1. Understanding the impact of Schizophrenia and Autism risk genes on brain development
Karun K. Singh, Ph.D.
Stem Cell and Cancer Research Institute
Department of Biochemistry and Biomedical Sciences
McMaster University

2. Genes and Neurodevelopment`
Normal Brain Development
Brain Development Disorder
SCHIZOPHRENIA
AUTISM
Genetic Mutation

3. Symptoms of Schizophrenia`
Positive symptoms
Hallucinations
Delusions
Thought disorder (confused thinking and speech)
Negative symptoms
Diminished emotional expression
Diminished motivation
Inability to experience pleasure
Cognitive symptoms
Poor concentration
Difficult to plan and organize
Poor memory
Treatments
Antipsychotic drugs (old and new generation)
Cognitive behavior therapy
When we think of SZ, many people associate what are the positive symptoms, such as the hallucinations… 3 3 3

4. Neuropathology of Schizophrenia`
Unaffected twin
Schizophrenic twin
N Eng J Med
So what do we know about the neuropathology of these disorders? Well we actually don’t know very much, mostly because there are many inconsistencies between studies. However, what is agreed upon by many is the reduction of grey matter.
Reduction of brain volume (Grey Matter)
Lateral Ventricle enlargement
Decreased GABAergic interneurons (GABA and GAD67 staining)
Less extensive arborization/neuronal complexity (dendritic/synapse)
OVERALL – Nothing absolutely conclusive from brain imaging 4 4 4

5. Genetics plays a major role in psychiatric disorders`
Genetics plays a major role in psychiatric disorders
And we know genetics play a major role in psychiatric disorders. This is a figure from one of Tom Insels papers last year describing the degree to which genetics plays a role.
**Complex genetics at play in psychiatric disorders**
(Tom Insel, Director NIMH, JCI, 2010) 5 5

6. Whole Genome Sequencing
2011
Loci for SZ
mir137
CSMD1
TRIM26
PCGEM1
Loci for BPD
ITIH3-ITIH4 region
Exome Sequencing
Whole Genome Sequencing
…more and more genes!
Here is a timeline of some of the major genetic discoveries since late 90’s early 2000’s. In the early years using linkage analysis, many genes were implicated. DISC1 was one of the first discovered genes, and I’ll talk more about that in a few slides.

7. Modeling the Genetics of Neurodevelopmental and Psychiatric Disorders`
Modeling the Genetics of Neurodevelopmental and Psychiatric Disorders
Brain Disease
Risk Gene
Developing
Mouse Brain
Human Cellular
Reprogramming
Patient Cells
Why do this?
Understand how genes impact brain structure/function during development
Understand core signaling pathways affected

8. Disrupted in Schizophrenia-1 (DISC1)
Chromosome (1; 11) translocation
disrupts the DISC1 locus. 8 8

10. `
Experimental model to examine DISC1/Dixdc1: The developing mouse cortex
Approach: In utero electroporation to study neural stem cells and neuronal differentiation/maturation
CP - neurons
Direction of migration
To examine whether Dixdc1 regulates DISC1 function in neural stem cells, we used IUE…
IZ – axon tracts
VZ – neural stem cells

11. Dixdc1 functionally interacts with DISC1 to regulate neural progenitor proliferation`
Singh et al., 2010

12. Dixdc1 and DISC1 regulate neuronal migration`
Dixdc1 and DISC1 regulate neuronal migration
DISC1
DIXDC1 X
Frag2
Singh et al., 2010

13. Dixdc1 is a critical regulator of DISC1 and embryonic brain development`
Early brain development
Mid-brain development
Singh et al., 2010

14. `
Does Dixdc1 functionally interact with DISC1 in neural connectivity development?
Dendrite Growth
Spine Structure
Synapse Formation
Penzes et al., Nat Neurosci 2011

15. `
Does a DISC1-Dixdc1 pathway regulate the growth of dendrites and synapses?
Control shRNA
DISC1 shRNA
Dixdc1 shRNA
DIV6
Dixdc1 Frag2
Vickie Kwan, preliminary results
Confirming this in vivo  in utero electroporation

16. `
Examining synapses with genetic tools: Trans-synaptic labeling using Rabies virus
Advantages
Single Cell Resolution
Watch dynamics over time
Ed Callaway Lab
Salk Institute

17. Trans-synaptic labeling in mouse cortical neurons`
Trans-synaptic labeling in mouse cortical neurons
IUE E15
Culture E17
DIV 10
DIV 14
LV-Syn-HTG
Infect:
Rabies
Fix cells
Rabies virus courtesy of Ian Wickersham and Heather Sullivan (Seung
How does DISC/Dixdc1 regulate synapse formation?

18. Studying the Genetics of Autism`
Not included:
Additional CNVs
Exome sequencing
(de novo mutations)
Whole genome sequencing
Aldinger et al., 2011 Neuron

19. Human cellular reprogramming to create patient-derived neural cells`
Human cellular reprogramming to create patient-derived neural cells
Advantages
Model autism genes
Behavior assays
Neural circuits and cell types involved
Some Disadvantages
Difficult to model disease gene networks, large CNVs, human SNPs
Gene disruption doesn’t completely mimic human genetics (eg. KO≠mutation)
Not patient brain tissue (human brain specific)

20. Human cellular reprogramming to create patient-derived neural cells`
Human cellular reprogramming to create patient-derived neural cells
Reprogramming
genes
Differentiate
Induced pluripotent stem (iPS) cells
patient skin
samples
Patient-derived
NEURAL CELLS
Induced Neuronal (iN) or
Neural Progenitor (iNP) cells
Direct conversion
ADVANTAGES of Direct Conversion:
Faster than iPS method
Epigenetic signature of patient cell is likely preserved
Specific Neuronal subtype generation (Spinal Motor, Dopaminergic)

21. Studying the Genetics of Autism`
Aldinger et al., 2011 Neuron

22. Tuberous Sclerosis Complex (TSC): a genetic disorder with high rates of autism`
Collaboration with Dr. Philippe Major, Sainte-Justine Hospital, Montreal
Benign tumors in vital organs including brain
Autism features (syndromic autism) 25-60% in ASD
Learning disabilities, developmental delay
Epilepsy
Kelleher, III and Bear, 2012 CELL
Mouse Models
Protein Translation
Plasticity (mGluR-LTD)

23. Using Cellular Reprogramming to Study TSC and Autism`
Using Cellular Reprogramming to Study TSC and Autism
Assay Development
Phenotype and
Direct Cellular
Reprogramming
Drug
Screen
Healthy or TSC Fibroblasts
Human
Neural
Cells
Synapse Function
Dendrite/Spine Growth
Trans-synaptic labeling
Automated Electrophysiology

24. In vivo Human Neuronal Model using Xenotransplantation`
In vivo Human Neuronal Model using Xenotransplantation
Transplant
Patient-derived neural cells
Fluorescent Label
In vivo profile of human cells:
How do patient neural cells functionally integrate into the developing
or adult brain?

25. Acknowledgements My lab: University of Montreal- Vickie Kwan
Shashwat Desai
Omar Shehab
University of Montreal-
Sainte Justine Hospital
Dr. Philippe Major (TSC)
Massachusetts Institute of Technology:
Dr. Li-Huei Tsai
Funding:
Ontario Research Fund