Anatomy and Cell Biology, East Carolina University Brody School of Medicine, Greenville, NC, USA.
J Neurosci Res. 2020 Nov;98(11):2148-2165. doi: 10.1002/jnr.24692. Epub 2020 Jul 26.
Excitatory synapse formation begins in mid-fetal gestation. However, due to our inability to image fetal synaptogenesis, the initial formation of synapses remains understudied. The recent development of human fetal brain spheroids provides access to this critical period of synapse formation. Using human neurons and brain spheroids, we address how altered actin regulation impacts the formation of excitatory synapses during fetal brain development. Prior to synapse formation, inhibition of RhoA kinase (ROCK) signaling promotes neurite elongation and branching. In addition to increasing neural complexity, ROCK inhibition increases the length of protrusions along the neurite, ultimately promoting excitatory synapse formation in human cortical brain spheroids. A corresponding increase in Rac1-driven actin polymerization drives this increase in excitatory synaptogenesis. Using STORM super-resolution microscopy, we demonstrate that actomyosin regulators, including the Rac1 regulator, α-PIX, and the RhoA regulator, p115-RhoGEF, localize to nascent excitatory synapses, where they preferentially localize to postsynaptic compartments. These results demonstrate that coordinated RhoGTPase activities underlie the initial formation of excitatory synapses and identify critical cytoskeletal regulators of early synaptogenic events.
兴奋性突触形成始于胎儿中期妊娠。然而，由于我们无法对胎儿突触发生进行成像，突触的初始形成仍未得到充分研究。最近开发的人类胎儿脑类器官为研究这一关键的突触形成时期提供了可能。利用人类神经元和脑类器官，我们研究了肌动蛋白调节的改变如何影响胎儿大脑发育过程中兴奋性突触的形成。在突触形成之前，抑制 ROCK 信号通路可促进轴突伸长和分支。ROCK 抑制不仅增加了神经元的复杂性，还增加了沿着轴突的突起的长度，最终促进了人类皮质脑类器官中兴奋性突触的形成。相应增加 Rac1 驱动的肌动蛋白聚合，从而增加兴奋性突触形成。使用 STORM 超分辨率显微镜，我们证明肌球蛋白调节剂，包括 Rac1 调节剂α-PIX 和 RhoA 调节剂 p115-RhoGEF，定位于新形成的兴奋性突触，优先定位于突触后区室。这些结果表明，协调的 RhoGTPase 活性是兴奋性突触初始形成的基础，并确定了早期突触发生事件的关键细胞骨架调节剂。
