lecture vi. making connections - nslc · bio 3411 lecture vi. making connections 9/17/12 2 what the...

Bio 3411 Lecture VI. Making Connections

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Lecture VI. Making Connections

Bio 3411 Monday

September 17, 2012 !!

September!17,!2012! 1!Lecture!VI.!Making!Connec<ons!

†Bentley, D., & Caudy, M. (1983). Nature, 304(5921), 62-65. †Dickson, B. J. (2002). Science, 298, 1959-1964. †Hannula-Jouppi, et al. (2005). PLoS Genet 14, e50. †Foty, R. A., & Steinberg, M. S. (2004). Int J Dev Biol, 48(5-6), 397-409. †Hayashi, T., & Carthew, R. W. (2004). Nature, 431(7009), 647-652. Jen, J. C., et al. (2004). Science, 304, 1509-1513. †Moscona, A., & Moscona, H. (1952). J Anat, 86(3), 287-301. †Myers, P. Z., & Bastiani, M. J. (1993). J Neurosci, 13(1), 115-126. †Sperry, R. W. (1963). Proc Natl Acad Sci U S A, 50, 703-710. †Tessier-Lavigne, M., & Goodman, C. S. (1996). Science, 274(5290), 1123-1133. †Townes, P. L., & Holtfreter, J. (1955). J Exp Zool, 128, 53–120. †Walter, J., Henke-Fahle, S., & Bonhoeffer, F. (1987). Development, 101(4), 909-913. †Wilson, H. (1907). Science, 25, 912-915.

Reading NEUROSCIENCE: 5th ed, pp!507?536!

4th ed, pp 577-609

2 Lecture VI. Making Connections September!17,!2012!

____________________________________________ †(pdfs on course website: [http://www.nslc.wustl.edu/courses/Bio3411/bio3411.html]


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What the last Lecture was about

•  Programmed cell death (apoptosis) is a physiological mechanism distinct from necrotic cell death.

•  Apoptosis occurs widely during normal development of the nervous system.

•  Isolation of specific molecules involved in promoting growth and survival – �trophism,� e.g., Nerve Growth Factor (NGF).

•  What is the �death mechanism� that NGF (and other neruotrophins) inhibit?

•  Broader implications: controlled cell death in neuroembryology vs uncontrolled cell growth of cancer.

•  Gene homologies between organisms - humans and worms (nematodes)

•  Molecular models for apoptosis

•  How do trophic factors connect to this cell death pathway(s)?

September!17,!2012! Lecture!VI.!Making!Connec<ons! 3!



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What this Lecture is about

•  General mechanisms for assembling neurons and groups of neurons

•  Diffusion vs Contact

•  Attraction vs Repulsion

•  Examples of impacts of contact

•  Examples of impacts of diffusion

•  Specification by growth factors

•  The chemoaffinity hypothesis



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Fertilization

Embryonic morphogenesis

Induction of Neuroectoderm

Neurulation

Differentiation: 1. Formation and placement of neuroblasts 2. Axonal outgrowth 3. Growth cones, selective migration 4. Selective fasciculation 5. Target selection 6. Synaptogenesis 7. Etc…(cell shape, neurotransmitter, ionic channels, receptors)

Adult neuronal plasticity (Activity-dependent?)

Segm

enta

tion

Outline of Neurodevelopment

Selective Adhesion Determines Specificity of Tissue and Cellular Associations



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Selective Aggregation of dissociated embryonic tissues (vertebrate and invertebrate) suggests ancient (surface)

Adhesion Molecules


1. Sponges (Wilson, 1907) 2. Amphibians (Townes and Holtfretter, 1955) 3. Chick (Moscona, 1952)

Epidermis + Mesoderm


Experimental recreation of morphogenesis by mixing cells expressing low and high levels of one surface

adhesion gene (N-cadherin)

Green = high N-cadherin Red = low N-cadherin

+4 hrs +24 hrs

(Foty and Steinberg, 2004)


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Growth Cones are Dynamic Sensory Organelles that Guide the Growth of Embryonic Axons!


(Forscher lab)

Sensing and Transducing: •  Diffusible Cues •  Contact-dependent Cues •  Trophic Factors •  Neurotransmitters

Extracellular Cues

Intracellular Signaling Pathways

Cytoskeletal Rearrangment

Ca+2

GTP cAMP

2nd

Messengers


Purves 5th ed, p. 510


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(Play GFP-Actin Growth Cone Movie) Dr. Andrew Matus

Friedrich Miescher Institute, Switzerland




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Effect/Proximity.

A0rac2on. Repulsion.

Distant.Contact.

Functional Classes of Axonal Guidance Molecules!

(Secreted)!

(Membrane Associated)!

(netrin)! (sema, slit)!

(fas)! (eph)!

Molecules may function for both: 1. Selective adhesion 2. Intracellular signaling!



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Conserved Structural Classes of Axonal Guidance Molecules: Modular Construction and Multifunctionality!

1. Laminin, fibronectin and extracellular matrix proteins.

2. Cadherins and catenins. (Ca+2 dependent)

3. Cell adhesion molecules (CAMs) (containing IgG domains).

4. Receptor tyrosine kinases and receptor phosphatases.!





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Pioneer Neurons and Guidepost Cells guide the initial path of peripheral nervetracts in embryonic grasshopper

limbs!

(Bentley and Caudy, 1983)!


Pioneer Neurons

Guidepost Cells Growth Cone

CT1 Photoablated Control


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Early Embryonic Insect Neurons form a Repeated Segmental Scaffold


Grasshopper embryo

Commissural Tracts

Longitudinal Tracts

Identified Neurons

Q1 pCC aCC

MP1

Q1

Q1

MP1

MP1

pCC aCC

Q1 (Meyers and Bastiani, 1993)

Molecules Mediating Axonal Guidance!



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Drosophila robo disrupts longitudinal

tract formation

Robo acts as a receptor for a midline repulsive cue


Human ROBO Mutation causes HGPPS (Horizontal Gaze Palsy with Progressive Scoliosis)

(reduced hindbrain volume) (scoliosis)

(horizontal gaze palsy)

Jen, et al., 2004


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Purves 5th ed, pp. 533-4


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Purves 5th p. 531

Retinotectal Mapping Visualized by Dye Injection in Zebrafish!


(Friche,et al. 2001)


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Zebrafish ROBO Mutant (astray) Disrupts Midline Retinotectal Axonal Projections


September!17,!2012!

Do Molecular Cues Determine the Retinotectal Spatial-topic Map?!!

34!Lecture!VI.!Making!Connec<ons!

A (T) D

V

T N

Retina

P (N)

M (D)

L (V)

Optic Tectum

Optic tectum

A

P

dorsal

ventral

temporal

nasal

D

T

N

V

M

L

anterior

medial

posterior

lateral

A (T) P (N)

M (D) L (V)


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Molecules Mediating Axonal Guidance!

1.  Biochemical approach: Friedrich Bonhoeffer, retinotectal culture assay.

35!

Temporal Nasal Functional Assay

Fractionate Native Factors

Observe Neuronal Specificity

Purify and Identify Factor

(Ephrins...) Temporal Axons Nasal Axons

September!17,!2012! Lecture!VI.!Making!Connec<ons!


Purves 5th p. 520


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Pioneer Neurons Create the Early Scaffold of the Adult Nervous System


pioneer neuron

guidepost cells

growth cone

selective fasciculation

Pioneer neuron and guidepost cells may die after pathway is pioneered,

by apoptosis

Axonal Guidance Cues


selective fasciculation

diffusible attractant

diffusible repellant

Contact-dependent attractant

Contact-dependent repellant

(Timing is critical)


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What this Lecture was about

•  Grouping neurons and processes

•  Partner selection

•  Some genetic foundations/correlates

•  Systematic organization of connections

•  Roles of contact and diffusion

•  Deja vu


Sequential Restrictions (Refinements) are the Bases for Development


pluripotent, stem cell

differentiated

genetic

environmental


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Finis

lecture vi. making connections - nslc · bio 3411 lecture vi. making connections 9/17/12 2 what the...

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