During Brain Development Final Destinations for Neurons and Glia Get Separated from Germinal Niches

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1 During Brain Development Final Destinations for Neurons and Glia Get Separated from Germinal Niches

2 Two mayor forms of neuronal migration: Radial and Tangential Leber & Sanes, 95

3 How do young neurons actually move?

4 Modes of Cell Translocation Govek et al, Dev Neurobiol. 2010

5 Migratory Behavior Varies According to Region

6 LGE

7 MGE

8 Radial Migration

9 Radial Glia Guided Migration Takes Young Neurons directly from the VZ to the Cortical Plate Guided by the Processes of Radial Glia; Maintains Topological Maps from thevz to the Cortical Plate Birth Migration Differentiation

10 Sequential Addition of Neurons During Development Desai and McConnell (2000)

11 Radial Glial Fibers as Guides for Neuronal Migration Rakic, 1972

12 Radial Glial Fibers as Guides for Neuronal Migration Rakic, 1972

13 Protomap Hypothesis Rakic, 1988

14 Radial Glia Guided Migration Continues in the Adult Brain of some Vertebrate Species

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16 Initiation, Adhesion, Locomotion, Termination -Very low density lipoprotein R -Low-density lipoprotein R-related protein 8 -Lis1 -Filamin 1 Marin and Rubenstein (2003)

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18 Initiation, Adhesion, Locomotion, Termination -Very low density lipoprotein R -Low-density lipoprotein R-related protein 8 -Lis1 -Filamin 1 Marin and Rubenstein (2003)

19 Reelin!3ß1R VLDLR LRP8 (internalization + P) CNR (Cadherin-related Neuronal Receptor) Cdk5+ P DAB1 (Drosophila Disabled, scrambler, yotari) NUDEL Microtubules Reorganization

20 Gap Junctions The adhesive, not the channel, properties of gap junctions are essential for neural migration (Elias and Kriegstein, 2008)

21 Asymmetrical Cx expression in bifurcated migrating neurons Cx43 Vimentin - radial glia Tuj1 - neurons Cx26 Vimentin - radial glia Tuj1 - neurons

22 Gap junction adhesions in radial migration 1) Gap junctions are necessary for neuronal migration. 2) CX26 and CX43 silencing by RNA-interference disrupts migration 3) Gap junction adhesions, but not channels, rescue the sirna induced migration defect. 4) Gap junctions can provide adhesion in cortical cells and interact with the actin cytoskeleton. 5) Gap junction adhesions act dynamically in migrating neurons to stabilize the leading processes and aid in the translocation of the nucleus. Elias and Kriegstein, Nature, 2008

23 Tangential Migration MGE migration SVZ-OB migration

24 MGE migration

25 Routes on Interneuron Migration from MGE into Cortex

26 MGE

27 Marin and Rubenstein, Nature Neuroscience Reviews, 2001 Tangential Migration of Local Circuit Neurons: GABA & ACh

28 Expression of LacZ by the Dlx5/6-Enhancer in the Embryonic Telencephalon: Shows Migration of Cells From the Subpallium (Subcortex) to the Pallium (Cortex)!"#$ From John

29 !"#$%&'()*+,*-'.+/0'1'2"345'6,'7+,80,9+"'(:8;+93,' %&'"()*&+"$+,-./+0112

30 Ultrasound Guided Transplantation Wichterle et al. Development 2001

31 A Remarkable Journey: MGE progenitors invading cortex in vivo Wichterle et al. Development 2001

32 MGE cells: source of GABAergic Cortical Interneurons Wichterle et al. Development 2001

33 Stereotyped dynamic behavior of tangentially migrating interneurons Martini, F. J. et al. Development 2009;136:41-50

34 Gad65-GFP E12.5, E13.5 Leading process branching is common to many neurons in the developing brain. (A-D) Serial sections through the brain (A,B) and spinal cord (D) of Gad65-Gfp embryos at E12.5 (D) and E13.5 (A,B). In Gad65- Gfp embryos, Gfp expression is restricted to GABAergic neurons, some of which have typical morphologies of migrating cells. The schematic drawinsg in C depict the approximate localization of sections (A,B,D). (E-H) High-magnification images of cells migrating in the developing cortex (E), thalamus (F), midbrain reticular formation (G) and ventral spinal cord (H). All these cells display similar morphologies characterized by the presence of a branched leading process (arrowheads). H, hippocampus; Dh, dorsal horn of the spinal cord; dth, dorsal thalamus; dra, dorsal raphe nucleus; FP, floor plate; GP, globus pallidus; Hyp, hippothalamus;

35 dsred electroporated; Cells following relatively straight paths make branches with relatively small angles. Movie 3. Branch dynamics in migrating neurons following relatively constant trajectories. Interneurons were imaged while migrating through the subpallium after 12 hours of electroporating the MGE of a E13.5 slice with

36 Gad65-GFP; Changing directions involves making new branches and necleokinesis into the angled branch. Movie 4. Branch dynamics in migrating neurons rapidly changing direction. An E16.5

37 Branch dynamics during tangential migration Martini, F. J. et al. Development 2009;136:41-50 Fig. 2. Branch dynamics during tangential migration. (A) A telencephalic slice from an E16.5 Gad65-Gfp embryo. GABAergic cells are observed in green in the different regions in which migration was studied: subpallium (lateral ganglionic eminence, LGE), cortical subventricular zone (SVZ) and cortical plate (CP). Owing to the massive accumulation of migrating neurons in the subpallium, analysis of neurons migrating through the LGE was performed in slice cultures in which the MGE was

38 Branch dynamics during Neuregulin 1 (Nrg1) induced chemotaxis Martini, F. J. et al. Development 2009;136:41-50 Fig. 3. Branch dynamics during Nrg1-induced chemotaxis. (A-A'') Images of a dsred-electroporated slice perfused with a micropipette containing recombinant EGF domain from Nrg1 (13 nm) and Alexa 488 (green channel). To induce drastic changes in direction, cortical interneurons (red channel) migrating through the LGE were confronted with the micropipette at an angle that is perpendicular to their normal trajectory. (B-B') Schematic representation of the trajectory change followed by the cell shown in C. White arrow indicates the micropipette. (C) Representative time-lapse sequences of a migrating cell that developed drastic trajectory changes in response to the chemoattractant. The cell generates a new leading process toward the pipette immediately before changing its trajectory (t=1:35). The angle generated before the most significant change in direction is the largest made by the neuron during this sequence (C). The cell chose the branch oriented towards the chemoattractant to continue migration. The gradient is also visualized in red owing to laser cross-contamination. (C'-C'') Drawings illustrate the morphology of the cell shown in C. Diagrams in C'' depict the movement of this cell. New branches are shown in green; chosen branch is tipped with a red arrowhead. The numbers indicate the angle formed by the branches.

39 Selective Guidance of different sub-populations of MGE cells Marin et al. Science 2001

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41 GABA+ Potassium-Chloride Cotransporter (KCC2) = You Have Arrived Bortone and Polleux, 2009 Neuron

42 Telencephalic Interneurons Multiple interneuron subtypes in cortex, OB and striatum. Most contains GABA (gamma-aminobutyric acid). GABA interneurons can be further subdivided: - Ca++ binding proteins(calbindin, calretinin and parvalbumin) - neuropeptides (eg, Somatostatin, Neuropeptide Y). Not all interneurons are GABAergic: - dopamine - acetylcholine Not all GABA neurons are interneurons.

43 Tangential Migration SVZ-OB migration

44 Routes on Interneuron Migration from MGE into Cortex

45 Lois and Alvarez-Buylla 1993 Lois and Alvarez-Buylla, 1994

46 Chain Migration Lois, García-Verdugo and Alvarez-Buylla, 1996

47 PSA-NCAM!1,!5, ß1, ß6*, ß3*, ϒ1 * adult ephrin-b2 & B3 :: EphB2, EphB3, EphA4

48 The Adult Brain Contains an Extensive Network of Pathways for Chain Migration Wichterle et al. Neuron 1998

49 The Adult Brain Contains an Extensive Network of Pathways for Chain Migration Wichterle et al. Neuron 1998

50 The Adult Brain Contains an Extensive Network of Pathways for Chain Migration cc NC SVZa M RMS Doetsch and Alvarez-Buylla PNAS. 1996

51 B SVZ Astrocyte C Amplifying Precursor A OB Interneurons B1 B2 A C E Doetsch et al, J. Neurosci Doetsch et al, Cell 1999

52 How do Cells Orient Over Such Large Territory?

53 Chemorepulsion guides SVZ neuroblasts? Wu, W., Wong, K., Chen, J., Jiang, Z., Dupuis, S., Wu, J.Y. and Rao, Y. Directional guidance of neuronal migration in the olfactory system by the protein Slit. Nature 400: , Hu, H. Chemorepulsion of neuronal migration by Slit2 in the developing mammalian forebrain. Neuron 23: , 1999.

54 Telling Direction of Migration Retrovirus encoding alkaline phosphatase (AP) Chain of neuroblasts (PSA-NCAM+) (Wichterle et al., 1997)

55 MIA (MIGRATION-INDUCING ACTIVITY) Netrin :: DCC (DELETED IN COLORECTAL CANCER) Slit1 & Slit2

56 MIA (MIGRATION-INDUCING ACTIVITY) NC M RMS Netrin :: DCC (DELETED IN COLORECTAL CANCER) Slit1 & Slit2

57 Map of Cell migration directions in the SVZ cell migration choroid plexus olfactory bulb RMS wall adhesion

58 Mapping the direction of CSF flow

59 Q: Is the CSF flow required for directional cell migration? cell migration cilia beating /CSF flow olfactory bulb choroid plexus site of wall adhesion RMS third ventricle

60 The Tg737orpk mutant has defective ependymal cilia Tg737 +/+ Tg737 orpk/orpk NoelMurcia

61 The Tg737orpk mice have small olfactory bulbs Volume of granule cell layer (mm3) Tg737 +/ Tg737orpk /orpk

62 The Tg737 orpk mutation disrupts normal network formation of migrating neuroblasts in SVZ Tg737 +/+ Tg737 orpk/orpk

63 slit1+/+;slit2+/+ slit1-/-;slit2-/- Slit proteins are required for repulsion of neurblasts by choroid plexus in collagen gel culture CP +/+ SVZ +/+ CP -/- SVZ +/+

64 cell migration olfactory bulb ectopic choroid plexus? choroid plexus Rostral migratory stream third ventricle

65 Ectopic CP GFP+ SVZ cells Ectopic choroid plexus inhibits migration of grafted SVZ cells into the olfactory bulb Control Choroid plexus-grafted Number of GFP+ cells in OB 5 days after transplantation (n=4) (n=5)

66 Slit proteins are partially involved in the repulsive activity of choroid plexus in vivo normal wild-type CP mutant CP

67 Slit2 protein secreted into CSF can reach SVZ and forms a concentration gradient

68 Sawamoto et al, 2006

69 70 Sawamoto et al, 2010

70 Sawamoto et al, 2010

71 Merkle et al, 2008

72 NSCs are: heterogeneous and restricted in potential organized in different domains Merkle et al, 2008

73 NSCs are: heterogeneous and restricted in potential organized in different domains NSC potential is: maintained during postnatal development determined by a cellautonomous mechanism Merkle et al, 2008

74 Sawamoto et al, 2010

75 Text 74 Sawamoto et al, 2010

76 Slit1=GET OUT OF MY WAY! Astrocytes forming migration path of new neurons express Robo receptor New neurons use Slit1 to interact with astrocytes in their path for rapid migration New neurons actively regulate distribution and morphology of astrocytes in their path 75 Sawamoto et al, 2010

77 Why Such a long Migration?

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