Lecture 23- Axon Regeneration

Question 1

Can neuron survive an axon injury?

Answer 1

-A neuron may survive axotomy (at least for a time) -axotomy= severed -the proximal part can survive, but it depends -only in the PNS in mammals

Question 2

Can mammals regenerate axons in the CNS?

Answer 2

-Mammalian CNS injury: no axon regeneration

Question 3

What happens when you have an injury in the CNS?

Answer 3

• Distal portion of severed axon degenerates
• Dystrophic neurons no growth cone, no crossing of lesion site
• Some compensatory axon sprouting
• only short distance
• the axon that is spared tries to compensate by more sprouting but this will only somewhat help functionality

Question 4

What is the aim in axon regenaration?

Answer 4

1: Survival of neuron
2: Generation of growth cone= through lesion site
3: Growth of axon=through lesion site
4: Extension of axon= beyond lesion site
5: Re-establishment of connection with target
6: Functional recovery
- most of the research is concentrating on the first 4 stages

Question 5

Where can spontaneous axon regeneration occur?

Answer 5

-Peripheral nervous system Neuron can have axon in both CNS and PNS, but only regenerates in PNS - CNS of invertebrates and lower vertebrates (dorsal root ganglion= have process both in pNS and CNS but only PNS can regenerate)

Question 6

What is often used as a model for axon injury?

Answer 6

-SCI and optic nerve crush are models of axon injury

Question 7

How was it shown that peripheral nerve grafts can support CNS axon regeneration?

Answer 7

• damage a nerve and then provide a peripheral graft, the axon regenerate in the PNS nerve graft but not in the CNS
• CNS axons are capable of regeneration when
• Regrowth stops at CNS -Same result seen in injured spinal cord

Question 8

How do the mature CNS and PNS environments differ?

Question 9

What happens to a neurite in vitro when it comes into contact with oligodendrocytes?

Answer 9

-In vitro: contact with oligodendrocytes and CNS myelin extracts prevents neurite outgrowth

Question 10

What is myelin?

Answer 10

• Myelin is part of the specialised oligodendrocyte membrane - Composed of lipids and proteins

Question 11

What is Nogo?

Answer 11

-a myelin associated inhibitor -one of the inhibitory factors in myelin -Nogo gene generates 3 isoforms: Nogo A, B and C -Nogo A also has inhibitory domains in its N-terminal sequence -Nogo66 is the inhibitory domain that is common to all nogo isoforms

Question 12

What is the receptor for Nogo?

Answer 12

-Nogo receptor (NgR1) on neurons throughout CNS

Question 13

Does NgR1 have an intracellular domain?

Answer 13

-NgR1 lacks intracellular domain

Question 14

What are the signalling subunits of NgR1?

Answer 14

• p75 / TROY and Lingo-1 act as signalling subunits -they activate Rho GTPase= causes collapse of growth cone (chnage in cytoskeleton) -Rho activation = growth cone collapse

Question 15

What are the inhibitory componenets of myelin?

Answer 15

• Nogo-A, MAG and OMgp have different structures but bind same receptor complex
• MAG and OMgp inhibit neurite outgrowth in vitro

Question 16

What happens in Nogo knockout mice?

Answer 16

• Results from Nogo knockout mice vary in different studies
• Treatment with antibody that blocks Nogo-A induces axon sprouting / regeneration

Question 17

What prevents axon regeneration?

Answer 17

-Extrinsic factors: the lesion evironment: Inhibitory components in myelin debris -Nogo, MAG, OMgp are components of myelin which inhibit axon regrowth -Inhibitory myelin components signal via receptors (NgR1, NgR2) and co- receptors (p75 / TROY, Lingo-1) on the neuronal membrane - Contact with myelin inhibitors induces cytoskeletal changes in the neuron:leads to growth cone collapse, prevents axon growth through lesion site

Question 18

What is the astrocyte response in injury in the CNS?

Answer 18

• astrocytes perform a variety of roles= important in support roles
• after disease or trauma to the CNS= respond and get astrogliosis = the astrocytes change, become bigger, and thicker

Question 19

What is the glial scar?

Answer 19

-glial scar= major component are the big astrocytes, but a avriet yof cells in the glial scar -physical barrier= astrocyte shave dense processes so the axons cannot penetrate -also chemical -Forms over time following injury -Physical and chemical barrier to axon regeneration

Question 20

What is elevated in the glial scar?

Question 21

What does the glial scar do?

Answer 21

-The glial scar limits the spread of immune cells -limits the inflammation to a localized area, thus it is good - The inflammatory response is necessary but inefficient - debris / other debris - Immune cells release both growth-promoting and neurotoxic substances - may increase area of damage -the immune cells can stick around for a long time, even years= chronic neuroinflammation= probably unhelpful

Question 22

Is the inflammatory response started by the astrocytes necessary and efficient?

Answer 22

• The inflammatory response is necessary but inefficient
• debris / other debris
• Immune cells release both growth-promoting and neurotoxic substances
• may increase area of damage
• when the glial scar is removed= the immune cells are released and damage more area

Question 23

What is the effect of guidance molecules in injury?

Answer 23

-Developmental axon guidance molecules are expressed in the injured CNS -Still present in mature CNS or re-upregulated following injury -Many repulsive guidance molecules - Expression on neurons, astrocytes, oligodendrocytes/myelin

Question 24

What happens at at a lesion site in the CNS?

Answer 24

-Multiple Eph / ephrin interactions at the lesion site -e.g. Interaction of EphA4 (neuron) with ephrin B3 (myelin) Interaction of EphA4 (astrocytes) with ephrins (on reactive astrocytesn as well) -some developmental guidance molecules are still expressed in the injured adult CNS -mostly repulsive guidance cues -have ephrin and semaphorin -eph receptors= tyrosine kinase receptors

Question 25

What happens in EphA4 knockout mice?

Answer 25

-EphA4 knockout mice have increased axon regeneration and functional recovery following SCI (spinal cord injury)

Question 26

What prevents axon regeneration (the long one)?

Answer 26

-Extrinsic factors: the lesion evironment 1.The glial scar & inflammation - glial scar is a chemical and physical barrier to axon regeneration - e.g. upregulation of CPSGs inhibitory ECM component - inflammatory response is inefficient and contributes to toxicity 2.Axon guidance molecules - repulsive axon guidance ligands and receptors are present / upregulated after injury (ephrin, semaphorin, slit, netrin families) - in glial scar, oligodendrocytes/myelin, neurons - causes growth cone collapse and prevents axon regrowth into lesion site

Question 27

What are the differences between regeneration and development environment?

Answer 27

1.Regeneration: -environment : Oligos and myelin Reactive astrocytes Inflammatory -guidance signals: Re-upregulated but inhibitory -growth signals: Not expressed in way that supports regrowth 2. Development: -non-myelinated, no astrocyte reactivity, non-inflammatory -guidance signals: re upregulated, but inhibitory -growth signals: abundant -comparison of the dev. plus adult environment = normally would only grow in development with regeneartion must deal with :

Question 28

What is also true of the regeneration vs development?

Answer 28

-must grow much longer way than in development as the adult is larger than the baby, -Mature CNS axons are capable of regeneration -How well can mature CNS neurons actually regrow?

Question 29

What are the intrinsic neuronal differences between the regeneration vs development?

Answer 29

-Developing and mature neurons respond differently to their environment inhibitory myelin components -Even in an optimal (non-inhibitory) environment, mature CNS -We need (lots of) regenerating axons to travel a long distance

Question 30

What do younger neurons have higher than the older neurons?

Answer 30

-Young neurons have higher levels of cAMP, how to boost the intrinsic ability of neurons to grow, the developing neurons= have - Acute effect: conversion of repulsive into attractive signal - Long term effect: activation of transcription factors

Question 31

What does cAMP do?

Answer 31

-cAMP promotes neurite outgrowth -cAMP and Nogo and MAG and OMGp go against each other

Question 32

Can the regeneration be helped with higher cAMP?

Answer 32

-yes -one way of improving the regeneration = prevent cAMP breakdown or put in cAMP homologue

Question 33

What is the mTOR/PTEN and protein synthesis?

Answer 33

- regenerating axons need increased protein synthesis - mTOR levels decrease in neurons with age, and following axotomy Phosphatase and tensin homolog (PTEN) is a negative regulator of cell growth PTEN deletion = increased protein synthesis - if you manipulate PTEN= negative regulator of mTOR/PTEN

Question 34

What happens to mice with PTEN deletion?

Answer 34

-PTEN deletion increases axon regeneration and sprouting following injury -PTEN deletion enhances growth cone protein synthesis -Allows extensive regeneration though inhibitory lesion site -surprisingly good at increasing sprouting and regeneration

Question 35

What are the intrinsic factors preventing axonal regeneration?

Answer 35

-Intrinsic factors: capacity of the neurons to regrow - mature CNS neurons have not lost the capacity to regenerate axons, but their intrinsic capacity for growth has diminished with age - mature neurons have lower levels of cyclic AMP (cAMP) than developing neurons, which alters the growth cone response increasing cAMP in mature neurons a potential treatment strategy - regenerating axons must travel longer distances than developing axons mice with PTEN deletion have enhanced protein synthesis, promoting axon growth through the inhibitory lesion environment

Question 36

Is the capacity of neurons diminished over time?

Answer 36

-cell intrinsic mechanisms: capacity of the neurons to regrow -diminished in mature compared to developing neurons

Question 37

What is the evolutionary explanation for the reduced regenerative ability of mature neurons?

Answer 37

-The types of injury that would damage the CNS would almost certainly lead to rapid demise in the wild before regeneration could take place -The most likely evolutionary explanation for the loss of CNS regenerative capacity is that this is an unselected by-product of gaining the increasingly complex nervous systems that selection pressures have favoured over time

Question 38

What treatments are developed to promote axon regeneration?

Answer 38

- Multiple interventions? - Neuroprotective factors - Block inhibitory factors in myelin - Block repulsive axon guidance molecules -Digest CSPGs in glial scar - Inhibition of inflammatory response - Promote re-myelination of surviving neurons - Neurotrophins to promote regeneration - Transplantation of cells - Olfactory ensheathing cells: neuroprotective - Human embryonic stem cells: re-myelination of spared axons - Induced pluripotent stem cells