Distribution Pattern Of Early Cells And Pioneer Fibers In Human Embryonic Forebrain

The forebrain is the processing center of the central nervous system and the critical events of forebrain development happen at early stages including the neuroepithelial(NE)stage when the predecessor neurons give birth,the prepalte(PP)stage when the early projecting neurons generate in the neocorex and the pioneer fibers form in different regions,and the early cortical plate(CP)stage when longitudinal projecting axons start to grow out and to find their proper targets.The present understanding of mammalian forebrain development is largely based on studies of rodents and,to a lesser extent,of other species such as carnivores and macaque monkey.Our recent work in mouse demonstrate that Isl1-positve cells in the forebrain is indispensible for brain wiring by providing the pioneer fibers and functioning as guidepost cells to steer later projecting axon at the PP and early CP stages,and Isl1-positive cells function is highly dependent on the expression of Celsr3 and Fzd3 in those cells.In this study,our scientific question is: Wheter the mechanisms involved in regulating mouse forebrain development are conserved during human forebrain development,such as the distribution pattern of predecessor neurons in the neocortex,pioneer fibers in the forebrain,the interaction of projecting axons with Isl1-positive guidepost cells? For this purpose,we collected human embryos from postconceptual week 5(Wpc5),when the primordium of the forebrain is composed mostly of neuroepithelial progenitors,to Wpc8,the end of the embryonic period when the initial CP appears in the cerebral cortex.Using immunohistochemistry studies,we confirmed the early appearance of predecessor neurons and tangential fibers in the primordium of the neocortex at the NE stage and provided the additional evidence that some of early telencephalic neurons originated from the olfactory placode and migrated tangentially in the telencephalic marginal zone.At Wpc6 and Wpc7,human forebrain maintained the typical identities of the PP stage as described in embryonic day(E)12.5 mouse,including no cortical plate by HE staining,Reelin-positive pioneer cortical neurons(Cajal-Retizius Cells)and the plexiform primodial layer(PPL)beneath the marginal zone,which were further confirmed by Di I tracing studies.Using cell-specific markers,such as PAX6 for the progenitors of the dorsal telencephalon,MASH1 for the progenitors of the ventral telencephalon,TBR2 for the intermediate progenitor cells and TBR1 for postmitotic neurons,we found that they have the similarexpression patterns between human and mouse at the PP stage.In addition,at the PP stage,we provided the evidences that two pools of Isl1-positive guidepost cells located in the ventral telencephalon and the prethalamus;the pioneer fibers crossed the diencephalon-telencephalon junction(DTJ)at both directions prior to the thalamocortical projecting axon arrival and the pioneer fibers from the prethalamus to the dorsal thalamus by Di I tracing studies and immunostaining,which pioneer fibers acted as a bridge as described in mice.At the early CP stage(Wpc8),we found projecting axons from early subplate neurons crossed the pallial-subpallial boundary(PSPB)and innervated the ventral telencephalon,where they followed the stream of Isl1-positive cells.Since it is difficult to directly test the role of Celsr3-and Fzd3-dependent Isl1-positive guidepost cells steering forebrain wiring reported in mice by knockout teniques in human embryos,we studied the expression pattern of CELSR3 and FZD3 m RNA in the human forebrain at Wpc6 and Wpc8,and found their expression was in the overlapping and complimentary manner,similar to that in mouse studies,indicating CELSR3 and FZD3 may play the similar roles in human forebrain development.In conclusions,cortical predecessor neurons originate from both the local proliferation zone and the olfactory placode at the human NE stage;Wpc6-7 human embryonic forebrains display the typical characteristics of cortical development at the PP stage and the cellular distribution pattern of different progenitor cells reported in mouse;Two pools of Isl1-positive cells in the ventral telencephalon and the ventral thalamus and the pioneer fibers across the DTJ also exist in human forebrain at the PP stage;Early corticofugal projecting axons make close contact with Isl1-positive guidepost cells in the ventral telencephalon at the early CP stage in human embryos.Thus,the mechanisms involved in the forebrain wiring including Celsr3-and Fzd3-dependent guidepost cells and the pioneer fibers in mice may be conserved in human forebrain development.