Visualizing septin and cell dynamics in mammalian brain slices.

Abstract

Correct neuronal migration is crucial for the brain architecture and function. During brain development, excitatory and inhibitory neurons generated in the ventricular zone (VZ) of the dorsal telencephalon and ganglionic medial eminence, respectively, move to their final destinations in tightly regulated spatiotemporal manners. While a variety of morphological methods have been applied to neurobiology, in utero electroporation (IUE) technique is one of the most powerful tools for rapid gain- and loss-of-function studies of brain development. This method enables us to introduce genes of interest into VZ progenitor and stem cells of rodent embryos, and to observe resulting phenotypes such as proliferation, migration, and cell morphology at later stages. In this chapter, we first summarize basic immunohistochemistry methods that are foundations for any advanced methodsand showed data on the distribution of Sept6, Sept9, and Sept14 as examples. Then, IUE method is described where functional analyses of Sept14 during brain development are used as examples. We subsequently refer to the in vivo electroporation (IVE)-mediated gene transfer, which is conceptually the same method as IUE, into granule cells of hippocampal dentate gyrus in neonatal mice. Finally, an IUE-based time-lapse imaging method is explained as an advanced technique for the analyses of cortical neuron migration. IUE and IVE methods and the application would contribute greatly to the morphological analyses of septins as well as other molecules to elucidate their neuronal functions and pathophysiological roles in various neurological and psychiatric disorders.