Generation and regeneration of Motor Neurons Flashcards

1
Q

where are MNs located in the CNS

A

HB
ventral horn of SC

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2
Q

what is local processing

A

activation of the flexor muscle inhibits the extensor muscle (antagonistic muscle)

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3
Q

where is Shh produced

A

floorplate and notochord

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4
Q

MN production

A

neural progenitor domain pMN in the ventral neural tube

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5
Q

Class I/II TF

A

class I TFs (Pax6/Dbx2) repressed by Shh
Class II TFs (Nkx2.2/Nkx6.1) activated by Shh

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6
Q

Evidence to show graded Shh signalling

Ericson et al., 1997

A

Titration of Shh forms different neuronal subtypes:
V1 - low conc
V2 - intermediate conc
MN - high conc of Shh

use reverse anti-Shh antibody - Shh conc is 0 (no V1/V2/MN)

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7
Q

motor column

A

PGC (FoxP1 low)
LMC (FoxP1 high)
MMC (Lhx3)
HMC

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8
Q

sub column

A

LMCl (projects to dorsal limb)
LMCm (projects to ventral limb)

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9
Q

motor pool

A

rf-MP (Lhx1)
vasti-MP (Lsl1)

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10
Q

role of Hox genes

A

conserved
determine positional identity of cell types along the rostral caudal axis
males have 4 hox clusters
females have 1 hox cluster

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11
Q

Hox gene cluster

A

secondary organising centres along the R-C axis
morphogens which induce Hox genes: Gdf11, RA, FGF

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12
Q

FGF manipulation in chick/mammals using electroporation

A

FGF8 causes gain of Hoxc9/loss of hoxc6

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13
Q

LMCm/LMCl projections

A

LMC m project to ventral limb contains ephrin A TF: Isl1 Receptor: EphB
LMC l project to dorsal limb contains ephrin B TF: Lhx Receptor: EphA

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14
Q

ectopically expressing LMCl/m determinants

A

GFP (control) 50/50 projection to d/v limb
GFP+lim1 (LMC l) - most projections to dorsal limb
GFP+Isl1 - mostly ventral projections

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15
Q

role of Pea3

A

marks 2 motor pools in LMC m

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16
Q

dendritic morphology of Pea3/non-Pea3 pool

A

Pea3 pool - cresent shaped, no sensory axons, no medial projections
non-pea3 pool: project medially

17
Q

pluripotent embryonic stem cells

A

derived from inner cell mass
forms all germ layers

18
Q

in-vitro formation of MN from embryonic stem cells

A

express morphogens: RA/Shh (provide positional information)

19
Q

ectoderm to MN formation

A

primative ectoderm - rostral neural - via RA - caudal neural - via Shh - MN

neural induction/caudalisation/ventralisation

20
Q

what do ES derived MNs form

A

MMC
HMC
not PGC (controls involuntary movement)

21
Q

role of doxycycline

A

chemical on/off switch
induces TFs: Ngn2/Isl1/Lhx3
produces transcriptional profile of NIL motor neurons generated by extrinsic factors (RA/shh)

22
Q

NIP vs NIL

A

NIP - exist dorsally, cranial MN
NIL - exist ventrally spinal MN

23
Q

NIL MN

A

express all markers of somatic MNs derived with extrinsic factors but lack expression of progenitor specific genes (Sox1/olig1/olig2)
transition from early embryonic identity to post mitotic MN (skip neural progenitor stage)

24
Q

generation of induced MNs

A

mouse embryonic fibroblasts (MEFs) - expose to late progenitors (sox1/Pax6/Nkx6.1/Olig2) - MN+glia

25
application of induced MNs
ALS patient fibroblast - direct conversion to MN (respond to chemicals) form NMJ
26
amyotrophic lateral sclerosis
selective degeneration of MNs/neurons of the motor cortex - progress muscle weakness and spasticity (most of CNS unaffected) risk of development: 1in 500 death of patient: 3-5 yrs after onset of symptoms (lose innervation of respiratory muscles) no effective treatment 10-15% genetic cause most commonly: defect of superoxidase dimutase 1 (SOD1) point mutation sporadic ALS is usually polygenic
27
mutant SOD1
extract SOD1 from astrocytes - coexpress with hESC MN - Hb9/GFP+ cells astro SOD1 mutant - degeneration of MNs
28
use of co-cultured astrocytes and MNs
ALS drug screening
29
limitation of induced MN
optogenetic activation of MN (blue light on ChR) replace input from spinal circuits do not reflect normal MN pathology
30
in-vitro neuromuscular circuits | Osaki et al., 2018
develop in-vitro NM device using human iPSC MN (form NMJs with myofibers) - stabilised by 2 flexible pillars in ALS, MN fail to induce myofiber contractions when stimulated with light