Selective vulnerability in motor

neuron disorders

Eva Hedlund, PhD

Ludwig Institute for Cancer Research
Karolinska Institutet, Sweden
 Selective vulnerability in motor neuron disorders

• Somatic motor neurons, but not autonomic

motor neurons

• Selective vulnerability among somatic motor

neurons:

Amyotrophic lateral sclerosis (ALS):

• Cortical, spinal and lower cranial nerve motor neurons
degenerate

Spinal bulbar muscular atrophy (SBMA):

• Spinal and lower cranial nerve motor neurons are lost
Spinal muscular atrophy (SMA):
• Spinal motor neurons degenerate
1. arm muscle
2. tongue - Higher cranial nerves damage late or are spared in ALS,
3. cortical (upper) motor neurons
4. brainstem (bulbar) motor neurons SMA and SBMA
5. axon bundles
6. rib muscles
7. spinal motor neurons
The reasons for this selective vulnerability
8. leg muscle are unknown
 Cell intrinsic and extrinsic disease mechanisms

Astrocyte
Interneuron

Microglia

? Activation
Toxic signal?
Signal
?

Motor neuron

Muscle
Schwann cells

Questions:
• Why are some motor neurons more vulnerable to degeneration than others?
• Can analysis of the normal intrinsic properties of different motor neuron subpopulations give
clues to mechanisms of relative vulnerability?

Methodology:
• Isolation of individual neurons using laser capture microdissection (LCM) and analysis of
gene expression differences using oligomicroarrays (Rat Genome 230 2.0 Array, Affymetrix)
 Isolation of motor neuron subpopulations with differential vulnerability
to degeneration in motor neuron disorders

Laser capture microdissection (LCM) of motor neurons

Hedlund et al. 2010 Brain

 Global gene expression analysis of motor neuron subpopulations

Differences in the number of

genes involved in transcription,
RNA -processing, -binding and -
splicing and regulation of
translation

• Motor neurons in CN12 and cervical spinal cord had more commonality in gene expression
levels than those in CN3/4
• All three motor neuron subpopulations displayed distinct profiles and exhibited genes with
unique expression
• Differential Hox gene expression validated the microarray data Hedlund et al. 2010 Brain
 Protein analysis confirmed differential expression in motor neurons of
CN3/4, CN12 and the cervical spinal cord

Differential expression of multiple proteins with

implications for motor neuron vulnerability and
protection

• Can exogenous delivery of CN3/4-restricted genes confer neuroprotection to vulnerable

motor neurons?
• Developed an in vitro toxicity assay to analyze possible effects on spinal motor neurons.
Hedlund et al. 2010 Brain
 The CN3/4-restricted gene IGF-II protected spinal motor neurons from
glutamate-induced toxicity
Selection criteria:
• IGF-II support regeneration of motor axons (Caroni and Grandes. 1990 J Cell Biol; Near et al.1992 PNAS)
• IGF-I can protect motor neurons in a mouse model of ALS (Kaspar et al. 2003 Science)
• We identified a preferential expression of IGF-I and IGF-II within CN3/4 motor neurons
In vitro assay of Glutamate overload
-Excitotoxicity

Astrocyte

L-trans-2,4-pyrrolidine-2,4-
dicarboxylic acid (PDC)

+ Glutamate

EAAT

NMDA
Glu AMPA
Ca2+
AMPA

ROS

Motor neuron

Hedlund et al. 2010 Brain

Guanine Deaminase, a CN3/4-preferential gene, protected motor neurons
Selection criteria:
• Guanine deaminase is important for synaptic function and dendritic branching (Firestein et al. 1999
Neuron) (Akum et al. 2004 Nat Neurosci) (in hippocampal neurons).
• ALS-associated mutations in TDP-43 attenuates the dendritic function of this protein (Lu et
al.2009 Mol Brain).
• We identified a restricted expression of guanine deaminase in CN3/4 motor neurons

Hedlund et al. 2010 Brain

 Conclusions and future perspectives

• Analysis of the intrinsic properties of different motor neurons subpopulations in

the normal animal can give important clues to mechanisms of relative vulnerability
in motor neuron disorders and may hopefully be used to develop treatments for
these diseases.

• Future studies will analyze if the identified molecules can protect neurons after
toxicity is initiated and evaluate the effect in other models of motor neuron
degeneration (in vitro and in vivo).

• Future studies aim to also analyze possible differences in mRNA splicing in

different motor neuron subpopulations
 Acknowledgements

Harvard Medical School/McLean Hospital

Ole Isacson (Director)
Martin Karlsson
Teresia Osborn
Wesley Ludwig

Supported by
ALS Association (EH)
ALS Research Program Therapeutic Development Award/DOD (OI)
The Consolidated Anti-Aging Foundation (OI)
National Institute on Aging (TO)