Please take two minutes to fill in the quick questionnaire
During the lecture, do interrupt with questions if you have any

1. Define an animal model and discuss why and how they are used.
2. Describe the neuropathology of spinal cord injury (SCI) in humans.
3. Give examples of different animal models of SCI.
4. Critically evaluate different animal models of SCI (pros, cons).
1. Write lecture notes! Read them soon, to refresh your memory.
2. I will cover the key issues but you need to read the recommended reviews and papers. Write notes. Test a classmate on their knowledge and understanding.
3. Be critical. Question what you are told!
4. Before any exams, find and read additional up-to-date papers (e.g. by authors on the reading list)
5. Think about how animal models are sufficient and where they fail.
6. Cite authors (e.g. Smith et al,. 2007) to substantiate written claims.

Whereas good looking humans can be supermodels, an “Animal
Model” is NOT a beautiful, photogenic pet.
Thus, the following are NOT examples of good animal models

An “animal model” refers to the use of a non-human animal to simulate a human disease or injury.
They are used where it is practically or ethically difficult to use humans.
They can be
Naturally occurring
In a normal animal, e.g. after road traffic accident
In an abnormal (spontaneous) mutant
Induced experimentally
Surgical
Genetically engineered

We want to discover safe and effective therapies for various diseases and injuries.
Many potential therapies require testing for safety and efficacy in animals before it is possible to move to a clinical trial.
If you understand the pros and cons of each model, you can better evaluate the research (e.g. criticise the papers)
Ethical implications.

Approval of Personal and Project Licence from Home Office (UK)
Training & supervision
Development of animal model if necessary
Genetic engineering / breeding of mutant, etc.
Pre-training / habituation
Surgery under anaesthetic
Spinal cord injury
Delivery of a therapy
Postoperative care (analgesia, antibiotics, etc.)
Post-injury behavioural testing
Electrophysiology / imaging
Terminal anaesthesia, removal of tissues (e.g. fixation)
Cutting of tissues
Staining of tissues to reveal injury site / regenerating axons, etc.

Introduction to spinal cord injury
Incidence, prevalence
Pathology
Types of animal model
Surgical / naturally occurring / genetic engineering
Species of animals used
Rats / mouse / cats / non-human primates / dogs
Outcome measures
Behavioural tests
Histology

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
Prevalence in USA 250,000
Incidence in USA 11,000
CAUSES:
SEQUELAE:
• Motor dysfunction below the injury site
• Loss of sensation below the injury site
• Pain
• Bladder, bowel, sexual dysfunction
I’ll limit discussion to animal models of locomotor dysfunction after SCI.


Some (v limited) spontaneous recovery / compensation

Few acute therapies
 steroids (SCI) – contraversial

Few chronic therapies
 rehabilitation (locomotor)
 adaptation (sexual, bladder, bowel)

None fully restorative

So we need to develop safe and effective therapies

emphasise CST and sensory axons quad v para

VARIANTS:
1.
Contusion
2.
Compression /
Maceration
3.
Laceration (cut)
4.
Solid core injuries



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Very few new neurons are born (neurogenesis)
Spontaneous failure of CNS axon regeneration
Limited endogenous repair (adult vs neonate)
Insufficient compensatory plasticity

Poor intrinsic axon growth

Pro-growth molecules down-regulated

Anti-growth pathways switched on

Inhospitable extrinsic environment

Cysts, cavities



Fibrotic scar
Growth-inhibitory molecules (intact & injured)
Lack of growth factors, permissive substrates

1.
2.
3.
4.
5.
Reduce cell death
(neuroprotection)
Promote regrowth of injured axons
(regeneration)
Promote compensatory regrowth by uninjured axons
(collateral sprouting)
Demyelination (?)
Rehabilitation

1.
i.
Model cell death (neuroprotection)
Induce characteristic pattern of cell death ii.
iii.
Sparing
Allow measurement of any neuroprotection
2.
3.
Model axon injury i.
Full transection of some axons ii.
iii.
iv.
Sparing of other axons
Allow measurement of any regeneration
Allow measurement of any collateral sprouting
Model demyelination (if any)
4.
Model locomotor (or other) deficit i.
Allow measurement of any recovery

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
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Different surgical methods
Species (rat, human, cat, dog, non-human primate)
Outcome measures
Other variables


Time until therapy is delivered
Level of spinal cord

Usually at midline, mid thoracic (T9) in rats or mice
Basso et al., 1996 Exp Neurol 139:244-256
Also unilateral in cervical spinal cord (C5)
Various impactors:
NYU
OSU
IH
Hill et al., 2001 Exp Neurol 171:153-169.

Usually midthoracic
Surgical knife cut
Advantages
True regeneration
Disadvantages
Very harsh
Scar
Postoperative care de Winter et al 2002

Cut or crush with forceps
Often done in rats / mice
There may be no ventral CST in mice
(Steward et al., 2007)

Advantages
Deficits are mild – loss of fine but not gross motor control
Easy to trace
Cuts ascending sensory fibers completely
Disadvantages
Does not cut corticospinal tract (CST) completely
Thus cannot be used rigorously to assess true axon regeneration
Steward et al., 2003

Freund et al., 2006 - primate
• Interrupts tracts unilaterally (e.g. CST in primate; RST in rodents)
• Primate CST is lateral
• Rat CST is mostly dorsal but also lateral and ventral
• Advantage
• Deficits are unilateral
• Contralateral tract / limb serves as within-animal control -> power


Rat

Most common; cheap, friendly



Anatomy well understood
Disadvantage: CST lesions aren’t very disabling
Quadruped; does this model us as bipeds?


Mice

Smaller – behaviour can be tougher to measure



Transgenics / knockouts exist

Zheng et al,. 2006
Anomaly - very little cyst formation
Disadvantage: CST lesions aren’t very disabling


Non-human primate

Similar anatomy to human


Similar pathology
Disadvantages: Expensive; ethically challenging

Cats

Used less often nowadays

Earliest trials based on cat studies


Naturally occurring injuries

Road traffic accidents

Spinal disc herniation (hernia / prolapse / “slipped”)

Chondrodystrophic dogs

Bassets, dachshunds, bulldogs

Commonly cervical or thoracolumbar


Compression / contusion

Use autopsy material to understand SCI

Test out new therapies in dogs?

Ethical opportunity

Jeffery et al,. 2006a,b

Disadvantages

Sporadic

Not controlled – variable (but this models human)

• Most species are not bipedal (but consider birds).
• Species can have different musculoskeletal arrangements
• e.g. rat has fused radius and ulnar bones
• Species can have different neuroanatomical arrangements
• e.g. amount of direct cortico-motorneuronal synapses
Lemon &
Griffiths (2005)


Behavioural testing

Locomotion (forelimb, hindlimb)

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Pain
Bladder function

Electrophysiology / imaging

Histology (Tissue processing)

Size of injury

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Axon growth
Myelination (traditional stains)
Transplant characteristics


Grid walk / beam – show Schallert video

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Rats / mice
Easy
Sensitive to deficits (e.g. after CST injury)
Quantitative (count faults)
Forelimb and / or hindlimb


Forelimb reaching

Non-human primates

Rats
Show rat reaching video from Tim Schallert


Open field locomotion

BBB test

Basso, Beattie, Bresnahan, 1995 A sensitive and reliable locomotor rating scale for open field testing in rats. J Neurotrauma. 12:1-21.


Contusion / weight drop
Transection


Forelimb and hindlimb
Rats (BMS for mice)




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Connectivity / electrical properties of axons
MEP – motor evoked potential
SSEP – somatosensory evoked potential
TCMS – transcranial magnetic stimulation
EMG - electromyography

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
CT – computed tomography
MRI – magnetic resonance imaging fMRI – functional MRI
PET – positron emission tomography

Allows repeated measurements (longitudinal)


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To study tissue / cells / molecules
Traditional histology

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H & E
Nuclei - cresyl violet
Solochrome cyanine – Myelin

Modern histology

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Tract tracing – anterograde / retrograde / transsynaptic
Protein expression

Immunolabelling
Western blotting / proteomics

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Gene expression

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In situ hybridisation
Northern blotting / microarray / real time PCR


What do I want to model or measure?


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Cell loss / Neuroprotection?

Contusion / compression best?
True axon regeneration?

Complete section of tract required
Collateral sprouting?

Partial sparing of tract required

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What animal should I use?
What axon tracts do I want to cut?

Primate CST is in different location to rodent CST


Animal models allow controlled simulation of a human SCI and testing of therapies

Different types exist to model different aspects of SCI

Majority use surgical, a few use naturally occurring

Pros and cons to each model.

Any questions?

Anatomy of human spinal cord: Kandel, Schwartz & Jessell, Principles of Neural Science
Spinal cord injury statistics: http://www.spinalcord.uab.edu/show.asp?durki=21446
Reviews on animal models for spinal cord injury
Courtine et al., 2007 Can experiments in nonhuman primates expedite the translation of treatments for spinal cord injury in humans? Nat. Med. 13(5):561-6
Moon & Bunge, 2005. From animal models to humans. Journal of Neurological Physical Therapy
29:55-70.
Brosamle & Huber, 2006 Cracking the black box. Drug Discovery Today: Disease Models.
Jeffery et al., 2006. Clinical canine spinal cord injury provides an opportunity to examine the issues in translating laboratory techniques into practical therapy. Spinal Cord. 44:584-593.
Zheng et al., 2006. Genetic models for studying inhibitors of spinal axon regeneration. TINS
29:640-6.
Steward et al., 2003. False resurrections. J Comparative Neurol. 459:1-8.
Lemon & Griffiths, 2005. Comparing the function of the corticospinal system in different species: organizational differences for motor specialization? Muscle and Nerve. 32:261-79
Key papers to read critically
Jeffery et al., 2006. Autologous Olfactory Glial Cell Transplantation Is Reliable and Safe in Naturally
Occurring Canine Spinal Cord Injury. J Neurotrauma 22:1282-1293
Freund et al., 2006. Nogo-A-specific antibody treatment enhances sprouting and functional recovery after cervical lesion in adult primates. Nature Medicine. 12:790-2 and pages 1220,
1231-1233. Include Supplementary Materials.

Articles on ethics of using animals in research (optional)
The animal ethics reader (eds. Susan Armstrong and Richard Botzler)
Articles on ethics of using non-human primates in research (very optional)
The Great Ape Project (eds. Paola Cavalieri and Peter Singer)