Genetic Manipulation of Gyrencephalic Carnivores Using In Utero Electroporation
Higher mammals including primates and carnivores have developed unique brain structures, which are believed to be associated with higher brain functions. However, our molecular understanding of the formation, function and diseases related to these structures is still limited, mainly because genetic manipulations that can be applied to higher mammals had been poorly available. Here we describe a rapid and efficient method that enables in vivo genetic manipulations in the brain of gyrencephalic carnivores using in utero electroporation. Using our method, expression of transgenes becomes detectable within a few days after electroporation and persists for at least 2 months after birth. Our method is useful for expressing transgenes in neural progenitors, superficial and deep cortical post-mitotic neurons, and for examining the morphologies and axonal trajectories of GFP-expressing individual progenitors and neurons in ferrets. Furthermore, multiple genes can be efficiently co-expressed in the same progenitors and neurons. Our method promises to be a powerful tool for investigating the mechanisms underlying the development, function, and pathophysiology of neuronal structures that are unique to higher mammals.
Key wordsFerret Gyrencephalic carnivore Cerebral cortex Higher mammals In utero electroporation Neurons Neural progenitors Outer subventricular zone Inner fiber layer Outer radial glia
We are especially thankful to the late Dr. Lawrence C. Katz for his advice at the initial phase of this project. We are grateful for Drs. Shoji Tsuji, Haruhiko Bito, Takashi Kadowaki, Eisuke Nishida, Yoshiki Sasai, and Shigetada Nakanishi for their continuous encouragement. This work was supported by Grant-in-Aid for Scientific Research from MEXT, PRESTO from JST. This work was also supported by Takeda Science Foundation and Takeda Medical Research Foundation.
- 4.Bernard A, Lubbers LS, Tanis KQ, Luo R, Podtelezhnikov AA, Finney EM, McWhorter MME, Serikawa K, Lemon T, Morgan R, Copeland C, Smith K, Cullen V, Davis-Turak J, Lee C-K, Sunkin SM, Loboda AP, Levine DM, Stone DJ, Hawrylycz MJ, Roberts CJ, Jones AR, Geschwind DH, Lein ES (2012) Transcriptional architecture of the primate neocortex. Neuron 73:1083–1099CrossRefPubMedCentralPubMedGoogle Scholar
- 14.Petros TJ, Rebsam A, Mason CA (2009) In utero and ex vivo electroporation for gene expression in mouse retinal ganglion cells. J Vis Exp 31:e1333Google Scholar