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Lecture VI. Making Connections
Bio 3411 Monday
September 20, 2010
September 20, 2010 1Lecture VI. Making Connections
†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.
_________________
ReadingNEUROSCIENCE: 4th ed, Chapter 23, pp
577-609
2Lecture VI. Making ConnectionsSeptember 20, 2010
†(pdfs on course websites: [http://artsci.wustl.edu/~bio3411/] & [http://www.nslc.wustl.edu/courses/Bio3411/bio3411.html]
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 20, 2010 Lecture VI. Making Connections 3
September 20, 2010 Lecture VI. Making Connections 4
September 20, 2010 Lecture VI. Making Connections 5
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
September 20, 2010 Lecture VI. Making Connections 6
September 20, 2010 7Lecture VI. Making Connections
Fertilization
Embryonic morphogenesis
Induction of Neuroectoderm
Neurulation
Differentiation:1. Formation and placement of neuroblasts2. Axonal outgrowth3. Growth cones, selective migration4. Selective fasciculation5. Target selection6. Synaptogenesis7. Etc…(cell shape, neurotransmitter, ionic channels, receptors)
Adult neuronal plasticity(Activity-dependent?)
Seg
men
tati
on
Outline of Neurodevelopment
Selective Adhesion Determines Specificity of Tissue and Cellular Associations
September 20, 2010 8Lecture VI. Making Connections
Selective Aggregation of dissociated embryonic tissues (vertebrate and invertebrate) suggests ancient (surface)
Adhesion Molecules
September 20, 2010 9Lecture VI. Making Connections
(Townes & Holtfretter, 1955)
1. Sponges (Wilson, 1907)
2. Amphibians (Townes and Holtfretter, 1955)
3. Chick (Moscona, 1952)
Epidermis + Mesoderm
September 20, 2010 10Lecture VI. Making Connections
Experimental recreation of morphogenesis by mixing cells expressing low and high levels of one surface
adhesion gene (N-cadherin)
Green = high N-cadherinRed = low N-cadherin
+4 hrs +24 hrs
(Foty and Steinberg, 2004)
Growth Cones are Dynamic Sensory Organelles that Guide the Growth of Embryonic Axons
September 20, 2010 11Lecture VI. Making Connections
(Forscher lab)
Sensing and Transducing:• Diffusible Cues• Contact-dependent Cues• Trophic Factors• Neurotransmitters
Extracellular Cues
Intracellular SignalingPathways
CytoskeletalRearrangment
Ca+2
GTP
cAMP
2ndMessengers
September 20, 2010 Lecture VI. Making Connections 12
September 20, 2010 Lecture VI. Making Connections 13
(Play GFP-Actin Growth Cone Movie)Dr. Andrew Matus
Friedrich Miescher Institute, Switzerland
September 20, 2010 Lecture VI. Making Connections 14
September 20, 2010 Lecture VI. Making Connections 15
September 20, 2010 Lecture VI. Making Connections 16
September 20, 2010 Lecture VI. Making Connections 17
Effect/Proximity
Attraction Repulsion
DistantContact
Functional Classes of Axonal Guidance Molecules
(Secreted)
(Membrane Associated)
(netrin)
(sema, slit)
(fas)
(eph)
Molecules may function for both:1. Selective adhesion2. Intracellular signaling
September 20, 2010 18Lecture VI. Making Connections
September 20, 2010 Lecture VI. Making Connections 19
September 20, 2010 Lecture VI. Making Connections 20
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.
September 20, 2010 21Lecture VI. Making Connections
September 20, 2010 Lecture VI. Making Connections 22
September 20, 2010 Lecture VI. Making Connections 23
Pioneer Neurons and Guidepost Cells guide the initial path of peripheral nervetracts in embryonic grasshopper
limbs
(Bentley and Caudy, 1983)
September 20, 2010 24Lecture VI. Making Connections
PioneerNeurons
Guidepost Cells Growth Cone
CT1 Photoablated Control
Early Embryonic Insect Neurons form a Repeated Segmental Scaffold
September 20, 2010 25Lecture VI. Making Connections
Grasshopperembryo
Commissural Tracts
Longitudinal Tracts
Identified Neurons
Q1 pCCaCC
MP1
Q1
Q1
MP1
MP1
pCCaCC
Q1(Meyers and Bastiani, 1993)
Molecules Mediating Axonal Guidance
September 20, 2010 26Lecture VI. Making Connections
September 20, 2010 Lecture VI. Making Connections 27
Drosophila robo disrupts longitudinal
tract formation
Robo acts as a receptor for a midline repulsive cue
September 20, 2010 Lecture VI. Making Connections 28
Human ROBO Mutation causes HGPPS(Horizontal Gaze Palsy with Progressive Scoliosis)
(reduced hindbrain volume) (scoliosis)
(horizontal gaze palsy)
Jen, et al., 2004
September 20, 2010 Lecture VI. Making Connections 29
September 20, 2010 Lecture VI. Making Connections 30
September 20, 2010 Lecture VI. Making Connections 31
Retinotectal Mapping Visualized by Dye Injection in Zebrafish
September 20, 2010 32Lecture VI. Making Connections
(Friche,et al. 2001)
Zebrafish ROBO Mutant (astray)Disrupts Midline Retinotectal Axonal Projections
September 20, 2010 33Lecture VI. Making Connections
September 20, 2010
Do Molecular Cues Determine the Retinotectal Spatial-topic Map?
34Lecture VI. Making Connections
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)
Molecules Mediating Axonal Guidance
1. Biochemical approach: Friedrich Bonhoeffer, retinotectal culture assay.
35
Temporal NasalFunctional Assay
Fractionate Native Factors
ObserveNeuronal Specificity
Purify and IdentifyFactor
(Ephrins...)Temporal AxonsNasal Axons
September 20, 2010 Lecture VI. Making Connections
September 20, 2010 Lecture VI. Making Connections 36
Pioneer Neurons Create the Early Scaffold of the Adult Nervous System
September 20, 2010 37Lecture VI. Making Connections
pioneer neuron
guidepost cells
growth cone
selectivefasciculation
Pioneer neuron and guidepost cells may die after pathway is pioneered,
by apoptosis
Axonal Guidance Cues
September 20, 2010 38Lecture VI. Making Connections
selectivefasciculation
diffusibleattractant
diffusiblerepellant
Contact-dependentattractant
Contact-dependentrepellant
(Timing is critical)
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
September 20, 2010 Lecture VI. Making Connections 39
Sequential Restrictions (Refinements) are the Bases for Development
September 20, 2010 40Lecture VI. Making Connections
pluripotent, stem cell
differentiated
genetic
environmental
September 20, 2010 Lecture VI. Making Connections 41
September 20, 2010 Lecture VI. Making Connections 42
Finis