Cellular Migration and Formation of Neuronal Connections: Comprehensive Developmental Neuroscience

Comprehensive Developmental Neuroscience: Cellular Migration and Formation of Neuronal Connections

Editors-in-Chief

Professor John L.R. Rubenstein

Department of Psychiatry, University of California at San Francisco, San Francisco, CA, USA

Professor Pasko Rakic

Duberg Professor of Neurobiology and Neurology, Director Kavli Institute for Neuroscience, Yale University School of Medicine, New Haven, CT, USA

Table of Contents

Cover image

Title page

Copyright

Editors-in-Chief

Section Editors

Contributors

Introduction to Comprehensive Developmental Neuroscience

I: Formation of Axons and Dendrites

Chapter 1. Development of Neuronal Polarity In Vivo

1.1 Introduction

1.2 Axon Initiation In Vitro Versus In Vivo

1.3 Distinction Between Cues Regulating Axon Specification Versus Axon Growth

1.4 Extracellular Cues Regulating Neuronal Polarization and Axon Initiation

1.5 Intracellular Pathways Underlying Neuronal Polarization

References

Chapter 2. Role of the Cytoskeleton and Membrane Trafficking in Axon–Dendrite Morphogenesis

2.1 Introduction

2.2 Developmental Stages

2.3 Role of Cytoskeleton in Establishment of Neuronal Polarity

2.4 The Role of (Membrane) Trafficking During Neuronal Polarization

2.5 Maintaining Neuronal Polarity

2.6 Future Work on Neuronal Morphogenesis

References

Chapter 3. Axon Growth and Branching

3.1 Introduction

3.2 Cell Biological Mechanisms

3.3 Extracellular Regulation During Development

3.4 Intracellular Signaling Mechanisms

3.5 Concluding Remarks

References

Chapter 4. Axon Guidance: Semaphorin/Neuropilin/Plexin Signaling

4.1 Introduction: General Features of Axon Guidance

4.2 Discovery of the Semaphorin, Neuropilin, and Plexin Families

4.3 The Class 3 Secreted Semaphorins in Vertebrates: Neuropilin/Plexin Receptors and Signal Transduction Cascades

4.4 Generating Diversity of Axon Responses to Sema3’s

4.5 Uncovering the in vivo Contribution of Sema3’s to Axon Guidance: Insights from Animal Models

4.6 Sema3 Signaling and Sensorimotor Projections

4.7 Sema3 Signaling and Connections of the Cerebral Cortex

References

Chapter 5. Roles of Eph–Ephrin Signaling in Axon Guidance

5.1 Introduction

5.2 Eph–Ephrin Signaling Is Essential for Axon Guidance in Many Contexts

5.3 Modes of Eph–Ephrin Signaling in Axon Guidance

5.4 Eph–Ephrin Signaling in Invertebrate Nervous Systems

5.5 Eph Signaling in Axon Regeneration

Acknowledgments

References

Chapter 6. Axon Guidance: Slit–Robo Signaling

6.1 Introduction

6.2 Slits and Their Receptors

6.3 Slit–Robo Function in Midline Crossing

6.4 Modulation of Slit–Robo Signaling

6.5 Signaling Downstream of Robo

6.6 Beyond the Midline: Additional Roles for Slit–Robo in the Nervous System

6.7 Slit–Robo Contributions to Axon Targeting in a Complex Target Field

6.8 Involvement of Slit–Robo in Disorders of the Nervous System

6.9 Slit–Robo: Players Outside the Nervous System

6.10 Conclusion

References

Chapter 7. Nonconventional Axon Guidance Cues

7.1 Introduction

7.2 Morphogens

7.3 Morphogens in Axon Guidance

7.4 Conclusion and Perspectives

Acknowledgments

Glossary

References

Chapter 8. Axon Regeneration

8.1 Introduction

8.2 Inhibitors of CNS Axon Regeneration

8.3 The Physiological Function of CNS Regeneration Inhibitors and Their Receptors

8.4 Therapeutic Implications and Future Directions

8.5 Conclusions

See also

Acknowledgments

References

Chapter 9. Axon Maintenance and Degeneration

9.1 Introduction

9.2 Essential of Axonal Transport in Axon Maintenance

9.3 Proteasome and Autophagy Pathways in Axon Homeostasis

9.4 Role of Glial Cells in Axon Maintenance

9.5 Maintaining Axon Track Positions

9.6 Axon Pruning and Axon Degeneration

References

Chapter 10. Dendrite Development: Invertebrates

10.1 Structure and Anatomy of Invertebrate Dendrites

10.2 Methods for Manipulating and Studying Dendrite Morphology in Drosophila

10.3 Anatomical Background for Key Model Systems in Which Dendritic Morphogenesis Is Studied in Invertebrates

10.4 Cell Biology of Dendritic Growth

10.5 Transcriptional Control of Dendritic Morphology

10.6 Posttranscriptional Control of Dendritic Development

10.7 Control of Dendritic Field Formation I: Guidance and Targeting

10.8 Control of Dendritic Field Formation II: Dendritic Self-Avoidance and Tiling

10.9 Dendritic Remodeling

See also

References

Chapter 11. Dendritic Development: Vertebrates

11.1 Introduction

11.2 Molecular and Cellular Control of Dendrite Development

11.3 Regulation of Growth Direction

11.4 Regulation of Extension and Branching

11.5 Local Adaptation and Rapid Turnover of Dendrites to Meet Synaptic Partners

11.6 Emergence of Dendritic Geometry and Structural Specializations for Integration and Computation

11.7 Dendrite Development and Neurodevelopmental Disorders

11.8 Conclusion

Acknowledgments

References

II: Migration

Chapter 12. Cell Polarity and Initiation of Migration

Abbreviations

12.1 Introduction

12.2 Migratory Behaviors During Radial Migration in the Developing Cerebral Cortex

12.3 Molecular Mechanisms That Regulate the Initiation of Migration and Cell Polarity During Migration

12.4 Conclusion

See also

Glossary

References

Chapter 13. Leading Process Dynamics During Neuronal Migration

13.1 Introduction

13.2 Diverse Leading Process Dynamics During Neuronal Migration

13.3 Cytoskeleton Dynamics in Migrating Neurons

13.4 General Perspective

Acknowledgments

References

Chapter 14. Nucleokinesis

14.1 Nucleokinesis: Overview

14.2 The Nucleus

14.3 Interactions Between the Nucleus and the Cytoskeleton

14.4 The Cytoskeleton

14.5 Cell Polarity

14.6 Conclusions and Future Directions

Acknowledgments

References

Chapter 15. Migration in the Cerebellum

15.1 Migration of Cerebellar Neurons in the Developing Cerebellum

15.2 Regulation of Granule Cell Migration by Calcium Signaling and Neuropeptides

15.3 Alcohol Impairs Granule Cell Migration

References

Chapter 16. Radial Migration in the Developing Cerebral Cortex

16.1 Introduction

16.2 Production of Cortical Projection Neurons

16.3 Organization of the Neocortex

16.4 Trajectory of Migrating Neurons in the Developing Brain

16.5 Modes of Migration

16.6 Radial Migration in the Developing Human Neocortex

16.7 Factors that Regulate the Radial Migration of Cortical Neurons

16.8 Summary

References

Chapter 17. Radial Migration of Neurons in the Cerebral Cortex

17.1 Overview of Neuronal Migration During Cerebral Cortical Development

17.2 Patterns and Cellular Mechanisms of Radial Migration

17.3 Signaling Mechanisms Regulating Radial Migration

17.4 Radial Neuronal Migration and Human Neurodevelopmental Disorders

References

Chapter 18. Migration in the Hippocampus

18.1 Overview of Hippocampal Structure and Lamination

18.2 Developmental Specification of Hippocampal Fields

18.3 Migration of Cajal–Retzius Cells in the Hippocampus

18.4 Migration of Hippocampal Pyramidal Neurons

18.5 Migration of Hippocampal Interneurons

18.6 Migration of Neural Progenitors and Granule Cells in the Dentate Gyrus During Development

18.7 Conclusions

References

Chapter 19. Hindbrain Tangential Migration

19.1 Introduction

19.2 Tangential Migration: A Historical Overview

19.3 Molecular Mechanisms Controlling the Tangential Migration of Precerebellar Neurons

19.4 Molecular Mechanisms Controlling the Tangential Migration of Facial Motor Neurons

19.5 Ending Tangential Migration

19.6 Conclusion

Acknowledgments

References

Chapter 20. Tangential Migration: The Forebrain

20.1 Introduction

20.2 The Origin and Trajectory of Tangentially Migrating Cells in the Forebrain

20.3 Mechanisms That Guide Tangential Migration

20.4 Switch from Tangential to Radial Migration

20.5 Migration Defects and Disease: Lissencephaly

20.6 Future Directions

References

Chapter 21. Transcriptional Regulation of Tangential Neuronal Migration in the Vertebrate Hindbrain

21.1 Introduction

21.2 Breaking Boundaries: Transcriptional Regulation of Tangential Neuronal Migration Across Rhombomeres

21.3 Conclusions and Perspectives

Acknowledgments

References

Chapter 22. Postnatal Neurogenesis of the Forebrain

22.1 Introduction

22.2 Migration of Neuroblasts: Cell Autonomous Versus Extrinsic Factors

22.3 Radial Migration of Adult-Born Neurons and Their Integration

22.4 Hijacking Migratory RMS Neuroblasts

22.5 Outlooks

Acknowledgments

References

Chapter 23. Migration of Myelin-Forming Cells in the CNS

23.1 Introduction

23.2 Migratory Paths Followed by Oligodendrocyte Progenitor and Precursor Cells

23.3 Chemokinetic Factors: The Motility of Oligodendrocyte Precursors

23.4 Adhesion and Chemotactic Mechanisms: How the Movement of Oligodendrocyte Precursors Is Guided

23.5 Concluding Remarks

Acknowledgments

References

Chapter 24. Neuronal Migration and Brain Patterning

24.1 Cortical Development

24.2 The Borders of the Developing Pallium and the Generation of Tangentially Migrating Glutamatergic Neurons

24.3 CR Subtypes and Extrinsic Control in Regionalization of the Cerebral Cortex

24.4 CP Neurons and Their Role in Corticogenesis

24.5 Other Migrating Cells Generated at the Borders and Brain Patterning: The Example of Neural Crest Cells

24.6 Growth and Cell Fate Specification in Brain Patterning: Migration and Long-Range Patterning

24.7 Progenitor Domains, Tangential Migration of Glutamatergic Neurons and Cortical Evolution

24.8 Conclusions

See also

Acknowledgments

References

Chapter 25. Neuronal Migration of Guidepost Cells

25.1 An Introduction to Guidepost Cells

25.2 Role of Neuronal Migration in the Formation of the LOT

25.3 Hippocampal Cajal–Retzius Cells in the Formation of Axonal Connections

25.4 Migration of Neuronal Guidepost Cells in the Formation of Thalamocortical Connections

25.5 Neuronal Migration of Guidepost Cells in the Formation of the Corpus Callosum

25.6 Neuronal Migration of Guidepost Cells and Evolution of Brain Wiring

25.7 Toward an Integration of Migrating Guidepost Neurons in Normal and Pathological Brain Development

25.8 Conclusions

References

Chapter 26. Neuronal Migration Disorders

26.1 Introduction

26.2 Types of Malformations in NMD

26.3 Type 1 Lissencephaly

26.4 Mutations in Tubulin Subunits TUBA1A, TUBB2B, and TUBA8

26.5 Subcortical Band Heterotopia

26.6 Periventricular Heterotopia

26.7 Polymicrogyria

26.8 Cobblestone Cortical Malformation

26.9 Focal Cortical Dysplasia

26.10 Summary

Acknowledgments

References

III: Synaptogenesis

Chapter 27. Molecular Composition of Developing Glutamatergic Synapses

27.1 Introduction

27.2 Molecular Composition of Glutamatergic Synapses

27.3 Cellular Mechanisms of Transport of Molecules to Developing Synapses

27.4 Final Thoughts

Acknowledgments

References

Chapter 28. In Vivo Imaging of Synaptogenesis

28.1 Introduction

28.2 In Vivo Analysis of Synapse Development in the Neuromuscular Junction

28.3 Small Vertebrate Model Systems: Xenopus and Zebrafish

28.4 Visualizing Synaptogenesis in Mammals

28.5 Future perspectives

References

Chapter 29. Genetic Analysis of Synaptogenesis

29.1 Introduction

29.2 Studying Synaptogenesis in Genetic Model Organisms

29.3 Genetic and Molecular Tools for Large-Scale Genetic Screens

29.4 Perspective

References

Relevant Websites

Chapter 30. Activity-Regulated Genes and Synaptic Plasticity

30.1 The Role of Activity in Circuit Formation

30.2 Signaling from the Synapse to the Nucleus

30.3 Activity-Dependent Gene Expression

30.4 Activity-Regulated Genes that Modulate Synaptic Strength

30.5 Activity-Regulated Genes that Act in Synapse Addition and Elimination

30.6 Other Activity-Regulated Effector Genes

30.7 Posttranscriptional Regulation

30.8 Conclusions

References

Chapter 31. New Imaging Tools to Study Synaptogenesis

31.1 Introduction

31.2 Conventional Fluorescence Microscopy and Electron Microscopy

31.3 Recent Advances in Fluorescence Microscopy: Toward Super-resolution Microscopy

31.4 Recent Advances in Electron Microscopy: 3D Sectioning Techniques and Correlative Microscopy

31.5 New Developments in Genetically Encoded Photosensitive Tools

31.6 New Physical Tools to Culture Neurons

31.7 Conclusion

References

Chapter 32. Wnt Signaling

Abbreviations

32.1 Introduction

32.2 Wnts and Their Signaling Pathways

32.3 Regulation of Presynaptic Differentiation

32.4 Regulation of Postsynaptic Organization

32.5 Wnt Proteins as Antisynaptogenic Factors

32.6 Wnt Signaling and Activity-Mediated Synaptic Remodeling

Glossary

Acknowledgments

References

Relevant Websites

Chapter 33. Neurotrophins and Synaptogenesis

33.1 Introduction

33.2 Neurotrophins and the Integration of Circuits

33.3 Neurotrophin Signaling

33.4 Translation of Neurotrophic Effects into Synaptogenesis

33.5 Effects of Neurotrophins on the Inhibitory/Excitatory Balance

33.6 Specificity of Neurotrophin Actions

33.7 Conclusion and Perspectives

References

Chapter 34. Ephrins and Eph Receptors – Synaptogenesis and Synaptic Function

34.1 Introduction

34.2 Eph Forward Signaling in Synapse Formation and Spine Morphogenesis

34.3 Ephrin Reverse Signaling in Spine and Synapse Formation

34.4 Eph–Ephrin Signaling in Synaptic Plasticity

34.5 The Actions of Presynaptic Ephrins and Postsynaptic Ephs

34.6 The Actions of Presynaptic Ephs and Postsynaptic Ephrins

34.7 Modulation of Spine Morphology and Synaptic Plasticity by Eph–Ephrin Signaling at the Neuron–Glia Interphase

34.8 Regulation of Neuromuscular Junction Function by Eph–Ephrin Signaling

34.9 Conclusions and Perspectives

References

Chapter 35. Neuroligins and Neurexins

Abbreviations

35.1 Introduction

35.2 Along Came a Spider: Discovery of NRX and NL Proteins

35.3 Extensive Diversification: Gene and Protein Structures

35.4 Crystal Clear: Structural Insights into NL–NRX Interactions

35.5 Dynamic Modifications: Regulation of Alternative Splicing

35.6 Nonexclusive Partners: Complexes with Additional Synaptic Proteins

35.7 Acting Locally: Polarized Transport and Synapse-Specific Localization

35.8 More than Glue: Synaptic Functions of NL–NRX Complexes

35.9 From Synapses to Behavior: Disruption of the NRX–NL Complex in Neurodevelopmental Disorders

35.10 Twenty Years and on: Outlook

References

Chapter 36. Circuit Assembly in the Developing Vertebrate Retina

36.1 Introduction

36.2 Retinal Synaptic Laminae

36.3 Axonal and Dendritic Development

36.4 Synaptogenesis

36.5 Functional Assembly

Acknowledgments

References

Chapter 37. Synaptogenesis in the Adult CNS – Neocortical Plasticity

37.1 Neuronal Structural Plasticity in the Adult?

37.2 Lessons from In Vivo Structural Imaging in the Adult Neocortex

37.3 Molecular Aspects of Synapse Formation in the Adult

37.4 Conclusion

References

Chapter 38. Synaptogenesis in the Adult CNS – Hippocampus

38.1 Overview of the Hippocampus and Adult Neurogenesis in the Dentate Gyrus

38.2 Synaptogenesis in the Adult Hippocampus – Newborn Granule Cells

38.3 Synaptogenesis in the Adult Hippocampus – CA1 and CA3 Pyramidal Cells and Other Neurons of the Hippocampus

38.4 Conclusions

See also

Acknowledgments

References

Chapter 39. Synaptogenesis in the Adult CNS–Olfactory System

39.1 Introduction

39.2 Olfaction – A Central Sense Driving Behavior

39.3 Basic Architecture of the OB

39.4 Synaptogenesis in the Adult OB

39.5 Mechanisms of Synaptogenesis in the Adult OB

39.6 Future Perspective

Acknowledgments

References

Chapter 40. Synaptogenesis and Recovery from Cortical Trauma

40.1 Introduction

40.2 Functional Consequences of Cortical Damage

40.3 Axonal Sprouting

40.4 New Synapse Formation and Ultrastructural Changes After Cortical Injury

40.5 Functional Significance of Postinjury Synaptogenesis

40.6 Early Postlesion Events Leading to Initiation of Axonal Sprouting and Synaptogenesis

40.7 Role of Neuron–Astrocyte and Neuron–Glia Signaling in Cortical Injury

40.8 Regulation of Neurite Outgrowth

40.9 Arborization: Presynaptic Elements

40.10 Postsynaptic Elements

40.11 Rehabilitative Therapies

40.12 Concluding Remarks

References

IV: Developmental Sequences in the Maturation of Intrinsic and Synapse Driven Patterns

Chapter 41. GABA: A Multifacet Device that Exerts a Crucial Role in Brain Development

41.1 Introduction

41.2 GABA Depolarizes, Occasionally Excites Immature Neurons, and Produces a Rise of [Ca²+]I

41.3 Excitatory Actions of GABA are Mediated by a Developmental Expression of Chloride Cotransporters NKCC1 and KCC2

41.4 gabaergic Signals Develop Before Glutamatergic Ones in Many Brain Structures

41.5 The GABA-Glutamate Sequence in Other Brain Regions

41.6 A Parallel Developmental Sequence of Brain Patterns

41.7 An Abrupt Shift of [Cl−]I During Delivery Illustrates the Importance of Modulating GABA Actions

41.8 Activity-Dependent Plasticity of EGABA and Dynamic Chloride Regulation

41.9 Nonsynaptic Trophic Actions of GABA on Neuron Migration and Growth

41.10 General Conclusions

References

Chapter 42. Lessons from Zebrafish: Ion Channels Guide Neuronal Development

Abbreviations

42.1 Introduction

42.2 Existing Experimental Obstacles/Advantages of Zebrafish System

42.3 Activity and Development of Zebrafish Spinal Cord

42.4 An Unexplored Mechanism

42.5 Future Directions and Future Challenges

References

Chapter 43. Regulation of AMPA-Type Glutamate Receptor Trafficking

43.1 Introduction

43.2 Structure and Functions of AMPA Receptors

43.3 Synapse Development and AMPA Receptor Trafficking

43.4 Actin and Microtubule Organization in Neurons

43.5 AMPA Receptor Trafficking Along Microtubules in Dendrites

43.6 AMPA Receptor Trafficking Along the Actin Cytoskeleton in Dendritic Spines

43.7 Surface Trafficking of AMPA Receptors

43.8 Conclusion

Acknowledgments

References

Chapter 44. Pre- and Postsynaptic Assembly and Maturation: Principal Mechanisms and Coordination

Abbreviations

44.1 Synaptic Modules and Their Ultrastructure

44.2 Synapse Assembly and Maturation

44.3 Synapse Dynamics and Plastic Changes

44.4 Conclusions

References

Chapter 45. Cajal–Retzius and Subplate Cells: Transient Cortical Neurons and Circuits

45.1 Introduction

45.2 Cajal–Retzius Neurons

45.3 Subplate Neurons

45.4 Developmental Destiny of Cajal–Retzius and Subplate Neurons: Preservation or Disappearance

45.5 Conclusions and Perspectives

See also

Acknowledgments

References

Chapter 46. Chloride Homeodynamics Underlying Pathogenic Modal Shifts of GABA Actions

Abbreviations

Symbols

46.1 Developmental and Dynamic Shifts of Cl− Homeostasis Change GABA Actions

46.2 Pathological Models of Dynamic Cl− Homeostasis Perturbation

46.3 Pathological Models of Developmental Cl− Homeostasis Disorders

46.4 Pathological Models of Regenerating Immature Cl− Homeostasis Induced by Injury

46.5 Perspectives of Possible Involvement of Cl− Homeodynamics in Pathogenesis

46.6 Conclusion

References

Chapter 47. GABAergic Signaling at Newborn Mossy Fiber–CA3 Synapses: Short- and Long-Term Activity-Dependent Plasticity Processes

Abbreviations

47.1 Mossy Fiber Synapses

47.2 Co-release of Glutamate and GABA from MF Terminals

47.3 Criteria for Identifying Single MF-Evoked Responses

47.4 GABA is the Main Neurotransmitter Released from Immature MF Terminals

47.5 Presynaptic Modulation of GABA Release

47.6 Activity-Dependent Changes in Synaptic Efficacy

47.7 Conclusions

Acknowledgments

References

Chapter 48. BDNF and the Plasticity of Brain Networks During Maturation

48.1 Introduction

48.2 A Short Introduction to Neurotrophin Signaling Pathways

48.3 BDNF is a Target-Derived Messenger

48.4 BDNF is a Target-Derived Messenger for Activity-Dependent GABAergic Synaptic Plasticity in the Developing Brain

48.5 Conclusions

See also

Acknowledgments

Glossary

References

Chapter 49. Retinal Waves: Underlying Cellular Mechanisms and Theoretical Considerations

49.1 Introduction

49.2 How do We Record Retinal Waves?

49.3 Cellular Mechanisms Underlying Wave Generation in the Developing Retina

49.4 Retinal Waves are Under Homeostatic Control

See also

References

Chapter 50. Multimodal GABAA Receptor Functions on Cell Development

Nomenclature

50.1 Developmental Shifts of GABAA Receptor Subunit Composition

50.2 GABAA Receptor Function in Neurogenesis

50.3 GABAA Receptor Function on Migration

50.4 GABAA Receptor Function on Synaptogenesis

50.5 GABAA Receptor Function on Excitatory Neurotransmission

50.6 When Does Inhibitory GABAA Receptor-Mediated Neurotransmission Emerge?

50.7 Perspectives for Possible Involvement of Perturbations of Multimodal GABAA Receptor Development in Pathogenesis

50.8 Conclusion

See also

References

Chapter 51. Retinal Waves and their Role in Visual System Development

51.1 Introduction

51.2 Spatiotemporal Patterns of Retinal Waves

51.3 Influence of Waves on Retinal Development

51.4 Retinal Waves and Retinotopic Refinement

51.5 Retinal Waves and Eye-Specific Segregation

51.6 Retinal Waves and ON/OFF Segregation in the dLGN

51.7 Retinal Waves Influence the Developing Visual Cortex

51.8 Conclusions

References

Chapter 52. The Maturation of Firing Properties of Forebrain GABAergic Interneurons

52.1 Introduction

52.2 Interneuron Diversity

52.3 Embryonic Origins of Cortical Interneuron Subtypes

52.4 Postnatal Maturation of Interneuron Electrophysiology and Gene Expression

52.5 Activity-Dependent Postnatal Maturation of Cortical Interneuron Subtypes

52.6 Concluding Remarks

References

Chapter 53. Multiple Roles of KCC2 in the Developing Brain

53.1 Introduction

53.2 KCC2 and the CCC Family

53.3 Spatiotemporal Pattern of KCC2 Expression

53.4 Functional Regulation of KCC2

53.5 Functional Role of KCC2

Acknowledgment

References

Chapter 54. NKCC1 and Brain Maturation

54.1 Introduction

54.2 NKCC1 is a Key Prerequisite for GABAergic Depolarization

54.3 Expression, Regulation, and Modulation of NKCC1

54.4 Effects of NKCC1 on Brain Development

54.5 NKCC1 and Neonatal Epilepsy

54.6 NKCC1 During Adult Neurogenesis

54.7 Outlook and Final Remarks

References

Chapter 55. Maturation of Inhibitory Synaptic Transmission in the Spinal Cord: Role of the Brain Stem and Contribution to the Development of Motor Patterns

Abbreviations

55.1 Introduction

55.2 Development of the Locomotor Pattern

55.3 Development of Spontaneous Activity

55.4 Maturation of GABAA- and Glycine-Receptor-Gated Chloride Channels

55.5 Maturation of Chloride Homeostasis

55.6 Conclusion

References

Chapter 56. Calcium Signals Regulate Neurotransmitter Phenotype

56.1 Introduction

56.2 Calcium Spiking

56.3 Neurotransmitter Phenotype Specification – Intrinsic Determinants

56.4 Neurotransmitter Phenotype Plasticity

56.5 Mechanisms and Relevance of NPP

56.6 Perspectives

56.7 Summary

References

Index

Copyright

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Editors-in-Chief

Professor John L.R. Rubenstein,     Department of Psychiatry, University of California at San Francisco, San Francisco, CA, USA

Professor Pasko Rakic,     Duberg Professor of Neurobiology and Neurology, Director Kavli Institute for Neuroscience, Yale University School of Medicine, New Haven, CT, USA

Section Editors

Professor Arturo Alvarez‐Buylla,     Department of Neurological Surgery and The Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research University of California, San Francisco, School of Medicine, San Francisco, CA, USA

Professor Yehezkel Ben‐Ari,     Institute of Neurobiology of the Mediterranean Sea (INMED) AND CEO of Neurochlore Company, INSERM (the French Institute of Health and Medical Research), Marseille, Department of the Bouches du Rhone, France

Professor Kenneth Campbell,     Divisions of Developmental Biology and Neurosurgery Cincinnati Children’s Hospital Medical Center University of Cincinnati College of Medicine Cincinnati, OH, USA

Professor Hollis T. Cline,     Department of Molecular and Cellular Neuroscience, The Scripps Research Institute, La Jolla, CA, USA

Dr. François Guillemot,     Division of Molecular Neurobiology MRC National Institute for Medical Research, London, UK

Professor Takao Hensch,     Department of Molecular and Cellular Biology Harvard University, Cambridge, MA, USA

Professor Pat Levitt,     Zilkha Neurogenetic Institute and Department of Cell and Neurobiology, Keck School of Medicine of University of Southern California, Los Angeles, CA, USA

Dr Oscar Marín,     Instituto de Neurociencias, CSIC and Universidad Miguel Hernández, Alicante, Spain

Professor Dennis D.M. O’Leary,     Vincent J. Coates Chair in Molecular Neurobiology, Molecular Neurobiology Laboratory, The Salk Institute, La Jolla, CA, USA

Professor Franck Polleux,     The Scripps Research Institute, Dorris Neuroscience Center, La Jolla, CA, USA

Dr David H. Rowitch,     University of California, San Francisco, CA, USA

Dr Gordon M. Shepherd,     Department of Neurobiology, Yale School of Medicine, New Haven, CT, USA

Professor Helen Tager‐Flusberg,     Department of Psychology and Department of Anatomy & Neurobiology, Boston University, Boston, MA, USA

Contributors

M.S. Airaksinen,     University of Helsinki, Helsinki, Finland

S.A. Anderson,     Weill Cornell Medical College, New York, NY, USA

E.S. Anton,     The University of North Carolina School of Medicine, Chapel Hill, NC, USA

S.L. Barrow,     University of California, Davis, CA, USA

F. Beaubien,     McGill University, Montréal, QC, Canada

R. Belvindrah,     Institut Pasteur, Paris, France,     CNRS, Paris, France,     INSERM UMR-S 839, Paris, France,     Université Pierre et Marie Curie, Paris, France,     Institut du Fer á Moulin, Paris, France

Y. Ben-Ari,     INMED, INSERM U901, Marseilles, France

F. Bielle,     Institut de Biologie de l’École Normale Supérieure (IBENS), Paris, France,     Institut National de la Santé et de la Recherche Médicale (INSERM) U1024, Paris, France,     Centre National de la Recherche Scientifique (CNRS) UMR 8197, Paris, France

K. Boekhoorn,     Utrecht University, Utrecht, The Netherlands

A.B. Booker,     University of Connecticut, Storrs, CT, USA

U. Borello,     CNRS-UMR 7592, Institut Jacques Monod, Université Paris Diderot, Paris, France

F. Bradke,     German Center for Neurodegenerative Diseases (DZNE), Axonal Growth and Regeneration, Bonn, Germany

V. Castellani,     University of Lyon, Lyon, France

M.V. Chao,     New York University, New York, NY, USA

F. Charron,     Institut de Recherches Cliniques de Montréal (IRCM), Montreal, QC, Canada,     University of Montreal, Montreal, QC, Canada,     McGill University, Montreal, QC, Canada

A. Chedotal,     Institut de la Vision, Paris, France

E. Cherubini,     International School for Advanced Studies (SISSA), Trieste, Italy

A.D. Chisholm,     University of California, San Diego, CA, USA

J.-F. Cloutier,     McGill University, Montréal, QC, Canada

C.L. Cunningham,     University of California, Davis, CA, USA

M.B. Dalva,     Thomas Jefferson University, Philadelphia, PA, USA

F. de Castro,     Hospital Nacional de Parapléjicos-SESCAM, Toledo, Spain,     Instituto Cajal-CSIC, Madrid, Spain

M. Demarque,     University of California San Diego, La Jolla, CA, USA,     CNRS UPR 3294, Gif-sur-Yvette, France

T.L. Dickendesher,     The University of Michigan, Ann Arbor, MI, USA

T. Di Meglio,     Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland

Y. Duan,     The University of Michigan, Ann Arbor, MI, USA

R. Eavri,     Massachusetts Institute of Technology, Cambridge, MA, USA

J. Falk,     University of Lyon, Lyon, France

D.A. Feldheim,     University of California, Santa Cruz, CA, USA

M.B. Feller,     University of California at Berkeley, Berkeley, CA, USA

A. Filosa,     Max-Planck Institute of Neurobiology, Munich-Martinsried, Germany

K.C. Flynn,     German Center for Neurodegenerative Diseases (DZNE), Axonal Growth and Regeneration, Bonn, Germany

K.D. Foote,     Lerner Research Institute, Cleveland, OH, USA

A. Fukuda,     Hamamatsu University School of Medicine, Hamamatsu, Japan

T. Furukawa,     Hamamatsu University School of Medicine, Hamamatsu, Japan

F.H. Gage,     Salk Institute for Biological Studies, La Jolla, CA, USA

J.-L. Gaiarsa,     INMED, INSERM U901, Université de Méditerranée, Université de La Méditerranée, Marseille, France

S. Garel,     Institut de Biologie de l’École Normale Supérieure (IBENS), Paris, France,     Institut National de la Santé et de la Recherche Médicale (INSERM) U1024, Paris, France,     Centre National de la Recherche Scientifique (CNRS) UMR 8197, Paris, France

G. Gerlitz,     NCI, NIH, Bethesda, MD, USA

D.A. Gibson,     University of Southern California, Los Angeles, CA, USA

R.J. Giger,     The University of Michigan, Ann Arbor, MI, USA

A. Griveau,     CNRS-UMR 7592, Institut Jacques Monod, Université Paris Diderot, Paris, France,     Howard Hughes Medical Institute, and Eli and Edythe Broad Institute for Stem Cell Research and Regeneration Medicine, University of California, San Francisco, CA, USA

W.B. Grueber,     Columbia University, New York, NY, USA

Z. He,     Harvard Medical School, Boston, MA, USA

M.H. Hennig,     University of Edinburgh, Edinburgh, UK

C.C. Hoogenraad,     Erasmus Medical Center, Rotterdam, The Netherlands

C.A. Hübner,     University Hospital Jena, Friedrich Schiller University Jena, Jena, Germany

L. Izzi,     Institut de Recherches Cliniques de Montréal (IRCM), Montreal, QC, Canada,     University of Montreal, Montreal, QC, Canada

F. Jeanneteau,     New York University, New York, NY, USA

D. Kerschensteiner,     Washington University School of Medicine, St. Louis, MO, USA

R. Klein,     Max-Planck Institute of Neurobiology, Munich-Martinsried, Germany

H. Komuro,     Lerner Research Institute, Cleveland, OH, USA

Y. Komuro,     Lerner Research Institute, Cleveland, OH, USA

A.R. Kriegstein,     University of California, Davis, CA, USA

T. Kumada,     Lerner Research Institute, Cleveland, OH, USA,     Hamamatsu University School of Medicine, Hamamatsu, Japan

J.H. Leslie,     Massachusetts Institute of Technology, Cambridge, MA, USA

G. Li,     University of California, San Francisco, CA, USA

O. Llano,     University of Helsinki, Helsinki, Finland

P.-M. Lledo,     Institut Pasteur, Paris, France; CNRS, Paris, France

C. Lohmann,     Netherlands Institute for Neuroscience, Amsterdam, The Netherlands

J.J. LoTurco,     University of Connecticut, Storrs, CT, USA

C.S. Lu,     Harvard Medical School, Boston, MA, USA

A. Ludwig,     University of Helsinki, Helsinki, Finland

H.J. Luhmann,     University Medical Center of the Johannes Gutenberg-University of Mainz, Mainz, Germany

L. Ma,     University of Southern California, Los Angeles, CA, USA

S.J. Le Marchand,     Thomas Jefferson University, Philadelphia, PA, USA

T.C. Martin,     University of Colorado Anschutz Medical Campus, Aurora, CO, USA

A.K. McAllister,     University of California, Davis, CA, USA

D. McNeal,     University of Kansas Medical Center, Kansas City, KS, USA

A. Mizrahi,     The Hebrew University of Jerusalem, Jerusalem, Israel

F. Moya,     Instituto de Neurociencias de Alicante (UMH-CSIC), Universidad Miguel Hernandez-Consejo Superior de Investigaciones Científicas, San Juan de Alicante, Spain

M. Munz,     McGill University, Montreal, QC, Canada

K. Nakajima,     Keio University School of Medicine, Tokyo, Japan

Y. Nakanishi,     Hamamatsu University School of Medicine, Hamamatsu, Japan

E. Nedivi,     Massachusetts Institute of Technology, Cambridge, MA, USA

S.B. Nelson,     Brandeis University, Waltham, MA, USA

S.C. Noctor,     University of California, Davis, CA, USA

R.J. Nudo,     University of Kansas Medical Center, Kansas City, KS, USA

N. Ohno,     Lerner Research Institute, Cleveland, OH, USA

B.W. Okaty,     Harvard Medical School, Boston, MA, USA

T.J. Petros,     Weill Cornell Medical College, New York, NY, USA

C.K. Pfeffer,     University of California San Diego, La Jolla, CA, USA

A. Pierani,     CNRS-UMR 7592, Institut Jacques Monod, Université Paris Diderot, Paris, France

S.J. Pleasure,     University of California, San Francisco, CA, USA

F. Polleux,     The Scripps Research Institute, La Jolla, CA, USA

J.E.A. Prince,     McGill University, Montréal, QC, Canada

O. Reiner,     The Weizmann Institute of Science, Rehovot, Israel

A.B. Ribera,     University of Colorado Anschutz Medical Campus, Aurora, CO, USA

F.M. Rijli,     Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland

C. Rivera,     University of Helsinki, Helsinki, Finland,     Université de la Méditerranée, Aix-Marseille, France

E.S. Ruthazer,     McGill University, Montreal, QC, Canada

P.C. Salinas,     University College London, London, UK

P. Scheiffele,     University of Basel, Basel, Switzerland

D. Schreiner,     University of Basel, Basel, Switzerland

K. Sekine,     Keio University School of Medicine, Tokyo, Japan

E. Sernagor,     Newcastle University, Newcastle upon Tyne, UK

S.J. Sigrist,     Free University Berlin, Berlin, Germany,     Charité Universitätsmedizin Berlin, Berlin, Germany

C. Sotelo,     Institut de la Vision, Paris, France,     Instituto de Neurociencias, Miguel Hernandez University and CSIC, Alicante, Spain

N.C. Spitzer,     University of California San Diego, La Jolla, CA, USA

A. Stanco,     University of California, San Francisco, CA, USA

M. Stiess,     German Center for Neurodegenerative Diseases (DZNE), Axonal Growth and Regeneration, Bonn, Germany

S.C. Suzuki,     University of Washington, Seattle, WA, USA

H. Tabata,     Keio University School of Medicine, Tokyo, Japan

N. Toni,     University of Lausanne, Lausanne, Switzerland

P. Uvarov,     University of Helsinki, Helsinki, Finland

M. Valdeolmillos,     Instituto de Neurociencias de Alicante (UMH-CSIC), Universidad Miguel Hernandez-Consejo Superior de Investigaciones Científicas, San Juan de Alicante, Spain

D. Van Vactor,     Harvard Medical School, Boston, MA, USA

L. Vinay,     Centre National de la Recherche Scientifique (CNRS) & Aix-Marseille Université, Marseille, France

F. Wang,     Duke University, Durham, NC, USA

C. Wichmann,     Charité Universitätsmedizin Berlin, Berlin, Germany,     University of Göttingen, Gottingen, Germany

R.O.L. Wong,     University of Washington, Seattle, WA, USA

T. Yoshimatsu,     University of Washington, Seattle, WA, USA

B. Zalc,     Université Pierre & Marie Curie, Paris, France

C. Zhao,     Salk Institute for Biological Studies, La Jolla, CA, USA

Introduction to Comprehensive Developmental Neuroscience

It is broadly accepted that understanding the genetic, molecular, and cellular mechanisms of neural development is essential for understanding evolution and disorders of neural systems. Recent advances in genetic, molecular, and cell biological methods have generated a massive increase in new information. By contrast, there is a paucity of comprehensive and up-to-date syntheses, references, and historical perspectives on this important subject. Therefore, we embarked on the formidable task of assembling a novel resource entitled ‘Comprehensive Developmental Neuroscience.’ We hope that the books in this series will serve as valuable references for basic and translational neuroscientists, clinicians, and students.

To help with this enormous task, we invited leading experts in various subfields to select the subjects and invite appropriate authors. We were gratified by the number of busy scientists who accepted the invitation to write their articles. All the chapters have been peer reviewed by the Section Editors to ensure accuracy, thoroughness, and scholarship.

In the resulting three volumes, we cover a broad array of subjects on neural development. We organized the volumes chronologically according to the ordered steps in neural development. In addition, each volume is subdivided into three to four sections, each edited by world experts in these areas. The sections have 10–20 chapters that are written and illustrated by leading scientists.

This Volume in the series has 56 chapters devoted to migration (cell and axonal), the formation of neuronal connections, and the maturation of neural functions. This volume is subdivided into four sections. The first is on mechanisms that control the formation of axons and dendrites. The second is on the mechanisms that regulate cell migration that disperses specific subtypes of cells along highly defined pathways to specific destinations. The third section is on the regulation of synapse formation and maintenance during development; in addition, it has chapters on synaptogenesis in the mature nervous system in response to neurogenesis, neural activity, and neural trauma. The final section is on the developmental sequences that regulate neural activity, from cell-intrinsic maturation to early correlated patterns of activity.

Volume 1 in the series has 48 chapters devoted mainly to patterning and cell type specification in the developing central and peripheral nervous systems (CNS and PNS). This volume is subdivided into three sections. The first is on the mechanisms that control regional specification, which generate subdivisions of the nervous system. The second is on mechanisms that regulate the proliferation of neuronal progenitors and that control differentiation and survival of specific neuronal subtypes. The third section addresses the mechanisms controlling development of non-neural cells: astrocytes, oligodendroyctes, Schwann cells, microglia, meninges, blood vessels, ependyma, and choroid plexus.

Volume 3 in the series has 40 chapters devoted to the anatomical and functional development of neural circuits and neural systems, as well as chapters that address neurodevelopmental disorders in humans and experimental organisms. This volume is subdivided into three sections. The first is on the mechanisms that control the assembly of neural circuits in specific regions of the nervous system, and as a function of neural activity and critical periods. The second section concentrates on multiple aspects of cognitive development, particularly in humans. The final section addresses disorders of the nervous system that arise through defects in neural development, building on the principles that are addressed in earlier sections of the book.

John L.R. Rubenstein

Pasko Rakic

I

Formation of Axons and Dendrites

Chapter 1 Development of Neuronal Polarity In Vivo

Chapter 2 Role of the Cytoskeleton and Membrane Trafficking in Axon–Dendrite Morphogenesis

Chapter 3 Axon Growth and Branching

Chapter 4 Axon Guidance

Chapter 5 Roles of Eph–Ephrin Signaling in Axon Guidance

Chapter 6 Axon Guidance

Chapter 7 Nonconventional Axon Guidance Cues

Chapter 8 Axon Regeneration

Chapter 9 Axon Maintenance and Degeneration

Chapter 10 Dendrite Development

Chapter 11 Dendritic Development

Chapter 1

Development of Neuronal Polarity In Vivo

F. Polleux,    The Scripps Research Institute, La Jolla, CA, USA

Outline

1.1 Introduction

1.2 Axon Initiation In Vitro Versus In Vivo

1.2.1 Axon Initiation In Vitro

1.2.2 Axon Initiation In Vivo

1.3 Distinction Between Cues Regulating Axon Specification Versus Axon Growth

1.4 Extracellular Cues Regulating Neuronal Polarization and Axon Initiation

1.4.1 Netrin-1 and Wnt Control Axon Initiation in C. elegans

1.4.2 Polarized Emergence of the Axon in Retinal Ganglion Cells of Xenopus

1.4.3 Extracellular Cues Underlying the Emergence of Axon and Dendrites in Mammalian Neurons

1.5 Intracellular Pathways Underlying Neuronal Polarization

1.5.1 Role of Local Protein Translation and Degradation for Axon Specification and Axon Growth

1.5.2 Role of Cytoskeletal Dynamics in Axon Initiation and Growth

1.5.3 Major Signaling Pathways Involved in Axon Initiation and Growth

1.5.3.1 LKB1 and its Downstream Kinases SAD-A/B and MARK1-4

1.5.3.2 PAR3–PAR6–APKC

1.5.3.3 Ras- and Rho-Family of Small GTPases

1.5.3.4 PI3-Kinase and PTEN Signaling during Axon Specification

1.5.3.5 AKT/Protein Kinase B

1.5.3.6 GSK3 and Axon Specification

References

1.1 Introduction

The ability of neurons to form a single axon and multiple dendrites underlies the directional flow of information transfer in the central nervous system (CNS). Dendrites and axons are molecularly and functionally distinct domains. Dendrites integrate synaptic inputs, triggering the generation of action potentials at the level of the soma. Action potentials then propagate along the axon that makes presynaptic contacts onto target cells. This chapter reviews what is known about the cellular and molecular mechanisms underlying the ability of neurons to polarize and form a single axon and multiple dendrites during development. This question has received much attention over the past three decades using mainly in vitro approaches, where neurons from distinct parts of the developing mammalian brain can be dissociated and cultured to get single cell resolution. Remarkably, neurons can polarize to form a single axon and multiple dendrites, and later establish functional synaptic contacts in these reductionist conditions. This approach became, and remains, the dominant model to study axon initiation and growth and has yielded the identification of many molecules that regulate axon formation in vitro. At present, only a few of the genes identified using in vitro approaches have been shown to be required for axon initiation and outgrowth in vivo. In vitro axon initiation and elongation is thought to reflect the intrinsic ability of neurons to polarize in the absence of relevant extracellular cues. However, extracellular cues have been shown to play an important role during neuronal polarization in vivo. In this chapter, we focus on our current understanding of the complex interplay between extracellular cues and intracellular signaling pathways underlying the emergence of axon and dendrite during neuronal polarization in vivo.

1.2 Axon Initiation In Vitro Versus In Vivo

1.2.1 Axon Initiation In Vitro

Historically, the advent of in vitro dissociated neuronal cultures provided an experimental template for improving our understanding of the cell biology of neuronal polarity, including the specification of the molecular identity of axon and dendrite. Pioneering work using these cultures established a paradigm in which isolated neurons in culture can adopt spatially and functionally distinct dendritic and axonal domains (Craig and Banker, 1994; Goslin and Banker, 1989). Careful analysis of these cultures led to the observation that cultured hippocampal neurons transition through several stages, from freshly plated stage 1 cells bearing immature neurites to stage 5 cells that exhibit mature axons, dendrites, dendritic spines, and functional synapses (Craig and Banker, 1994; Dotti et al., 1988). It should be noted that in the classical E18 rat hippocampal cultures, most plated cells were polarized postmitotic neurons before dissociation; therefore, neuronal polarization using this in vitro model likely corresponds to the repolarization of previously polarized neurons in vivo. It is therefore important to keep in mind that molecular manipulations in this in vitro model act on previously polarized neurons that may retain some aspects of polarization, which can be critical for interpreting the results. Recent advances in the techniques allowing the manipulation of gene expression more specifically in neural progenitors, such as in utero or ex utero cortical electroporation (Hand et al., 2005; Hatanaka and Murakami, 2002; Saito and Nakatsuji, 2001; Tabata and Nakajima, 2001), provide a paradigm to (a) manipulate gene expression in progenitors, that is, before neuronal polarization occurs upon cell cycle exit and (b) visualize the earliest stages of neuronal polarization in a contextual cellular environment, that is, in organotypic slices or intact embryonic brain (Barnes et al., 2007; Calderon de Anda et al., 2008; Hand et al., 2005).

1.2.2 Axon Initiation In Vivo

Neuronal polarization can be divided into several specific steps in vivo. On cell cycle exit, mammalian neurons usually migrate over a long distance before reaching their final destination. In vivo, most neurons undergo axon–dendrite polarization during migration. While migrating, neocortical pyramidal neurons (PN) form a leading process and a trailing process, each becoming the axon or the dendrite (Figure 1.1). Careful examination of the morphological transition between neural progenitors and postmitotic neurons reveals that neurons can inherit their axon and dendrite polarity directly from the apicobasal polarity of their progenitors. This is the case for retinal ganglion cells and bipolar cells in the developing vertebrate retina (Hinds and Hinds, 1978; Morgan et al., 2006; Zolessi et al., 2006, reviewed in Barnes and Polleux, 2009). In other neuronal subpopulations undergoing long-range migration, neuronal morphogenesis undergoes extensive stereotypical changes, leading to polarized outgrowth of their axon and dendrites. This is the case for cerebellar granule neurons (CGN) as well as cortical and hippocampal PN, two of the best-studied models of neuronal polarization (Gao and Hatten, 1993; Hatanaka and Murakami, 2002; Komuro et al., 2001; Noctor et al., 2004; Rakic, 1971, 1972; Shoukimas and Hinds, 1978). Both CGN and PN acquire their axon–dendrite polarity from the polarized emergence of their trailing and leading processes, respectively, during migration (reviewed in Barnes and Polleux 2009). Precise examination of the process dynamics occurring shortly after cell cycle exit in dorsal telencephalic progenitors suggest that there is often a slight delay between the trailing process formation (axon initiation), which frequently precedes leading process formation (Calderon de Anda et al., 2008; Hand and Polleux, 2011). However, one thing is clear for both PN and CGN: axon formation starts before or during radial migration. Interestingly, different neuronal populations display distinct modes of axon formation, which reflect their mode of migration, lineage, and type of axon projection. For example, cortical interneurons, which will form axons projecting only locally within the cortex, originate from the ventral telencephalon and have to migrate over very long distances before initiating their axon after reaching their final destination in the cortex (Bortone and Polleux, 2009; Cobos et al., 2007; Yamasaki et al., 2010). Even though the precise mechanisms underlying the emergence of the axon of cortical interneurons are currently unknown, the striking difference with radially migrating pyramidal cortical neurons, which initiate axon formation during migration, leads to the hypothesis that their ability to form an axon is inhibited during tangential migration and/or that axon initiation in cortical interneurons depends on factor(s) present only in their final environment, the cortex.

Figure 1.1 Axon development in the neocortical pyramidal neurons. (a) During mouse corticogenesis, from embryonic day E11 to P1, neuronal polarization occurs rapidly on cell cycle exit (see more details in Figure 1.2) when neurons (green) transition from a multipolar morphology to a bipolar morphology by forming a leading process (future apical dendrite), which drives radial migration and a trailing process, which becomes the axon. Once neurons reach their final position, they undergo extensive dendritic branching. (b) At early postnatal stages (P1–P7), the axon of pyramidal neurons are guided either laterally (for corticofugal projections of layer 5/6) or medially (for callosal projection of layer 2/3 and 5). (c) On reaching their final target, axons undergo extensive, layer-specific branching concomitant with synaptogenesis. For example, callosal axons branch both ipsilaterally and contralaterally in layer 5 and layer 2/3 (green axons).

As discussed later in the chapter, an emerging concept from recent work done primarily in C. elegans suggests that in vivo, the ‘symmetry-breaking’ events that lead to the emergence of the dendrite and the axon require the ability of postmitotic neurons to sense gradients of extracellular cues, leading to the asymmetric activation of signaling pathways underlying the emergence of the axon. This data is supported by recent evidence in mammals showing that extracellular cues such as TGFß plays a role in axon specification in vivo by triggering noncanonical Par6-dependent signaling downstream of TGFß-receptor activation (Yi et al., 2010).

1.3 Distinction Between Cues Regulating Axon Specification Versus Axon Growth

Most studies published over the past two decades in this field have been performed using in vitro approaches. The classic paradigm for confirming the regulatory role of a gene in neuronal polarity is to show that downregulation of its expression using shRNA technology or gene knockout technology is required for axon formation. These experiments are typically done using both staining with axon-specific makers and measurement of neurite length because the axon usually grows 5–10 times faster than do neurites becoming dendrites. However, this type of evidence may not be sufficient to distinguish unambiguously an effector of axon specification from a molecule simply required for axon growth (Jiang et al., 2005). Conversely, showing that overexpression or overactivation of a candidate molecule leads to the emergence of multiple neurites, displaying the molecular identity of an axon is generally used to suggest that this molecule is sufficient to confer axon identity. However, this approach is limited by the fact that it relies on overexpression, which can be complicated by abnormal activation of a pathway normally not involved in axon specification or neuronal polarity. Recent technical advances allow the manipulation of gene expression in vivo by in utero cortical electroporation in rodent cortex or cerebellum (Famulski et al., 2010; Saito and Nakatsuji, 2001) or transgenic approaches in Xenopus (Zolessi et al., 2006). Therefore, a more biologically relevant validation of the function of a candidate gene during neuronal polarization often includes testing its requirement using gene knockout or shRNA-mediated knockdown technologies or the analysis of conventional or conditional knockout in combination with in utero electroporation allowing single cell resolution analysis of axon formation (Barnes et al., 2007; Shelly et al., 2007; Yi et al., 2010).

1.4 Extracellular Cues Regulating Neuronal Polarization and Axon Initiation

1.4.1 Netrin-1 and Wnt Control Axon Initiation in C. elegans

Is there any in vivo evidence for the role of extracellular cues in the specification of neuronal polarity? Important progress in our understanding of the molecular and cellular mechanisms specifying axon initiation during neuronal polarization has been made using the C. elegans model. This pioneering work has markedly enhanced our understanding of how extracellular cues instruct axon initiation in vivo. The neurons of the nematode have a stereotyped morphology, for example, specific projections along the dorsoventral and anterior–posterior body axes. Elegant experiments have identified an extracellular cue, UNC-6 (Netrin), along with its receptor UNC-40 (DCC), as a critical gene orchestrating axon initiation in vivo (Adler et al., 2006). This work also identified downstream proteins in this pathway, including (mammalian orthologs are shown in parenthesis when established) AGE-1 (phosphoinositide-3 kinase – PI3K), DAF-18 (PTEN), UNC-34 (Enabled), CED-10 (Rac), UNC-115/AbLIM, and MIG-10/lamellipodin. The current model for the relationship between these genes and UNC-6/netrin signaling involves DAF-18’s limitation of AGE-1 activity following UNC-40/DCC stimulation and the asymmetric recruitment of MIG-10/lamellipodin to the plasma membrane. This recruitment requires activated CED-10/Rac direct binding to MIG-10/lamellipodin and the involvement of the PAK-like kinase, Pak-1 (Adler et al., 2006). The involvement of a kinase in cytoskeletal rearrangement is consistent with the similar role of MIG-10/lamellipodin and the likely mechanism under which it operates once recruited to the plasma membrane to stimulate directed neurite outgrowth. Another regulator thought to act in concert to drive filopodial formation with MIG-10/lamellipodin is the Enabled homolog, UNC-34 (Chang et al., 2006). Finally, SLT-1 (Slit) is another extracellular cue that likely acts through MIG-10 recruitment (Chang et al., 2006) to control neuronal polarization.

Two other studies have identified the diffusible signal Wnt and its receptor as critical regulators of axon specification and neuronal polarity (Hilliard and Bargmann, 2006; Prasad and Clark, 2006). In addition to identifying loss of function for Lin-44 (Wnt) and its receptor Lin-17 (Frizzled – Fzl), one of the screens identified VPS-35, a component of retromer complex that regulates vesicular traffic and is required for proper Wnt secretion (Pan et al., 2008; Prasad and Clark, 2006).

1.4.2 Polarized Emergence of the Axon in Retinal Ganglion Cells of Xenopus

Careful live imaging experiments of Xenopus retinal ganglion cell polarization revealed that polarized axon outgrowth requires some unidentified extracellular cues present in the basal lamina (Randlett et al., 2011; Zolessi et al., 2006). The axon of developing RGCs normally grows on the basal side of the neuron. In a mutant called Nok, characterized by the absence of retinal pigmented epithelium, some postmitotic RGC neurons show a defective polarized outgrowth of their axon on the apical side along the now-exposed basal lamina. In this context, the polarized emergence of the axon on the basal side of the RGC is correlated with the position of the centrosome, Par3, and the apical complex (containing at least atypical protein kinase C (aPKC), β-catenin, and F-actin) on the apical side of the cell where the dendrite will emerge. Taken together, this work strongly suggests that (a) the basal lamina contains some important extracellular cues playing a role in the polarized emergence of the axon of RGC neurons and that (b) RGC neurons inherit the intrinsic apicobasal polarity of their progenitor at least with regard to the Par3/aPKC components of the polarity complex.

Recently, a signaling cascade has been linked to potential extracellular cues regulating axon initiation in vivo (Barnes and Polleux, 2009; Barnes et al., 2008). Conditional deletion of LKB1 in pyramidal cortical neurons (also called Par4 or STK11) demonstrated that LKB1 is required for axon initiation in cortical neurons but does not impact their radial migration (Barnes et al., 2007). Structure/function analysis indicates that phosphorylation of LKB1 at Serine 431 is required for its function in axon specification (Barnes et al., 2007), and Shelly et al. linked this phosphorylation to the ability of extracellular cues such as BNDF to stimulate cAMP production and protein kinase A (PKA)-dependent phosphorylation of S431 in the nascent axon (see below for details; Shelly et al., 2007, 2010, 2011).

1.4.3 Extracellular Cues Underlying the Emergence of Axon and Dendrites in Mammalian Neurons

Several lines of evidence suggest that extracellular cues can direct the polarized emergence of the axon and the dendrites both in vitro and in vivo. One paradigm involves dissociated cortical or hippocampal PN plated on striped substrates coated with two different cell adhesion molecules (e.g., laminin and NgCAM, reviewed in Barnes and Polleux, 2009). The first immature neurite of E18 hippocampal neurons, which contacts the boundary between two stripes, systematically becomes the axon. This occurs regardless of the fact that the initial outgrowth of immature neurites occurred on laminin or NgCAM, suggesting that immature neurites can detect changes in the nature of the extracellular substrate rather than the absolute nature of the novel substrate they are encountering (Esch et al., 1999). Using a similar approach, Shelly and colleagues showed that neurites of immature hippocampal neurons growing on a patterned substrate can detect the presence of brain-derived neurotrophic factor (BDNF), which plays an instructive role in axon specification because the first neurite contacting a BDNF stripe systematically becomes the axon (Shelly et al., 2007). The effect of BDNF on axon specification requires cAMP-dependent PKA activation and phosphorylation of LKB1 in position 431 by PKA (Shelly et al., 2007, 2010), suggesting that LKB1 phosphorylation on S431 acts as a detector of neuronal symmetry breaking by extracellular cues such as BDNF in this in vitro context.

The overlay assay is an in vitro assay developed to detect the existence of putative extracellular cues playing a role in cortical axon guidance and neuron polarization. This rather simple assay involves plating fluorescently labeled dissociated cortical neurons onto cortical slices to test whether polarized axon emergence in vivo is mainly the result of asymmetric activation of intracellular effectors (maybe inherited by progenitors) or if extracellular cues can play a role in axon specification. Polleux et al. have demonstrated that scenario 2 is the most likely because only a couple of hours after plating, the vast majority of cortical neurons displayed a single, short axon directed ventrally toward the ventricle (Polleux et al., 1998) as observed for radially migrating neurons in vivo. These authors went on to demonstrate that the class 3 secreted semaphorin, Sema3A, which is enriched in the most superficial part of the cortical wall (the top of the cortical plate), plays a role in repulsing axon initiation ventrally toward the ventricle (Polleux et al., 1998). More recently, Sema3A was shown to also regulate the polarized emergence of the leading process/apical dendrite both in the overlay assay, that is, independently of radial migration where it requires cGMP production and PKG activation (Polleux et al., 2000) and also in vivo during radial migration (Chen et al., 2008). Interestingly, Sema3A can play a role in the specification of dendritic identity by activating a cGMP-dependent pathway involving activation of cGMP-gated calcium channels (Nishiyama et al., 2011) and also by repressing axonal identity in a LKB1-dependent manner (Shelly et al., 2007, 2010, 2011; see Figure 1.2).

Figure 1.2 Extracellular signals regulate axon and dendrite specification in neocortical pyramidal neurons (PN). Time-lapse analysis has revealed that polarization of axodendritic polarity in PN occurs in vivo in a stepwise manner on cell cycle exit: following asymmetric division of radial glial cells (step 1) or intermediate progenitors in the subventricular zone (SVZ) (not shown) recently generated postmitotic neurons first form transient, dynamic neurites (multipolar stage; step 2). Axon specification occurs when a trailing process (TP) is stabilized (red) either before (step 3) or after the formation of a leading process (LP) in neurons engaging radial translocation (step 4). On reaching their final destination at the top of the cortical plate (CP) (step 5), neurons detach from the radial glial scaffold and start an extensive program of axon growth (see Figure 1.1) and dendritic branching concomitant with synaptogenesis (step 6).

Recently, TGFß signaling was shown to be required for the polarized emergence of the axon of radially migrating PN in vivo (Yi et al., 2010; Figure 1.3). TGFß ligands are expressed in the germinal zone of the cortex, where they could act as an instructive ‘ventral’ cue for the polarized emergence of the axon in multipolar neurons before engaging radial migration. In vitro experiments demonstrated that local application of TGFß on a single neurite in immature Stage 1 cortical neurons is sufficient to trigger fast axonal extension (Yi et al., 2010). Importantly, conditional genetic deletion of TGFß receptor 2 expression leads to the production of neurons without trailing process/axon in vivo. One noticeable difference with the conditional ablation of LKB1 (see below), which also leads to the absence of trailing process/axon formation but not radial migration defects, is that TGFß receptor 2 conditional deletion leads to retardation of radial migration in a subset of cortical neurons (Yi et al., 2010).

Figure 1.3 Intracellular pathways underlying axon specification in response to extracellular cues in vivo. As shown in Figure 1.2, recent data (Yi et al. 2010) provided evidence that TGFß receptor activation is required for axon specification/trailing process stabilization in vivo. This work also suggested that TGFß receptor signaling in the context of axon specification required a ‘noncanonical’ branch of TGFß involved in EMT, which triggers Par6 phosphorylation by TGFß receptor 2 and recruits the ubiquitin ligase Smurf1 and locally degrades RhoA, three steps previously shown to be required for axon specification. Another kinase previously shown to be required in vivo for axon specification is LKB1 (Par4), which is phosphorylated on S431 specifically in the axon in response to several signaling pathways (PKA, p90RSK, or aPKC). LKB1 phosphorylates and activates several other kinases, including SAD-A/B kinases, which have been shown to be required for axon specification in vivo. The actual link between TGFß receptor/Par6/Smurf1/RhoA and the LKB1/SAD kinase pathway is currently unknown but might involve direct interaction between Par6 and LKB1 or phosphorylation of LKB1 by aPKC, which is known to form a complex with Par6. The downstream effectors of SAD kinases and other LKB1-dependent kinases are currently unknown but might involve local phosphorylation of microtubule-associated proteins such as Tau or MAP1b but probably involve many other effectors, all involved in axon specification. Proteins indicated in red have been shown to be required in vivo for axon specification, whereas proteins indicated in orange have only been studied in dissociated neuronal cultures in vitro.

The authors went on to demonstrate that TGFß receptor function during axon specification requires the phosphorylation of Par6 on S435 previously shown to mediate epithelial to mesenchymal transition (EMT; Ozdamar et al., 2005). As discussed later in the chapter, this ‘noncanonical’ TGFß receptor-dependent signaling represents an attractive in vivo signaling pathway for axon specification since it is known to involve recruitment of the ubiquitin ligase Smurf1 that induces local degradation of RhoA during EMT, which have both previously been involved in axon specification (Schwamborn et al., 2007a,b). Therefore, these results suggest that neuronal polarization might have coopted signaling pathways regulating EMT for the purpose of axon specification.

Overall, this work suggests that the polarized emergence of a single axon/trailing process and the apical dendrite/leading process are controlled at least in part by extracellular cues such as TGFß and Sema3A, respectively, expressed in a graded manner along the neuron’s migratory path (Figure 1.2). More work needs to clarify if other extracellular cues are required for the polarized emergence of axons and dendrites in diverse neuronal cell types and how these extracellular cues mediate their effects on the specification of the unique molecular identity of axons and dendrites.

1.5 Intracellular Pathways Underlying Neuronal Polarization

1.5.1 Role of Local Protein Translation and Degradation for Axon Specification and Axon Growth

Spatial regulation of protein expression by selective degradation has been demonstrated in several contexts during neuronal development, including axonal pruning (Watts et al., 2004), various aspects of axon guidance (Bloom et al., 2007; Campbell and Holt, 2001; DiAntonio et al., 2001; Lewcock et al., 2007), synapse formation (DiAntonio et al., 2001; Nakata et al., 2005), synapse maintenance (Aravamudan and Broadie, 2003; DiAntonio et al., 2001; Ehlers, 2003; Speese et al., 2003), and synapse elimination (Ding et al., 2007, reviewed in DiAntonio and Hicke, 2004). Acute treatment with the proteosome inhibitor lactacystin blocks axogenesis in dorsal root ganglion cells (Klimaschewski et al., 2006). Furthermore, more prolonged inhibition of protein degradation with lactacystin leads to the formation of multiple axons (Yan et al., 2006). The protein kinase AKT, which we previously described as critical for neuronal polarity, appears to undergo selective degradation (Yan et al., 2006). In fact, this degradation selectively targets the inactive pool of AKT in neurites, resulting in a net enrichment of AKT in a single process that contains active AKT, the nascent axon. This phenomenon is consistent with the negative feedback signal model proposed by Kaibuchi and colleagues (Arimura and Kaibuchi, 2007) to explain axon specification of a single axon during neuronal polarization. Schwamborn et al. showed that the small GTPase Rap1b is regulated by a similar scheme because the active form of Rap1b is spared from degradation and ultimately enriched in the axon (Schwamborn et al., 2007b). In this case, the ubiquitin ligase acting on Rap1b is Smurf2, whereas the related Smurf1 appears to affect only neurite outgrowth. Additional work has demonstrated that an interaction between Smurf2 and the polarity scaffold PAR3 must exist for proper neuron polarization (Schwamborn et al., 2007a). This finding appears to be related to PAR3 targeting of Smurf2 to the axon because perturbation of the interaction of either PAR3–Smurf2 or PAR3–KIF3A results in Rap1b increase in all neurites. The converse situation exists for LIM kinase (LIMK) because levels of this protein must be reduced for axon initiation in vitro (Tursun et al., 2005). Future experiments should clarify the molecular mechanisms regulating the function of Smurf1/2 ubiquitin ligases in axon specification in vivo especially in the context of TGFß signaling (Yi et al., 2010).

1.5.2 Role of Cytoskeletal Dynamics in Axon Initiation and Growth

Appropriate regulation of the actin and microtubule cytoskeleton is critical for neuronal polarization and has been the focus of numerous studies. Experiments using the actin-destabilizing agents, lactrunculin B and cytochalasin D, indicate that remodeling of the actin-based cytoskeleton is an important regulatory step in axon formation (Bradke and Dotti, 1997). Specifically, actin depolymerization localized to a single neurite in unpolarized stage 2 hippocampal neurons is sufficient to confer axonal identity. A proposed mechanism is that loose actin filaments allow the egress of microtubules and lead to rapid elongation of a given neurite, perhaps outpacing the transport of negative regulators of axonal identity. The idea of cellular asymmetries being reinforced by localized microtubule stabilization and invasion proposed by Kirschner and Mitchison (1986) was elegantly demonstrated using a photoactivatable form of the tubulin-stabilizing compound taxol, which can direct axonal specification to a single immature neurite (Witte et al., 2008). Collectively, these results suggest that a dynamic equilibrium between actin depolymerization and microtubule stabilization might play a role in specifying axonal initiation and axonal identity. Future investigations will need to identify the effectors regulating this balance locally and also characterize how specific upstream signaling pathways might regulate the spatial and temporal regulation of these cytoskeletal dynamics.

1.5.3 Major Signaling Pathways Involved in Axon Initiation and Growth

1.5.3.1 LKB1 and its Downstream Kinases SAD-A/B and MARK1-4

A pioneering genetic screening performed by Kemphues and colleagues in the late 1980s identified six Par genes encoding distinct protein families. Many studies have since demonstrated that invertebrate and vertebrate Par genes play critical roles in epithelial cell polarity during development as well as in the context of cell transformation and metastasis (Goldstein and Macara, 2007; Kemphues et al., 1988). Although this pathway is critical to polarity in many species, the signal linking this pathway to extracellular cues has remained elusive. The furthest upstream component known in this cascade is an evolutionarily conserved kinase named LKB1 or PAR4. LKB1 translocates from the nucleus and is activated by heterodimerization with one of two related pseudokinases known as Strad-α and -ß (Dorfman and Macara, 2008). In addition to binding Strad, LKB1 function in neuronal polarity requires its phosphorylation at S431, a target of both PKA and p90RSK kinases (Collins et al., 2000; Sapkota et al., 2001), and this phosphorylation can be triggered by extracellular cues such as BDNF (Shelly et al., 2007; Figure 1.2). This event might be mediated partly by cues providing chemotactic attraction of radially migrating neurons toward the cortical plate such as Sema3A (Chen et al., 2008; Polleux et al., 2000) or by other extracellular cues, including neurotrophins (NTs) such as BDNF/NT4/NT3 (Shelly et al., 2007), Netrin (Adler et al., 2006), FGFs, or any other cues that can activate cAMP-dependent PKA or p90 RSK (RSK1-3). At this point, we can speculate that any other presently uncharacterized serine/threonine PKA able to phosphorylate S431 on LKB1 might mediate the polarizing function of extracellular cues found in different developing brain regions (Barnes and Polleux, 2009). Future investigations will identify the relevant extracellular cues and the corresponding signaling pathways triggering phosphorylation of LKB1 in position S431, thereby specifying the axon in developing cortical PN in vivo.

Once LKB1 is activated by binding to its necessary coactivator Strad-α and S431 phosphorylation occurs (only in the neurite becoming the axon), LKB1 phosphorylates SAD-A/B kinases (and probably microtubule affinity-regulated kinases, MARK1-4; Matenia and Mandelkow, 2009), which are required for axon specification partly by phosphorylating microtubule-associated proteins (MAPs) such as Tau. On the basis of the function of SAD kinases in presynaptic vesicular clustering in C. elegans (Crump et al., 2001), we can hypothesize that SAD-A/B kinases might also specify axon identity by directing presynaptic vesicular trafficking in the neurite becoming the axon. Most important, genetic deletion of LKB1 in cortical PN prevents axon formation, whereas overexpression of LKB1 and its coactivator Strad in neural progenitors or LKB1 alone in postmitotic cells is sufficient to lead to the formation of multiple axons (Barnes et al., 2007; Shelly et al., 2007).

Experiments in Xenopus laevis suggested that LKB1 may regulate aPKC inactivation of glycogen synthase kinase 3 (GSK3)ß (Ossipova et al., 2003), two proteins involved in neuronal polarity (see below). However, at this point, the exact contribution of LKB1 in adenomatous polyposis coli (APC)/GSK3 function in neuronal polarity is poorly understood. LKB1 also phosphorylates and activates a family of 13 PKAs related to the C. elegans PAR1 protein (Lizcano et al., 2004). To date, three of these have been implicated in regulating axon formation: SAD-A and SAD-B kinases as well as MARK-2 (microtubule affinity regulating kinase-2; also called Par1b). RNAi knockdown of SAD kinases partially abrogates the ability of LKB1 overactivation to induce multiple axon formation in cortical neurons, indicating that