BDNF is altered in a brain-region specific manner and rescues deficits in Spinocerebellar Ataxia Type 1.

Spinocerebellar ataxia type 1 (SCA1) is an adult-onset, dominantly inherited neurodegenerative disease caused by the expanded polyQ tract in the protein ATAXIN1 (ATXN1) and characterized by progressive motor and cognitive impairments. There are no disease-modifying treatments or cures for SCA1. Brain-derived neurotrophic factor (BDNF) plays important role in cerebellar physiology and has shown therapeutic potential for cerebellar pathology in the transgenic mouse model of SCA1, ATXN1[82Q] line that overexpress mutant ATXN1 under a cerebellar Purkinje-cell-specific promoter. Here we demonstrate decreased expression of brain derived neurotrophic factor (BDNF) in the cerebellum and medulla of patients with SCA1. Early stages of disease seem most amenable to therapy. Thus, we next quantified Bdnf expression in Atxn1154Q/2Q mice, a knock-in mouse model of SCA1, during the early symptomatic disease stage in four clinically relevant brain regions: cerebellum, medulla, hippocampus and motor cortex. We found that during the early stages of disease, Bdnf mRNA expression is reduced in the hippocampus and cerebellum, while it is increased in the cortex and brainstem. Importantly, we observed that pharmacological delivery of recombinant BDNF improved motor and cognitive performance, and mitigated pathology in the cerebellum and hippocampus of Atxn1154Q/2Q mice. Our findings demonstrate brain-region specific deficiency of BDNF in SCA1 and show that reversal of low BDNF levels offers the potential for meaningful treatment of motor and cognitive deficits in SCA1.

Altered Capicua expression drives regional Purkinje neuron vulnerability through ion channel gene dysregulation in spinocerebellar ataxia type 1.

Selective neuronal vulnerability in neurodegenerative disease is poorly understood. Using the ATXN1[82Q] model of spinocerebellar ataxia type 1 (SCA1), we explored the hypothesis that regional differences in Purkinje neuron degeneration could provide novel insights into selective vulnerability. ATXN1[82Q] Purkinje neurons from the anterior cerebellum were found to degenerate earlier than those from the nodular zone, and this early degeneration was associated with selective dysregulation of ion channel transcripts and altered Purkinje neuron spiking. Efforts to understand the basis for selective dysregulation of channel transcripts revealed modestly increased expression of the ATXN1 co-repressor Capicua (Cic) in anterior cerebellar Purkinje neurons. Importantly, disrupting the association between ATXN1 and Cic rescued the levels of these ion channel transcripts, and lentiviral overexpression of Cic in the nodular zone accelerated both aberrant Purkinje neuron spiking and neurodegeneration. These findings reinforce the central role for Cic in SCA1 cerebellar pathophysiology and suggest that only modest reductions in Cic are needed to have profound therapeutic impact in SCA1.

Mapping SCA1 regional vulnerabilities reveals neural and skeletal muscle contributions to disease.

Spinocerebellar ataxia type 1 (SCA1) is a fatal neurodegenerative disease caused by an expanded polyglutamine tract in the widely expressed ataxin-1 (ATXN1) protein. To elucidate anatomical regions and cell types that underlie mutant ATXN1-induced disease phenotypes, we developed a floxed conditional knockin mouse (f-ATXN1146Q/2Q) with mouse Atxn1 coding exons replaced by human ATXN1 exons encoding 146 glutamines. f-ATXN1146Q/2Q mice manifested SCA1-like phenotypes including motor and cognitive deficits, wasting, and decreased survival. Central nervous system (CNS) contributions to disease were revealed using f-ATXN1146Q/2Q;Nestin-Cre mice, which showed improved rotarod, open field, and Barnes maze performance by 6-12 weeks of age. In contrast, striatal contributions to motor deficits using f-ATXN1146Q/2Q;Rgs9-Cre mice revealed that mice lacking ATXN1146Q/2Q in striatal medium-spiny neurons showed a trending improvement in rotarod performance at 30 weeks of age. Surprisingly, a prominent role for muscle contributions to disease was revealed in f-ATXN1146Q/2Q;ACTA1-Cre mice based on their recovery from kyphosis and absence of muscle pathology. Collectively, data from the targeted conditional deletion of the expanded allele demonstrated CNS and peripheral contributions to disease and highlighted the need to consider muscle in addition to the brain for optimal SCA1 therapeutics.

Increased intrinsic membrane excitability is associated with hypertrophic olivary degeneration in spinocerebellar ataxia type 1.

One of the characteristic areas of brainstem degeneration across multiple spinocerebellar ataxias (SCAs) is the inferior olive (IO), a medullary nucleus that plays a key role in motor learning. In addition to its vulnerability in SCAs, the IO is also susceptible to a distinct pathology known as hypertrophic olivary degeneration (HOD). Clinically, HOD has been exclusively observed after lesions in the brainstem disrupt inhibitory afferents to the IO. Here, for the first time, we describe HOD in another context: spinocerebellar ataxia type 1 (SCA1). Using the genetically-precise SCA1 knock-in mouse model (SCA1-KI; both sexes used), we assessed SCA1-associated changes in IO neuron structure and function. Concurrent with degeneration, we found that SCA1-KI IO neurons are hypertrophic, exhibiting early dendrite lengthening and later somatic expansion. Unlike in previous descriptions of HOD, we observed no clear loss of IO inhibitory innervation; nevertheless, patch-clamp recordings from brainstem slices reveal that SCA1-KI IO neurons are hyperexcitable. Rather than synaptic disinhibition, we identify increases in intrinsic membrane excitability as the more likely mechanism underlying this novel SCA1 phenotype. Specifically, transcriptome analysis indicates that SCA1-KI IO hyperexcitability is associated with a reduced medullary expression of ion channels responsible for spike afterhyperpolarization (AHP) in IO neurons - a result that has a functional consequence, as SCA1-KI IO neuron spikes exhibit a diminished AHP. These results reveal membrane excitability as a potential link between disparate causes of IO degeneration, suggesting that HOD can result from any cause, intrinsic or extrinsic, that increases excitability of the IO neuron membrane.

Disruption of the ATXN1-CIC complex reveals the role of additional nuclear ATXN1 interactors in spinocerebellar ataxia type 1.

Spinocerebellar ataxia type 1 (SCA1) is a paradigmatic neurodegenerative disease in that it is caused by a mutation in a broadly expressed protein, ATXN1; however, only select populations of cells degenerate. The interaction of polyglutamine-expanded ATXN1 with the transcriptional repressor CIC drives cerebellar Purkinje cell pathogenesis; however, the importance of this interaction in other vulnerable cells remains unknown. Here, we mutated the 154Q knockin allele of Atxn1154Q/2Q mice to prevent the ATXN1-CIC interaction globally. This normalized genome-wide CIC binding; however, it only partially corrected transcriptional and behavioral phenotypes, suggesting the involvement of additional factors in disease pathogenesis. Using unbiased proteomics, we identified three ATXN1-interacting transcription factors: RFX1, ZBTB5, and ZKSCAN1. We observed altered expression of RFX1 and ZKSCAN1 target genes in SCA1 mice and patient-derived iNeurons, highlighting their potential contributions to disease. Together, these data underscore the complexity of mechanisms driving cellular vulnerability in SCA1.

Decreasing mutant ATXN1 nuclear localization improves a spectrum of SCA1-like phenotypes and brain region transcriptomic profiles.

Spinocerebellar ataxia type 1 (SCA1) is a dominant trinucleotide repeat neurodegenerative disease characterized by motor dysfunction, cognitive impairment, and premature death. Degeneration of cerebellar Purkinje cells is a frequent and prominent pathological feature of SCA1. We previously showed that transport of ATXN1 to Purkinje cell nuclei is required for pathology, where mutant ATXN1 alters transcription. To examine the role of ATXN1 nuclear localization broadly in SCA1-like disease pathogenesis, CRISPR-Cas9 was used to develop a mouse with an amino acid alteration (K772T) in the nuclear localization sequence of the expanded ATXN1 protein. Characterization of these mice indicates that proper nuclear localization of mutant ATXN1 contributes to many disease-like phenotypes including motor dysfunction, cognitive deficits, and premature lethality. RNA sequencing analysis of genes with expression corrected to WT levels in Atxn1175QK772T/2Q mice indicates that transcriptomic aspects of SCA1 pathogenesis differ between the cerebellum, brainstem, cerebral cortex, hippocampus, and striatum.

Delineating regional vulnerability in the neurodegenerative disease SCA1 using a conditional mutant ATXN1 mouse.

Spinocerebellar ataxia type 1 (SCA1) is a fatal neurodegenerative disease caused by an expanded polyglutamine tract in the widely expressed ATXN1 protein. To elucidate anatomical regions and cell types that underlie mutant ATXN1-induced disease phenotypes, we developed a floxed conditional knockout mouse model ( f-ATXN1 146Q/2Q ) having mouse Atxn1 coding exons replaced by human exons encoding 146 glutamines. F-ATXN1 146Q/2Q mice manifest SCA1-like phenotypes including motor and cognitive deficits, wasting, and decreased survival. CNS contributions to disease were revealed using ATXN1 146Q/2Q ; Nestin-Cre mice, that showed improved rotarod, open field and Barnes maze performances. Striatal contributions to motor deficits were examined using f-ATXN1 146Q/2Q ; Rgs9-Cre mice. Mice lacking striatal ATXN1 146Q/2Q had improved rotarod performance late in disease. Muscle contributions to disease were revealed in f-ATXN1 146Q/2Q ; ACTA1-Cre mice which lacked muscle pathology and kyphosis seen in f-ATXN1 146Q/2Q mice. Kyphosis was not improved in f-ATXN1 146Q/2Q ;Nestin - Cre mice. Thus, optimal SCA1 therapeutics will require targeting mutant ATXN1 toxic actions in multiple brain regions and muscle.

Spatial and Temporal Diversity of Astrocyte Phenotypes in Spinocerebellar Ataxia Type 1 Mice.

While astrocyte heterogeneity is an important feature of the healthy brain, less is understood about spatiotemporal heterogeneity of astrocytes in brain disease. Spinocerebellar ataxia type 1 (SCA1) is a progressive neurodegenerative disease caused by a CAG repeat expansion in the gene Ataxin1 (ATXN1). We characterized astrocytes across disease progression in the four clinically relevant brain regions, cerebellum, brainstem, hippocampus, and motor cortex, of Atxn1154Q/2Q mice, a knock-in mouse model of SCA1. We found brain region-specific changes in astrocyte density and GFAP expression and area, early in the disease and prior to neuronal loss. Expression of astrocytic core homeostatic genes was also altered in a brain region-specific manner and correlated with neuronal activity, indicating that astrocytes may compensate or exacerbate neuronal dysfunction. Late in disease, expression of astrocytic homeostatic genes was reduced in all four brain regions, indicating loss of astrocyte functions. We observed no obvious correlation between spatiotemporal changes in microglia and spatiotemporal astrocyte alterations, indicating a complex orchestration of glial phenotypes in disease. These results support spatiotemporal diversity of glial phenotypes as an important feature of the brain disease that may contribute to SCA1 pathogenesis in a brain region and disease stage-specific manner.

Antisense Oligonucleotide Therapeutic Approach for Suppression of Ataxin-1 Expression: A Safety Assessment.

Spinocerebellar ataxia type 1 (SCA1) is a lethal, autosomal dominant neurodegenerative disease caused by a polyglutamine expansion in the ATAXIN-1 (ATXN1) protein. Preclinical studies demonstrate the therapeutic efficacy of approaches that target and reduce Atxn1 expression in a non-allele-specific manner. However, studies using Atxn1-/- mice raise cautionary notes that therapeutic reductions of ATXN1 might lead to undesirable effects such as reduction in the activity of the tumor suppressor Capicua (CIC), activation of the protease ß-secretase 1 (BACE1) and subsequent increased amyloidogenic cleavage of the amyloid precursor protein (APP), or a reduction in hippocampal neuronal precursor cells that would impact hippocampal function. Here, we tested whether an antisense oligonucleotide (ASO)-mediated reduction of Atxn1 produced unwanted effects involving BACE1, CIC activity, or reduction in hippocampal neuronal precursor cells. Notably, no effects on BACE1, CIC tumor suppressor function, or number of hippocampal neuronal precursor cells were found in mice subjected to a chronic in vivo ASO-mediated reduction of Atxn1. These data provide further support for targeted reductions of ATXN1 as a therapeutic approach for SCA1.

Antisense oligonucleotide-mediated ataxin-1 reduction prolongs survival in SCA1 mice and reveals disease-associated transcriptome profiles.

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited ataxia caused by expansion of a translated CAG repeat encoding a glutamine tract in the ataxin-1 (ATXN1) protein. Despite advances in understanding the pathogenesis of SCA1, there are still no therapies to alter its progressive fatal course. RNA-targeting approaches have improved disease symptoms in preclinical rodent models of several neurological diseases. Here, we investigated the therapeutic capability of an antisense oligonucleotide (ASO) targeting mouse Atxn1 in Atxn1154Q/2Q-knockin mice that manifest motor deficits and premature lethality. Following a single ASO treatment at 5 weeks of age, mice demonstrated rescue of these disease-associated phenotypes. RNA-sequencing analysis of genes with expression restored to WT levels in ASO-treated Atxn1154Q/2Q mice was used to demonstrate molecular differences between SCA1 pathogenesis in the cerebellum and disease in the medulla. Finally, select neurochemical abnormalities detected by magnetic resonance spectroscopy in vehicle-treated Atxn1154Q/2Q mice were reversed in the cerebellum and brainstem (a region containing the pons and the medulla) of ASO-treated Atxn1154Q/2Q mice. Together, these findings support the efficacy and therapeutic importance of directly targeting ATXN1 RNA expression as a strategy for treating both motor deficits and lethality in SCA1.