神经发生的分子机制

1. 神经发生的分子机制 景乃禾 中国科学院上海生命科学研究院 生物化学与细胞生物学研究所

2. 人脑的构成

3. 神经元的种类

4. 中枢神经系统 (脑＋脊髓) 细胞数量：1012 (1万亿) 其中： 神经细胞（神经元）1011 神经胶质细胞：9 X 1011 细胞种类：神经元 (Neuron，许多种类) 神经胶质细胞 (Glia) 星型胶质细胞 (Astrocyte) 少突神经胶质细胞 (Oligodendrocyte) 细胞联系：神经元－神经元 神经元－神经胶质细胞 1012 X (102-103)=1014-1015

5. 人脑的发育 (Gilbert, 1991)

6. 中枢神经系统发育的基本过程 一、神经系统的诱导 (Neural Induction) 主要研究：早期胚胎的神经外胚层(神经干细胞，Neural Stem Cell)是如何产生的？多潜能干细胞是如何分化为神经干细胞的？ 二、神经系统的发生 (Neurogenesis) 主要研究：神经干细胞是如何分化为各种神经元和神经胶质细胞的？ 三、神经联系的建立 (Axon Guidance, Synapse Formation) 主要研究：神经细胞是如何与其靶细胞建立神经联系的？其中包括：轴突的靶向生长和突触联系的建立。 四、神经系统的可塑性 (Neural Plasticity, Adult Neural Stem Cell) 主要研究：成年动物神经系统的可塑性和神经系统损伤后的修复；成体神经干细胞分化的机制。

7. 神经诱导 (Neural Induction) 多潜能干细胞 神经干细胞

8. Major Steps in Neural Differentiation Competence: Cells have the ability to become neural precursors if they are exposed to the right combination of signals. Specification: Cells have received the signals to become neural precursor cells but will still respond to signals that repress a neural character (not fully committed). Commitment: Cells have received the signals to become neural precursor cells and will progress to become neurons even in the presence of signals that repress a neural character. Differentiation:Neural precursor cells exit the cell cycle to become post-mitotic neurons.

9. History of Neural Induction Hypothesis • Spemann Organizer (1924-1990) • Default Model (1992-1997) • Neural Induction in Chick (2001) • Neural induction in Mouse (2007)

10. Induction of Embryonic Primordia by Implantation of Organizers from a Different Species Hans Spemann and Hilde Mangold Arch. Mikr. Anat. Entw. Mech. 100, 599-638, 1924

11. Classical Transplantation Experiment by Spemann and Mangold The donor tissues could recruit the host cells to become the secondary neural tube. Dorsal blastopore lip (Hemmati-Brivanlou & Melton, 1997)

12. “Spemann Organizer” Spemann named the dorsal blastopore lip the “organizer”, and proposed that in normal development this region induces and organizes a correctly patterned nervous system in neighboring dorsal ectoderm. In the absence of this influence, as on the ventral side, the ectoderm differentiates as epidermis. Epidermis: “Default” fate for gastrula ectoderm Neural specification: needs a positive signal from neighboring cells (Neural Induction). “Default”: Cell autonomous.

13. This hypothesis dominated the developmental biology field for several decades. A considerable effort over several decades failed to identify the gene products responsible for neural induction in the embryo.

14. “Default Model” BMP inhibitors: Noggin, Chordin, Follistatin Wilson & Edlund, 2001

15. “Default Model” (Hemmati-Brivanlou & Melton, 1997)

16. Chordin Noggin Follistatin BMP4 Ectoderm Neural The “default model” in Xenopus Question: How do these BMP inhibitors antagonize BMPs’ function? Stern, Development, 2005

17. Default Model in Chick and Mouse Early Development • Questions unsolved: • In Foxa2 (HNF3b) KO mice, there is no node, but the embryos have the neural tissues (Node = Organizer in Xenopus). • Neural induction is initiated before gastulation. • BMP antagonists are not required for neural induction.

18. Gastrulation in Chicken Embryo

19. The status of Wnt signaling regulates neural and epidermal fates in the chick embryo Nature, 2001, 411, 325-330 Wilson et al.,

20. Summary of Experiments

21. An unifying mechanism of “neural induction” ? FGF, WNT and BMP play important roles in neuralization of amniote embryos (humans, rodents and birds) • Question: • How does FGF induce neural? • What about BMP inhibitors? Wilson et al., Nature Neurosci., 2001

22. Neural induction in chick embryos ---Embryologist’s view Stage XIII-2 Stage XI-XII Stage 3+-4 End of Stage 4 ???

23. Neural induction in chick embryos ---Genetic cascade

24. Models of neural induction Xenopus Chick

25. Default Model in Chick and Mouse Early Development Questions unsolved: Why Xenopus and Chick or Mouse have used different mechanisms for neural induction?

26. Neural induction in mouse embryos

27. Early mouse development Preimplantation

28. Early mouse development E3.5 ICM E4.5

29. Early mouse development Postimplantation (early) Epiblast (late) Epiblast (Primitive ectoderm) VE: visceral endoderm AVE: anterior VE DVE: distal VE Anterior neuroectoderm

30. Pluripotent cell lineages in mouse embryo E3.0 E3.5 E4.5 E5.5 Niwa, Development, 2007

31. Neuroectodermal fate of epiblast cells in the distal region of the mouse egg cylinder: implication for body plan organization during early embryogenesis Development 121, 87-98 (1995)

32. Fate-mapping of the distal cap epiblast by carbocyanine dye labeling E6.5 Dye injection

33. The distal cap epiblast cells migrate to the anterior neuroectoderm

34. Cell movements in early mouse embryos Epiblast to posterior Extraembryonic ectoderm to posterior epiblast Epiblast to posterior Distal VE to AVE

35. 6.25 dpc 6.0 dpc 5.75 dpc 5.5 dpc Otx2 VEcis-lacZ Otx2 KI-lacZ Movement of Otx2-positive cells from DVE to AVE DVE: distal visceral endoderm; AVE: anterior visceral endoderm

36. Cell movements in early mouse embryos

37. Cell movements in early mouse embryos

38. Early post-implantation development in the mouse Nature Rev Gen, 8, 368, 2007

39. Mouse Gastrulation and Germ Layer Formation Cell 132, 661–680, 2008

40. Default Model in Chick and Mouse Early Development • Questions unsolved: • In Foxa2 (HNF3b) KO mice, there is no node, but the embryos have the neural tissues (Node = Organizer in Xenopus). • Neural induction is initiated before gastulation. • BMP antagonists are not required for neural induction. Why do Xenopus and Chick or Mouse use different mechanisms for neural induction???

41. New findings BMP signaling inhibits premature neural differentiation in the mouse embryo Development 134, 3359-3369 (2007)

42. BMP-Smads Signaling Pathway

43. BMPR1a is essential for BMP signaling in the early mouse embryo pSmad1/5/8: BMP pathway activated

44. Premature neural differentiation of the epiblast occurs in BMPR1a-/- embryo WT Pluripotent markers: Oct4, Nanog and Fgf5 Bmpr1a-/- Neural stem cell markers: Six3, Hesx1 and Sox1 WT Bmpr1a-/-

45. Suppression of mesoderm in BMPR1a-/- mouse embryo E6.5 Mesoderm markers and mesoderm-inducing signals: FGF8, Eomes, T, Nodal, Cripto, Wnt3 Note: Ectopic neural differentiation occurred in the same embryo

46. BMP signaling is required in the epiblast for mesoderm specification and to inhibit neural differentiation E5.5 E7.5 E6.5 WT Bmpr1a epiblast-specific KO at E6.5 WT Bmpr1a epiblast-specific KO at E6.5 E6.5 E7.5

47. Inhibition of FGF signaling does not block neural specification in BMPR1a-/- mouse embryo Hesx1 (neural marker) E5.5 E6.5 Epiblast KO Control Control Epiblast KO Bmpr1a-/- FGFs are not acting as direct neural inducers in the early post-implantation mouse embryo.

48. Model for BMPs maintain epiblast pluripotency in mouse Node • BMP signaling is required to inhibit epiblast neural differentiation • BMP2/4 signal via Bmpr1a to maintain epiblastpluripotency

49. Signaling centers and molecules implicated in neural induction AVE: Anterior visceral endoderm; MGO: Mid-gastrula organizer; EE: Extra-embryonic region; PS: Primitive streak

50. Tissues implicated in mouse neural induction Signaling factors Necessary Sufficient Correct time/place No Yes Lefty, Cerberus AVE No Node Yes No No Chordin, Noggin GO Yes Yes Yes Chordin