Brain development and repair

Question 1

What are the consequences of failure for neural tube to close completely?

Answer 1

1. Failure at rostral end: Anencephaly
2. Failure at caudal end: Spina bifida

Question 2

What are the types of spina bifida?

Answer 2

1. Spina bifida occulta: Hidden spina bifida. The spinal cord is not fully enclosed within the vertebrae, but the overlying skin is normal. These cases are often asymptomatic.
2. Meningocele: Gap in vertebrae allows the protrusion of meninges outside to create herniation. No protrusion of spinal cord means no neurological symptoms.
3. Meningomyelocele: Gap in vertebrae allows the protrusion of meninges and spinal cord outside, resulting in complications such as bladder dysfunction and hydrocephaly.

Question 3

What determines anterior → posterior patterning of the neural tube?

Answer 3

1. Expression of different Hox genes along the length of the neural tube (Hox code).
2. Secretion of factors from specific “signalling centres” along neural tube.

Question 4

What is the role of the midbrain-hindbrain boundary (isthmus) in mediating neural tube development?

Answer 4

Secretes Fgf8, inducing development of the tectum of midbrain rostro-dorsally and substantia negra rostro-ventrally, cerebellum of the hindbrain caudo-dausally and motor neurones of cranial nerve [III] & [V] caudo-ventrally.

Question 5

What is the process of ventral → dorsal patterning of neural tube?

Answer 5

1. Notochord secretes SHH.
2. Neural tube directly dorsal to notochord receive relatively high [SHH] and develop into floor plate.
3. As SHH diffuses more dorsally, concentration decreases, resulting in development of motor neurones and inhibition of floor plate and sensory neurone development.
4. SHH is also secreted by the floor plate itself and creates concentration gradient that supports further development of the neural tube, including the roof plate.

Question 6

What is the process of radial patterning of the neural tube?

Answer 6

1. As the neural tube develops, it is organised into stratified neuroepithelium, with multi-potent stem cells dividing in the ventricular (innermost) layers.
2. Subsequent layers form in an inside-out fashion, with the layer VI developing first, and layer I last.
3. Multipotent stem cells differentiate into radial glial cells that send processes all the way to the most superficial aspects of the neuroepithelium.

Developing neurones (also from ventricles) follow the processes of the glial cells in order to reach surface of neuroepithelium.

Question 7

What is the structure and function of growth cones?

Answer 7

Structure:

• At the distal end of every developing neurone is a growth cone, responsible for guiding the growing axon.
• The growth cone is made up of lamellipodium and filopodium.

Function:

• It depends on attraction, adhesion and repulsion in order to guide growing axons roughly to their synapsing targets.

Question 8

What are the different mechanisms by which growing axon can be guided?

Answer 8

1. Diffusible attractive molecule
2. Diffusible repulsive molecule
3. Contact attraction
4. Contact repulsion

Question 9

What are the types of diffusible attractive molecules?

Answer 9

1. Neurotrophic: Promotes axon survival and further growth
2. Neurotropic: Attracts growth cones towards source of factor

Question 10

What are examples of diffusible attractive molecules?

Answer 10

1. NGF
2. Netrin

Question 11

What are diffusible repulsive molecules?

Answer 11

They repel the growth cones away from their source

Question 12

What are examples of diffusible repulsive meolecules?

Answer 12

1. Semaphorins
2. Netrins
3. Slit

Question 13

What is the mechanism of contact attraction?

Answer 13

Growth cones make contact with adjacent cells via CAM molecules. Adhesion of cones onto CAMs on nearby cells causes activation of signalling cascades resulting in growth of axon towards the cells.

Question 14

What are examples of contact attractive molecules?

Answer 14

CAMs

Question 15

What is the mechanism of contact repulsion?

Answer 15

Contact with another cell causes growth cone to steer away from cell.

Question 16

What are examples of contact repulsive molecules?

Answer 16

1. Semaphorins
2. Ephrins
3. Some proteoglycans

Question 17

What is the process of growth of spinal cord commissural (pain/temperature) fibres?

Answer 17

1. Dorsal spinal neurone axons are attracted ventrally by netrin, an attractive factor produced by floor plate.
2. Once the axons reach the floor plate, they decussate to the other side of the spinal cord (facilitated by TAG-1 binding to Nr-CAM).
3. Once axon crosses floor plate, it turns to grow rostrally. A number of other change occur including:
   - Replacement of TAG-1 with L1-CAM to promote fasciculation (bundling together) in formation of spinothalamic tract.
   - Decreased sensitivity to netrin and increased sensitivity to inhibitory factors (e.g. Sema3b, Slit2). These changes ensure the axons don’t cross back.

Question 18

What is the structure of the fish/amphibian retinotopic map in the tectum?

Answer 18

Temporal retina → Anterior tectum

Nasal retina → Posterior tectum

Dorsal retina → Ventral tectum

Ventral retina → Dorsal tectum

Question 19

What evidence is there for the fact that development of the retinotopic map in fish/amphibians is mediated by interactions between axons and tectal tissue?

Answer 19

1. Severing eyeballs of frog and rotating them 180^o resulted in retinal fibres regenerating to innervate same places of tectum, resulting in displacement of visual reflexes by 180^o (Roger Sperry).
2. Excising half of the retina resulted in other half regenerating to innervate correct portion of tectum (Roger Sperry).
3. Creating arrangement whereby strips of membrane was alternated between posterior tectal and non-posterior tectal tissue results in nasal retinal axons growing on both membranes but temporal axons growing on non-posterior tectal membrane only (they were repelled by posterior tectal strips). (Friedrich Bonhoeffer).

Question 20

What are examples of neurotrophic factors?

Answer 20

1. Insulin-like neurotrophins (NGF, BDNF, NT3, NT4)
2. GDNF

Question 21

What is the mechanism of neurotrophic action of NGF?

Answer 21

NGF binds to TrkA is taken up by growth cone and transported to soma of developing neurone by retrograde transport mechanisms. In soma, they inhibit pro-apoptotic factors.

Question 22

What are the mechanisms by which axonal selection occurs?

Answer 22

1. Competition for neurotrophic support
2. Selection by correlated firing (Hebb’s law)

Question 23

How are generic (temproary) synapses formed before Hebbian selection occurs?

Answer 23

Interactions between pre-synaptic neurexin and post-synatic neuroligin.

Question 24

What are the principles of Hebb’s law?

Answer 24

1. Selection against uncorrelated inputs
2. Selection for correlated inputs
3. Selection for correlating neighbouring inputs

Question 25

What is the overall process of axon development?

Answer 25

1. Developing axons are guided towards their target organs by a number of attraction/repulsion mechanisms. 2. As the axons approach the target, competition for secreted neurotrophic factors ensure that only a certain number of axons survive and form synapses. 3. Formation of generic (temporary) synpases. 4. Activity-dependent selection refines synapses formed so that only the useful ones remain.

Question 26

What are the types of injuries that can be sustained by the peripheral nervous system?

Answer 26

1. Avulsion: Nerve root completely torn from spinal cord. This type of damage may not be repairable. 2. Stretch (neuropraxia): This type of injury doesn’t result in transection of any nerve fibres. It simply leads to the transient loss of conduction in fibres (due to compression of nerve fibres as result of swelling) that usually recovers by itself. 3. Rupture: Forceful stretch results in peripheral fibre becoming torn partially/fully. Can sometimes be repaired depending on the extent of damage.

Question 27

What is Wallerian degeneration?

Answer 27

Degeneration of axon and associated myelin distal to site of lesion, following lesion.

Question 28

What are the conditions that need to be satisfied in order for axon generation in the PNS to occur?

Answer 28

1. Tract of schwann cells remain across gap created by lesion (endoneurial tube). 2. Gap is \< 1cm.

Question 29

How do schwann cells support axon regeneration in the PNS?

Answer 29

1. Schwann cells secrete growth factors and cytokines to guide and promote axon growth (e.g GDNF, NGF, BDNF…). 2. Schwann cells provide a tract (endoneurial tube) for regenerating axon to follow.

Question 30

What happens if conditions for PNS axon regeneration is not met?

Answer 30

Cut axon end swells and forms a neuroma, which may be responsible for neuropathic pain.

Question 31

What is the overall sequence of events that occur following lesion of peripheral axon?

Answer 31

1. Following cut, axon any myelin distal to the site of cut degenerate and become fragmented (Wallerian degeneration). 2. Macrophages and schwann cells phagocytose fragmented axons/myelin. 3. New growth cone forms at the cut end of axon. 4. Axons begin to grow following secretion of neurotrophic substances by schwann cells that remain lining the space degenerated axon used to fill (endoneurial tract). 5. Axons regenerate and make contact with innervated structures. 6. Remyelination occurs.

Question 32

What is the accuracy of axon regeneration dependent on?

Answer 32

1. Type of lesion (crushed axons regenerate with higher accuracy compared to cut axons) 2. Distance of lesion (the greater the size of gap formed during lesion, the less accurate the regeneration)

Question 33

What are the types of neuronal injuries that can occur in the CNS?

Answer 33

1. Axon lesion 2. Neuronal loss

Question 34

What are the factors in the CNS that inhibit axon regeneration?

Answer 34

1. Extracellular matrix lacks growth-promoting molecules such as laminin. 2. Astrocytes fill area of lesion, along with macrophages and other inflammatory cells. This results in deposition of scar tissue. Astrocytes in this scar produce growth-inhibiting factors such as chondroitin sulphate proteoglycans (CSPGs). 3. Oligodendrocytes produce growth-inhibitory factors such as nogo. 4. Surrounding cells may also express repellent factors such as ephrins and semaphorins.

Question 35

What may be the functional purpose of axon growth inhibition in the CNS?

Answer 35

Inaccurate axon regeneration may disrupt the compex circuitry in the CNS.

Question 36

What are the strategies that may be utilised to promote axon growth in the CNS?

Answer 36

1. Introduction of neurotrophic factors promote axon sprouting, but not long distance growth. 2. Provide favourable surface to promote growth (especially across gaps): - Peripheral nerve/Schwann cell graft - Olfactory ensheathing cell graft 3. Blocking inhibitory Nogo with monoclonal antibodies seems to promote CNS axon regeneration. 4. Blocking inhibitory proteoglycans (e.g. chondroitinase ABC). 5. Grafting neural progenitor cells to site of lesion may promote regeneration of axons, even of fully transected spinal cord. 6. Combination therapy (some combination of above therapies, e.g. Schwann cell graft + neurotrophic factor).

Question 37

What are the advantages of using olfactory ensheathing cells in providing favourable surface to promote CNS axon growth compared to schwann cells?

Answer 37

One benefit of OEC grafts over Schwann cell grafts is that Schwann cells negatively interact with astrocytes in CNS (reactive gliosis), causing secretion of inhibitory factors. OECs on the other hand, don’t cause this effect.

Question 38

What are the major causes of CNS neuronal loss?

Answer 38

1. Ischaemia (i.e. stroke): Via excitotoxic mechanism leading to increased [Ca2+]i and Ca2+ toxicity. 2. Neurodegenerative diseases (e.g. MS, Parkinson’s disease, Alzheimer’s disease): Degeneration of neurones due to autoimmune attack or other factors.

Question 39

How can CNS neuronal loss injuries be treated?

Answer 39

1. Grafting foetal neurones 2. Grafting pleuripotent neuronal stem cells

Question 40

What are teh problems with fetal graft transplants?

Answer 40

1. Ethical issues with using foetal tissue 2. Logistical issues with obtaining/preserving samples 3. Death of grafted neurones 4. Inability for axons to grow out of graft 5. No regulation of grafted tissue 6. Additional side-effects

Question 41

What are stem cells?

Answer 41

Pleuripotent cells capable of division and self-renewal. They differentiate to form variety of cell types.

Question 42

What are progenitor cells?

Answer 42

Unipotent cells capable of division and self-renewal, but can only differentiate into one type of cell.

Question 43

What are induced pleuripotent stem cells (iPSCs)?

Answer 43

Differentiated cells that are converted to pleuripotent stem cells by expression of Yamanaka factors.

Question 44

What are the Yamanaka factors?

Answer 44

1. Oct4 2. Sox2 3. c-Myc 4. Klf4