Keynote Speaker: Dr. Giulio Tononi · 21st Annual NMHI Research Day March 5, 2020 Bernard Snell...

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Keynote Speaker: Dr. Giulio Tononi

March 5, 2020 Edmonton, AB

The University of Alberta acknowledges that we are located on Treaty 6 territory, and respects the histories, languages, and cultures of First Nations, Metis, Inuit, and all First Peoples of Canada,

whose presence continues to enrich our vibrant community.

Keynote Address – Dr. Giulio Tononi Credentials: MD, PhD, Professor of Psychiatry, Director of the Wisconsin Institute for Sleep and Consciousness

Consciousness: From Theory to Practice

What is consciousness, and what is its neural substrate in the brain? Why are certain parts of the brain important for consciousness, but not others that have even more brain cells and are just as complicated? Why does consciousness fade with dreamless sleep even though the brain remains active? Does consciousness always fade when patients become unresponsive after brain damage, during generalized seizures, during general anesthesia, or even in deep sleep? And are newborns, animals, and intelligent computers conscious? Integrated Information Theory (IIT) is an attempt to answer these and other questions in a principled manner. IIT starts not from the brain, but from consciousness itself - the world of experience – and derives from it what it takes for a system to be conscious. The results of this exploration can account for many empirical findings, lead to counterintuitive predictions, and has motivated the development of promising new tests for the practical assessment of consciousness in non-communicative subjects.

21st Annual NMHI Research Day

March 5, 2020 Bernard Snell Hall, University of Alberta Hospital

SCHEDULE

8:30 – 9:00 Registration and Coffee 9:00 – 9:15 Welcome and Introduction

Dr. Douglas Zochodne -- NMHI Director Rebecca Long -- NGSA President

9:15 – 11:45 Student Oral Presentations

9:15 Alice Atkin -- supervised by Dr. Anthony Singhal Act fast! Or slow? Perceptual representation of visual reach targets is minimally

available after a delay of 3 seconds

9:30 Caylin Chadwick -- supervised by Drs. Anna Taylor & Bradley Kerr Pain and affect in a mouse model of multiple sclerosis: Contributions of the

kappa opioid system 9:45 Joseph Kamtchum Tatuene -- supervised by Dr. Glen Jickling

High-risk plaques and risk of stroke in asymptomatic carotid stenosis: Meta- analysis of prospective observational studies and therapeutic implications

10:00 Nicholas Batty -- supervised by Dr. Karim Fouad

Bigger bang for your buck: Simultaneous stimulation of two cAMP downstream effectors

10:15 Rachel Ward-Flanagan -- supervised by Dr. Clayton Dickson Assessing the efficacy of analgesia in chloral hydrate anesthesia

10:30 BREAK 10:45 Lion Budrass -- supervised by Dr. Sue-Ann Mok Chaperone DNAJA2 as a tau protein regulator in neurons

11:00 Ewen Lavoie -- supervised by Dr. Craig Chapman What's limbs got to do with it? Limb visualization in VR changes movement

behaviour and feelings of embodiment during object interactions

11:15 Brittany Fedor -- supervised by Dr. Frederick Colbourne Early rehabilitation does not reduce impairment after intra-striatal hemorrhagic

stroke in rats

11:30 Andrew Schmaus -- supervised by Dr. Satyabrata Kar Thermal response of amyloidogenic elements in cultured N2a cells: potential relevance to Alzheimer’s Disease pathology

11:45 – 12:00 Guest Speaker Jasmeen Saini -- Neuro Nexus 12:00 – 13:00 LUNCH 13:00 – 14:00 Panel Discussion: Career Options for Science Graduates Dr. Anastassia Voronova (Academia) Dr. Jason Acker (Industry) Dr. Valerie Sim (Medicine) 2:00 – 3:00 Poster Session

Presenters: Rebecca Long, Aislinn Maguire, Matthew Doan, Krista Metz, Charbel Baaklini, Behdad Parhizi, Sebastian Caballero, Mischa Bandet, Pedram Parnianpour, Komal Bharti, Brian Marriott, Nicole Dittmann, Shane Nicholls, Joseph Kamtchum Tatuene, Avyarthana Dey, Emma Schmidt, Adrianne Watson, Krishnapriya Hari, David Roszko, Tejal Aslesh, Wojciech Pietrasik, Somnath Gupta, Shihao Lin, Ryan Moukhaiber, Brandon E. Hauer, Vaibhavi Kadam, Hailey Pineau, Abhishek Dahal, Jennifer Bertrand, Bahareh Behroozi Asl, Timo Friedman, Zoe Dworsky-Fried, John Monyror, Sucheta Chakravarty

3:00 – 4:00 Keynote Presentation

Dr. Giulio Tononi -- MD, PhD, Professor of Psychiatry, Director of the Wisconsin Institute for Sleep and Consciousness Consciousness: From Theory to Practice

4:00 – 4:15 Closing Remarks

Panelist Biographies

Dr. Anastassia Voronova, PhD

Dr. Anastassia Voronova is an Assistant Professor at the University of Alberta, and she holds Canada Research Chair Tier II in Neural Stem Cell Biology. After studying the regulation of embryonic stem cells during her PhD, she then went on to make seminal discoveries during her post-doctoral fellowship about the epigenetic regulation of neural stem cells during development in autism spectrum disorder, and about the role of cell-cell communication in instructing neural stem cells to become oligodendrocytes. Today, her research focuses on translating these discoveries on neural stem cells in development to brain regeneration from adult neural stem cells.

Dr. Jason Acker, MBA, PhD

Dr. Jason Acker is the Senior Development Scientist with the Canadian Blood Services’ Centre for Innovation, and a Professor in the Department of Laboratory Medicine and Pathology here at the University of Alberta. His research focuses on understanding the response of cells and tissues to ex vivo storage and the development of methods for their preservation and use as therapeutic products. Dr. Acker’s blood services laboratory has responsibility for developing scientific and technical evidence to support innovative changes in blood product manufacturing, storage and utilization at Canadian Blood Services. He is also actively involved in consulting with and advising companies and organizations developing biobanking and cell therapy programs and is co-founder of two University start-up companies.

Dr. Valerie Sim, MD, PhD

Dr. Valerie Sim self-identifies as a scientist who practices medicine to support her music habit. By day she is a prion scientist at the Centre for Prions and Protein Folding Diseases at the University of Alberta and a clinical neurologist consultant for rapidly progressive dementia cases locally and throughout Canada. By night she is a violinist and fiddler. In her lab, Dr. Sim grows prion-infected brain slices in a dish in order to ask: 1) how a prion’s size and shape can influence patterns of disease and risks of transmission; and 2) how targeting multiple steps along the disease pathway might produce more effective treatments. Clinically, she is medical director of the Canadian CJD Association and co-founder of the Edmonton Cognitive Neurology clinic.

Oral Presentation 1

Act fast! Or slow? Perceptual representation of visual reach targets is minimally available after a delay of 3 seconds

Alice Atkin* (1) & Anthony Singhal (1,2)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Psychology, University of Alberta, Edmonton, Alberta, Canada

Previous research suggests that the N170 ERP component represents ventral stream activity during reach-to-target tasks. Typically peaking over lateral occipital and temporal sites, N170 amplitude has been shown to be larger when reaches are planned using memory rather than online vision, and this amplitude decays over short periods of time (1-3s) as the fidelity of the memory begins to decay. fMRI data, however, indicates that despite this decay, perceptual memory for the target can be re-recruited for successful action even at long delays (up to 18s). Whether this decay continues steadily beyond 3s or eventually reaches a floor is not currently known. To investigate this question, we created a delayed action experiment with delays of 0, 1, 3, and 5 seconds. N170 amplitude in response to an auditory Go-cue was analyzed for each delay length at 4 electrode clusters: left temporal, right temporal, occipital, and parieto-occipital. Behavioural measures of performance were also analyzed. The results of the experiment affirmed that the N170 is generated within the ventral stream, with occipital and temporal clusters producing larger N170 amplitudes compared to the parieto-occipital cluster. While N170 amplitude did decrease from the 1s to 3s delay, in line with previous research, it then increased again at the 5s delay. Behavioural data indicated that increased delay length was associated with decreased reach accuracy only up to 3s, suggesting that perceptual memory decay reaches a floor at around 3s, however this pattern was not perfectly reflected by the N170 for reasons which are currently unclear.

Oral Presentation 2

Pain and Affect in a Mouse Model of Multiple Sclerosis: Contributions of the Kappa Opioid System

Caylin I. Chadwick* (1), Zoe Dworsky-Fried (2), Bradley J. Kerr (1,2,3), Anna M.W. Taylor (1,2,3)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Pharmacology, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Anesthesiology and Pain Medicine, University of Alberta, Edmonton, Alberta, Canada

Multiple Sclerosis (MS) is a debilitating disease in which inflammation and autoimmune reactions result in demyelination within the central nervous system (CNS). Neuropathic pain and negative affect are two symptoms commonly experienced with MS. The kappa opioid receptor (KOR) is involved in modulating mood and pain sensation, and changes in KOR function have been described in several models of chronic pain. My project aims to characterize changes in KOR expression and function in a model of MS. To accomplish this, male and female C57Bl/6 mice are induced with experimental autoimmune encephalomyelitis (EAE), which models the inflammation, autoimmunity, and demyelination that occurs in human MS. The mice then undergo behavioural testing 1) to establish the presence of a lowered pain tolerance, 2) to determine the analgesic efficacy of the KOR agonist U50,488H (1.6-30 mg/kg, i.p.), and 3) to evaluate the aversive qualities of the drug. Post-mortem tissue analysis will occur via fluorescent in situ hybridization for mRNA quantification, and via western blotting for protein quantification. Specifically, I will be probing for 1) KOR mRNA, 2) dynorphin mRNA (KOR endogenous ligand), 3) KOR total protein, and 4) KOR phosphorylated (i.e., activated) protein, in the spinal cords and brains of these animals and their non-diseased controls. This project is particularly interested in the EAE-induced changes to the kappa opioid system in the CNS, and whether an individual’s mRNA and protein findings correlate with their behavioural reaction to KOR agonism. Preliminary findings have shown that KOR analgesia is intact in EAE, while KOR agonism-associated aversion is diminished. This suggests that these effects are dissociable, and warrants future investigation into the downstream targets of KOR agonism.

Oral Presentation 3

High-risk plaques and risk of stroke in asymptomatic carotid stenosis: meta-analysis of prospective observational studies and therapeutic implications

Joseph Kamtchum-Tatuene* (1), Jean Jacques Noubiap (2), Alan H. Wilman (3), Gina Sykes (4),

Maher Saqqur (4), Ashfaq Shuaib (4), Glen C. Jickling (4) 1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Medical Research Institute, University of Adelaide, Adelaide, Australia 3 - Department of Biomedical Engineering, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Medicine, University of Alberta, Edmonton, Alberta, Canada BACKGROUND AND AIMS: Imaging features of high-risk plaques may identify patients at increased risk of stroke and optimize the risk-benefit ratio of carotid revascularization. We aimed to assess the relevance and feasibility of such an approach by summarizing data on prevalence of high-risk plaque features and associated risk of stroke in asymptomatic carotid stenosis. METHODS: Prospective observational studies reporting prevalence of high-risk plaque features and associated risk of stroke in asymptomatic carotid stenosis, were identified by searching PUBMED and Ovid-EMBASE, from inception to July 31, 2019. Prevalence of high-risk plaque features and annual incidence of ipsilateral ischemic events were pooled using random-effect meta-analysis. RESULTS: A total of 21205 participants from 67 studies were included, age 29-95 years, from 15 countries. The prevalence of high-risk plaques was 26.6% (95% CI: 22.9-30.5). Ulceration was present in 13.1% (3.5-27.1), microembolic signals in 14.3% (10.0-19.2), intraplaque hemorrhage in 19.1% (13.8-25.0), silent brain infarcts in 23.1% (14.2-33.3), thin or ruptured fibrous cap in 24.1% (12.0-38.7), impaired cerebrovascular reserve in 29.2% (15.1-45.7), AHA plaque type IV-VI in 30.8% (15.6-48.4), lipid-rich necrotic core in 36.3% (27.7-45.2), echolucency in 42.3% (32.2-52.8), and neovascularization in 43.4% (31.4-55.8). The incidence of ipsilateral ischemic events was 3.2 per 100 person-years in 10381 participants from 20 cohorts, after a mean duration of follow-up of 2.8 years. Incidence was higher in patients with high-risk features, 4.3 per 100 person-years (95% CI: 2.5-6.5) compared to those without, 1.2 per 100 person-years (95% CI: 0.6 – 1.8), with an odds ratio of 3.0 (95% CI: 2.1-4.3). CONCLUSIONS: High-risk features are common in asymptomatic carotid stenosis and the associated risk of ipsilateral ischemic events is higher than the periprocedural risk of stroke. Routine assessment of asymptomatic carotid stenosis beyond the grade of stenosis should be implemented to improve stroke risk stratification and optimize medical therapy. Trials using multimodal neurovascular imaging for risk stratification before randomization are warranted to determine the optimal strategy for stroke prevention in asymptomatic carotid stenosis. FUNDING: JK-T is supported by a Banque of Montreal Financial Group Graduate Scholarship. JJN is supported by an Adelaide Scholarship International, University of Adelaide, Australia. ACKNOWLEDGEMENTS:The authors are grateful to the University of Alberta Library for providing access to resources, and especially, to the Interlibrary Loan/Document Delivery Unit for promptly supplying the full-texts of all articles not in the collection or available in open access repositories.

Oral Presentation 4

Bigger bang for your buck: Simultaneous stimulation of two cAMP downstream effectors

Nicholas J. Batty* (1), Romana Vavrek (2), Pamela Raposo (2), & Karim Fouad (1,2)

1 - Neuroscience and Mental Health Institute, Faculty of Medicine and Dentistry, Edmonton, Alberta 2 - Department of Physical Therapy, Faculty of Rehabilitation Medicine, Edmonton, Alberta

cAMP has long been studied in CNS injury for its role in neuronal growth through its downstream effector: protein kinase A (PKA). In recent years, a number of studies have demonstrated that many of the growth promoting effects attributed to PKA are potentially instead the result of a different downstream effector: exchange protein activated by cyclic AMP (EPAC). The exact separate roles and mechanisms of action of PKA and EPAC have yet to be determined. We aim to study the mechanisms of cAMP based neurite extension and compare the outcomes of selective agonism of either downstream effector on their own or in combination. In order to initially compare the effects of selective agonism of PKA versus EPAC, neurite outgrowth was measured in primary cortical neuron cultures. To create these cell cultures, P0-P1 rat pup cortices were processed and plated. After 3 days, cultures were treated with either a selective PKA agonist or a selective EPAC agonist. Cultures were fixed on the tenth day after plating and immunocytochemistry was performed to allow for analysis of neurite length and thickness. These results triggered in vivo studies, which were performed in conjunction with rehabilitative motor therapy in rats with a C4 dorsolateral quadrant transection injury. Immediately following injury, Alzet mini-osmotic pumps containing a selective EPAC agonist was placed between the shoulder blades with a cannula directly over the primary motor cortex innervating the injured side of the spinal cord. Results demonstrate a significant increase in axonal collateralization into the grey matter above the site of injury, though no changes in functional recovery. In order to reach functional effects we went back to in vitro experiments utilizing both agonists on the same culture. Results indicate that whilst both EPAC agonism and PKA agonism separately increase neurite length of primary cortical neurons in culture, both agonists together result in neurite length and thickness increases that go beyond the expected additive effects of the agonists, suggesting a synergistic effect that demonstrates that EPAC and PKA do not converge solely on the same molecular signalling pathway as previously thought. In vivo experiment to follow up on these findings are currently underway. Funding: CIHR

Oral Presentation 5

Assessing the efficacy of analgesia in chloral hydrate anesthesia

Rachel Ward-Flanagan* (1) & Clayton T. Dickson (1,2,3) 1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Psychology, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Anesthesiology & Pain Medicine

In both research and clinical applications, anesthesia is typically defined by a series of discrete endpoints: unconsciousness, reduced behavioural responsiveness, amnesia, and analgesia. Specifically, anesthesia should give a researcher or clinician the ability to ensure the subject or patient will have no memory of a potentially painful surgical manipulation through pharmacological control of unconsciousness and the elimination of sensitivity to pain. By this definition, the use of chloral hydrate anesthesia in animal research remains an ongoing topic of debate, despite its continued use. The conflict surrounding chloral hydrate arises due to assertions that it does not provide adequate analgesia to meet the requirements of a sole anesthetic agent and that it causes tissue damage when administered interstitially via either intraperitoneal or subcutaneous routes. However, our current data suggests that in an acute experimental model using rats, a continuous intravenous infusion of chloral hydrate induces a stable brain state analogous to the slow oscillatory activity typically observed during non-REM (deep) sleep. Furthermore, in this state, an aversive stimulus applied to the toe pad does not elicit either a reflexive response or a spike in the electrographic recording, both of which are typically associated with nociception. Consequently, chloral hydrate may represent a reliable, cheaper alternative anesthetic to ketamine-xylazine for producing a stable, slow oscillatory brain state. Funding: RWF – NSERC PGS-D3; AGES CTD – NSERC 2016-06576

Oral Presentation 6

Chaperone DNAJA2 as a tau protein regulator in neurons

Lion Budrass* (1), Sue-Ann Mok (1,2) 1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada Molecular chaperones are some of the most important regulators of proteostasis in all organisms. In many cases regarded as Heat Shock Proteins (HSPs), they play a role in folding, refolding, degrading and disaggregating proteins. Molecular chaperones tend to form complexes with co-chaperones for correct binding to proteins. One example would be the HSP70/HSP40 complex. Various neurodegenerative diseases manifest pathologies that may be directly or indirectly related to aberrant protein aggregation or misfolding. Alzheimer’ Disease (AD) and Frontotemporal Dementia (FTD) both express pathologies that include Tau tangles, an intracellular aggregate formed from phosphorylated Tau, a microtubule associated protein. Tau tangles are presumed to be neurotoxic, leading to disruption of cellular homeostasis and eventual apoptosis. It has previously been shown that a HSP40 chaperone, DNAJA2 inhibits tau aggregation and is increased in some human brains showing tau pathology. The proceeding proposed studies may not only help us understand regulation of DNAJA2 in tau pathologies in e.g. AD, but also the mechanisms behind its regulation. Hypothesis: DNAJA2 up- or downregulation on various levels affects tau expression and folding. Using RT-PCR, I want to study mRNA levels of DNAJA2 in TG2541 (tau) mouse fore- and hindbrain at various ages. If this correlates with previous findings of upregulation of DNAJA2 on a protein level, I could use the data to more precisely determine the regulation of DNAJA2. A recent publication suggests that DNAJA2 mRNA can be regulated by the microRNA (miRNA), miR-155-5p, in immune cells and we now want to test the potential regulation of DNAJA2 by miR-155-5p in neurons. We will utilize a luminescent reporter construct containing the 3’UTR of DNAJA2 to determine if miR155-5p could regulate DNAJA2 expression and if this miRNA is induced by cellular stress conditions such as tau aggregation. Finally, through a Mass Spectrometry screening study, I want to elucidate similarities and differences between expression levels of >200 molecular chaperones in 3 different human cell types (iPSCs, Cardiomyocytes and iNeurons). By knocking down the expression of various chaperones, including DNAJA2 I then want to analyze changes in the protein folding capacity of those cells. The relevance of DNAJA2 in protein folding has not yet been probed at an in-depth level and a further exploration of its function may shed some light on the chaperone’s involvement in tau pathology and AD. Funding bodies: NSERC

Oral Presentation 7

What's limbs got to do with it? Limb visualization in VR changes movement behaviour and feelings of embodiment during object interactions

Ewen Lavoie* (1,2) & Craig S. Chapman (1,2)

1 - Kinesiology, Sport, and Recreation, University of Alberta, Edmonton, Alberta, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada

Commercially available virtual reality (VR) hardware and software are changing the landscape of education, industry, gaming, and human behavioural research. As this occurs, it is important to test whether behaviour in VR is similar to the real-world, and assess what sensory modalities contribute most to a person's immersive experience. We translated an object interaction task from the real-world (Lavoie et al., 2018) into VR so that we could dissociate a movement from its visual appearance. Participants completed at least 20 trials in two conditions: Controllers - where they saw a visual representation of the VR controller, and Arms - where they saw a set of virtual limbs. We found participants seeing Arms moved more awkwardly in order to make the virtual limbs look similar to how they would if they were interacting with a real-world object. These movement changes were accompanied by an increase in self-reported feelings of ownership over the limbs as compared to the controllers. Overall this suggests our movements are planned to provide optimal visual feedback, even at the cost of less efficient movements. There is something about seeing a set of limbs in front of you, doing your actions, that affects your moving, and in essence, your thinking. Although many studies report the positive impacts of VR technology, until studies are completed assessing just how "real" VR is, any results should be taken with a grain of salt.

Oral Presentation 8

Early rehabilitation does not reduce impairment after intra-striatal hemorrhagic stroke in rats

Brittany A. Fedor* (1), Anna J. Kalisvaart (2), Shivani Ralhan (2), Tiffany Kung (1), Frederick Colbourne (1,2)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, AB, Canada

2 - Department of Psychology, University of Alberta, Edmonton, AB, Canada

Background. Intracerebral hemorrhage (ICH) is a devastating stroke. Blood released into the brain’s parenchyma causes rapid mechanical damage, while initiating numerous secondary injury mechanisms, such as from toxic by-products of blood (clot) degradation. These events contribute to a very protracted course of cell death after ICH, which has proven to be particularly difficult to prevent with drug therapies. Remarkably, rehabilitative treatments (rehab) can mitigate much of that late cell death. One recent hypothesis suggests rehab helps accelerate the clearance of the old blood clot (hematoma) thereby minimizing neurotoxicity. Unlike other brain injuries such as ischemic stroke or traumatic brain injury, there are few studies that look at the modifiable factors (e.g. timing, duration, intensity) that may impact treatment efficacy after ICH. As we do not know which treatment factors have the greatest impact on recovery, the exact mechanisms by which rehab attenuates chronic cell death after ICH remain unknown. Owing to the uniqueness of ICH, we must study these issues in ICH models and not solely rely upon data from ischemic stroke or traumatic brain injury.

Objective. Here we manipulated enriched rehabilitation (ER) duration in the first month after moderate ICH to determine the impact on behavioural recovery and chronic cell death (60-day survival).

Hypothesis. We hypothesized that both rehab groups would perform better on measures of skilled reaching and have smaller lesions compared to controls, but longer treatment was expected to confer greater neuroprotective and behavioural benefits.

Methods. Rats were randomly assigned to one of three groups: CONTROL, ER10 (10 days ER), or ER20 (20 days ER). Animals were trained on a skilled reaching task, then received a collagenase induced striatal ICH. D5 through D14, animals in the ER10 and ER 20 groups completed four 15 minute reaching sessions each day followed by six hours of environmental enrichment daily for 10 days. All animals completed mid-treatment assessment on a skilled reaching task at D16 & D17. Animals in the ER20 group completed a second 10 day treatment regimen starting on D19 and ending D28. All animals completed a post-treatment assessment of skilled reaching on D30 & D31. Animals recovered in standard housing until euthanization on D60. Brains were perfusion fixed and extracted for histological processing.

Results. As expected, all animals displayed significant impairment following ICH. Interestingly, there was no significant difference in reaching ability at the mid- or post-treatment assessment (p > 0.05), regardless of treatment condition. Histological processing and analysis for assessment of lesion volume and white matter damage is currently in progress.

Implications. Clinical evidence suggests that earlier intervention may be the key to improving patient outcomes after stroke, but some populations may not benefit. Recent unpublished findings from our lab showed little behavioural benefit of rehab after ICH when started at D5. In addition, this protocol did not impact hematoma volume when assessed in the sub-acute phase (14 days). Together, these findings suggest that rehab does not provide much benefit – additional study is needed to identify predictors of poor outcome (ex: intervention time, CST integrity).

Oral Presentation 9

Thermal response of amyloidogenic elements in cultured N2a cells: potential relevance to Alzheimer’s Disease pathology

Andrew Schmaus* (1, 2) & Satyabrata Kar (1-4)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Center for Prions and Protein Folding Diseases, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Medicine, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Psychiatry, University of Alberta, Edmonton, Alberta, Canada

Alzheimer’s Disease (AD), the most prevalent cause of dementia afflicting the elderly, is characterized by the accumulation of tau-positive neurofibrillary tangles (NFTs) and amyloid-beta (Aβ) containing extracellular plaques within the diseased brain. Production and aggregation of these proteins correlates to the neurodegeneration and loss of cognitive function seen in AD patients. While a small subset of cases originates intrinsically via mutations in genes encoding proteins such as the amyloid precursor protein (APP), the vast majority of disease arises sporadically and potentially due to extrinsic or environmental influences. Recent studies have indicated that perturbations in body temperature may have a role in the development of pathological features associated with AD. Thus, our hypothesis is that hypothermic conditions enhance the levels of AD related proteins, such as Aβ peptides, in neuron-like cells. We used wild-type murine neuroblastoma (N2a) and Swedish APP mutant N2a cells to investigate the effects of ambient temperature on the production, secretion, and degradation of APP-derived products. Cells grown in a normothermic (37°C) environment or subjected to either hypothermic (27°C) or hyperthermic (40°C) conditions were assayed for amyloidogenic markers by western blot, ELISA, and fluorescent immunocytochemistry. Additionally, we evaluated how these conditions influence cell viability, which may be relevant to the neurodegeneration seen in AD. Our results show an inverse relationship between temperature and APP processing in both wild-type and mutant cells, indicated by increasing α/β C-terminal fragment (CTF) levels in cells exposed to a hypothermic temperature. This effect is accompanied by a decrease in cellular viability at 27°C. Endolysosomal constituents are altered as well, with an increase in the lysosomal marker LC3, as well as an increased colocalization of APP and Aβ with endolysosomal markers (LAMP1 and LC3) at 27°C. We also show a linear relationship between temperature and the secretion of Aβ peptides. Clearance mechanisms may play a role in these effects, as we see a decreased rate of α/β CTF clearance in cells exposed to hypothermic temperature through a cycloheximide pulse chase assay. The results that we have obtained so far indicate that a cellular thermal response may directly influence AD-related pathology by altering the production and secretion of AD-associated molecules. These results may contribute to an understanding of environmental influences on AD pathogenesis and the development of alternate therapeutic avenues which are inclusive of both intrinsic and extrinsic factors. Funding: QEII MSc Award, FoMD 75th Anniversary Award, SynAD Trainee Award.

Poster Presentation 1

What are the consequences of simplifying a sensorimotor task for translation to the clinic?

Jennifer K Bertrand* (1,2), Brea Chouinard (2,3), Alona Fyshe (1,3,4) & Craig S Chapman (1,2)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Faculty of Kinesiology, Sport and Recreation, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Computing Science, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Psychology, University of Alberta, Edmonton, Alberta, Canada

Assessment of sensorimotor behaviour allows researchers and clinicians a unique vantage point on the complex neurological system involved in human cognition and its translation to movement. When these integral systems experience disease or disorder, recording the dynamics of the sensory and motor systems can afford rich diagnostic insights, improving the understanding of a system’s dysfunction. Our lab has successfully implemented a novel sensorimotor recording and analysis procedure for functional movement tasks. Dubbed Trifecta-Lab, we established the infrastructure to simultaneously and wirelessly record 3 rich forms of data – eye movements, body movements, and the brain’s electrical activity (using electroencephalography, or EEG). Despite Trifecta-Lab’s immense promise, as it stands, it is not a clinically feasible sensorimotor assessment tool. The research tools are prohibitively expensive ($50,000+) and the complexity of administering these tools is a massive barrier. Therefore, the proposed project, referred to as Trifecta-Lite, aims to develop and validate an accessible, consumer-grade version of the established laboratory-grade sensorimotor procedure for a clinical setting. We will create a computer-based version of the functional movement task, substituting consumer-grade products (<$500 total) for each recording stream, using a Muse EEG headset, a Tobii Pro Fusion eye tracking bar, and recording computer mouse movements with an open source software toolbox. We will run 70 participants in Trifecta Lite to compare to the already-collected 70 participant dataset from Trifecta Lab. Specifically, we will compare two types of sensorimotor measures: eye-hand coordination and movement intention.The primary objective of this project is to understand how the sensitivity in our sensorimotor measures change when hardware and protocols are simplified for eventual translation to applied settings. For eye-hand coordination measures, we predict this measure to be highly preserved between Lab and Lite, though remapping hand movement to a mouse in 2D space might result in novel eye-hand dynamics. When exploring how well we can predict movement intention from EEG data, we might see great differences in this measure between Lab and Lite due to the quality and density of the EEG sensors. If we are able to validate our Lite data quality, we imagine a future where a person going into a clinic can perform a brief computer based task and receive detailed measurements about how quickly and accurately they move, where they think it’s important to look, and in real-time, what their brain is doing.


Aging of Prefrontal White Matter: Insights from Diffusion Tensor Imaging

Wojciech Pietrasik* (1,2), Ivor Cribben (2,3), Fraser Olsen (1), Yushan Huang (1), Nikolai Malykhin (2,4)

1 - Department of Biomedical Engineering, University of Alberta, Edmonton, Alberta, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 3 - Finance and Statistical Analysis, Alberta School of Business, Edmonton, Alberta, Canada 4 - Department of Psychiatry, University of Alberta, Edmonton, Alberta, Canada Health research has been increasingly focusing on understanding the mechanisms and patterns of brain aging. Cognitive declines that occur with age, such as deficits in processing speed and attention, implicate the prefrontal cortex (PFC) as a key region affected by aging. Previous diffusion tensor imaging (DTI) studies confirmed the degeneration of PFC white matter (WM) with age; however, these studies did not investigate differential aging of the distinct functional subdivisions comprising the PFC. This study used DTI to determine whether healthy aging differently affects the subregions of the PFC. 140 cognitively healthy participants aged 18-85 (62 men, 78 women) were scanned on a 1.5T MRI scanner. PFC WM was segmented into medial (MOFC) and lateral (LOFC) orbitofrontal, medial prefrontal (MPFC), and dorsolateral prefrontal (DLPFC) components using reliable protocols. Linear and non-linear regressions were tested to find the model best describing the relationship between DTI parameters and age. All subregions were affected by age; fractional anisotropy (FA), tract volume, fiber count, and fiber length declined with age, while mean (MD), axial (AD), and radial (RD) diffusivities increased with age. Subregions varied in the onset and patterns of decline, the MOFC and MPFC experienced robust linear declines in FA compared to weaker declines in the DLPFC and LOFC while tract volume decreased non-linearly in all subregions with the greatest decline in the MPFC. This study provides insight into the regional differences in PFC WM during aging; these differences may help explain why certain cognitive deficits are observed as we age. Wojciech Pietrasik was supported by a Canadian Institutes of Health Research operating grant (MOP11501) and the Natural Sciences and Engineering Research Council of Canada operating grant (06186) to Nikolai Malykhin; and by the Alberta Synergies in Alzheimer’s and Related Disorders (SynAD) program which is funded by the Alzheimer Society of Alberta and Northwest Territories through their Hope for Tomorrow program and the University Hospital Foundation. SynAD operates in partnership with the Neuroscience and Mental Health Institute at the University of Alberta.


Carotid plaque with high-risk features in embolic stroke of undetermined source: systematic review and meta-analysis

Joseph Kamtchum-Tatuene* (1), Alan H. Wilman (2), Maher Saqqur (3), Ashfaq Shuaib (3), Glen

C. Jickling (3)

1- Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2- Department of Biomedical Engineering, University of Alberta, Edmonton, Alberta, Canada 3- Department of Medicine, University of Alberta, Edmonton, Alberta, Canada

BACKGROUND AND AIMS: An ipsilateral mild carotid stenosis, defined as plaque with <50% luminal narrowing, is identified in nearly 40% of patients with embolic stroke of undetermined source (ESUS) and could represent an unrecognized source of athero-embolism. We aimed to summarize data regarding the frequency of mild carotid stenosis with high-risk features in ESUS. METHODS: We searched Pubmed and Ovid-Embase for studies reporting carotid plaque imaging features in ESUS. The prevalence of ipsilateral and contralateral mild carotid stenosis with high-risk features was pooled using random-effect meta-analysis. RESULTS: Eight studies enrolling 323 participants were included. The prevalence of mild carotid stenosis with high-risk features in the ipsilateral carotid was 32.5% (95% CI: 25.3 – 40.2) compared to 4.6 % (95% CI: 0.1 – 13.1) in the contralateral carotid. The odds ratio of finding a plaque with high-risk features in the ipsilateral versus the contralateral carotid was 5.5 (95% CI: 2.5 – 12.0). CONCLUSIONS: Plaques with high-risk features are five times more prevalent in the ipsilateral compared to the contralateral carotid in ESUS, suggesting a relationship to stroke risk.


Characterizing Strains of Neurodegenerative Disease in Whole Brain Organotypic Slice Culture

Hailey Pineau* (1,2), Grant Norman (1,2), David Westaway (1,2) & Valerie Sim (1,2)

1 - Centre for Prions and Protein Folding Diseases, University of Alberta, Edmonton, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Canada

Background: A limitation of current neurodegenerative disease models is their inability to recapitulate the range of phenotypic diversity typical of living patients. In neurodegenerative diseases such as prion disease and Alzheimer’s disease (AD), such phenotypic diversity is thought to be related to different “strains” or pathological conformations taken on by the culprit proteins that misfold. One technique that has been used to model strain-specific pathology in prion disease is the prion organotypic slice culture assay (POSCA). In this technique, mouse cerebellar brain slices are cultured and infected with prions and develop pathology as seen in vivo. Objective: Because prion disease is not exclusive to the cerebellum, we propose to assess the utility of this technique for whole-brain coronal slices using well characterized strains of scrapie. Because Aβ and tau, the proteins that misfold in AD, have prion-like infectivity, we also aim to assess the usefulness of POSCA for modeling strains in AD. Methods: We have prepared whole-brain coronal sections from Tga20 mice and infected them with brain homogenates containing 22L, RML, or ME7 strains of mouse-adapted scrapie. After a 56-day culturing period, the slices were either homogenized for western blot analysis, or underwent fixation and immunostaining. Using confocal microscopy, we are characterizing the distribution of PrP from these different scrapie strains. We have also prepared slice cultures from mice that develop Aβ pathology and exposed them to brain homogenates from AD patients who had either a typical or rapidly progressive disease course (i.e. slow or fast phenotypes or strains). Using confocal microscopy, we are characterizing the distribution, size, number, and morphology of aggregates that form. Additionally, we are using several biochemical techniques to compare the fast and slow AD brain homogenates. Specifically, western blots have been performed to quantify levels of total and phosphorylated tau, and ELISAs to assess levels of Aβ40 and Aβ42. Additionally, conformational stability of Aβ has been quantified using guanidinium denaturation curves. Furthermore, Asymmetric Flow Field Flow Fractionation will be used to assess the size distribution of Aβ aggregates, and Real Time Quaking Induced Conversion (RT-QuIC) will be done to analyze aggregation kinetics of the Aβ from these fast and slow AD samples. Conclusion: The development of a whole-brain strain-specific ex vivo model will be a huge advance in the field of AD and prion research, as it will not only allow a window into disease pathogenesis, but provide a system open to treatment studies.


Chemokine fractalkine directly regulates CNS precursor survival, migration and oligodendroglial differentiation

A. Watson* (1), Y. Li (1), K. Goodkey (1), T. Footz (1), A. Voronova (1,2)

1 - Medical Genetics, University of Alberta, Edmonton, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Canada

Oligodendrocytes produce myelin, an essential component of the central nervous system. In the adult brain, oligodendrocytes are generated in via a 2-step process: neural precursor cells (NPCs) in the subventricular zone are committed to oligodendrocyte precursor cells (OPCs), which then differentiate into oligodendrocytes. Formation of oligodendrocytes from their multipotent progenitors is regulated by signals present in the NPC niche. Fractalkine (FKN), an NPC niche ligand that signals via its sole receptor (CX3CR1), increases oligodendrocyte formation from cortical NPCs during embryonic development (Voronova, Neuron, 2017). FKN receptor is also expressed in postnatal and adult NPCs and OPCs (Ji, Neurosci Lett, 2004; Krathwohl, Stem Cells, 2004 Watson, Neurosci Lett 2020). However, the role of FKN signalling in postnatal NPCs and OPCs is not currently known. To address this gap, I cultured postnatal primary murine NPCs and OPCs in the presence or absence of FKN. My results indicate ~65% of postnatal NPCs bind FKN directly conjugated to fluorophore Alexa-647 (FKN-647) in vitro, which indicates NPCs express functional FKN receptor. When FKN is added to NPC cultures, it enhances their differentiation into OPCs and oligodendrocytes, but does not affect NPC proliferation. With regards to OPCs, which were induced from NPCs by culturing dissociated neurosphere cells in OPC media containing PDGF-AA, they exhibited enhanced migration and decreased apoptotic death when cultured in the presence of FKN. However, exogenous FKN did not affect OPC differentiation. My results demonstrate exogenous FKN directly mediates postnatal NPC/OPC function for enhanced oligodendrogenesis, migration and/or cell survival. I am currently investigating whether FKN signalling is necessary for precursor function by inhibiting FKN signalling via function-blocking antibodies or knocking out FKN receptor using CRISPR technology. Funding: University of Alberta, NSERC, Multiple Sclerosis Society of Canada


Epigenetics: New Concepts for the Study of Substance Use Disorders and Behavioral Addictions

Bahareh Behroozi Asl* (1), Katherine J. Aitchison (1,2)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, AB, Canada

2 - Departments of Psychiatry and Medical Genetics, University of Alberta, Edmonton, AB, Canada The definition of addiction has expanded to include “behaviors” in addition to “substances” in the Diagnostic and Statistical Manual for Mental Disorders, fifth edition DSM-V (American Psychiatric Association, 2013). Specifically, a “Substance Use and Addictive Disorder” section replaced the “Substance-related Disorder” section of previous versions. Pathological gambling is listed under the title of “Substance Use and Addictive Disorder” in the (DSM-5). Further studies are needed to include other behavioral addictions, many of which are currently considered as impulse control disorders. Behavioral addictions have neurobiological and psychosocial commonalities with substance use disorder (SUDs). Repetition of compulsive behavior despite adverse consequences, reduced control of problematic behaviors, appetitive urge and hedonic quality (after a drug is taken/ the behavior is performed) are other common features of SUDs and behavioral addictions. Such common features of SUDs and behavioral addictions may be studied at different levels. The results from each study can be considered as a starting point for the others. For example, while personality features are measured through clinical questionnaires, the results may help neuroscientists to study brain regions relevant to those manifestations, which in turn help geneticists to make hypotheses about relationships between genetic structures and expression of potentially relevant neurotransmitter receptors. The etiology of addiction has been a focus of research for many decades from different approaches. Environmental determinants are important factors that can influence the probability of developing an addiction. However, the reaction to the same environment varies among different people. Molecular approaches are another way of shedding light on the underlying mechanism of addiction and finding similarities between SUDs and behavioral addictions. Although research into behavioral addictions is in its early stages, in addition to SUDs, genetic studies have been done in these addictions to predict the susceptibility to addiction and understand its etiology. Addiction like other polygenic conditions is the product of interactions between genes and the environment. More recently, epigenetics have been added as a complementary approach to genetic studies. Epigenetics is a mechanism for the environment having potentially lasting effects on gene expression. It is defined as chemical changes of DNA structure or non-sequence based heritable factors that without changing the sequence which can, in turn, affect the expression of the respective genes. Among the various epigenetic mechanisms (which also include histone modification and miRNA effects), DNA methylation is one of the most important epigenetic variations, and can definitely drive lasting changes in gene expression. In this review, a brief exploration of the similarities between substance and non-substance addictions will be followed by an introduction to epigenetic regulatory mechanisms with an emphasis on DNA methylation. Finally, recent epigenetic studies in addictions are reviewed and the limitations that should be addressed to improve the addiction study are discussed.


Clustered protocadherins restrict neurite outgrowth during peripheral nerve regeneration

Rebecca M. Long* (1), Ambika Chandrasekhar (1), Douglas W. Zochodne (1, 2)

1 - Neuroscience and Mental Health Institute, Faculty of Medicine and Dentristry, University of Alberta 2 - Department of Neurology, Faculty of Medicine and Dentistry, University of Alberta Peripheral nerves are at greater risk of damage than the brain or spinal cord. In addition, a range of common conditions generally titled ‘neuropathies’ render axon damage that is disabling and often irreversible. The peripheral nervous system (PNS) has a limited capacity to regenerate but new understanding of its biology may yield insights into better regrowth. During regeneration of sensory axons, fibres must navigate to their target region while coordinating with existing intact axons. In the developing central nervous system (CNS) growing neurons use self-recognition strategies, which in mammals is driven by the clustered protocadherins (Pcdh; Lefebvre et al. 2012). Neurons lacking either the α or γ cluster show dendritic trees with high instances of self-overlap and little complexity (Suo et al. 2012, Ing-Esteves et al. 2018). This effect has been demonstrated in multiple types of CNS neurons, but its expression and function are unknown in the PNS. We suggest that Pcdh clusters participate in the patterning of epidermal re-innervation, and contribute to regenerative success. Pcdh-α and -γ proteins are expressed in the dorsal root ganglion (DRG) cell body, alongside low level axonal expression. We collected DRGs at three timepoints (0, 36, 72h) following a sciatic nerve axotomy and found that the Pcdh-γ mRNA levels decrease in the DRG at 36h, followed by a return to baseline by 72h (p<0.05) with a similar trend in protein (p=0.28), suggesting sensory neurons may act quickly to restore Pcdh levels following an injury. We knocked down Pcdh-α and/or -γ using siRNA in dissociated adult mouse DRG neuron cultures and conducted a neurite extension analysis after 72h. We observed increased outgrowth following Pcdh-γ knockdown (p<0.05), as well as when both clusters were knocked down simultaneously (p<0.05) compared to a scrambled control. We observed a similar trend in the Pcdh-α knockdown (p = 0.09). These preliminary results indicate that the Pcdh protein may act as a regenerative “brake”, with self-recognition restricting outgrowth of sensory neurons during regeneration in order to facilitate structured patterning of skin re-innervation. Taken together, we demonstrate Pcdh expression in the PNS, primarily localized to the DRG sensory neuron perikarya , and these levels respond to peripheral axotomy injury. When Pcdh clusters are knocked down, there is an increase in total neurite outgrowth. This suggests that Pcdh may act as a restrictor for unwarranted sprouting, and may have important implications in manipulating the extent and patterning of peripheral axon regeneration.


Contributions of the amygdala to multiple sclerosis-related pain

Zoë Dworsky-Fried* (1), Bradley J Kerr (1,2,3), Anna MW Taylor (1,2,3)

1 - Department of Pharmacology, University of Alberta, Edmonton, Alberta, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Anesthesiology & Pain Medicine, University of Alberta, Edmonton, Alberta, Canada

Introduction: Chronic pain is a highly prevalent and disabling symptom associated with multiple sclerosis (MS). Recent data indicate that alterations within limbic brain circuitry are critical regulators of pain hypersensitivity in chronic pain states. However, we know very little about how these brain circuits change in MS and how these pathological adaptations might contribute to MS-related pain. The amygdala is a small nucleus within the limbic brain that integrates sensory and affective information. The central nucleus of the amygdala (CeA), in particular, plays an integral role in pain modulation. Amygdala processing of noxious stimuli is altered in chronic pain states, which contributes to pathological pain hypersensitivity. The amygdala is also involved in endogenous pain control via the opioid system. Chronic pain induces a loss of opioid function in the amygdala, which further influences pain hypersensitivity. Here, we describe how experimental autoimmune encephalomyelitis (EAE), the most frequently used animal model of MS, alters amygdala function and how these changes contribute to pain hypersensitivity and loss of endogenous opioid control. Methods: We employed the myelin oligodendrocyte glycoprotein (MOG)-induced EAE model in male and female C57BL/6 mice. The reinforcing properties of the μ-opioid receptor agonist, morphine, were assessed with the conditioned place preference paradigm. Morphine antinociception was measured using the tail withdrawal assay and the formalin test. Following behavioural testing, animals were euthanized and tissue was harvested. Neuronal activity and microglial activation in the CeA were analyzed with immunohistochemistry, and CeA μ-opioid receptor gene (OPRM1) expression was analyzed with fluorescence in situ hybridization. Results: EAE induction led to significant inflammation and T-cell infiltration in the spinal cord, which was accompanied by a reduction in thermal pain thresholds. Tissue analysis revealed increased levels of the immediate early gene, c-FOS, and robust microglial activation in the CeA at disease onset, while OPRM1 expression levels remained unchanged compared to controls. CeA neurons of EAE mice were less responsive to morphine, despite the higher basal activity. This decreased morphine response was correlated with lower pain thresholds, reduced morphine analgesia, and impaired morphine reward. Conclusions: Our data suggest that dysfunction of the μ-opioid system within the amygdala might contribute to altered pain control and perception in EAE. Our experiments provide insight into why opioids are less effective in treating MS-related pain and identify inflammation within the amygdala as a potential target to improve analgesic efficacy in this patient population. Financial support was provided by a Canada Graduate Scholarship from the Canadian Institutes of Health Research.


Differential effects of target height on immediate and delayed pointing actions: an ERP study

Ryan Moukhaiber*(1), Leanna Cruikshank (1), Jeremy B. Caplan (1,2), Anthony Singhal (1,2)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Psychology, University of Alberta, Edmonton, Alberta, Canada

An influential model of perception and action suggests that immediate, visually guided actions are driven by the dorsal visual stream, whereas delayed, memory-guided actions are also driven by the ventral stream (Goodale and Milner, 1992). Visually guided actions are those that we complete while using available visual information, such as reaching for a cup of coffee while looking at it. In the absence of visual information, our hands actions are guided by previously stored visual information; these are known as memory-guided actions. Consequently, memory-guided actions are less accurate and slower than visually guided actions (Cruikshank et al., 2012). Behavioral and Electroencephalography (EEG) studies investigating these action types have only compared hand action performance to objects in the lower visual field, even though many everyday hand actions are initiated toward objects in the upper visual field (i.e., reaching for a book on a shelf). The present experiment investigated hand action performance as a function of the height of the target, comparing target locations in the upper and lower visual fields. The N170 ERP was measured and used as a perceptual marker as it has been shown to be larger for memory-guided actions than visually guided actions (Cruikshank et al., 2012). We tested the hypothesis that since memory-guided actions place more demands on perception, they should take longer to execute in the upper compared to the lower visual field and be associated with a greater amplitude N170 ERP versus visually guided actions. Our results indicated that memory-guided actions were initiated faster than visually guided actions but were more erroneous overall. Move over, N170 ERPs exhibited a greater amplitude difference between conditions in the left hemisphere versus the right hemisphere, which replicates previous visuomotor literature. Contrary to our hypothesis, there was not an interaction between reach-height and either reaching type. This suggests that hand-arm kinematics and the neural circuitry underlying visually- and memory-guided actions may be more similar for targets in upper visual field.


Dynamic remodeling of an intrinsic inhibitory response

Shane Nicholls* (1) & Ray W Turner (1)

1 - Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada

A key mechanism to control neuronal activity is to generate afterhyperpolarizations (AHPs) following spike discharge, with the current study focused on the medium AHP (mAHP) and slow AHP (sAHP) of CA1 hippocampal pyramidal cells in vitro. Both AHPs reflect activation of the voltage and calcium-gated potassium channel (Kv7) and a calcium-gated IKCa potassium channel. The phosphatidylinositol PIP2 modulates Kv7 current amplitude, but the potential for PIP2 to interact with IKCa channels has not been reported. This is important in consideration of the ability for cholinergic inputs to modify PIP2 levels and the mAHP/sAHP. HYPOTHESIS: The Kv7 and IKCa channels that underlie the mAHP/sAHP in CA1 pyramidal cells exhibit modulation by the levels of PIP2. OBJECTIVES: Test the ability for PiP2 to modulate IKCa and/or Kv7 channels and the mAHP/sAHP using recordings in tsA-201 cells and CA1 pyramidal cells in vitro. IKCa and Kv7 currents will be pharmacologially isolated and the level of PIP2 raised using the agonist dIC8-PIP2 or reduced using blockers/phosphatases by direct infusion through the electrode during whole-cell recordings. RESULTS: Recordings of Kv7.2/7.3 co-expressed in tsA-201 cells confirm reports that an increase in PIP2 using 10µM dIC8-PIP2 increases Kv7 current amplitude, while infusion of 10 µM oxotremorine to lower PIP2 levels reduced or blocked Kv7 current. Kv7 current could be pharmacologically isolated in CA1 pyramidal cells and evoked as a tail current of ~200 msec using a step command from -70 mV to 0 mV (100 msec) to trigger intrinsic calcium currents. As found in tsA-201 cells Kv7 current was increased by internal infusion of dIC8-PIP2 and reduced by a calcium channel blocker. By comparison, changing the levels of PIP2 evoked an opposite response for IKCa channels expressed in tsA-201 cells or when isolated in CA1 pyramidal cells, with dIC8-PIP2 infusion decreasing and wortmannin increasing IKCa current amplitude. Voltage clamp recordings in pyramidal cells revealed an increase in the mAHP/sAHP duration with depletion of PIP2 using oxotremorine. CONCLUSIONS: PIP2 asserts opposing effects on IKCa and Kv7 current output in both cultured tSA-201 and in vitro CA1 hippocampal neurons, providing a potential means to dynamically modulate the ionic basis of the mAHP/sAHP. This work was supported by an NSERC Discovery grant (RWT) and an NSERC Canada Graduate Studentship (SN).

Poster Presentation 11 Effects of gangliosides on the secretion of extracellular vesicles in health and in Huntington’s

disease

Vaibhavi Kadam* (1), John Monyror (2), Jing Huang (3), Aislinn Maguire (1), Anissa Viveiros (2), Luis Carlos Morales (2), Qian Wang (2), Nitya Khetarpal (2), Aja Rieger (3), Elena Posse de

Chaves (1,2) and Simonetta Sipione (1,2) 1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Pharmacology, University of Alberta, Edmonton, Alberta, Canada 3 – Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada 4 –Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada Background: Gangliosides are glycosphingolipids containing one or more sialic acid residues, highly enriched in the brain and with vital roles in intercellular communication, cell signalling and calcium homeostasis. Many studies, including ours, have shown that levels of gangliosides, especially ganglioside GM1, are decreased in ageing and in neurodegenerative diseases such as Huntington’s disease (HD) and Parkinson’s disease (PD). Previous work in our laboratory has shown that restoring normal levels of GM1 has disease-modifying and therapeutic effects in HD mouse models, and reduces levels of the toxic protein, mutant huntingtin (mHTT), which causes the disease. These effects of GM1 are not due to changes in the transcription of the Htt gene, indicating the involvement of proteostatic mechanisms. Furthermore, proteomics analysis of HD mice brains (compared to untreated controls) suggests the involvement of extracellular vesicles as a potential pathway to eliminate mHTT. Extracellular vesicles (EV) are membrane-enclosed nanoparticles secreted by most cells, with roles in cell-cell communication, immune-modulation, and proteostasis. They are also known to carry misfolded proteins. Thus, we hypothesize that GM1 may exert its beneficial activity by stimulating vulnerable neurons to secrete EVs packaged with mHTT, thus decreasing cellular proteotoxic stress. Results: Secretion of EVs was measured in primary embryonic neurons, human fibroblasts, Neuro2a cells, as well as a neuronal cell line transfected with mHTT-eGFP by fluorescence, imaging flow cytometry, nanoparticle tracking analysis and immunoblotting for EV markers (Flotillin-1, ALIX, CD9, CD81). Cells were pre-treated with the lipophilic dye Vybrant™ DiI or DiD in order to label membranes, including EVs. EVs were isolated from cell-conditioned media by either ultracentrifugation or size exclusion chromatography. We have found that increasing cellular levels of GM1 by its exogenous administration results in increased secretion of EVs, while inhibition of the ganglioside biosynthetic pathway in cells produces the opposite effect. We further show that, in HD cells, administration of GM1 increases the secretion of mHTT in EVs, thus contributing to explain, at least in part, the neuroprotective activity of this ganglioside in HD models. Future studies will confirm these findings in animal models.


Evaluating the subsequent memory effect as predictive of memory

Sucheta Chakravarty* (1), Yvonne Y. Chen (2) & Jeremy B. Caplan (1,3)

1 - Department of Psychology, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Neurosurgery, Baylor College of Medicine, Houston, Texas, USA 3 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada

To isolate brain activity that reflects effective processes during the study phase of a memory task, cognitive neuroscientists commonly contrast brain activity during study of later-remembered versus later-forgotten items. This 'subsequent memory effect' method has been described as identifying brain activity 'predictive' of memory outcome. However, decades of behavioural research has told us that memory success depends not only on cognitive processes during study of an item, but on many processes that occur at other times (e.g., competition from other studied items, study-test compatibility, etc.). We show that conventional event-related potential 'subsequent memory effect' signals are predictive, but indeed, only to a small degree (N=59, 225 items/participant). This improves when machine-learning classifier methods developed are applied, but the predictive effects are still modest. These findings suggest that the term 'predictive' is, at a minimum, over stating the standard subsequent memory effect. For an approach to study-related brain activity to be more predictive may require integrating the myriad and interesting other factors known to influence memory outcome, with standard univariate, as well as classifier-based approaches.


Facilitation of reflex by GABA

Krishnapriya Hari*, Shihao Lin, Ana M. Lucas-Osma, Keith K. Fenrich, David J. Bennett

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Faculty of Rehabilitation Medicine, University of Alberta, Edmonton, Alberta, Canada

GABA is a major inhibitory neurotransmitter in the central nervous system and critically important for propriospinal reflex circuitry. Classically, GABA is thought to be involved in the inhibition of sensory neurotransmission to motoneuron via PAD circuit (Primary Afferent Depolarization). However, using immunohistochemistry techniques we recently identified GABAA receptors near nodes and at many branch points along afferent axons while being absent at afferent terminals. This nodal GABAA receptors aids sodium spike initiation and facilitates action potential propagation beyond the branch points where as GABAB receptors at terminals are engaged in presynaptic inhibition. We used genetically modified mice that express optogenetic actuators (Channelrhodopsin-2) in GAD2 neurons which contact the afferents and release GABA. With the use of electrophysiological techniques, we recorded monosynaptic EPSP which gives the measure of afferent to motoneuron signaling from ventral roots by stimulating the dorsal roots of spinal cord in-vitro in response to blue light (excites GAD2 neurons) as well as pharmacological agents. Activation of GAD2 neurons with light facilitated the monosynaptic reflex. This facilitation was almost doubled with the application CGP55845 (GABAB receptor antagonist) while it was inhibited in Gabazine (GABAA receptor antagonist) showing the excitatory action of GABA via GABAA receptors for the first time. In summary, the release of GABA prime the reflex via GABAA receptors and modulates the sensory afferent transmission. This novel finding opens the possibility of new pathways for the regulation of nerve conduction.


Fluctuating Cognition in Neurodegenerative Disease: Comparing Reaction Time Mean Rate and Inconsistency

Sebastian Caballero* (1), G. Peggy McFall (1,2), Myrlene Gee (3), Krista Nelles (3), Jacqueline

Burt (3), Stuart MacDonald (4), Roger A. Dixon (1,2), and Richard Camicioli (1,3)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Psychology, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Medicine, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Psychology, University of Victoria, Victoria, British Columbia, Canada Fluctuating cognition is characterized by marked intra-individual variations in attention and alertness. Elevated fluctuations may distinguish Lewy Body Dementia (LBD) and Parkinson’s Disease Dementia (PDD) from Alzheimer’s Disease (AD) and related disorders. The Canadian Comprehensive Assessment of Neurodegeneration and Dementia includes LBD-PDD, AD, Parkinson’s disease (PD), mild cognitive impairment (MCI), and PD-MCI cohorts. Two multi-trial reaction time (RT) tests provide measures of mean rate and variability in speeded performance. Objective: We compared RT performance (mean rate) and inconsistency (intraindividual variability) across all cohorts. The three research goals were: (1) examine group differences in latency, (2) examine group differences in inconsistency of latency, and (3) test the relative discriminative potential of latency and inconsistency. Method: The sample included the following clinically diagnosed subjects: 10 LBD-PDD (M age = 75.01), 21 AD (M age = 75.80), 12 PD (M age = 67.22), 61 MCI (M age = 71.74), and 9 PD-MCI (M age = 74.60). Two RT tasks were: simple reaction time (SRT, 50 trials) and four-choice reaction time (CRT4, 60 trials). Individual mean rates were calculated as average latency. Inconsistency was indexed by computing the raw intraindividual standard deviations. Prediction analyses were performed with logistic regression. Results: First, significant group differences in latency were observed: SRT, F(4, 108) = 6.84, p < .001; CRT4, F(4, 108) = 11.97, p< .001. For both tasks, follow-up analyses showed: (a) LBD-PDD and AD exhibited slower RT than MCI and (b) LBD-PDD exhibited slower RT than AD, PD, and PD-MCI. Second, significant group differences in intra-individual inconsistency were observed: SRT, F(4, 108) = 7.01, p < .001; CRT4, F(4, 108) = 7.17, p < .001. For both tasks, follow up analyses showed that LBD-PDD was more inconsistent than PD and MCI. For SRT, LBD-PDD was more inconsistent than AD and PD-MCI. Third, latency significantly discriminated group membership between (a) MCI vs. AD, (b) MCI vs. LBD-PDD, (c) PD vs. LBD-PDD, and (d) PD vs. AD. Inconsistency significantly discriminated (a) MCI vs. LBD-PDD, (b) PD vs. LBD-PDD, (c) AD vs. LBD-PDD, (d) MCI vs. AD, (e) PD vs. AD, and (f) MCI vs. PD-MCI. Conclusions: Overall, these neurocognitive speed tasks discriminated the LBD-PDD group from the other cohorts in both average rate (slower) and greater fluctuations (inconsistency). Specifically, these measures discriminated LBD-PDD from AD. Furthermore, prediction analyses revealed that latency and inconsistency significantly discriminated the more cognitively impaired groups from the less cognitively impaired groups.


Functional Connectivity Changes of Resting State Networks in ALS: A Multicenter Study

Pedram Parnianpour, Sanjay Kalra 1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Canada 2 - Department of Biomedical Engineering, University of Alberta, Edmonton, Canada 3 - Division of Neurology, University of Alberta, Edmonton, Canada Introduction Amyotrophic lateral sclerosis (ALS) is a rare neurodegenerative disease characterized by the selective degeneration of upper and lower motor neurons. Even though ALS is categorized as a motor neuron disease, studies have illustrated degeneration in extra-motor regions. This study aims to investigate disease-related changes on the baseline function of the brain in the resting-state networks (RSNs) and to explore clinical associations with these changes. Method Data from seventy-one ALS patients and fifty-four healthy subjects who participated in the Canadian ALS Neuroimaging Consortium (CALSNIC) was analyzed. Preprocessing and group independent component analysis were applied to raw data and twelve RSNs were identified. Then, functional connectivity (FC) of the networks and inter-network connections were assessed and compared between patients and the control group. Finally, correlations between FC and behavioral and clinical scores were evaluated in ALS patients. Results Significant FC changes were demonstrated for ten networks (p < 0.001, cluster size threshold = 20 voxels). Inter-network connectivity exhibited decreased FC between sensorimotor and executive control networks and increased FC between sensorimotor and medial visual networks. Decreased FC between the sensorimotor and executive control networks was associated with decreased walking performance of patients. Increased FC between the sensorimotor and medial visual network was associated with increased upper motor neuron signs on neurological examination. Conclusion In summary, FC changes exist between the sensorimotor network and non-motor networks. The presence of clinical correlations with these FC changes suggest abnormalities in these networks play a role in behavioral manifestations in ALS.


Ganglioside GM1-dependent uptake and clearance of neuronal extracellular vesicles by microglia in health and Huntington’s disease

John Monyror* (1), Danny Galleguillos (1,2), Vaibhavi Kadam (2), Matthew Doan (3),

Christopher Power (2,3,4), Nicolas Touret (5), Matthew Macauley (4,6), Simonetta Sipione (1,2)

1 - Department of Pharmacology, University of Alberta, Edmonton, Alberta, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Medicine, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Medical Microbiology & Immunology, University of Alberta, Edmonton, Alberta, Canada 5 - Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada 6 - Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada

Gangliosides are sialic acid-containing glycosphingolipids that are most abundant in the central nervous system. These lipids are anchored into the plasma membrane via ceramide and expose a headgroup composed of sugar groups and sialic acid residues to the extracellular space. Gangliosides are therefore poised to participate in several cell-cell interactions and have been shown to modulate immune cell signalling. Several of these interactions appear to be mediated by sialic acid residues, often through binding to sialic acid-binding (siglec) proteins. In Huntington’s disease, a protein misfolding neurodegenerative disease caused by the expansion of an N-terminal polyglutamine stretch in the protein huntingtin, levels of ganglioside GM1 are greatly reduced. In mouse models of Huntington’s disease, administration of GM1 reverses motor and non-motor symptoms. Strikingly, these changes are accompanied by a significant decrease in levels of both soluble and aggregated mutant huntingtin (mHTT) through a mechanism independent of transcription. We have found that treatment with GM1 facilitates neuronal secretion of mHTT within extracellular vesicles (EVs) - membrane-enclosed nanoparticles that range between 30-1000 nm in diameter – thus alleviating the intracellular toxic burden on neurons. Secretion of mHTT within EVs, however, could contribute to disease spreading in a prion-like manner, if EVs and their toxic cargo are not cleared from the brain. Here, we investigate how microglia may eliminate these neuronal EVs and mHTT packaged within them in a ganglioside-dependent manner, to reduce the overall burden of mHTT in the brain. We hypothesize that GM1 decorating the surface of EVs promotes uptake of EVs and preferential sorting of toxic cargoes such as mHTT into lysosomes for degradation. We further hypothesize siglec-1 as the putative receptor for neuronal EVs on microglia. Our data suggest that EVs derived from cells treated with GM1 are enriched with this ganglioside and are taken up more efficiently by murine and human microglia compared to GM1-poor EVs. Furthermore, we have found that microglia derived from siglec-1 KO mice show impaired uptake of EVs, in spite of normal phagocytic activity towards latex beads and macropinocytosis of dextran. Finally, enrichment of GM1 on EVs appears to increase localization of EV-bound mHTT to microglial lysosomes. Our data provides new insights on how gangliosides can modulate EV clearance to enhance brain proteostasis in health and disease. This work also sheds light on the mechanisms underlying the disease-modifying effects of GM1 in Huntington’s disease mice. Funding support: CIHR, APRI, GlycoNET, SynAD


Human Pegivirus is neurotropic and associated with neuroinflammation

Matthew Doan* (1), William Branton (2), Brienne McKenzie (3), Tom C. Hobman (4), Benjamin Gelman (5), Christopher Power (1,2,3)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Medicine, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Cell Biology, University of Alberta, Edmonton, Alberta, Canada 5 - Department of Pathology, University of Texas Medical Branch, Houston, Texas, USA Viral infections are among the most commonly identified causes of encephalitis in humans. Human pegivirus (HPgV) is a positive sense, single-stranded RNA virus that is a member of the Flaviviridae family. We recently reported HPgV infection in the brains of two patients with fatal leukoencephalitis (Balcom, Doan et al., 2018). HPgV antigen (NS5A) was detected chiefly in glial cells (astrocytes and oligodendrocytes) in cerebral white matter. To further investigate HPgV infection of human glial cells, we transfected astrocytoma U251 cells with a molecular clone of HPgV from which viral stocks were prepared. Primary human astrocyte cultures were then infected using these stocks and immunodetection of NS5A was performed 4, 7 and 14 days post-infection (PI). The mean viral spread (HPgV+ immunodetection/μm 3 ) peaked at Day 7 PI and droplet digital PCR (ddPCR) analyses of astrocyte culture supernatants showed 7577, 10834 and 2905 viral RNA copies/mL at Days 4, 7 and 14 PI, respectively. Viral antigen detection in astrocytes was associated with cell lysis and cell death, evidenced by increased lactose dehydrogenase in the media and a decrease in DAPI intensity. To determine the in vivo neuroinflammatory effects of HPgV infection, we identified an additional 11 (of 109) patients with HPgV brain infection (700-11,000 HPgV RNA copies/gm). The expression of proinflammatory cytokine genes (IL1B, TNFA, IL6) was markedly increased in the brains from patients with HPgV co-infected with HIV (n=6) compared to HPgV mono- infected (n=5) patients. The present studies indicate that HPgV is a neurotropic virus with the capacity to infect and spread in primary human glial cells. Moreover, HPgV might act synergistically with HIV-1 to promote neuroinflammation in vivo. These findings highlight the importance of considering HPgV as a potential cause of encephalitis in humans and exploring its associated neuroinflammation implicated in other neurological diseases.


Large-Scale functional connectivity alterations in Amyotrophic Lateral Sclerosis: A multicenter study

K. Bharti* (1), M. Khan (1), P. Parnianpour (1), S. Kalra (1), L. Korngut (2), R. Frayne (2), H. Bremberg (3), C. Shoesmith (4), A. Genge (5), A. Donne (6), N. Dupre (6), S. Graham (7), L.

Zinman (7), M. Benatar (8), S. Gibson (9), R. C. Welsch (9)

1 - University of Alberta, Edmonton, AB, Alberta, Canada; 2 - University of Calgary, Calgary, Alberta, Canada; 3 - University of British Columbia, Vancouver, BC, Canada; 4 - Western University, London, Ontario, Canada; 5 - McGill University, Montreal, Quebec, Canada; 6 - Université Laval, Quebec, Quebec, Canada; 7 - University of Toronto, Toronto, Ontario, Canada; 8 - University of Miami, Miami, FL, Canada; 9 - University of Utah Utah, UT, Canada Amyotrophic lateral sclerosis is a multisystem neurodegenerative disorder characterized by progressive upper motor neuron, lower motor neuron, and frontotemporal lobar degeneration resulting in muscular weakness and cognitive impairment. Functional magnetic resonance imaging based studies have reported several cortical and subcortical brain structures implicated in the pathophysiology of Amyotrophic lateral sclerosis. Resting-state functional magnetic resonance imaging studies using an independent component analysis approach are very few in number and most importantly have been restricted to a limited number of resting-state networks. In order to address this gap in the literature, we investigated functional abnormalities in the motor and cognitive domains of multiple resting-state networks and their importance in the pathophysiology of amyotrophic lateral sclerosis. In this study, we examined the intra-network resting-state functional connectivity using an independent component analysis approach, and its clinical correlations with behavioral data in a large multi-centered cohort. A total cohort of 235 subjects (115 healthy subjects; 120 patients with amyotrophic lateral sclerosis) were recruited across North America through the Canadian Amyotrophic Lateral Sclerosis Neuroimaging Consortium. Structural and functional data analysis was performed using FSL. Preprocessing steps include brain extraction of the T1 weighted-3D images using the brain extraction tool, head motion correction using the MCFLIRT tool, slice timing correction and spatial smoothening at full width half maximum of 5 mm. The group melodic output of 13 independent components representing the best resting-state networks were selected according to the previous literature. Intra-network resting-state functional connectivity differences were assessed for the 13 resting-state networks by means of two-sample unpaired t-test using the threshold free cluster enhancement technique and 5000 nonparametric random permutations in FSL 24. Statistical differences were assessed within the mask of respective resting-state networks using general linear model and covariates of no interest (age, gender, motion parameters, sites of data acquisition, and types of scanner). Group differences and clinical correlation results were corrected for multiple comparisons using family wise error approach at a significance level of p<0.05. As compared to healthy controls, patients with Amyotrophic lateral sclerosis displayed higher intra-network resting-state functional connectivity in 11 out of 13 resting-state networks. Clinical disability negatively correlated with higher intra-network resting-state functional connectivity in 11 out of 12 resting-state networks. Additionally, significant clinical correlations were also observed with disease progression rate, symptom duration, upper motor neuron burden, tapping scores, and cognitive performance. The present study demonstrates that Amyotrophic lateral sclerosis is associated with abnormal intrinsic functional activity in large scale resting-state networks which subserves motor and cognitive abilities.


Locomotion-related V3 neurons regulate sensory transmission

Shihao Lin*, Ana M. Lucas-Osma, Yaqing Li, Krishnapriya Hari, Marilee J. Stephens, Leo Sanelli, Karim Fouad, Keith F. Fenrich, David J. Bennett

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Faculty of Rehabilitation Medicine, University of Alberta, Edmonton, Alberta, Canada After spinal cord injury (SCI), the locomotion-related V3 interneurons mediate an NMDA dependent long-lasting sensory reflexes known as spasms. Here, we examined the role of V3 neurons on sensory transmission to understand spasms. Confocal microscopy revealed that V3 neurons form a circuit that allow sensory inputs to generate primary afferent depolarization (PAD) in the afferents. Intracellular recording revealed that optogenetic activation of V3 neurons generate a depolarization in afferents. We found that NMDA is involved in the synaptic transmission by which V3 neurons drive PAD. This NMDA component is crucial in supporting V3-mediated PAD in amplifying sensory reflexes. After SCI, V3 neurons become NMDA dependent in generating PAD suggesting greater potential in reflex-amplification. We think spasms occur due to excessive amplification of reflexes by V3 PAD. Therefore, these results suggest a mechanisms underline spasms and provide a therapeutic target for controlling spasms.


Longitudinal post-stroke two-photon calcium imaging of the limb-associated somatosensory cortex in head-fixed mice within the Neurotar homecage

Mischa V. Bandet* (1,2) and Ian R. Winship (1,2,3)

1 – Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 – Neurochemical Research Unit, University of Alberta, Edmonton, Alberta, Canada 3 – Department of Psychiatry, University of Alberta, Edmonton, Alberta

Ischemic stroke leads to alterations in the balance of excitation/inhibition as cortical plasticity re-wires lost connections and compensates for loss of function within damaged tissue. Previous studies have largely used acute imaging preparations and anesthetized animals to study ongoing changes in the response selectivity of individual neurons post-stroke. Unfortunately, acute preparations do not allow for the tracking of cell populations across multiple imaging timepoints and anesthetized preparations result in a lack of self-generated corollary signals occurring during self generated movement. Here we used in vivo two-photon calcium imaging within the somatosensory cortex of GcAMP6S mice with the aim of measuring activity within large neuronal networks as the mouse moved around within the Neurotar mobile homecage. Neuronal activity within the peri-infarct region was measured weekly prior to, and for two months following photothrombotic stroke directed to the forelimb somatosensory cortex. Behavioral recovery was measured using an automated tapered beam task and string-pulling task, and correlated with changes in cell activity and remapping of individual neurons over time. Our data suggest a decrease in the movement elicited somatosensory activity within the first several weeks post-stroke that re-emerged coincident with behavioral recovery. Ongoing analysis will identify local network changes in peri-infarct cortex that correlate with good or poor recovery.


Microglia regulate remyelination through myelin debris clearance

Baaklini CS*(1), Lopez M(2), Ho M(3), Kerr BJ(4), Plemel JR(1)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Biological Sciences, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Cell Biology, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Pharmacology, University of Alberta, Edmonton, Alberta, Canada Demyelination of axons leaves them vulnerable to degeneration. Boosting remyelination can protect axons and may, therefore, be a new neuroprotective strategy to treat white matter diseases such as Multiple Sclerosis. Phagocytes in the CNS clear inhibitory myelin debris to promote an environment permissive for remyelination. However, the extent to which microglia and CNS-infiltrating macrophages contribute to this process is unclear. It is also unclear whether these cells have other roles during remyelination. I hypothesize that microglia regulate remyelination through oligodendrocyte progenitor cells recruitment and differentiation and myelin debris phagocytosis. To differentiate between these cell types, we fate-mapped microglia and other CNS macrophages by labeling them with TdTomato using CX3CR1CreER; RosatdTom mice. To determine the contributions of these cells to remyelination, injected the gliotoxin lysolecithin into the ventral spinal cord white matter. At the peak of phagocytosis, we measured the volume of myelin inside microglia (tdTom+ve) and infiltrating macrophages (TdTom-ve). We find that although microglia outnumber CNS-infiltrating macrophages, the total volume of phagocytosed myelin is equivalent between them because each macrophage phagocytosed more myelin. To determine if microglia are necessary for different steps of remyelination, we ablate microglia by injecting diphtheria toxin into transgenic mice that express the diphtheria toxin receptor on microglia. Preliminary data show that when microglia numbers are reduced, there is decreased oligodendrocyte differentiation suggestive of delayed remyelination. Understanding the immune cell involvement in remyelination will aid in the development of therapies.


Modulation of the spinal cord circuits and cortico-spinal tract via transcutaneous spinal cord stimulation

Behdad Parhizi* (1), Trevor Barss (2), Vivian Mushahwar (3)

1 - Neuroscience and Mental Health Institute, University of Alberta 2 - Department of Medicine in the Faculty of Medicine and Dentistry, University of Alberta 3 - Department of Medicine, Division of Physical Medicine and Rehabilitation, University of Alberta

Introduction: Coupling between cervical and lumbar spinal networks (cervico-lumbar coupling) is well-established in humans. We demonstrated that after incomplete spinal cord injury, cervico-lumbar coupling and spinal reflexes are impaired. Arm and leg cycling training restores these impairments to pre-injury levels. The outcomes of this rehabilitation training can be further enhanced using transcutaneous spinal cord stimulation (tSCS), by modulating the circuitry of the spinal cord non-invasively. In this project, we probed the effect of tSCS on cervico-lumbar coupling and on the excitability of the corticospinal tract. Methods: Neurologically-intact volunteers were seated in a recumbent leg cycling system. The activity of the flexor carpi radialis (FCR) muscle (H-reflex, motor evoked potential (MEP)) was assessed under 8 conditions with FCR activated to 10% maximal voluntary contraction: 1) legs static, 2) legs static with cervical tSCS applied to C3-4 and C6-7; 3) legs static with lumbar tSCS applied to T11 and L1; 4) legs static with simultaneous cervical and lumbar tSCS (combined tSCS). These conditions were repeated with the legs cycling. Results: While cervical tSCS non-significanlty increased the FCR H-reflex amplitude by 6.8% relative to the legs static, no tSCS condition, lumbar tSCS and combined tSCS significantly facilitated the FCR H-reflex by 11.1% and 19.6%, respectively. This facilitation may be a result of direct activation of posterior root afferents by tSCS and indirect activation of propriospinal networks. Leg cycling significantly suppressed the FCR H-reflex relative to the legs static, no tSCS condition. The suppression was present even when tSCS was applied to the cervical-, lumbar- or both cervical and lumbar regions simultaneously. Thus, leg cycling canceled the facilitatory impact of tSCS on the FCR H-reflex. Cervical tSCS as well as combined tSCS significantly increased MEP amplitude in FCR by 4.9% and 19.7%, respectively, compared to the legs static, no tSCS condition. This may provide novel evidence that tSCS allows spinal cord networks to become more accessible to supraspinal centers. Cycling also increased the amplitude of MEP by 18.6%; however, there were no further increases in MEP amplitude with the addition of tSCS to the cervical-, lumbar-, or cervical and lumbar regions simultaneously. Significance: Neuromodulation of spinal circuitry improves functions such as standing, posture, stepping, and mitigates spasticity. This project demonstrates, for the first time, that tSCS could play an important role in regulating cervico-lumbar connectivity and enhancing descending drive. Coupled with rehabilitation paradigms, it may further improve arm function and walking after neurological conditions.


Pleiotrophin as a modulator of neurite outgrowth, neuroinflammation and OPC differentiation

Somnath Gupta* (1,2), Matthew A Chruchward (1,2), Ian R. Winship (1,2), Kathryn G. Todd (1,2)

1 - Neurochemical Research Unit, Department of Psychiatry, University of Alberta, Edmonton, Alberta, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada

After CNS injury such as ischemic stroke, chondroitin sulphate proteoglycans (CSPGs) are produced by glial cells in extracellular matrix surrounding the injury. CSPGs are growth inhibitory, reducing axonal sprouting growth and migration of OPCs and thereby impairing recovery. Pleiotrophin (PTN), a growth factor and a cytokine, is upregulated in the central nervous system (CNS) during development and after injury. It has been proposed that PTN binds to CSPGs in the extracellular matrix to reduce CSPG inhibition of neuron and OPCs growth. However, the effect of CSPGs and PTN on different classes of glial cells is not well described. Here, we investigated the effect of PTN and PTN signaling on primary neuronal and glial cultures. First, neurons were plated on growth inhibitory CSPG matrices or growth permissive laminin matrices and treated with varying concentrations of PTN. PTN increased neurite outgrowth in CSPG matrices by binding to ALK receptor and activating Akt signaling pathway, suggesting that PTN can induce growth even in inhibitory environments such as the CSN after injury. Next, OPCs, microglia and astrocytes were isolated from mixed mouse glia cultures and plated on growth inhibitory CSPG matrices or growth permissive laminin matrices and treated with varying concentrations of PTN. PTN increased the proliferation of OPCs and their differentiation to mature oligodendrocytes in CSPGs matrices. Microglia plated on CSPGs had enhanced release of BDNF and IL 10 cytokines relative to laminin controls. PTN potentiated the release of IL 10 cytokine and BDNF while suppressing the release of NO, TNF, IL 1 and IL 6 from microglia in these CSPGs matrices. Combined, these data suggest that PTN signaling modulates axonal growth, remyelination process and promotes anti-inflammatory response even in inhibitory environments, thus may have potential as a pro-plasticity therapy following CNS injury.


Potential role of Tau in Kainic acid-induced Animal Model of Temporal Lobe Epilepsy

Abhishek Dahal* (1,2) and Satyabrata Kar (1-3)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Center for Prions and Protein Folding Diseases, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Medicine (Neurology), University of Alberta, Edmonton, Alberta, Canada

Introduction Kainic acid (KA) is an analog of the excitatory neurotransmitter glutamate that when injected systemically into adult rats can trigger seizures and neuronal loss in a manner that mirrors the neuropathology of human mesial temporal lobe epilepsy (mTLE). However, mechanisms responsible for the neuronal loss remains elusive. Some earlier studies have shown that the tau protein, which is critical in the development of Alzheimer’s disease, may have a role in triggering the seizures and/or loss of neurons associated with mTLE. However, the cellular site or the mechanism by which tau can influence mTLE pathology is still unclear. Thus, we are currently evaluating the potential role of tau in the KA-induced animal model of mTLE. Methods Adult male rats were injected intraperitoneally with either KA (12mg/kg) or normal saline. Control and KA-treated rats were euthanized at 12 hours, 2 and 12-days following treatment and their hippocampal brain regions were processed to evaluate the loss of neurons, activation of astrocytes and level/expression of total, phospho- and cleaved-tau using western blotting and immunohistochemistry. Results We observed that KA administration evoked seizures and progressive loss of neurons and activation of astrocytes in the hippocampal region of the brain. Interestingly the level/expression of astrocytic markers GFAP and Vimentin, but not Glutamine synthetase (GS) or S100β, are increased in KA-treated rats. This is supported by our double labelling studies which showed an increase in GFAP+Vimentin labelled astrocytes but no alterations in GFAP+GS labelled astrocytes in KA-treated rats suggesting different subsets of astrocytes are deferentially affected following KA treatment. In addition, we also observed an increase in GFAP+S100β labelled astrocytes following KA treatment. Furthermore, the level/expression of total and to some extent phospho-tau are elevated after 12-hour post-treatment and then decreases to the normal level in KA-treated rats. Additionally, we detected that the level and expression of caspase-cleaved tau in astrocytes were markedly increased in the hippocampus of KA-treated rats. Conclusion Our results, obtained so far, indicate that KA-induced seizures not only deferentially affect astrocytes but also increase the levels/expression of tau that can influence the development of mTLE pathology.


Presynaptic Control of Primary Afferent Conduction in Humans

Krista Metz* (1, 2), Dave Bennett (1, 3), Monica Gorassini (1, 4) 1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada 3 - Faculty Rehabilitation Medicine, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Biomedical Engineering, University of Alberta, Edmonton, Alberta, Canada Sensory inputs conveying proprioceptive and kinematic information are critical in shaping and modulating motor behaviour. GABA receptor activation is highly involved in the control of action potential propagation along Ia afferents, and therefore, the control of sensory information in the spinal cord. Activation of GABAA receptors along Ia afferents causes a primary afferent depolarization (PAD) because high intra-cellular chloride concentrations rush out of the cell when the receptors are opened. Historically, it was thought that activation of GABAA receptors on the Ia afferent terminal could only reduce sensory transmission traveling along Ia afferents, termed presynaptic inhibition. Recently, however, animal data has shown that activation of GABAA receptors along the Ia afferent far from the terminal can secure action potential propagation at branch points, where sensory transmission tends to fail. Thus, activation of GABAA receptors along the Ia afferent can secure sensory transmission, which results in greater activation of motoneurons and enhanced motor output, termed presynaptic facilitation. A longer-lasting tonic PAD, due to a build-up of extracellular GABA and activation of α5GABAA receptors, can also facilitate afferent transmission for 10s of seconds after the initial PAD stimulation. In light of this, we examined the human nervous system to find evidence of presynaptic facilitation in human participants and re-examined classical experiments investigating presynaptic inhibition in human participants. The H-reflex, which is a measure of sensory-evoked motor output in humans, was conditioned by a variety of inputs that putatively activate PAD. Modulation of the H-reflex from the conditioning stimulation was then compared to whole EMG and single motor unit recordings to dissociate presynaptic effects on the Ia afferent and postsynaptic effects on the motoneurons. The probability of discharge of a single motor unit before and after the putative PAD-producing input was also measured. Experiments were conducted in both healthy control participants and individuals with spinal cord injury. It appears that presynaptic facilitation of primary afferents exists in human participants, as evidenced by an increase in the size of the H-reflex from the conditioning input without a subsequent depolarization of the motoneurons. Classical presynaptic inhibition experiments appear to be contaminated by post-synaptic effects on the motoneurons. After a spinal cord injury, both inhibition of the H-reflex and tonic facilitation of the H-reflex were reduced. While further investigation is needed to determine the exact mechanisms, this presents a potential novel treatment target for sensory-related dysfunction and spasticity.


Sense and Sensitivity: Sex Differences in Macrophage Responses to TNFa

Timothy N. Friedman* (1), Bradley J. Kerr (2, 3, 4)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Anesthesiology and Pain Medicine, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Psychiatry, University of Alberta, Edmonton, Alberta, Canada 4 - Department of Psychology, University of Alberta, Edmonton, Alberta, Canada

Autoimmune diseases like Multiple Sclerosis and rheumatoid arthritis display sex biases towards women in addition to higher rates of chronic pain in women. Using the EAE animal model of MS, we have recently described significant alterations in the transcriptomic profile of the dorsal root ganglia (DRG). Significantly, miR-21 was upregulated in both sexes. miR-21 has been implicated in immune cell activation. However, the rate or extent at which this activation occurs is unclear. We hypothesize that female immune cells are intrinsically more sensitive to inflammatory signals through pathways mediated by miR-21. To address this, we treated bone-marrow derived macrophages (BMDMs), isolated from male and female mice, with escalating doses of the inflammatory cytokine tumor necrosis factor alpha (TNFa) for 24 hours. Macrophage activation was assessed by qPCR and ICC for miR-21, TLR7, TLR8, iNOS, and ARG1. We observed an increased sensitivity to TNFa stimulation in female BMDMs. While miR-21 was upregulated by TNFa in both sexes, female BMDMs had increased levels of iNOS and TLR7 and TLR8, endogenous receptors for miR-21. Our findings demonstrate that miR-21 expression is stimulated by TNFa exposure in vitro and that this shift in expression is associated with a more inflammatory phenotype primarily in female macrophages. The increased expression of miR-21 coupled with the upregulation of its endogenous receptor system in female macrophages may underlie a more aggressive, inflammatory phenotype that has implications for their interactions with sensory neurons in the DRG.


Sex-based ER and Mitochondrial changes in peripheral sensory neurons in response to inflammatory stimuli

Aislinn Maguire*, Fajr Haq, Bradley Kerr

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Anaesthesiology and Pain Medicine, University of Alberta, Edmonton, Alberta, Canada 3 - Department of Physiology, University of Alberta, Edmonton, Alberta, Canada Sensory neurons in the dorsal root ganglion (DRG) have been shown to become hyper-excitable/sensitized in response to chronic inflammatory stimuli as often occurs in autoimmune disorders like rheumatoid arthritis or Multiple Sclerosis (MS). This sensitization in response to inflammation has been postulated to be an underlying cause of the chronic pain associated with these disorders. The molecular pathways mediating sensitization, however, remain to be fully elucidated. We hypothesize that endoplasmic reticulum (ER) stress coupled with morphological and functional changes in the mitochondria of nociceptors (pain sensing sensory neurons) are involved in this process. Given the well-established sex bias of painful autoimmune disorders, we also predict that these changes are more prominent in female nociceptors. We have begun to test these hypotheses by stimulating male and female DRG sensory neurons in vitro with increasing doses of the inflammatory cytokine TNFα, and measuring markers of ER stress (pPERK and peIF2a), as well as examining subsequent changes in mitochondrial morphology and function between the sexes. We find that 24hrs of TNFα stimulation leads to an increase in peIF2α levels in both sexes but significant changes in mitochondrial morphology only occur in female sensory neurons. Ongoing studies are evaluating other arms of the integrated stress response to better elucidate the mechanisms mediating nociceptor, ER-mitochondrial crosstalk under chronic inflammatory conditions. These pathways may represent novel targets to treat pain associated with chronic, neuroinflammatory disorders.


Structural and neurometabolic correlates of motor neuron impairment in Amyotrophic Lateral Sclerosis (ALS): a multimodal fMRI-MRS-DTI-Texture study

Avyarthana Dey*, Ojas Srivastava, Abdullah Ishaque, Daniel Ta, Chris Hanstock, Dennell Mah,

and Sanjay Kalra

1 - Neuroscience and Mental Health Institute, University of Alberta 2 - Division of Neurology, Faculty of Medicine and Dentistry BACKGROUND: ALS is a neurodegenerative disorder characterized by upper and lower motor neuron loss resulting in limb, speech and swallowing paralysis, respiratory impairment, and death within 3-5 years of diagnosis. Magnetic resonance imaging (MRI) has revealed widespread alterations in cortical functional connectivity (FC), abnormal texture in the gray matter, reduced fractional anisotropy (FA) in the white matter, and altered neurochemical levels (reduced N-acetylaspartate to creatine ratio [NAA/Cr]). The contribution of structural and neurochemical changes to FC is unknown. We hypothesize that the FC between brain regions and its impairment in disease even in a resting or no-task condition is explained by the underlying structural and neurochemical properties of these brain regions. OBJECTIVE: To determine if impaired FC of the primary motor cortex (PMC) in ALS is associated with underlying structural damage and neurochemical deficits. METHODS: Fifty-nine patients with ALS and forty-six healthy controls (HC) were recruited from four Canadian centers (University of Calgary, University of Alberta, McGill University, University of Toronto) as part of a nationwide study in the Canadian ALS Neuroimaging Consortium (CALSNIC). Multimodal MRI data (T1, magnetic resonance spectroscopy, diffusion tensor imaging, and resting state functional MRI) was used in a statistical full factorial model to analyze the extent of alterations in FC, texture, microstructural white matter integrity, and neurochemical levels within the PMC. Multivariate linear regression analyses were performed to examine whether any alterations in the microstructural and neurochemical properties explain the functional activation patterns in the PMC. RESULTS: Reduced FA values and NAA/Cr levels in the PMC was observed in patients compared to HCs. However, there were no significant texture changes. Furthermore, the PMC exhibited an increase in FC with the superior parietal lobule and a decrease in FC with the middle frontal gyrus. These FC alterations showed significant associations with reduced FA and NAA/Cr values in the PMC. CONCLUSION: Underlying structural and neurochemical deficits of the PMC in ALS play an important role in explaining FC alterations.


Subcutaneous injection of a novel peptide-conjugated morpholino oligomer DG9-PMO improves symptoms with enhanced SMN2 exon 7 inclusion in a severe mouse model of spinal

muscular atrophy

Tejal Aslesh* (1), Rika Maruyama (2) Hong Moulton (3), Toshifumi Yokota (2,4)

1- Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2-Department of Medical Genetics, University of Alberta, Edmonton, Alberta, Canada 3-Department of Biomedical Sciences, College of Veterinary Medicine, Oregon State University, Corvallis, OR 4-The Friends of Garrett Cumming Research & Muscular Dystrophy Canada HM Toupin Neurological Science Endowed Research Chair, Edmonton, Alberta, Canada

Objective: Spinal muscular atrophy (SMA) is an autosomal recessive disorder caused by homozygous deletion of the survival motor neuron 1 (SMN 1) gene. SMA majorly affects infants and children and is characterized by muscle weakness resulting from the degeneration of α-motor neurons. Humans possess SMN2 gene, a paralog of SMN1 that is unable to produce SMN protein efficiently due to the loss of exon 7 caused by a single nucleotide transition in exon 7. Antisense therapy for SMA uses a ""knock-up"" approach. Splice-switching oligonucleotides (SSOs) that bind to SMN2 intron 7 splicing silencer number 1 (ISS-N1) induce exon 7 inclusion and restore the production of proper (FL) full-length SMN2 mRNA. Nusinersen (Spinraza), the first drug for the treatment of SMA, is an 18-mer SSO with phosphorothioate backbone and 2'-O-methoxyethyl modification (2'MOE) targeting ISS-N1 that induces exon inclusion and the production of SMN protein. Although there is much hope for nusinersen, the cost of the treatment, the requirement for repeated invasive intrathecal injections and the associated toxicity remain significant challenges. To overcome these challenges, we developed a next-generation antisense oligonucleotide (AO) and examined effects in a severe mouse model. Methodology To improve the in vivo efficacy, we developed a cell-penetrating peptide called DG9-a novel moiety modified from a human T-cell peptide. We employed DG9 in chemical synthesis to make a conjugate with an AO called phosphorodiamidate morpholino oligomer (PMO/morpholino) targeting SMN2. DG9 can enhance the cellular uptake of AOs through a peripheral injection, eliminating the need for intrathecal injections. We injected DG9-PMO into Taiwanese SMA mice subcutaneously on postnatal day 0 to determine the in vivo efficacy. Results DG9-PMO induced efficient FL-SMN2 expression following a single peripheral administration. RT-qPCR data revealed that DG9-PMO-treated SMA mice exhibited higher expression of the FL-SMN2 gene compared to NT (non-treated) control and unmodified PMO-treated mice. Western blots revealed increased levels of SMN protein in DG9-PMO treated mice. Additionally, these mice showed significantly increased body weight and survival over time when compared to NT and unmodified PMO-treated mice. DG9-PMO treated mice also exhibited improved muscle pathology. Conclusions DG9-PMO is a promising therapeutic option to treat SMA, which can overcome the necessity for invasive injections with a single peripheral administration. We are currently evaluating the long-term effectiveness and toxicity of treatments.


The Design and Fabrication of Flexible, Intraspinal Multisite Microelectrodes

David A. Roszko* (1,3) & Vivian K. Mushahwar (1,2,3)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Division of Physical Medicine and Rehabilitation, Department of Medicine, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada 3 - Sensory Motor Adaptive Rehabilitation Technology (SMART) Network, University of Alberta, Edmonton, Alberta, Canada

Introduction: Spinal cord injury (SCI) is a debilitating condition which can result in paralysis. Intraspinal microstimulation (ISMS) is a neural prosthetic technique which aims to restore the ability to stand and walk after severe SCI by applying electrical stimulation within the lumbar spinal cord. Previous ISMS implants were either flexible or had multiple stimulation sites – but not both. Both of these features are desirable: flexible implants are more biocompatible, but multisite implants are more efficacious. Here, we show the preliminary design and fabrication of a new polymer-based microelectrode for the spinal cord that is both flexible and has multiple stimulation sites. Methods: A 4-channel, polymer-based microelectrode was designed in AutoCAD (Autodesk, CA, USA). Based on this design, a spin coater (Laurell Technologies Corporation, PA, USA) and nanosecond laser (Coherent, CA, USA) were used to create prototype microelectrodes. The soft polymer, polydimethylsiloxane (PDMS), was used to construct early prototypes, and aluminum foil was used as the conductor. PDMS was deposited onto a glass substrate, and an aluminum foil layer was added to the PDMS. The aluminum foil layer was machined into a multisite microelectrode using the nanosecond laser. Another layer of PDMS encapsulated the aluminum foil, and the electrode active sites were exposed using the laser. The completed microelectrode structures were then removed from the glass substrate. The minimum feature size was measured to determine how small laser prototypes could be fabricated. Electrical short circuits were assessed to ensure proper electrical functionality. Results: The microelectrode was refined over 7 design iterations. Fine-tuned laser parameters of 47.27 J/cm2 and 2.36 J/cm2 were capable of selective metal and polymer layer ablation. Minimum feature sizes of 7 μm were achieved. Microelectrode short circuits were eliminated by increasing conductor separation distances from 7 μm to 10 μm. Conclusions: Flexible, multisite microelectrode prototypes were fabricated using laser micromachining. Laser micromachining was a rapid and cost effective way to create these devices. Small feature sizes were achievable and initial electrical tests indicate that the microelectrodes function properly. Preliminary characterization with aluminum metal derisks future tests with more expensive and biocompatible conductors such as platinum-iridium (Pt-Ir) alloys. Future microelectrode prototypes will test Pt-Ir micromachining and implant functionality. These implants may be translated to humans in the future to restore mobility in those with severe SCI. This work was funded by: NSERC-CGSM, CIHR, CFI and the University Hospital Foundation (Nancy & Jim Hutton and Leo Bock).


The dichotomous role of inflammation on recovery following spinal cord injury in rats

Emma Schmidt*(1), Abel Torres-Espin(2), Pamela Raposo(2), Romana Vavrek(2), Karim Fouad(2)

1 - Neuroscience and Mental Health Institute, University of Alberta; Edmonton, Canada 2 - Department of Physical Therapy, Faculty of Rehabilitation Medicine, University of Alberta; Edmonton, Canada The inflammatory response following spinal cord injury (SCI) is associated with increased tissue damage and impaired functional recovery. However, in certain environments inflammation can promote plasticity and the secretion of growth-promoting substances. Previous work in our laboratory has shown that inducing inflammation with a systemic injection of lipopolysaccharide (LPS) in the chronic stages of SCI reopens the window of opportunity for plasticity (and thus rehabilitative training) within the spinal cord and enhances functional recovery. In the present study we aimed to determine what effect a sub-acute (10 days post-SCI) injection of LPS (when levels of inflammation are still relatively high) has on rehabilitative training. To test this, rats were trained in a single pellet reaching task that targets their impaired forearm. 10 days following a unilateral cervical SCI, rats received a single intraperitoneal injection of LPS or saline. LPS treatment induced a transient sickness response that lasted up to 3 days. Rats then underwent 6 weeks of high-intensity rehabilitative training in the single pellet reaching task. LPS treated rats showed improved functional recovery by 4 weeks of rehabilitative training. Since inflammation is associated with increased anxiety-like behaviour, we tested the rats in the elevated plus maze 4 weeks following LPS or saline injection. LPS treated rats showed increased anxiety-like behaviour, indicating that a single injection of LPS induces chronic changes in anxiety-like behaviour. At the end of training, the contralesional motor cortex was traced to label the corticospinal tract (CST). Histological analysis did not reveal group differences in the lesion size nor did we find differences in the sprouting of the CST. Therefore, improved functional recovery may be due to other mechanisms such as increased excitability of uninjured CST axons or compensation by other descending spinal tracts. In summary, we show that eliciting inflammation, whether in the chronic or sub-acute stages of SCI, has a beneficial effect on the efficacy of rehabilitative training. However, sub-acute LPS treatment has a dichotomous effect such that it induces anxiety-like behaviour in parallel to improved training success.


The Organization of Projection Patterns of Claustro-cortical Neurons in the Mouse

Brian Marriott (1), Alison Do (2), Jesse Jackson (1,2)

1 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2 - Department of Physiology, University of Alberta, Edmonton, Alberta, Canada

While most of the claustrum’s (CLA) vast inputs and outputs have been identified, the organization of mouse

claustro-cortical connections remain poorly understood; particularly to what extent individual CLA cells may

innervate collateral targets along their projections. To quantify both the topographical organization of CLA

projection neurons and determine how they may collateralize in mice, we simultaneously injected four spectrally

separated retrograde tracers (Fast Blue, retro Adeno Associated Virus 2 encoding GFP or TdTomato, and Alexa

Fluor 647-conjugated Cholera Toxin B) across a selection of 14 different brain regions. Rather than using

parvalbumin (PV) immunohistochemistry to mark the canonical CLA domain and orient each injection, all brains

were injected in the Retrosplenial Cortex (RSC). The retrograde staining from the RSC consistently labels a tight

core of CLA cells which overlaps with the area labelled by PV staining. The mouse CLA exhibits a large dorsal-

ventral gradient between regions, with different brain regions labelling distinct patterns of CLA cells. There are

clear topological distinctions between a dorsal motor-projecting group, a core frontal-midline-projecting group,

and a ventral temporal-lobe-projecting group of CLA cells. Injections made in the frontal midline had high rates

of colocalization (up to 50% of CLA cells labelled), while injections made in a group of sensory or motor regions

had lower rates of colocalization. Most regions exhibited weak contralateral connections labelling a subset of

CLA cells with some colocalization between regions, but some regions did not have any contralateral labelling.

Collectively, this data demonstrates a heterogeneous population of projection cells in the CLA, topographically

organized according to projection target. In addition, this quadruple-retrograde methodology allows insight into

how individual CLA projection neurons may modulate several distinct regions simultaneously.


The role of striatal neurons in regulating subventricular zone neural stem cell function

Nicole Dittmann* (1,2) & Anastassia Voronova (1,2)

1 - Department of Medical Genetics, University of Alberta, Edmonton, Alberta, Canada 2 - Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada Mammalian brain contains two neural stem cell niches: the subventricular (SVZ) and subgranular (SGZ) zones. Neural stem cells (NSCs) located in the SVZ niche produce neurons, astrocytes and oligodendrocytes throughout life and can participate in regenerative processes, such as remyelination. We previously showed oligodendrocyte genesis from embryonic NSCs in the developing cortex is regulated by factors secreted by inhibitory neurons (Voronova et al. 2017 Neuron). I hypothesize postnatal SVZ NSCs are regulated by striatal GABAergic neurons, which are located in close proximity to the SVZ niche. To address this question, I developed a co-culture system of postnatal SVZ NSCs with striatal neurons. Striatal cultures showed enrichment in neurons that expressed GABA and Gad67. Postnatal SVZ NSCs were cultured as neurospheres, where the only cells that propagate are NSCs, and other cells such as microglia, neurons, oligodendrocytes and astrocytes do not survive. Neurosphere cells were then dissociated and plated as a monolayer to allow co-culture with striatal neurons or as secondary neurospheres, which were cultured in media conditioned by striatal neurons. My initial results indicate SVZ NSCs proliferation is increased in co-culture with striatal neurons. Moreover, there was an increase in the number of secondary neurospheres in the presence of media conditioned by striatal GABAergic neurons, which is indicative of increased proliferation. My results suggest that striatal GABAergic neurons may have a role in regulating NSCs in the SVZ niche of the brain. As GABA is known to inhibit NSC proliferation in the developing and adult brain (LoTurco et al., Neuron, 1995; Fernando et al., PNAS, 2011), we believe that pro-proliferative effect is due to a different repertoire of secreted factors. Future work intends to expand this analysis to neuronal, astroglial or oligodendroglial differentiation of SVZ NSCs and to discover the signalling molecules responsible for the observed changes.


The thalamic nucleus reuniens is an essential mediator of slow oscillatory synchrony between prefrontal cortex and hippocampus

Brandon E. Hauer* (1), Silvia Pagliardini (1,2), Clayton T. Dickson (1,2,3)

1: Neuroscience and Mental Health Institute, University of Alberta, Edmonton, Alberta, Canada 2: Department of Physiology, University of Alberta, Edmonton, Alberta, Canada 3: Department of Psychology, University of Alberta, Edmonton, Alberta, Canada

Synchronized activity across the brain is a hallmark of sleep, a phenomenon which has physiological relevance for a host of functions. In particular, coordinated activity between medial prefrontal cortex (mPFC) and hippocampus (HPC) has been suggested to be critical for some mnemonic processes during both wakefulness and sleep. One of the most ubiquitous and physiologically relevant activity patterns occurs during slow-wave states, a ~1 Hz rhythm known as the slow oscillation (SO). However, it is currently unclear how the SO is coordinated between mPFC and HPC. Here, we examined the nucleus reuniens (RE), a midline thalamic body with direct, reciprocal projections to both mPFC and HPC, for its possible role in synchronizing the SO between these disparate sites. We used a rat model under urethane anesthesia because of how closely and dynamically it models natural sleep, in terms of alternations of forebrain state and corresponding physiological measures. We first characterized RE neuronal activity using single- and multi-unit recordings, determining that RE units rhythmically fire during, and are phase synchronized with, the mPFC SO. Optogenetic excitation of the RE reliably produced an evoked response in the HPC. Similarly, exciting the mPFC produced a similar evoked response in the HPC, with longer latency. Chemogenetically inhibiting the RE blocked this mPFC-mediated evoked potential in the HPC, suggesting that the RE is a critical and functional excitatory intermediary between mPFC and HPC. Importantly, during RE inactivation, we observed a robust decrement in mPFC-HPC coherence at SO frequencies. Critically, this diminished synchrony was not a consequence of decreased SO power, which was unchanged pre- and post-RE inhibition. Our results demonstrate a decisive role for the RE in at least partly mediating mPFC-HPC SO coordination, a phenomenon which has marked implications for sleep-dependent memory consolidation. Funding provided by NSERC grant #2016-06576 awarded to C. Dickson; NSERC grant #435843 awarded to S. Pagliardini; NSERC PGS-D3 awarded to B. Hauer

Thank you for attending NMHI Research Day 2020

This event was brought to you by the Neuroscience Graduate Students’ Association

(NGSA) and the Neuroscience & Mental Health Institute (NMHI).

NGSA Executive Committee Members:

President: Rebecca Long

Vice President: Caylin Chadwick

VP Treasurer: Timo Friedman

VP Internal Communications: Brandon Hauer VP External Communications: Lion Budrass

VP Social: Rachel Ward-Flanagan

VP Outreach: Aislinn Maguire

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Keynote Speaker: Dr. Giulio Tononi · 21st Annual NMHI Research Day March 5, 2020 Bernard Snell...

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