MECHANISMS OF AXON GUIDANCE Flashcards
How does specific neuronal activity arise in the adult organism?
There are two hypotheses: The Weiss Resonance Theory and the Sperry Chemoaffinity Hypothesis.
What is the Weiss Resonance Theory?
Random and diffuse neuronal outgrowth occurs to all targets followed by elimination of non functional connections.
What is the Sperry Chemoaffinity Hypothesis?
Directed specific outgrowth occurs through axons following individual identification tags carried by the cells and fibres in the embryo.
How did we find out if the Weiss Resonance Theory or the Sperry Chemoaffinity Hypothesis was correct?
By testing the visual system of amphibia.
The optic nerve was cut and the temporal retina was removed.
If Weiss was correct there would be random and large outgrowth followed by elimination to the end product of regrowth to the tectum. If Weiss was correct, the optic nerve would grow back to the end product straight away.
Weiss was correct.
Axon pathways in embryos are highly stereotyped. What has also been shown in chick embryos?
That motor axons are guided specifically to their targets.
By cutting and replacing or reversing segments of the neural tube you still get correct motor axon outgrowth to their normal muscle targets. This suggests that axons are actively navigated.
From experiments of those such as reversal of the neural tube in the chick embryo, it can be concluded that there are factors in the environment axons use to find their correct targets. What are these called?
Guidance cues.
Why are insects good examples of model organisms?
Simple nervous systems
Embryos easy to observe and manipulate
Individual cells can be ablated (in larger insects)
How have guidance cues been mapped?
In the grasshopper, a detailed analysis resulted in the indentification of almost every neuron in the embryonic nerve cord allowing a map of axon projections to be made. There were highly stereotypes embryo to embryo, segment to segment.
Where are guidance cues found?
Reproducibility suggests that growth cones are responding to cues in the environment. Cues can be found on axons. Pathways seems to change when specific axons are encountered. If certain cells are ablated, most axons continue normally whilst others stall. Therefore, it must be that this axon growth cone is looking for specific cues from the ablated axon. This is the labelled pathway hypothesis.
What is the labelled pathway hypothesis?
Axons can selectively fasciculate with other axons.
Axon surfaces carry cues.
Different axon growth cones express different sets of receptors fo r such cues.
Early axons from an axon scaffold on which later axons can extend.
Establishes axon surfaces as one potential source of guidance cues.
Give an example of an axon scaffold in vertebrates.
Subplate neurons in the mammalian cortex.
Project from the cortex to thalamus prior to innervation of cortex by LGN.
Ablation of the subplate leads to failure of LGN.
How do the first axons find their way in an apparently featureless environment?
Pioneers follow a stereotyped pathway also. Growth cones of pioneers react at specific points of their route.
Give an example of a pioneer growth cone reacting to something in its route.
In the grasshopper limb bud, pioneer Ti1 makes a specific turn at the limb boundary and again as it approaches a specific cell, Cx1. Cells like this in the pathway are called guidepost cells.
What do the presence of guidepost cells imply?
That there are molecular difference in environment.
Patterning in the DV axis will predict where fibre tracts will form.
Axon guidance cues are located, not just on other axons but in many cell types too.
What are the four forces of axon guidance?
Chemoattraction
Chemorepulsion
Contact Attraction
Contact Repulsion
How are neurons different at each end?
Neurons come in many shapes and sizes but all exhibit polarity.
Axons have highly polarised microtubules (+ end towards growth cone) whilst dendrites have microtubules in mixed orientations.
The different microtubule organisation is due to localisation of different types of microtubule associated proteins which determine how the microtubules are cross linked.
Axons express Tau
Dendrites express MAP2
How is polarity determined?
By neurite selection. This seems to be a random choice as GFP shows ends being 'tried out'. Neurites contain dynamic microtubules (tyrosinated). Stabilited microtubules (acetylated) are present only in the newly polarised axons. Artificial stabilisation of microtubules by localised taxol application to one neurite selects for axon formation. By contrast, removal of a newly selected axon leads to another neurite being selected. This suggests there is competition between axons to stabilise microtubules and a feedback loop to prevent other neurites being selected.
In vivo, polarity is establishes as neurons are born. Radial glial cells are already polarised and polarity is preserved as cells undergo division. What does this imply about neurite selection?
Neurite selection is likely to be biased by the cellular environment.
Describe the anatomy of a growth cone.
A growth cone is made up of central, transitional and peripheral domains. The central region is formed of microtubules. The transitional and peripheral regions are made up of F actin.
In the peripheral regions, lamella and filopodia make up different kinds of F actin. In filopodia, the actin bundles are polarised to form larger bundles. In lamella, the actin bundles are cross linked into a net.
What happens in a resting growth cone?
F actin treadmills in a resting growth cone ie moves from hte periphery to the centre where it breaks down and joins the tip. In the resting growth cone, tubulin is dragged sporadically into the filopodia.
What happens in the presence of an attractive cue?
Tubulin is dragged into the filopodia more drammatically and the growth cone reorganises.
F actin treadmilling slows and stabilises the filopodium, dragging microtubules to the back of the filopodium via an actin tubulin link. The cue acts as a molecular clutch driving the growth cone towards the cue.
Growth cones can be repelled as well as attracted. How was this discovered?
When mixtures of neurons were in culture, they were found to fasciculate with their own kind are being repelled by different axon types. This is called growth cone collapse.
How does growth cone collapse occur?
A repellant cue causes growth cone collapse by destabilising F actin. (the opposite to an attractive cue)
Name a family of inhibitory guidance cues.
Semaphorins
How were semaphorins discovered?
Biochemical purification of the factor from the retina responsible for causing collapse of sensory axons led to identification of this family of inhibitory guidance cues.
Was is the primary effect of a growth cone of semaphorins?
Causes primarily F actin collapse
What flavours do semaphorins come in?
Many flavours but in most cases, membrane bound and secreted.
Axons cannot growth where they cannot attach. What is the link between strength of adhesion and amount of axon growth?
There is no simple link between adhesion and axon growth. Growth cones also need substrates permissive for growth, attachment is not enough.
This can be shown in the case of outgrowth of extra-cellular matric components: whilst collagen is more adhesive than laminin, there is more outgrowth from laminin.
How can you measure adhesion?
The speed of centrifuge that causes detachment.
What must occur for a growth cone to be repelled?
It still must attach to a repellant axon.
Permissive factors can define substrate paths in the embryo. Laminin promotes growth in the optic nerve but it does not dicatate the direction of growth, only that it can grow there. What can be say about laminin?
Laminin is permissive but not instructive.
