L20: Cell interactions in neural development Flashcards
Neural tube formation
- Notochord forms from mesoderm cells soon after gastrulation is complete
- Signals form notochord cause inward folding of ectoderm at the neural plate
- Ends of neural plate fuse and disconnect to form an autonomous neural tube
-> Neural fates induced by Shh gradient from floor plate (induced by notochord)
Stages of neuronal development
- Specification of neurons (commisural interneurons, dorsal root ganglion neurons, motor neurons)
- Axon outgrowth
- Making synapses
Notch and neural fate - overview + evidence
- Notch inhibits the neural fate in the neuroectoderm of Drosophila
- Notch mutants have too many neurons; expansion of the neuroectoderm dorsally
Example system for notch function in Drosophila…Phenotypes for different levels of expression
- Sensory bristles
- All four sensory bristle cells (hair, socket, sheath, sensory neuron) derived from one sensory organ precursor cell (SOP)
-
Notch expression level phenotypes…
NORMAL: Wt
HIGH: missing bristles
LOW: extra bristles
Notch basic info and structure
- Notch (type I rec.), DSL (delta/serrate/lag2 ligand)
- Highly conserved; heterodimeric proteins linked covalently
- Many EGF-like repeats in extracellular domain of both ligand and rec. which interact; rec. also has ankyrin repeats in intracellular domain (NICD)
-> involved in many developmental processes, can regulate cell proliferation (cancer)
Notch signalling
- Binding of DSL ligand to Notch rec. (via both of their EGF repeat sections) causes proteolytic cleavage of the Notch intracellular ‘ankyrin repeat’ domain (NICD)
- Translocated to nucleus, binds to CSL (CBD-1/Su(H)/lag1)
-> target genes expressed
Proneural clusters (process of lateral inhibition)
- A single SOP is specified from the cells in a proneural cluster (through lateral inhibition)
- One cell adopts a fate and inhibits surrounding cells from adopting the same fate
- Initially all cells express equal Notch and Delta
- One cell stochastically accumulates more delta (suppressor-of-hairless->enhancer-of-split proteins->achaete-scute-proteins)
- As a result, less delta transcribed in lateral cells (suppression of SOP fate), corresponding increase in delta in SOP (positive feedback loop)
-> at threshold, activation of bHLH transcription factors in SOP
-> neuronal fate specified - Limits concentration of bristles that eventually form
Growth cone structure and function
- Growth cones migrate towards correct target; dynamic structures driven by m.tubs and actin filaments…
- Filopodia: actin microfilaments, actin polymerisation/depolymerisation determines direction of movement
- Lamellipodia ( meshwork of actin microfilaments)
- Receptors on cell surface (operate by chemotaxis; long and short range, attractive and repulsive)
Examples of molecules involved in chemotaxis of growth cones
Long range…
- Netrins (chemoattraction)
- Semaphorins (chemorepulsion)
Short range…
- Cadherin (contact attraction)
- Ephrins (contact repulsion)
Commissural neurons
- Connect neurons to other neurons across the two sides of the body
- Grow towards the floor plate (from dorsal side)
- Floor plate is chemoattractive to these neurons
- Crossing the floor plate alters the chemotaxis of commissural neurons (transplantation experiments; involvement of Unc-5, no longer respond to netrin, proceeding to next step)
Netrin and its rec.s
-Similar to Unc-6
- Vertebrates have netrin 1 (expr. in floor plate cells) and netrin 2
- Acts as chemoattractants and repellants (trochlear motoneurons)
Receptors…
- Unc-40: Large TM protein, Ig and fibronectin repeats, expr. in commissural neurons, Abs to Unc-40 prevent axons from growing towards netrin
- Unc-5: Large TM protein. Ig repeats, large IC domain, may modify the response of Unc-40
Evidence for interaction of Unc-5 and Unc-40
- Neuron expressing Unc-40 in culture grows towards source of netrin (released from micropipette)
BUT this activity can be prevented by Abs against Unc-40 - Neuron expressing Unc-40 and Unc-5 no longer grows towards source of netrin
Role of the 3 types of neurons in dev
- Commissural: Connect neurons to other neurons across the two sides of the body
- Dorsal root ganglion: Sensory neurons make connections w/ rec.s in periphery (e.g. pain, temperature)
- Motor: Make direct connections w/ muscles at the NM junction
Evidence for changing response to netrin
- Transplantation expts; in explant, neurons grow towards floor plate (FP) but once they cross it, no longer respond to FP in another explant (ectopic FP placed contralaterally)
-> crossing floor plate alters chemotaxis of neurons - Continue to respond to ectopic FP in absence of FP in explant itself
Axons at target tissue (example)
-> making connections
- In muscle, axon forms synapse at neuromuscular junction (NMJ)
- Axon terminal, synaptic cleft (EC rich environment containing lots of fibers of basal lamina), cell
- Presynaptic cell releases vesicles of neurotransmitter, exocytosis into cleft, diffuse across to signal muscle cell
- Key neurotransmitter: Acetylcholine
