FENS 2022 - Abstracts of the speakers

Yehezkel Ben-Ari's talk abstract

The Neuroarcheology concept: predicting and treating brain disorders

The Neuroarcheology concept (Ben-Ari, 2008) posits that an in-utero insult leads to misconnected /misplaced neurons that remain immature and are the direct cause of deleterious sequels. Therefore, to treat disorders born in the womb, we must determine these deviations & use specific antagonists to block immature activity & attenuate the deleterious sequel. Relying on our earlier discovery of the evolutionary preserved developmental GABA polarity shift, we have determined these deviations in animal models and clinical trials to test their relevance. With my colleagues, I shall summarize the work done since my departure from academia & INMED to start-ups & privately funded research to illustrate how UpToDate basic science combined with conceptual innovation can increase cognitive research and at the same time lead to advances in treatments of medically orphan disorders.

Maternity and brain disorders: since ASD is born in utero, we succeeded with Machine Learning analysis of maternity data to identify at birth almost 100% of babies who will not have an ASD diagnosis later and almost half who will (Caly et al). If validated, Genesis ML will enable an early identification of babies at risk who can receive psycho-educative treatments that are known to be more efficient in early ages. Interestingly, fetal head circumference is bigger in a subpopulation of human fetuses already during the 2nd trimester suggesting that the reported bigger brains of toddlers and adolescents with ASD in fact starts in utero.

Bumetanide-a universal treatment ? : the GABA developmental shift (Ben-Ari et al 1989) is abolished in a large list of developmental, neurodegenerative disorders and traumatic insults. We validated this in autism, Fragile X, Rett syndrome and Maternal Immune Activation. We then used Bumetanide – the most specific antagonist of the chloride importer- in double blind clinical trials to treat Autism, Parkinson Disease and pilot cases of schizophrenia and Fragile X (Lemonnier et al 2017). In spite of many positive phases 2 performed in China, Holland and France, our phase 3 unfortunately failed. I shall briefly discuss why I think that this is a failure of the phase 3 not of Bumetanide.

A 3D developmental atlas: to better understand the alterations occurring in utero, using the clearing technique and light-sheet fluorescence microscopy, we have constructed a quantitative developmental 3D atlas of all Choline Acetyltransferase and Tyrosine Hydroxylase positive neurons of the rodent brains from E12 to P8. Combined with automated python scripts, this atlas is now used to investigate early changes in animal models of brain disorders.

Collectively, these observations stress the need to study brain development in order to comprehend and treat brain disorders.

References

1. Ben-Ari, Y. Neuro-archaeology: pre-symptomatic architecture and signature of neurological disorders. Trends in Neurosciences (2008) doi:10.1016/j.tins.2008.09.002.
2. Ben-Ari et al. Giant Synaptic Potentials in immature CA3 hipocampal neurons J Physiol 1989 416: 303-325
3. Lemonnier E, et al. Effects of bumetanide on neurobehavioral function in children and adolescents with autism spectrum disorders." Translational psychiatry 7.3 (2017): e10563.
4. Caly et al. Machine learning analysis of pregnancy data enables early identification of a subpopulation of newborns with ASD. 2021, Sci Rep Mar 25;11(1):6877.
5. Cloarec et al. Pyramidal growth and increased hippocampal volume during labor and birth in Autism. Sci AD. 2019, 23;5(1):eaav0394. doi: 10.1126/sciadv.aav0394.

Y. Ben-Ari
CEO Neurochlore, B&A Biomedical and B&A Therapeutics

Frédérique Bonnet-Brilhault's talk abstract

Autism: an early developmental disorder

Autism and Autism Spectrum Disorder (ASD) cover a large variety of clinical profiles which share two main dimensions : social and communication impairment and repetitive behaviors or restricted interests, which are present during childhood. There is now no doubt that genetic factors are a major component in the etiology of autism but precise physiopathological pathways are still being investigated. Furthermore, developmental trajectories combined with compensatory mechanisms will lead to various clinical and neurophysiological profiles from childhood to adulthood, which together constitute this Autism Spectrum Disorder. To better understand the pathophysiology of autism, comprehension of key neurophysiological mechanisms and brain circuits underlying the different bioclinical profiles is thus crucial. Adopting a translational approach which includes both thorough clinical description and a large variety of neurophysiological investigations our group has identified in children and adults with autism that
I) disturbances of sensory-perceptive information processing, from information capture to cortical treatment, stemming from both social and non-social environments, are fundamental cues in the pathophysiology of ASD,
II) both cortical, sub-cortical and Autonomic Nervous System (ANS) networks are involved,
III) individual profile can be identified and related to a combination of clinical parameters,
IV) pathophysiological cascade starts in-utero. Altogether these results emphasized the complexity of the key stages of the neurodevelopmental cascade leading to the specific combination of symptoms characterizing Autism Spectrum Disorder.

Frédérique Bonnet-Brilhault
Centre Hospitalier Regional et Universitaire de Tours, Tours, France

Charles Bourques's talk abstract

Clock time regulation by excitatory GABA

Under most conditions in the adult brain, neurons maintain a low level of intracellular chloride to allow efficient synaptic inhibition mediated by the ionotropic receptor for gamma amino butyric acid (GABA). However previous work has shown that GABA can be depolarizing during early post natal life and under various pathological conditions. Although rare, and often controversial, reports have suggested that GABA can also be depolarizing under physiological conditions. Neurons in the suprachiasmatic nucleus (SCN, the brain’s master circadian clock) display a 24 hour cycle in the their rate of action potential discharge whereby firing rates are high during the light phase and lower during the dark phase. Although it is generally agreed that this cycle of activity is a key mediator of the clock’s circadian output, surprisingly little is known about how non-photic signals can alter this activity and the timing (phase) of the clock. Using AAV-mediated targeting and opto- and chemogenetic approaches in mice we have shown that the SCN receives an excitatory GABAergic input from a subset of sodium sensing neurons in the OVLT. Activation of these neurons by a systemic salt load delivered at ZT19 stimulated the electrical activity of vasopressinergic SCN neurons which are normally silent at this time. Remarkably, this effect induced an acute reduction in non-shivering thermogenesis and body temperature, which is an adaptive response to the salt load. These results show that some neurons can maintain high intracellular chloride to enable GABA-mediated excitation under physiological conditions. This work is supported by the Canadian Institutes of Health Research.

Charles W. Bourque.
Centre for Research in Neuroscience, Research Institute of the McGill University Health Centre. Montreal General Hospital, 1650 Cedar Avenue, Montreal QC, Canada H3G1A4.

Hilgo Bruining's talk abstract

Precision medicine approach to bumetanide implementation in neurodevelopmental disorders

Several animal models of ASD have shown an abolished maturational shift in chloride downregulation resulting in neuronal hyperexcitability that could be corrected by bumetanide. These results were followed by several phase 2 clinical trials showing promising results in ASD. Due to specific actions in the heterogeneous ASD population, efficacy of bumetanide may be more pronounced in subpopulations and act on specific clinical endpoints. Findings and challenges from three innovative bumetanide trials in ASD and associated conditions will be presented. Collectively, these studies focused on the development of diagnostic companions and stratification strategies for bumetanide application, including a proof-of-concept EEG-assisted prediction model.

Hilgo Bruining
Child Psychiatrist and Full Professor in neurobiological developmental disorders at the Amsterdam UMC and Levvel

György Buzsáki's talk abstract

Neuroarcheology: uncovering the embryonic past of adult neurons

The incorporation of novel information into the hippocampal network is likely to be constrained by its innate architecture and internally generated activity patterns. However, the origin, organization, and consequences of such patterns remain poorly understood. Here, we show that hippocampal network dynamics are affected by sequential neurogenesis. We birthdated CA1 pyramidal neurons with in-utero electroporation over 4 embryonic days encompassing the peak of hippocampal neurogenesis, and compared their functional features in freely moving, adult mice. Neurons of the same birthdate displayed distinct connectivity, coactivity across brain states, and assembly dynamics. Same birthdate neurons exhibited overlapping spatial representations, which were maintained across different environments. Overall, the wiring and functional features of CA1 pyramidal neurons reflected a combination of birthdate and the rate of neurogenesis. These observations demonstrate that sequential neurogenesis during embryonic development shapes the preconfigured forms of adult network dynamics.

György Buzsáki
New York University School of Medicine

Enrico Cherubini's talk abstract

Alterations of GABAergic signaling at immature mossy fibers-CA3 synapses in an animal model of Autism

Alterations of synaptic plasticity processes may represent a convergent mechanism underlying several neuro-developmental disorders including Autism Spectrum Disorders. Here, we tested this hypothesis on mice carrying the human R451C mutation of the Nlgn3 gene (NLG3R451C KI mice), found in some families with autistic children. NLG3R451C KI mice were subjected to spike time dependent (STD) potentiation (LTP), at immature MF-CA3 connections, that at birth exhibit a GABAergic phenotype. Respect to littermate controls, NLG3R451C KI mice failed to express STD-LTP, an effect that persisted in adulthood when these synapses became glutamatergic. Similar results were obtained in NLG3 KO mice, suggesting a loss of function. In addition, while in controls STD-LTP was associated with an increased phosphorylation of TrkB receptors at potentiated synapses, this did not occur in NLG3R451C KI and NLG3 KO mice. Furthermore, in transgenic animals, immature MF-CA3 synapses exhibited hyperperpolarizing responses to GABA, indicating that an early GABA switch from the depolarizing to the hyperpolarizing direction, possibly consequent to a dysfunction of BDNF/TrkB signaling pathway, leads to persistent structural alterations of neuronal circuits.

Enrico Cherubini
European Brain Research Institute (EBRI), Roma, Italy

Eric Courchesne's talk abstract

The ASD living biology: from cell proliferation to clinical phenotype

ASD pathobiology begins prenatally and social affective and communication symptoms appear in the first years of life, giving rise to pervasive, lifelong challenges to patients and their parents, therapists, and educators, yet to differing degrees. These signature symptoms are much more than diagnostic markers. They signal dysfunction in human-specific, human-universal, likely genetic, experience-expectant neural and molecular process that normally drive social and affective communication development and learning in typical babies. Therefore, the central goal of our Center is to identify prenatal and postnatal molecular, cellular, neural, and behavioral subtypes of the ASD core social affect symptoms to enable very early-age detection, diagnosis, outcome prediction and social subtype molecular targeting. A corollary of this single overarching goal is that we aim to do so for the widest possible range and number of ASD toddlers, rather than for a rare genetic subtype. To accomplish this goal, we study many modalities and levels, from cell models and transcriptomics to social fMRI imaging, social eye tracking and early age social interactions. Thus, transcriptomics and cell models, such as cortical organoids, are two among many valuable sources of indirect, inferential and computational information that can illuminate the pathobiology underlying social subtypes of ASD. To accomplish this goal, large samples must be studied in order to accurately parse significantly different transcriptomic, neural and clinical social subtypes. Moreover, since different subtypes have different age-related changes in clinical and neural measures and different age-appropriate measurements, mixing wide ages, different measurement procedures, and subtypes in small sample studies risk reporting non-replicable cell/molecular-clinical correlations. So, patients in large samples must have comparable ascertainment, ages and clinical and neural phenotyping to enable linking biological, neural, behavioral and clinical data. Large sample studies of leukocyte gene expression in living ASD 1- and 2-year-olds, do reveal a gene network correlated with social symptom severity, and the activity of that exact DE-ASD Network is significantly greater in ASD neural progenitor cells and neurons derived from ASD toddlers. Leukocyte expression signatures are also diagnostically highly accurate and reveal ASD social subtypes and underlying molecular pathobiology. They are also correlated with ASD dysfunction in response to social speech, cortical patterning and treatment response. Thus, large multiscale study designs can link ASD cell models to the most important and complex core ASD dysfunction: social affect dysfunction.

Eric Courchesne
University of California, San Diego School of Medicine and Director of the UCSD Autism Center located in La Jolla, California

Eric Delpire's talk abstract

The Na-K-2Cl cotransporter-1 (NKCC1): a disease-causing transporter in humans

From the cloning of NKCC1 in 1994 it took more than 20 years for SLC12A2 to be recognized as a disease-causing gene. Following complete exome sequencing, a patient with multi-system dysfunction and a truncation mutation in NKCC1 was first described in 2016. The mutation causes dominant negative effects in Cl- secreting epithelia, as it leads to mistargeting of the cotransporter to the apical membrane of the epithelial cells. Children with complete absence of NKCC1 were also identified. Like the NKCC1 knockout mouse, they demonstrated sensorineural deafness, mental retardation, respiratory and intestinal deficits. We also identified individuals with single allele variants in SLC12A2 and sensorineural deafness, intestinal deficits, and autism. Interestingly, the mutations related to deafness involve a small alternatively spliced exon with unknown function. All these individuals indicate haploinsufficiency of the SLC12A2 gene. In in vitro work in intestinal and kidney epithelial cells, we showed that elimination of NKCC1 expression leads to changes in tight junctions. Expression of claudin-2, a leak junctional protein, is significantly upregulated in NKCC1 knockout cells. This is accompanied by a drop in the transepithelial transmembrane resistance and appearance of a leak of small molecular size molecules. The change is due to NKCC1 function as application of bumetanide or exposure to low Cl- media result in increased claudin-2 expression. Our data indicate that absence of NKCC1 leads to significant changes in epithelial barrier function and raise the possibility that similar changes occur at the blood brain barrier. This work is funded by NIH grants GM118944 and DK093501.

Eric Delpire, Ph.D.
Department of Anesthesiology, Vanderbilt University Medical School, Nashville, TN

Alexandra DURR & Sandrine HUMBERT's talk abstract

Developmental alterations in Huntington disease

Huntington Disease belongs to the family of polyglutamine diseases, a group of neurological disorders characterized by abnormal polyglutamine tract expansion in different proteins specific to each disease. As with all these ‘proteopathies,’ symptoms in Huntington Disease typically do not appear until mid-life or later. Yet huntingtin (HTT), the protein mutated in Huntington Disease, and mutant HTT are expressed from the very beginning of life and HTT is essential for mouse development. We will discuss how the HTT protein regulates several steps of mouse corticogenesis which are impaired in Huntington Disease mice. We will also show that the biology of progenitors is modified in human Huntington Disease mutation carrier fetuses and consider the viewing of Huntington Disease as an adult disorder with a developmental component.

Sandrine Humbert
Inserm U1216, GIN Grenoble Institute Neurosciences
Alexandra Durr
Paris Brain Institute/Sorbonne Université

Arnold Kriegstein's talk abstract

Genomic Insights into Human Brain Development and Malformations Associated with Epilepsy

The human cerebral cortex is more than three times expanded compared to our closest non-human primate relatives. The cortex emerges from an initially pseudostratified neuroepithelium that gives rise to radial glia, the neural stem cells of the cortex. A number of subtypes of radial glia have been identified, and single cell RNA sequencing (scRNAseq) has contributed to a novel model of primate corticogenesis, highlighted human-specific features of cortical development, suggested a relationship between outer radial glia (oRG) cells and brain tumors, and provided a benchmark for in vitro organoid models of brain development and disease. Our single-cell transcriptomic and in-situ data suggest that early cortical areal patterning is strongly defined by the mutual exclusion of strong frontal or occipital gene expression signatures, supporting the existence of a cortical protomap at the extremities, but also support for the protocortex hypothesis to refine spatial identity between the poles. Using velocity analysis, we find that major signaling pathways including Notch, Wnt, and mTOR drive the specification and maintenance of neuroepithelial stem cells and radial glia. Comparison of these progenitor populations to cortical malformations suggests a potential role of oRG cells. mTOR mediated malformations of cortical development like Focal Cortical Dysplasia (FCD), Hemimegalencephaly (HME) and Tuberous Sclerosis (TSC) are the most common causes of medication-resistant pediatric epilepsy. We used single nucleus sequencing strategies in donated patient surgical specimens to characterize the transcriptomic identity and relationships of cell types present within the malformations. We identified key developmental signatures normally present in progenitor cells, including oRG cells, within populations of dysmorphic neurons, balloon and giant cells present in FCD, HME, and TSC lesions, highlighting shared molecular and developmental origins of these disease-associated cell types. We also identified transcriptomic changes in mitochondrial oxidative phosphorylation pathways in ictal pyramidal neurons and interneurons, suggesting distinct modulation of glycolysis and energy metabolism pathways in epileptic neurons. The common molecular changes to gene expression observed in hallmark abnormal cells reflect a shared underlying biology in patients with mTOR-linked cortical malformations.

Arnold Kriegstein
Department of Neurology, Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, California

Heiko Luhmann's talk abstract

Transient cortical activity match sensory driven developmental activity

During late prenatal and early postnatal stages, the developing neocortical network shows a rich repertoire of spontaneous (ongoing) and sensory evoked oscillatory activities. These activity patterns, consisting of spindle bursts (delta brush) and early gamma oscillations, play important roles in the maturation of thalamocortical connectivity and cortical networks, e.g. columnar architecture (Molnár et al 2020). In preterm human infants, cortical burst activity recorded with EEG represents a biomarker for the clinical diagnosis of early brain damage and for the prediction of further development (Luhmann et al 2022).
Furthermore, during distinct developmental periods spontaneous activity patterns control the rate of programmed neuronal cell death (apoptosis) in a region- and cell-type specific manner. Current data indicate that pre-/neonatal pathophysiological noxes, induced for example by hypoxia, inflammation or drug exposure, may alter the spontaneous activity patterns and thereby disturb the maturation of the cerebral cortex. Cortical disorganization, abnormal cell death and pathophysiological circuits may cause neurological and psychiatric disorders.

References

• Luhmann HJ, Kanold PO, Molnar Z, Vanhatalo S. 2022. Early brain activity: Translations between bedside and laboratory. Prog Neurobiol 213: 102268
• Molnár Z, Luhmann HJ, Kanold PO. 2020. Transient cortical circuits match spontaneous and sensory-driven activity during development. Science 370

Heiko J. Luhmann
Institute of Physiology, Univ. Medical Center Mainz, Germany

Marina Sirota's talk abstract

Leveraging Clinical and Molecular Data to Advance Drug Discovery for AD in the Era of Precision Medicine

Alzheimer’s disease (AD) is a multifactorial neurodegenerative disorder caused by interactions among multiple genetic and environmental factors. The genetic complexity and multifactorial nature of AD pose unique challenges for developing effective therapies and suggest the need for a precision medicine approach that takes into account individual variability. The approach of drug repositioning has a number of advantages over the development of new drugs and has been done successfully for many disease conditions. Here, we applied a computational drug-repurposing strategy to identify drugs to treat apolipoprotein E4 (APOE4)-related AD based on gene expression. We first established APOE genotype-specific transcriptomic signatures of AD by analyzing publicly available human brain databases. We then queried these signatures against the Connectivity Map database, which contains transcriptomic perturbations of more than 1,300 drugs, to identify those that best reverse APOE genotype-specific AD signatures. Bumetanide was identified as a top drug for APOE4-related AD. Treatment of APOE4-knock-in mice without or with amyloid β (Aβ) accumulation using bumetanide rescued electrophysiological, pathological or cognitive deficits. Single-nucleus RNA sequencing revealed transcriptomic reversal of AD signatures in specific cell types in these mice, a finding confirmed in APOE4 induced pluripotent stem cell (iPSC)-derived neurons. In humans, bumetanide exposure was associated with a significantly lower AD prevalence in individuals over the age of 65 years in two independent electronic health record databases, suggesting the effectiveness of bumetanide in preventing AD. We are further extending these approaches to identifying combination therapies for AD and also leveraging real world data to better understand disease heterogeneity. Combining clinical and molecular data is a powerful tool to advance drug discovery in the era of precision medicine.

Marina Sirota
University of California, San Francisco

Harald Stontheimer's talk abstract

Loss of GABAergic inhibition contributes to tumor-associated epilepsy

Seizures are often the presenting symptom for patients ultimately diagnosed with a primary brain tumor (glioma). One third of patients go on to develop tumor-associated epilepsy that is refractory to anti-epileptic medication. Understanding the biology of tumor-associated epilepsy will elucidate mechanisms underlying other acquired seizure syndromes. We have been able to reproduce the human disease by implanting patient-derived glioma tissue into immune-suppressed mice. If implanted tissue was derived from a patient suffering from seizures, the animals also developed seizures; conversely, if tissue was derived from glioma patients who did not present with seizures, mouse implants typically lacked seizures. Comparing these mouse models we found that seizure-eliciting gliomas show assiduous Glutamate (Glu) release causing peritumoral inhibitory neurons to die by Glu-excitotoxicity. We find up to 40% loss of PV positive interneurons are permanently lost. Furthermore, the remaining 60% of surviving PV neurons showed depolarizing, excitatory GABA responses owing to altered expression of KCC2 and NKKC1 respectively, reversing the transmembrane Cl- gradient. Lastly, the release of proteolytic enzymes, namely MMP2, 3, & 9, destroys extracellular matrix structures encasing PV neurons in the form of perineuronal nets (PNN). Loss of the negatively charged PNNs causes an increase in membrane capacitance in PV neurons slowing their discharge frequency resulting in reduced GABA release. Taken together, changes in the tumor-associated brain suffer from a number of insults that each reduce GABAergic inhibition. Some of these are recapitulated in other forms of epilepsy and suggest novel disease etiology and new therapeutic targets.

Harald Sontheimer, Ph.D.
University of Virginia, Charlottesville VA

Nick Spitzer's talk abstract

Neurotransmitter switching contributes to two environmental models of neurodevelopmental disorders

Neurodevelopmental disorders including autism spectrum disorder are characterized by varying degrees of enhanced stereotypic behaviors, deficits in social interaction, and defects in communication. Early environmental factors play a key role and cortical neurotransmitter imbalance is a convergent phenotype seen across different types of disorders. Neurotransmitter switching involves loss of one transmitter and gain of another. It often changes the sign of the synapse from excitatory to inhibitory or vice versa and causes changes in behavior (Spitzer, 2017). We are testing the hypothesis that transmitter switching contributes to these disorders in mouse models. We treat pregnant females with either Poly Inosine:Cytosine (Poly I:C) or valproic acid (VPA) at embryonic day E12.5.
We find a transient decrease of 5000 GAD67+ neurons and an equal increase of VGLUT1+ neurons in the medial prefrontal cortex (mPFC) at postnatal day P10 in experimental mice compared to saline-treated controls, that has disappeared by P90. These changes do not result from altered number of precursors (Dlx2+ and Tbr1+) or apoptosis and occur specifically in PV+ and CCK+ neurons. P90 animals demonstrate enhanced stereotypy and deficits in social interaction compared to controls. We then generated PV-Cre/CCK-Cre mice to label neurons with Cre-dependent Herpes Simplex Virus (HSV)-GFP and search for VGLUT1 expression in GFP+ neurons. Stereotaxic injection of P9 PV-Cre/CCK-Cre Poly I:C-treated embryos with Cre-dependent HSV-GFP revealed VGLUT1+GFP+ neurons in the neonatal mPFC at P13 demonstrating expression of VGLUT1 in a subset of PV-Cre/CCK-Cre neurons in response to maternal Poly I:C treatment in both male and female mice, providing strong evidence for transmitter switching.
To test the role of the transmitter switch in emergence of behavioral disorders, we restored GAD67 specifically in PV and CCK neurons in the mPFC. Previous work showed that overriding one of the two transmitter changes in a switch is sufficient to prevent the change in behavior (Li & Spitzer, 2020). We stereotactically injected Cre-dependent HSV-GAD1-GFP in Poly I:C- treated male and female PV-Cre/CCK-Cre mice at P10 and scored them at P13. Overexpressing GAD67 in PV and CCK cells restored GAD67+GFP+ neurons in experimental mice and prevented gain of VGLUT1 in GFP+ neurons unlike Poly I:C-treated controls injected with HSV-GFP. Restoring GAD67 in the mPFC of PV-Cre/CCK-Cre mice rescued altered behavior in experimental male mice. Our results add to growing evidence that alterations in signaling in the nervous system during the early stages of its construction can be detrimental to the function of the mature brain.

Swetha K. Godavarthi & Nicholas C. Spitzer
Neurobiology Department and Kavli Institute. for Brain & Mind University of California San Diego, USA

Melanie Woodin's talk abstract

Neuronal potassium-chloride cotransporter 2 (KCC2) function is altered in the indirect pathway of the basal ganglia in Huntington’s Disease

Huntington’s disease (HD) is an inherited neurodegenerative disorder characterized by progressive motor dysfunction. Motor impairments result from extensive neurodegeneration of the basal ganglia (BG), a highly interconnected assembly of subcortical nuclei best-known for their role in movement-related functions. The primary site of degeneration is the striatum, the input structure of the BG which sends inhibitory projections to the surrounding nuclei of the BG. The striatum consists primarily of D1 and D2 dopamine receptor-expressing neurons which make up the direct pathway (dSPNs) and indirect pathway (iSPNs), respectively. dPNs and iSPNs are known to exert opposing effects on motor output, whereby activation of the direct pathway promotes movement, while the indirect pathway inhibits unwanted movement. In HD, there is a preferential loss of iSPNs, biasing BG output towards the direct pathway, though the cause of this enhanced susceptibility remains unclear. Accumulating evidence suggests that impaired synaptic inhibition within the BG circuitry may underlie this specific pattern of degeneration. Synaptic inhibition in the mature brain is largely mediated by the neurotransmitter gamma-aminobutyric acid (GABA). GABA exerts fast hyperpolarizing inhibition by binding to chloride-permeable GABAA receptors. GABAergic inhibition requires low intracellular chloride levels which is maintained by the potassium-chloride cotransporter, KCC2. However, when KCC2 function is reduced, intracellular chloride increases, subsequently weakening the GABAergic driving force and synaptic inhibition. Using a combination of electrophysiology, biochemical and behavioural assays, we determined that KCC2 function was reduced and GABAergic signaling was impaired in the indirect pathway of symptomatic R6/2 mice, a transgenic mouse model HD. In R6/2 mice, iSPNs demonstrated a depolarization in the reversal potential for GABA compared to dSPNs. In addition, the neuronal response to GABA was altered, from inhibitory to excitatory in the Globus Pallidus externa, the output structure of the indirect pathway. Interestingly, pharmacological reduction of [Cl-]i with the FDA-approved diuretic bumetanide effectively delayed the onset of motor impairments in R6/2 compared to controls. These findings provide evidence that the early impairments in GABAergic transmission in the indirect pathway may be a key mechanism contributing to circuitry and motor defects in HD. As therapeutic treatments for HD are limited, this work demonstrates the use of KCC2 as a novel potential therapeutic target in the treatment of HD.

Melanie Woodin
Dean, Faculty of Arts & Science at University of Toronto