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Cerebellar cortex in BTBR mice was expanded with increased foliation since post-natal. (A) DAPI stained cerebella of WT and BTBR mice at adult stage (P90). (B) Quantification of the sagittal area of cerebella and each component at adulthood (Student’s t-test; n = 6, 7 mice). (C) Quantification of the average lobe number showing increased foliation of adult BTBR mice (Student’s t-test; n = 6, 7 mice). (D) Whole mount images and sagittal section of brain in WT and BTBR mice at adult stage (P90). White and black dotted line delimit the lobule outline. Black arrowheads indicate the lobule fissures. (E) Schema graph illustrating the determination of perimeter and area. (F) Hematoxylin-eosin (HE) staining of middle sagittal cerebellar section at postnatal day 3, 7, and 14 (additional lobes highlighted in red in the counterdraw). (G) Quantification of the average lobule number in WT and BTBR mice at indicated stage (Student’s t-test; P3 n = 6,5; P7 n = 6,6; P14 n = 5,6). (H) Quantification of the average sagittal cerebellar area in WT and BTBR mice at indicated stage (Student’s t-test; P3 n = 6, 5; P7 n = 6, 6; P14 n = 5, 6). (I) Quantification of the average sagittal cerebellar section perimeter in WT and BTBR mice at indicated stage (Student’s t-test; P3 n = 6,5; P7 n = 6,6; P14 n = 5,6). All data are displayed as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001. Scale bar: (A,F) 200 μm; (D) 1 mm.
Source publication
Motor control and learning impairments are common complications in individuals with autism spectrum disorder (ASD). Abnormal cerebellar development during critical phases may disrupt these motor functions and lead to autistic motor dysfunction. However, the underlying mechanisms behind these impairments are not clear. Here, we utilized BTBR T⁺ Itpr...
Citations
... This receptor plays a critical role in regulating intracellular calcium levels (Mangla et al., 2020) and as calcium signaling is directly linked to ER homeostasis and eIF2α signaling, it is reasonable to speculate that the combination of calcium signaling dysregulation and Prkra lear-5J mutation may result in the enhanced dystonia phenotype, especially since we have previously linked PACT/RAX mediated PKR regulation to play a central role in cellular fate in response to ER stress Vaughn et al., 2015;Burnett et al., 2019). Previous reports demonstrate that the BTBR mice exhibit abnormal cerebellar development leading to motor dysfunction (Xiao et al., 2020). This study noted an increased proliferation of cerebellar granule neurons with enhanced cerebellar foliation, and hypertrophy of Purkinje cells with increased dendritic spines in BTBR mice. ...
Mutations in Prkra gene, which encodes PACT/RAX cause early onset primary dystonia DYT-PRKRA, a movement disorder that disrupts coordinated muscle movements. PACT/RAX activates protein kinase R (PKR, aka EIF2AK2) by a direct interaction in response to cellular stressors to mediate phosphorylation of the α subunit of the eukaryotic translation initiation factor 2 (eIF2α). Mice homozygous for a naturally arisen, recessively inherited frameshift mutation, Prkra lear-5J exhibit progressive dystonia. In the present study, we investigate the biochemical and developmental consequences of the Prkra lear-5J mutation. Our results indicate that the truncated PACT/RAX protein retains its ability to interact with PKR, however, it inhibits PKR activation. Furthermore, mice homozygous for the mutation have abnormalities in the cerebellar development as well as a severe lack of dendritic arborization of Purkinje neurons. Additionally, reduced eIF2α phosphorylation is noted in the cerebellums and Purkinje neurons of the homozygous Prkra lear-5J mice. These results indicate that PACT/RAX mediated regulation of PKR activity and eIF2α phosphorylation plays a role in cerebellar development and contributes to the dystonia phenotype resulting from this mutation.
Summary Statement
This study shows, for the first time, a role of reduced eIF2α phosphorylation in DYT-PRKRA and the cerebellum development in a mouse model.
... Similarly, a study conducted by Xiao et al. focused on BTBR T+ Itprtf/J (BTBR) mice as a model to explore abnormal cerebellar development associated with motor impairment in autism spectrum disorder (ASD). Through their transcription analysis, they identified TRPC as a novel risk gene implicated in motor dysfunction in ASD [58]. Conversely, Emily et al. employed Neurexin 1α knock-out mice to investigate the behavioural outcomes and motor learning deficits. ...
Developmental coordination disorder (DCD) is a developmental disorder characterized by impaired motor coordination, often co-occurring with attention deficit disorder, autism spectrum disorders, and other psychological and behavioural conditions. The aetiology of DCD is believed to involve brain changes and environmental factors, with genetics also playing a role in its pathogenesis. Recent research has identified several candidate genes and genetic factors associated with motor impairment, including deletions, copy number variations, single nucleotide polymorphisms, and epigenetic modifications. This review provides an overview of the current knowledge in genetic research on DCD, highlighting the importance of continued research into the underlying genetic mechanisms. While evidence suggests a genetic contribution to DCD, the evidence is still in its early stages, and much of the current evidence is based on studies of co-occurring conditions. Further research to better understand the genetic basis of DCD could have important implications for diagnosis, treatment, and our understanding of the condition’s aetiology.
... Loss-of-function mutations in the manganese transporters SLC30A10 or SLC39A14 lead to manganese-induced neurotoxicity, which is characterized by childhood-onset parkinsonism-dystonia (Quadri et al., 2012;Tuschl et al., 2012Tuschl et al., , 2016. While Slc30a10 knockout mice displayed severe hyperthyroidism without any obvious motor phenotypes (Hutchens et al., 2017), Slc39a14 knockout mice exhibited early-onset dystonic-like movements and motor coordination deficits which progressively worsened over time (Jenkitkasemwong et al., 2018;Rodichkin, Edler, McGlothan, & Guilarte, 2021;Rodichkin, Edler, McGlothan, & Guilarte, 2022;Xin et al., 2017). Manganese levels were significantly increased in the cerebellum, globus pallidus, midbrain, and cortex (Jenkitkasemwong et al., 2018;Xin et al., 2017). ...
... While Slc30a10 knockout mice displayed severe hyperthyroidism without any obvious motor phenotypes (Hutchens et al., 2017), Slc39a14 knockout mice exhibited early-onset dystonic-like movements and motor coordination deficits which progressively worsened over time (Jenkitkasemwong et al., 2018;Rodichkin, Edler, McGlothan, & Guilarte, 2021;Rodichkin, Edler, McGlothan, & Guilarte, 2022;Xin et al., 2017). Manganese levels were significantly increased in the cerebellum, globus pallidus, midbrain, and cortex (Jenkitkasemwong et al., 2018;Xin et al., 2017). Interestingly, treatment with the metal chelator Na 2 CaEDTA improved motor coordination (Xin et al., 2017). ...
... Manganese levels were significantly increased in the cerebellum, globus pallidus, midbrain, and cortex (Jenkitkasemwong et al., 2018;Xin et al., 2017). Interestingly, treatment with the metal chelator Na 2 CaEDTA improved motor coordination (Xin et al., 2017). Although the nigrostriatal dopaminergic system appeared structurally intact, potassium-stimulated dopamine release in the striatum was markedly inhibited compared to controls (Rodichkin et al., 2021(Rodichkin et al., , 2022. ...
Dystonia is currently ranked as the third most prevalent motor disorder. It is typically characterized by involuntary muscle over- or co-contractions that can cause painful abnormal postures and jerky movements. Dystonia is a heterogenous disorder-across patients, dystonic symptoms vary in their severity, body distribution, temporal pattern, onset, and progression. There are also a growing number of genes that are associated with hereditary dystonia. In addition, multiple brain regions are associated with dystonic symptoms in both genetic and sporadic forms of the disease. The heterogeneity of dystonia has made it difficult to fully understand its underlying pathophysiology. However, the use of animal models has been used to uncover the complex circuit mechanisms that lead to dystonic behaviors. Here, we summarize findings from animal models harboring mutations in dystonia-associated genes and phenotypic animal models with overt dystonic motor signs resulting from spontaneous mutations, neural circuit perturbations, or pharmacological manipulations. Taken together, an emerging picture depicts dystonia as a result of brain-wide network dysfunction driven by basal ganglia and cerebellar dysfunction. In the basal ganglia, changes in dopaminergic, serotonergic, noradrenergic, and cholinergic signaling are found across different animal models. In the cerebellum, abnormal burst firing activity is observed in multiple dystonia models. We are now beginning to unveil the extent to which these structures mechanistically interact with each other. Such mechanisms inspire the use of pre-clinical animal models that will be used to design new therapies including drug treatments and brain stimulation.
... Motor dysfunction in BTBR mice has been previously shown by Xiao et al. (2020) evaluating it from 2 weeks to 5 months of age. Researchers showed that abnormal cerebellar development could be an important factor related to motor dysfunction as well as dystonia-like behavior in these animals. ...
... A previous report of the same test performed with 22 days old BTBR mice showed that they traveled more than B6 controls in the first 15 min of the test session that was performed under red light (McFarlane et al., 2008). Furthermore, another study showed that 8 weeks old BTBR mice present hyperactivity compared to wild type control mice by an increased distance moved in all zones in the open field test (Xiao et al., 2020). However, another study measured the distance moved in the open field test in 15 months old male BTBR mice for 30 min under room lightening, in which BTBR mice moved a comparable distance than B6 controls (Jasien et al., 2014). ...
Introduction
Autism spectrum disorder (ASD) is a persistent neurodevelopmental condition characterized by two core behavioral symptoms: impaired social communication and interaction, as well as stereotypic, repetitive behavior. No distinct cause of ASD is known so far; however, excitatory/inhibitory imbalance and a disturbed serotoninergic transmission have been identified as prominent candidates responsible for ASD etiology.
Methods
The GABAB receptor agonist R-Baclofen and the selective agonist for the 5HT7 serotonin receptor LP-211 have been reported to correct social deficits and repetitive behaviors in mouse models of ASD. To evaluate the efficacy of these compounds in more details, we treated BTBR T⁺ Itpr3tf/J and B6.129P2-Fmr1tm1Cgr/J mice acutely with R-Baclofen or LP-211 and evaluated the behavior of animals in a series of tests.
Results
BTBR mice showed motor deficits, elevated anxiety, and highly repetitive behavior of self-grooming. Fmr1-KO mice exhibited decreased anxiety and hyperactivity. Additionally, Fmr1-KO mice’s ultrasonic vocalizations were impaired suggesting a reduced social interest and communication of this strain. Acute LP-211 administration did not affect the behavioral abnormalities observed in BTBR mice but improved repetitive behavior in Fmr1-KO mice and showed a trend to change anxiety of this strain. Acute R-Baclofen treatment improved repetitive behavior only in Fmr1-KO mice.
Conclusion
Our results add value to the current available data on these mouse models and the respective compounds. Yet, additional studies are needed to further test R-Baclofen and LP-211 as potential treatments for ASD therapy.
... For these reasons, we quantified the expression of NMDAR1s and CB1Rs in the cerebellar cortex. In particular, we measured these changes in lobule VII because changes in the structure and physiology of this area correlate with abnormal behaviors such as compulsive rituals, stereotypical performance, and difficulty to understand social cues [15], which are replicated in the abnormal behavioral phenotype of the BTBR strain [16,17]. The BTBR and C57 groups of each sex were exposed to a standard or enriched environmental conditions. ...
Objective
Environmental enrichment is used to treat social, communication, and behavioral deficits and is known to modify the expression of synaptic receptors. We compared the effects of environmental enrichment in the expression of glutamate and endocannabinoid receptors, which are widely expressed in the cerebellar cortex. These two receptors interact to regulate neuronal function and their dysregulation is associated with behavioral changes. We used BTBR + Itpr3tf/J mice, a strain that models behavioral disorders, and C57BL/6 mice for comparison. We studied the effects of genetic background, sex, environmental conditions, and layer of the cerebellar cortex on the expression of each receptor.
Results
The influence of genetic background and environmental enrichment had the same pattern on glutamate and endocannabinoid receptors in males. In contrast, in females, the effect of environmental enrichment and genetic background were different than the ones obtained for males and were also different between the glutamate and endocannabinoid receptors. Furthermore, an analysis of both receptors from tissue obtained from the same animals show that their expression is correlated in males, but not in females. Our results suggest that environmental enrichment has a receptor dependent and sexual dimorphic effect on the molecular expression of different receptors in the cerebellar cortex.
... Our finding of significant motor learning impairments in the BTBR mouse model is consistent with previous literature. One prior work by Xiao and colleagues noted a disruption of rotarod performance in male BTBR mice [82]. The present study confirms that finding, while also adding that female mice have a similar-albeit less severe-deficit. ...
... Our findings may apply more narrowly to one of these subpopulations. In addition, we have discovered abnormal foliation in both male and female mice, confirming one previous study in male mice [82] and notable because the earlier literature had rejected the notion of anatomical abnormality in the cerebellum [93]. This foliation defect is indicative of disruption of the maturation of the cerebellum early in postnatal murine brain development [94]; the observation that early granule cell layer development as well as Purkinje cell migration defects [82] might account for this disrupted foliation is in line with our observation of regional differences in the area of the granule cell layer and molecular cell layer. ...
... In addition, we have discovered abnormal foliation in both male and female mice, confirming one previous study in male mice [82] and notable because the earlier literature had rejected the notion of anatomical abnormality in the cerebellum [93]. This foliation defect is indicative of disruption of the maturation of the cerebellum early in postnatal murine brain development [94]; the observation that early granule cell layer development as well as Purkinje cell migration defects [82] might account for this disrupted foliation is in line with our observation of regional differences in the area of the granule cell layer and molecular cell layer. However, further investigation is required to understand the significance of hyperfoliation of some lobules and not others. ...
Recently, there has been increased interest in the role of the cerebellum in autism spectrum disorder (ASD). To better understand the pathophysiological role of the cerebellum in ASD, it is necessary to have a variety of mouse models that have face validity for cerebellar disruption in humans. Here, we add to the literature on the cerebellum in mouse models of autism with the characterization of the cerebellum in the idiopathic BTBR T + Itpr3tf/J (BTBR) inbred mouse strain, which has behavioral phenotypes that are reminiscent of ASD in patients. When we examined both male and female BTBR mice in comparison to C57BL/6J (C57) controls, we noted that both sexes of BTBR mice showed motor coordination deficits characteristic of cerebellar dysfunction, but only the male mice showed differences in delay eyeblink conditioning, a cerebellum-dependent learning task that is known to be disrupted in ASD patients. Both male and female BTBR mice showed considerable expansion of, and abnormal foliation in, the cerebellum vermis—including a significant expansion of specific lobules in the anterior cerebellum. In addition, we found a slight but significant decrease in Purkinje cell density in both male and female BTBR mice, irrespective of the lobule. Finally, there was a marked reduction of Purkinje cell dendritic spine density in both male and female BTBR mice. These findings suggest that, for the most part, the BTBR mouse model phenocopies many of the characteristics of the subpopulation of ASD patients that have a hypertrophic cerebellum. We discuss the significance of strain differences in the cerebellum as well as the importance of this first effort to identify both similarities and differences between male and female BTBR mice with regard to the cerebellum.
... Our finding of significant motor learning impairments in the BTBR mouse model is consistent with literature on ASD mouse models. One prior work, by Xiao and colleagues, had noted a disruption of rotarod performance in BTBR male male (Xiao et al., 2020). The present study confirms that finding, while also adding that female mice have a similar--albeit less severe--deficit. ...
... Our findings may apply more narrowly to one of these subpopulations. In addition, we have discovered abnormal foliation in both male and female mice, confirming one previous study in male mice (Xiao et al., 2020) and notable because earlier literature had rejected the notion of anatomical abnormality in the cerebellum (Stephenson et al., 2011). This foliation defect is indicative of disruption of the maturation of the cerebellum early in postnatal murine brain development (van der Heijden et al., 2021); the observation that early granule cell layer development as well as Purkinje cell migration defects (Xiao et al., 2020) might account for this disrupted foliation is in line with our observation of regional differences in the area of the granule cell layer and molecular cell layer. ...
... In addition, we have discovered abnormal foliation in both male and female mice, confirming one previous study in male mice (Xiao et al., 2020) and notable because earlier literature had rejected the notion of anatomical abnormality in the cerebellum (Stephenson et al., 2011). This foliation defect is indicative of disruption of the maturation of the cerebellum early in postnatal murine brain development (van der Heijden et al., 2021); the observation that early granule cell layer development as well as Purkinje cell migration defects (Xiao et al., 2020) might account for this disrupted foliation is in line with our observation of regional differences in the area of the granule cell layer and molecular cell layer. However, further investigation is required to understand the significance of hyperfoliation of some lobules and not others. ...
Recently, there has been increased interest in the role of the cerebellum in autism spectrum disorders (ASD). In order to better understand the pathophysiological role of the cerebellum in ASD, it is necessary to have a variety of mouse models that have face validity for cerebellar disruption in humans. Here, we add to the literature on the cerebellum transgenic and induced mouse models of autism with the characterization of the cerebellum in the BTBR T+Itpr3 tf /J (BTBR) inbred mouse strain, which has behavioral phenotypes that are suggestive of ASD in patients. When we examined both male and female adult BTBR mice in comparison to C57BL/6J (C57) controls, we noted that both mice showed motor coordination deficits characteristic of cerebellar function, but only the male mice showed differences in delay eyeblink conditioning, a cerebellum-dependent learning task that is also disrupted in ASD patients. Both male and female BTBR mice showed considerable expansion of and abnormal foliation in the cerebellum vermis--including significant expansion of specific lobules in the anterior cerebellum. In addition, we found a slight but significant decrease in Purkinje cell density in both male and female BTBR mice, irrespective of lobule. Furthermore, there was a marked reduction of Purkinje cell dendritic spines density in both male and female BTBR mice. These findings suggest that, for the most part, the BTBR mouse model successfully phenocopies many of the characteristics of the subpopulation of ASD patients that have a hypertrophic cerebellum. We discuss the significance of sex differences--revealed for the first time in BTBR mice, and present in only a small number of cerebellum studies--and the importance of concordance on other metrics between male and female BTBR mice.
Graphical Abstract
Summary of differences between BTBR mice (left) and C57 mice (right) demonstrated by this study, separated by sex.
... Interestingly, the mRNA expression of BOD1 was consistently downregulated in dystonia model mice from RNA-seq analyses data (GSE98839) (Fig. 3c). 21 However, the role of BOD1 in the brain is largely unexplored. Immunofluorescence staining revealed that BOD1 was highly expressed in PCs and colocalized with calbindin (Fig. 3d, e). ...
Cerebellar ataxias are characterized by a progressive decline in motor coordination, but the specific output circuits and underlying pathological mechanism remain poorly understood. Through cell-type-specific manipulations, we discovered a novel GABAergic Purkinje cell (PC) circuit in the cerebellar IV/V lobe that projected to CaMKIIα+ neurons in the fastigial nucleus (FN), which regulated sensorimotor coordination. Furthermore, transcriptomics profiling analysis revealed various cerebellar neuronal identities, and we validated that biorientation defective 1 (BOD1) played an important role in the circuit of IV/V lobe to FN. BOD1 deficit in PCs of IV/V lobe attenuated the excitability and spine density of PCs, accompany with ataxia behaviors. Instead, BOD1 enrichment in PCs of IV/V lobe reversed the hyperexcitability of CaMKIIα+ neurons in the FN and ameliorated ataxia behaviors in L7-Cre; BOD1f/f mice. Together, these findings further suggest that specific regulation of the cerebellar IV/V lobePCs → FNCaMKIIα+ circuit might provide neuromodulatory targets for the treatment of ataxia behaviors.
... In particular, among the brain regions that Heo and colleagues [219] examined, the cerebellum exhibited significantly higher expression of IL-33, IL-18 and IL-6 in BTBR mice than in control, suggesting that it could be a crucial area for neuroinflammation in humans with ASD. Finally, a recent study revealed an abnormal cerebellar development (enhanced foliation and PC hypotrophy with altered dendritic spine formation) concomitant with the progression of motor impairments in BTBR mice [223]. In summary, although there is a growing body of evidence supporting the relationship between cytokine alterations and ASD, systematic and large scale investigations are needed to better clarify the role of cerebellar inflammation on the emergence of ASD and the contribution to its etiological heterogeneity. ...
Autism spectrum disorders (ASD) are pervasive neurodevelopmental disorders that include a variety of forms and clinical phenotypes. This heterogeneity complicates the clinical and experimental approaches to ASD etiology and pathophysiology. To date, a unifying theory of these diseases is still missing. Nevertheless, the intense work of researchers and clinicians in the last decades has identified some ASD hallmarks and the primary brain areas involved. Not surprisingly, the areas that are part of the so-called “social brain”, and those strictly connected to them, were found to be crucial, such as the prefrontal cortex, amygdala, hippocampus, limbic system, and dopaminergic pathways. With the recent acknowledgment of the cerebellar contribution to cognitive functions and the social brain, its involvement in ASD has become unmistakable, though its extent is still to be elucidated. In most cases, significant advances were made possible by recent technological developments in structural/functional assessment of the human brain and by using mouse models of ASD. Mouse models are an invaluable tool to get insights into the molecular and cellular counterparts of the disease, acting on the specific genetic background generating ASD-like phenotype. Given the multifaceted nature of ASD and related studies, it is often difficult to navigate the literature and limit the huge content to specific questions. This review fulfills the need for an organized, clear, and state-of-the-art perspective on cerebellar involvement in ASD, from its connections to the social brain areas (which are the primary sites of ASD impairments) to the use of monogenic mouse models.
... For these reasons, we quanti ed the expression of NMDAR1s and CB1Rs in the cerebellar cortex. In particular, we measured these changes in lobule VII because changes in the structure and physiology of this area correlate with abnormal behaviors such as compulsive rituals, stereotypical performance, and di culty to understand social cues (15), which are replicated in the abnormal behavioral phenotype of the BTBR strain (16,17). The BTBR and C57 groups of each sex were exposed to a standard or enriched environmental conditions. ...
Objective: Environmental enrichment is used to treat social, communication, and behavioral deficits and is known to modify the expression of synaptic receptors. We compared the effects of environmental enrichment in the expression of glutamate and endocannabinoid receptors, which are widely expressed in the cerebellar cortex. These two receptors interact to regulate neuronal function and their dysregulation is associated with behavioral changes. We used BTBR +Itpr3tf/J mice, a strain that models behavioral disorders, and C57BL/6 mice for comparison. We studied the effects of genetic background, sex, environmental conditions, and layer of the cerebellar cortex on the expression of each receptor.
Results: The influence of genetic background and environmental enrichment had the same pattern on glutamate and endocannabinoid receptors in males. In contrast, in females, the effect of environmental enrichment and genetic background were different than the ones obtained for males and were also different between the glutamate and endocannabinoid receptors. Furthermore, an analysis of both receptors from tissue obtained from the same animals show that their expression is correlated in males, but not in females. Our results suggest that environmental enrichment has a receptor dependent and sexual dimorphic effect on the molecular expression of different receptors in the cerebellar cortex.
