The Guidance of Axons to Their Targets

1. The Guidance of Axons to Their Targets 서울대학교 어린이병원 신경외과 왕 규 창

2. Axon Growth: Two Views • molecular view • random growth with selective survival • stereotropism • mechanical guidance • along scratches, blood vessels or cartilage • resonance • congruent activity

3. Retina-Tectum Experiment • Roger Sperry, 1940s • lower vertebrates • regeneration of cut retinal axons • frog, cut optic nerve, rotation of the eye

4. Retina-Tectum Experiment • chemical matching rather than functional validation of random connection • chemospecificity hypothesis • recognition molecules • Molecular matching predominates during embryonic development. • Activity (experience) modifies the circuits once they have been established.

5. Axon Guidance Cues • from the target (wrong) • series of discrete steps • retina – optic fiber layer – retinal basal lamina and end-feet of glia – optic nerve head – optic stalk (‘pioneer’ axons) – optic chiasm- ventral diencephalon – superior colliculus at different subregions – - radial glial cells - synaptic partner – a specific layer – specific area of dendrites

6. Axon Guidance Cues • optic chiasm • different responses to special midline cells • intermediate targets • terminal arbor • interaction with target • patterns of neural activity

7. Positional Cues • correction of ‘mistaken’ fibers • ephrins • rotation of neural tube between the chiasm and the tectum • markers of position or polarity

8. Motor Axon Guidance • series of discrete steps • motor pool in the spinal cord – segmental ventral roots by barriers in the somites – rearrangement in plexus region – large nerve – target muscle – synapse on a muscle fiber • similar pattern to the retinal fibers

9. Axon Growth: Options • grow / turn / stop • positive and negative cues • finer control over the direction of growth

10. Pioneers • short distance in a small embryo • the first axons to exit the retina

11. Intermediate Targets • decision points • optic chiasm, limb plexus

12. Gradients • cell surface molecules • soluble molecules

13. Ranges of Cues • short-range cues • cell membranes • extracellular matrix • precise contact guidance • long-range cues • soluble molecules • less precise guidance

14. Growth Cone • Santiago Ramon y Cajal, 1890s • “both a sensory structure and a motor structure” • transduces positive and negative cues into signals that regulate the cytoskeleton and thereby determine the course and rate of axon outgrowth • coupling between the sensory and motor capabilities

15. Growth Cone • central core • microtubules, mitochondria, etc • lamellipodia • motile, ruffled appearance • filopodia • long slender extensions

16. Filopodia • sensory capability • rod-like, actin-rich, membrane-limited • highly motile: advance, retract, turn • length • rapid movement • flexibility

17. FilopodiaSecond Messengers • calcium • set point: optimal concentration • gradient of calcium: change of direction • cyclic nucleotides • modulate protein kinases, protein phosphatases, rho-family GTPases

18. Pathway Guidance Cues • promotion / inhibition • cell surface / extracellular matrix / soluble form

19. Pathway Guidance Cues • extracellular matrix adhesion • cell surface adhesion • fasciculation • chemoattraction • contact inhibition • chemorepulsion

20. ECM Adhesion • collagen, fibronectin, proteogylcans, etc • laminins • heterotrimer, at least 14 trimers, unique distribution, position- or stage-dependent signals • integrins • heterodimer, at least 16 alpha and 8 beta chains, specific ligands • all cells in the body: at least one integrin

21. Cell-Cell Adhesion • selective adhesive interaction • cadherin: calcium dependent • immunoglobulin-like adhesion molecules: calcium independent • cell-cell binding • short-range promoter of neurite growth

22. Cadherins • at least 100 related membrane-spanning glycoproteins • extracellular calcium binding segments • N-cadherin, proto-cadherins, cadherin-related neural receptors • cells throughout the body

23. Cadherins • homophilic interaction • prefers to bind to its own kind • selective adhesion • Cytoplasmic domain binds catenins, then affects cytoskeletal elements.

24. Adhesion Molecules • adhesion assay • Initial adhesion triggers a cytoplasmic reaction that strengthens the adhesion. • not just adhesion molecules but ‘signaling molecules’ activated by membrane receptors

25. Ig Superfamily • disulfide bridges • less ligand-specificity than cadherins • intracellular domain: protein tyrosine phosphatase or protein tyrosine kinase

26. Chemoattractants • soluble growth factors • trophic factor • chemotaxis: tropism • no clear examples of trophic and tropic factors in vivo • neuronal chemoattractants • two glycoproteins: netrins

27. Vertebrate vs. C. elegans • netrin unc-6 • unc-5H unc-5 • DCC, neogenin unc-40 • Ig superfamily • remarkably conserved during evolution

28. Ephrins • stripe assay • axons from temporal retina to the anterior tectum • heat treatment of each membranes • heat treatment of posterior membrane: random growth • presence of inhibitory material in posterior membranes

29. Ephrins • repulsive axon guidance signal (RAGS) = ephrin A5 • eph kinases: receptor tyrosine kinases • major group of inhibitory ligands and receptors in the developing nervous system

30. Ephrins • ephrin A2 and ephrin A5 • low-to high gradients in the rostral direction in the tectum • eph A3 (a kinase receptor which binds ephrin A2 and A5) • low-to-high gradient in the temporal direction in retinal ganglion cells

31. Somaphorins • an inhibitory molecule • at least 15 somaphorins • in distinct types of neurons and nonneural cells • key receptor: neuropillins, plexins

32. Chemorepulsion • semaphorins • some: membrane-bound • others: soluble factor • netrins • DDC or neogenin: attraction • unc-5H: repellant

33. Chemorepulsion • attract or inhibit according to the receptors or the level of intracellular messengers • some neurotransmitters • One synaptic transmission inhibits formation of another.