cre仅在特定的发育时间点由CSN BC-lat特异性表达。我们在Klhl14-T2A-Cre小鼠中建立条件病毒标记,作为可靠研究CSN bc - late轴突靶向的新方法,并确认该方法确定了CSN轴突靶向的已知分子调节因子。因此,Klhl14-T2A-Cre小鼠可以作为一种新的工具,以相对高通量的方式在体内鉴定CST轴突引导的分子调节因子。最后,我们证明了交叉病毒标记能够精确靶向成人中枢神经系统中的Klhl14-Cre+ CSN bc - late。总之,我们的研究结果表明,SCPN亚群的发育分子描述可以用于选择性和特异性地研究它们的发育,以及成熟时的解剖和功能组织。意义说明:皮层通过脑下投射神经元(subcerebral projection neurons, SCPN)连接脑干和脊髓靶点,这些神经元在发育过程中根据其新皮层位置和轴突靶点表现出分子多样性。我们创造了一个新的Klhl14-Cre小鼠系来利用这种发育描述并驱动Cre在特定SCPN亚群中的表达。这种发育特异性使得研究1)Klhl14+ SCPN在成熟皮层中的区域位置,2)成熟时它们的轴突侧支,以及3)哪些基因可以控制它们的轴突靶向。使用交叉工具,我们还可以选择性地标记成人中枢神经系统中的这些神经元。因此,SCPN的发育分子描述不仅提供了前瞻性的鉴定,而且可以在发育过程中进行分子分析,以及在成年期进行解剖和功能研究。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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