Canada

Perineuronal nets are highly organized components of the extracellular matrix which inhibit cellular plasticity and support cellular function. Depletion of perineuronal nets in rodents via developmental models of disease or via direct pharmacological interventions (e.g. chondroitinase) can impact cognitive function. In our current study, we utilized injections of an immune-evasive dual vector system, where the chondroitinase gene is under a doxycycline regulatory control, to induce bilateral secretion of chondroitinase, with temporal control, in the medial prefrontal cortex and retrosplenial cortex. These two brain regions are known to be involved in the function of working memory. We subjected mice to 30 days of activated chondroitinase-expression to deplete their perineuronal nets, followed by 30 days without to allow for a window of recovery. Prior to treatment, after treatment, and after 30 days without doxycycline-induced chondroitinase expression, animals were tested in a cross-modal object recognition task and an oddity task, which have been shown to be sensitive to manipulations of the medial prefrontal and retrosplenial cortex. Preliminary data indicates that controlled depletion of perineuronal nets in the medial prefrontal and retrosplenial cortex impacts performance of working memory. While there is some recovery in their working memory performance after 30 days without continuing enzymatic depletion, they do not recover to baseline behavioural performance. This data suggests that perineuronal net degradation in the medial prefrontal and retrosplenial cortex impacts working memory performance, and that these changes are long-lasting.

CAN-ACN 2021

Cognitive impairments in mice after targeted perineuronal net depletion in the medial prefrontal or retrosplenial cortex.

perineuronal nets

extracellular matrix

cognition

Colleagues and Friends:

I am very happy to welcome you to the 14th annual Canadian Neuroscience Meeting, and to the first virtual edition of our meeting. I want to highlight the important work the CAN meeting chair, **Roger Thompson** and co-chair, **Alyson Fournier**, did to put together the great program we are offering this year. Moving to a virtual format required a complete re-design of the program, and both Roger and Alyson did a splendid job in providing us with an appealing and well adapted program for our first virtual event. I wish to thank them for this very important effort.

Highlights of this year’s scientific meeting include invited speakers **Michael Hausser, Franck Polleux, Gwyneth Card, Magdalena Götz**and Brain Prize winner **Huda Zoghbi**. Plenary and parallel symposia, proposed by our members complete our diverse [scientific program.](https://can-acn.org/meeting-2021/program-2021/)

The 2021 Advocacy session, chaired and organized by CAN advocacy committee chair **Karun Singh**, will feature a special lecture and question period with Member of Parliament and past Science Minister **Kirsty Duncan**. This will be followed by a short overview of CAN’s advocacy activities over the last year. We invite you to participate in this session for insight from Dr. Duncan on the workings of Parliament Hill, and to learn more about what CAN has been doing to promote neuroscience research in Canada.

We are also excited to host **Lisa Willis** in the Equity Diversity and Inclusion session, organized by EDI committee chair **Jibran Khokhar**.

A special highlight of our meeting will be the lectures by award-winning young neuroscientists. We are please to host the CAN 2020 Young Investigator Award winner **Mihaela Iordanova**, and CAN 2021 Young Investigator Award winners **Aaron Phillips** and **Tabrez Siddiqui**. Highlights will also include presentation by the top three winners of the CAN-CIHR-INMHA Brain Star Award winners, **Claire Gizowski, Nuria Daviu Abant** and **Lukas Tamayo-Orrego**.

Independently organized satellite symposia complement and complete this exciting program, check them out here: https://can-acn.org/meeting-2021/2021-satellite-symposia/

Some of the most engaging sessions are always the poster sessions featuring the work of our trainees, so make sure to log in to Gather Town during the poster sessions to chat with the presenters.
We hope you enjoy this program, and look forward to welcoming you, virtually.

Shernaz Bamji

President of the Canadian Association for Neuroscience

![](https://assets.underline.io/uploads/markdown_image/1/image/b9aea25343a87d5aac3b9367dc780974.jpg)



Please head to https://can-acn.org/meeting-2021/registration-2021/ to register. 

14th Canadian Neuroscience Meeting from August 23rd-25th

poster

The ability of organisms to sense and respond to temperature in their ambient environment as well as in their own bodies is critical for survival. Thermoregulation occurs as an essential homeostatic process Currently there exists a paucity of information on the involvement of the afferent mechanism of thermoregulation in ectotherms. Here we investigate thermoregulation and the involvement of a family of thermosensing proteins known as the transient receptor potential (TRP) channels in the ectotherm Xenopus laevis. Using Xenopus larvae as an in vivo model, and a melanophore cell line as an in vitro model, our data indicate that within skin melanophores pigments known as melanosomes aggregate in cool temperatures and disperse at warmer temperatures. Our in vivo model suggests that this aggregation of skin pigments occurs as a systemic response. We identified TRP family genes in the Xenopus genome via RT-PCR, including a candidate (TRPM8) from the TRPM subfamily, which has been characterized previously as a cold sensor. Using immunohistochemistry, we visualized TRPM8 expression in Xenopus melanophores and the skin, and found that TRPM8 is co-localized with melanophores using a melanophore-specific marker. To investigate the function of TRPM8, we treated cultured melanophores and Xenopus larvae with an agonist (WS12), which resulted in aggregation of pigment in the absence of cool temperature. Our results suggest that in Xenopus larvae temperature alterations regulate skin pigmentation, a process that depends on a member(s) of the TRPM channel family. (Support from the Natural Sciences and Engineering Research Council, Canada)

The coordination of thermoregulation by circadian and thermosensory mechanisms

Weakly electric fish actively sense their environment using a self-generated electric field. Objects with electrical properties differing from those of the surrounding water cause small perturbations that are detected by electroreceptors distributed over their skin surface. When sensing conditions are unfavourable, fish appear to increase several movements (tail-bending and back-and-forth swimming) that are thought to improve electrosensory acquisition. However, these movements are variable and not reliably identified. Recently, unbiased data-driven approaches have been used to extract fundamental poses, combinations of which can be used to describe any animal behaviour. Here, we apply these approaches as a first step towards testing the hypothesis that specific movements can enhance electrosensory information acquisition. We find that fish behaviour can be explained by combinations of only a few fundamental basis shapes. Then, along with a newly developed model for quantifying the electric field, we characterize the spatiotemporal properties of natural electrosensory inputs and identify how they are affected by body pose and movement.

The effect of self-motion on electrosensory inputs

To learn more on the factors that govern the development of neural circuits, both in structural and functional terms, we use whole brain two-photon imaging on larval zebrafish that express a pan-neuronal genetically-encoded calcium indicator GCaMP6s. Using a resonant scanner and piezo driven objective, we record neural activity (GCaMP6 fluorescence) from up to ~50% of the whole neuronal population (~100,000 neurons), while simultaneously conveying visual stimulation using a screen oriented towards the head-restrained larva in agarose. This experimental paradigm leverages the early-developing visual system of the zebrafish to evoke reproducible neuronal responses and behavioral outputs across individuals. Abrupt changes in illumination induce navigational tail movements, which are monitored using a high-speed camera to identify distinct behavioral modules and their neural correlates. By varying the temporal properties of visual stimuli, we also probe the neural mechanisms of habituation and anticipation. Using graph theory, functional networks are generated from spontaneous brain activity recordings, which are then paired with the zebrafish structural connectome (Kunst et al., Neuron, 2019) in order to gain fundamental insight on the interaction between structure and function in vertebrate brain networks. Our dual spontaneous/stimulus-evoked experimental framework will be used to compare fish across different developing conditions, namely germ-free fish, to observe the impact of gut microbiota on brain connectivity, sensorimotor integration and behavior.

Whole-brain optical imaging in zebrafish larvae to investigate neural circuit development and connectivity

The subventricular zone neurogenic niche receives neuronal inputs from many areas of the brain. Although these inputs are known to regulate neurogenesis through neurotransmitter release at synapse-like connections between neuronal axon terminals and neural stem and progenitor cells (NPCs), few studies have examined how neuronal modulators at these synapse-like connections affect SVZ NPC function. Our recent work identified monoacylglycerol lipase (Mgll), a hydrolase that breaks down endocannabinoid 2-AG, as an extrinsic factor to regulate SVZ NPC function in culture. 2-AG is a well-known retrograde modulator that controls neurotransmitter release. To date, it remains unknown how Mgll-modulated 2-AG signaling in the SVZ niche regulate SVZ NPC behavior in vivo. In this regard, we propose to examine the role of neuronal Mgll activity from distinct brain region in regulating SVZ NPC function. Using retrograde monosynaptic rabies virus tracing and anterograde tracing, we identified direct synapse-like connections between hypothalamic arcuate nucleus (ARC) neuron axon terminals and a subpopulation of NPCs residing in the ventral SVZ microdomain as previously reported. Intriguingly, specific removal of Mgll expression in ARC neurons using virally transfected Cre recombinase in Mgll-floxed mice resulted in a decrease of SVZ NPCs in this microdomain and a subsequent reduction of proliferating cells in this microdomain. These findings provide an understanding of how long-range neuronal Mgll activity regulates adult SVZ NPC function in a microdomain-specific fashion.

Long-range hypothalamic neuronal projections to the subventricular zone neurogenic niche regulates neural stem cell function through endocannabinoid signaling

Melanin-concentrating hormone (MCH) is produced in the lateral hypothalamus. It has important roles in homeostatic and motivated behaviour. As MCH projections and receptors are widely distributed, the brain regions underlying MCH functions are not well understood. We showed that MCH neurons send dense projections to the lateral septum (LS) and innervate LS neurons by glutamate release. Consistently, retrograde injections into the LS showed that MCH neurons that innervate the LS are largely distributed medial to the fornix. However, it is not known if MCH can regulate the activity of LS neurons. In order to characterize MCH action in the LS, we first mapped the distribution of MCH-immunoreactive fibers and MCH receptor mRNA (Mchr1) throughout the LS. We observed the highest expression of both MCH fibers and Mchr1 toward the lateral and ventral borders of the intermediate subregion of the LS, and we selected LS cells for whole-cell patch clamp recordings from these regions. Bath application of MCH directly hyperpolarized LS neurons and elicited a membrane current that reversed at the equilibrium potential for chloride ions. This MCH-mediated hyperpolarization and current were blocked by the GABAA receptor antagonist bicuculline and indicated that GABAA-dependent mechanisms underlie the inhibitory actions of MCH in the LS. By contrast, MCH does not alter the frequency of glutamatergic or GABAergic input to LS neurons. In aggregate, the postsynaptic actions of MCH in the LS suggests that in addition to glutamate, MCH neurons may also regulate the LS by MCH release.

Melanin-concentrating hormone mediated inhibition of lateral septum neurons

Melanin concentrating hormone (MCH) is an orexigenic neuropeptide located in the lateral hypothalamus. Cells expressing MCH are heterogenous and can be marked by their coexpression of cocaine- and amphetamine-regulated transcript (CART) or the neurokinin 3 receptor (NK3R). Transcriptomic studies show that one-third of MCH neurons coexpress both CART and NK3R. However, the distribution of this MCH subpopulation has not yet been mapped in the mouse brain. We determined the percentage of MCH neurons that coexpress CART and/or NK3R and mapped their distribution to Allen Reference Atlas mouse brain templates. We identified MCH neurons by native EGFP fluorescence (EGFP-f) in Mch-cre;L10-Egfp mice and then labeled CART and NK3R immunoreactivity. We found that 96% of EGFP-f cells were MCH-positive. Within the lateral hypothalamus, EGFP-f cells may be clustered medial or lateral to the fornix. Nearly half (49%) of the EGFP-f cells counted expressed EGFP-f only and were most commonly found lateral to the fornix. In contrast, 47% of EGFP-f neurons coexpressed CART, and these cells were more common medial to the fornix. Of the CART-positive EGFP-f cells, half of them also coexpressed NK3R, which appeared evenly throughout the lateral hypothalamus. Lastly, 4% of EGFP-f neurons coexpressed NK3R only. Our results indicate a robust heterogeneity within MCH neurons, and we will expand our analysis to determine if these MCH subpopulations also express distinct electrical fingerprints based on their neurochemical properties.

Neurochemical characterization of melanin-concentrating hormone neurons in the mouse lateral hypothalamus

Melanin-concentrating hormone (MCH) activates MCH receptor 1 (MCHR1) to promote positive energy balance. MCH or MCHR1 deletion, in part at GABAergic cells, leads to a hyperdopaminergic state and hyperactivity, as MCH can inhibit dopamine release. Here we assessed if MCH can directly regulate dopaminergic or GABAergic neurons in the ventral tegmental area (VTA). We labelled mouse Mchr1 mRNA by RNAscope in situ hybridization throughout the VTA and quantified its expression on putative dopaminergic or GABAergic cells by colocalization with tyrosine hydroxylase (TH) immunoreactivity or Slc32a1 (Vgat) mRNA hybridization, respectively. Half of all Mchr1 cells expressed TH, which comprised 25% of TH VTA cells, while only 13% of Mchr1 cells expressed Vgat, which represented 13% of Vgat VTA cells. We then mapped the expression of Mchr1 in TH and Vgat VTA neurons onto Allen Reference Atlas brain templates to assess their spatial distribution. Mchr1 expression was most prevalent in the posterior VTA (Bregma -3.28 mm to -3.68 mm) and concentrated within the medial aspect of the VTA. Vgat expression and its colocalization with Mchr1 increased in more posterior levels but TH and Mchr1 colocalization was relatively higher posteriorly. Interestingly, 36% of Mchr1 cells were neither TH nor Vgat positive, and future experiments will determine if they may be glutamatergic VTA cells. Importantly, we provide evidence for Mchr1 expression in the VTA and provide the basis for electrophysiological recordings to assess MCH-mediated action at TH and Vgat VTA cells.

Spatial distribution of melanin-concentrating hormone receptor 1 in the ventral tegmental area

Object category recognition and discrimination may rely on the use of generalized category representations formed through exposure to various category exemplars. Presumably, to refine category representations, extraneous object features must be inhibited. Parvalbumin-containing GABAergic interneurons (PVINs) have been implicated in visual stimulus learning, including the inhibition of irrelevant stimuli. To determine the role of GABAergic transmission in the refinement of object category representations, C57/BL6 mice were administered a GABAA receptor antagonist (bicuculline) prior to exposure to various category exemplars. When tested on a rodent object category recognition (OCR) task, mice administered bicuculline failed to perform a 1-h retention delay which requires pre-exposure to category objects. Then, to specifically examine the role of GABAergic PVINs in the perirhinal cortex (PRh), a region necessary for object recognition, we infused male PVcre mice intracranially with adeno-associated virus containing inhibitory designer receptors exclusively activated by designer drugs (DREADDs) to control the activity of PVINs in the PRh. The DREADD agonist 21 (11-(1-piperazinyl)-5H-dibenzo[b,e][1,4]diazepine; C21) was administered systemically prior to exposure to object category exemplars. Mice administered C21 prior to pre-exposure sessions were not able to perform the OCR task with the 1-h retention delay. Thus, GABAergic transmission, and specifically PVIN activity in the PRh, may play an essential role in the refinement of object category representations.

The role of GABAergic inhibition in object category recognition

Cancer is a multi-stage disease caused by sequential mutations; however, the molecular mechanism generating cancer-initiating mutations for many cancers is not well understood. Using the developing cerebellum as model system, here we delineate a molecular mechanism for tumor initiation in medulloblastoma (MB), the most frequent malignant pediatric brain tumor. Activation of the Sonic hedgehog (SHH) pathway is frequent in MB, with mutations in the tumor suppressor PTCH1 (a negative regulator of SHH signaling) being the most common initiating event for SHH-MB. However, how SHH as a developmental mitogen promotes early carcinogenesis in the cells of origin, granule cerebellar progenitors (GCPs), remains to be determined. 
Here we report that physiological exposure of GCPs to Shh causes a distinct form of DNA replication stress, altering DNA replication dynamics to increase both origin firing and fork velocity. Shh promotes DNA helicase loading and activation in GCPs, with increased levels of Cdc7-dependent replication origin firing. S-phase duration is reduced and hyper-recombination consequently occurs. Such elevated recombination causes copy-number neutral LOH, an event seen frequently at the PTCH1/ptch1 locus. Moreover, Cdc7 inhibition to attenuate origin firing in a MB mouse model is sufficient to reduce somatic recombination and preneoplastic tumor formation. We therefore establish that tissue-specific replication stress induced by Shh acts to promote LOH, which in tumor-prone Ptch1+/- GCPs can result in loss of this tumor suppressor, as an early cancer initiating event.


Sonic hedgehog causes replication stress promoting cancer initiation in medulloblastoma

In humans and rodents, the perception of control during stressful events has lasting behavioral consequences. But how the brain links prior stressful experience to subsequent behaviors was poorly understood. We showed that hypothalamic CRH neurons increase their activity prior to the initiation of innate escape behavior. This anticipatory increase is sensitive to stressful stimuli that have high or low levels of outcome control. Specifically, experimental stress with high outcome control increases CRH anticipatory activity, increasing future escape behavior. By contrast, stress with no outcome control prevents the emergence of this anticipatory activity and decreases subsequent escape behavior. These observations indicate that CRH neurons encode stress controllability and contribute to shifts between active and passive innate defensive strategies.

Cognitive impairments in mice after targeted perineuronal net depletion in the medial prefrontal or retrosplenial cortex.

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The coordination of thermoregulation by circadian and thermosensory mechanisms

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