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BioRxiv : the Preprint Server For... Mar 2024The Cre-lox system is an indispensable tool in neuroscience research for targeting gene deletions to specific cellular populations. Here we assess the utility of several...
The Cre-lox system is an indispensable tool in neuroscience research for targeting gene deletions to specific cellular populations. Here we assess the utility of several transgenic lines, along with a viral approach, for targeting cerebellar Purkinje cells. Using a combination of a fluorescent reporter line () to indicate -mediated recombination and a floxed line () we show that reporter expression does not always align precisely with loss of protein. The commonly used line exhibits a gradual mosaic pattern of recombination in Purkinje cells from P7-P14, while loss of Dag1 protein is not complete until P30. drives recombination in precursor cells that give rise to GABAergic neurons in the embryonic cerebellum, including Purkinje cells and molecular layer interneurons. However, due to its transient expression in precursors, results in stochastic loss of Dag1 protein in these neurons. , which is often described as a "pan-neuronal" line for the central nervous system, does not drive -mediated recombination in Purkinje cells. We identify a line that drives efficient and complete recombination in embryonic Purkinje cells, resulting in loss of Dag1 protein before the period of synaptogenesis. -mediated delivery of at P0 results in gradual transduction of Purkinje cells during the second postnatal week, with loss of Dag1 protein not reaching appreciable levels until P35. These results characterize several tools for targeting conditional deletions in cerebellar Purkinje cells at different developmental stages and illustrate the importance of validating the loss of protein following recombination.
PubMed: 38585758
DOI: 10.1101/2024.03.28.587263 -
Neurobiology of Disease Jun 2024We performed a comprehensive study of the morphological, functional, and genetic features of moonwalker (MWK) mice, a mouse model of spinocerebellar ataxia caused by a...
We performed a comprehensive study of the morphological, functional, and genetic features of moonwalker (MWK) mice, a mouse model of spinocerebellar ataxia caused by a gain of function of the TRPC3 channel. These mice show numerous behavioral symptoms including tremor, altered gait, circling behavior, impaired motor coordination, impaired motor learning and decreased limb strength. Cerebellar pathology is characterized by early and almost complete loss of unipolar brush cells as well as slowly progressive, moderate loss of Purkinje cell (PCs). Structural damage also includes loss of synaptic contacts from parallel fibers, swollen ER structures, and degenerating axons. Interestingly, no obvious correlation was observed between PC loss and severity of the symptoms, as the phenotype stabilizes around 2 months of age, while the cerebellar pathology is progressive. This is probably due to the fact that PC function is severely impaired much earlier than the appearance of PC loss. Indeed, PC firing is already impaired in 3 weeks old mice. An interesting feature of the MWK pathology that still remains to be explained consists in a strong lobule selectivity of the PC loss, which is puzzling considering that TRPC is expressed in every PC. Intriguingly, genetic analysis of MWK cerebella shows, among other alterations, changes in the expression of both apoptosis inducing and resistance factors possibly suggesting that damaged PCs initiate specific cellular pathways that protect them from overt cell loss.
Topics: Animals; Mice; Disease Models, Animal; Phenotype; Cerebellum; Purkinje Cells; TRPC Cation Channels; Genotype; Spinocerebellar Ataxias; Mice, Neurologic Mutants; Mice, Inbred C57BL; Mice, Transgenic
PubMed: 38575093
DOI: 10.1016/j.nbd.2024.106492 -
Memantine suppresses the excitotoxicity but fails to rescue the ataxic phenotype in SCA1 model mice.Biomedicine & Pharmacotherapy =... May 2024Spinocerebellar ataxia type 1 (SCA1) is a debilitating neurodegenerative disorder of the cerebellum and brainstem. Memantine has been proposed as a potential treatment...
Spinocerebellar ataxia type 1 (SCA1) is a debilitating neurodegenerative disorder of the cerebellum and brainstem. Memantine has been proposed as a potential treatment for SCA1. It blocks N-methyl-D-aspartate (NMDA) receptors on neurons, reduces excitotoxicity and decreases neurodegeneration in Alzheimer models. However, in cerebellar neurodegenerative diseases, the potential value of memantine is still unclear. We investigated the effects of memantine on motor performance and synaptic transmission in the cerebellum in a mouse model where mutant ataxin 1 is specifically targeted to glia. Lentiviral vectors (LVV) were used to express mutant ataxin 1 selectively in Bergmann glia (BG). In mice transduced with the mutant ataxin 1, chronic treatment with memantine improved motor activity during initial tests, presumably due to preserved BG and Purkinje cell (PC) morphology and numbers. However, mice were unable to improve their rota rod scores during next days of training. Memantine also compromised improvement in the rota rod scores in control mice upon repetitive training. These effects may be due to the effects of memantine on plasticity (LTD suppression) and NMDA receptor modulation. Some effects of chronically administered memantine persisted even after its wash-out from brain slices. Chronic memantine reduced morphological signs of neurodegeneration in the cerebellum of SCA1 model mice. This resulted in an apparent initial reduction of ataxic phenotype, but memantine also affected cerebellar plasticity and ultimately compromised motor learning. We speculate that that clinical application of memantine in SCA1 might be hampered by its ability to suppress NMDA-dependent plasticity in cerebellar cortex.
Topics: Animals; Memantine; Disease Models, Animal; Spinocerebellar Ataxias; Phenotype; Mice; Ataxin-1; Motor Activity; Cerebellum; Purkinje Cells; Receptors, N-Methyl-D-Aspartate; Mice, Transgenic; Mice, Inbred C57BL; Neuroglia; Male; Neuronal Plasticity
PubMed: 38574621
DOI: 10.1016/j.biopha.2024.116526 -
PLoS Computational Biology Apr 2024According to the motor learning theory by Albus and Ito, synaptic depression at the parallel fibre to Purkinje cells synapse (pf-PC) is the main substrate responsible...
According to the motor learning theory by Albus and Ito, synaptic depression at the parallel fibre to Purkinje cells synapse (pf-PC) is the main substrate responsible for learning sensorimotor contingencies under climbing fibre control. However, recent experimental evidence challenges this relatively monopolistic view of cerebellar learning. Bidirectional plasticity appears crucial for learning, in which different microzones can undergo opposite changes of synaptic strength (e.g. downbound microzones-more likely depression, upbound microzones-more likely potentiation), and multiple forms of plasticity have been identified, distributed over different cerebellar circuit synapses. Here, we have simulated classical eyeblink conditioning (CEBC) using an advanced spiking cerebellar model embedding downbound and upbound modules that are subject to multiple plasticity rules. Simulations indicate that synaptic plasticity regulates the cascade of precise spiking patterns spreading throughout the cerebellar cortex and cerebellar nuclei. CEBC was supported by plasticity at the pf-PC synapses as well as at the synapses of the molecular layer interneurons (MLIs), but only the combined switch-off of both sites of plasticity compromised learning significantly. By differentially engaging climbing fibre information and related forms of synaptic plasticity, both microzones contributed to generate a well-timed conditioned response, but it was the downbound module that played the major role in this process. The outcomes of our simulations closely align with the behavioural and electrophysiological phenotypes of mutant mice suffering from cell-specific mutations that affect processing of their PC and/or MLI synapses. Our data highlight that a synergy of bidirectional plasticity rules distributed across the cerebellum can facilitate finetuning of adaptive associative behaviours at a high spatiotemporal resolution.
Topics: Neuronal Plasticity; Animals; Models, Neurological; Cerebellum; Computer Simulation; Conditioning, Eyelid; Purkinje Cells; Blinking; Conditioning, Classical; Synapses; Computational Biology; Mice; Cerebellar Cortex
PubMed: 38574161
DOI: 10.1371/journal.pcbi.1011277 -
Nature Neuroscience May 2024Supervised learning depends on instructive signals that shape the output of neural circuits to support learned changes in behavior. Climbing fiber (CF) inputs to the...
Supervised learning depends on instructive signals that shape the output of neural circuits to support learned changes in behavior. Climbing fiber (CF) inputs to the cerebellar cortex represent one of the strongest candidates in the vertebrate brain for conveying neural instructive signals. However, recent studies have shown that Purkinje cell stimulation can also drive cerebellar learning and the relative importance of these two neuron types in providing instructive signals for cerebellum-dependent behaviors remains unresolved. In the present study we used cell-type-specific perturbations of various cerebellar circuit elements to systematically evaluate their contributions to delay eyeblink conditioning in mice. Our findings reveal that, although optogenetic stimulation of either CFs or Purkinje cells can drive learning under some conditions, even subtle reductions in CF signaling completely block learning to natural stimuli. We conclude that CFs and corresponding Purkinje cell complex spike events provide essential instructive signals for associative cerebellar learning.
Topics: Animals; Purkinje Cells; Mice; Optogenetics; Association Learning; Conditioning, Eyelid; Male; Mice, Inbred C57BL; Cerebellum; Nerve Fibers; Mice, Transgenic; Cerebellar Cortex; Female
PubMed: 38565684
DOI: 10.1038/s41593-024-01594-7 -
Disease Models & Mechanisms Jun 2024Purkinje cell dysfunction disrupts movement and causes disorders such as ataxia. Recent evidence suggests that Purkinje cell dysfunction may also alter sleep regulation....
Purkinje cell dysfunction disrupts movement and causes disorders such as ataxia. Recent evidence suggests that Purkinje cell dysfunction may also alter sleep regulation. Here, we used an ataxic mouse model generated by silencing Purkinje cell neurotransmission (L7Cre;Vgatfx/fx) to better understand how cerebellar dysfunction impacts sleep physiology. We focused our analysis on sleep architecture and electrocorticography (ECoG) patterns based on their relevance to extracting physiological measurements during sleep. We found that circadian activity was unaltered in the mutant mice, although their sleep parameters and ECoG patterns were modified. The L7Cre;Vgatfx/fx mutant mice had decreased wakefulness and rapid eye movement (REM) sleep, whereas non-REM sleep was increased. The mutants had an extended latency to REM sleep, which is also observed in human patients with ataxia. Spectral analysis of ECoG signals revealed alterations in the power distribution across different frequency bands defining sleep. Therefore, Purkinje cell dysfunction may influence wakefulness and equilibrium of distinct sleep stages in ataxia. Our findings posit a connection between cerebellar dysfunction and disrupted sleep and underscore the importance of examining cerebellar circuit function in sleep disorders.
Topics: Animals; Purkinje Cells; Wakefulness; Ataxia; Sleep; Sleep, REM; Mice; Circadian Rhythm; Disease Models, Animal; Male
PubMed: 38563553
DOI: 10.1242/dmm.050379 -
Journal of Neuropathology and... Apr 2024Two aspects of the neuropathology of early Huntington disease (HD) are examined. Neurons of the neostriatum are counted to determine relative loss in striosomes versus...
Two aspects of the neuropathology of early Huntington disease (HD) are examined. Neurons of the neostriatum are counted to determine relative loss in striosomes versus matrix at early stages, including for the first time in preclinical cases. An immunohistochemical procedure is described that tentatively distinguishes early HD from HD mimic disorders in postmortem brains. Counts of striatal projection neurons (SPNs) in striosomes defined by calbindin immunohistochemistry versus counts in the surrounding matrix are reported for 8 Vonsattel grade 0 (including 5 premanifest), 8 grade 1, 2 grade 2 HD, and for 8 control postmortem brains. Mean counts of striosome and matrix SPNs were significantly lower in premanifest grade 0 versus controls, with striosome counts significantly lower than matrix. In 8 grade 1 and 2 grade 2 brains, no striosomes with higher SPN counts than in the surrounding matrix were observed. Comparing dorsal versus ventral neostriatum, SPNs in dorsal striosomes and matrix declined more than ventral, making clear the importance of the dorsoventral site of tissue selection for research studies. A characteristic pattern of expanded polyglutamine-immunopositive inclusions was seen in all HD cases. Inclusions were always present in some SPNs and some pontine nucleus neurons and were absent in Purkinje cells, which showed no obvious cell loss.
Topics: Humans; Huntington Disease; Corpus Striatum; Neostriatum; Neurons; Calbindins
PubMed: 38553027
DOI: 10.1093/jnen/nlae022 -
Brain Research Jul 2024A Disintegrin And Metalloprotease 10 (ADAM10), is able to control several important physiopathological processes through the shedding of a large number of protein...
A Disintegrin And Metalloprotease 10 (ADAM10), is able to control several important physiopathological processes through the shedding of a large number of protein substrates. Although ADAM10 plays a crucial role in the central nervous system (CNS) development and function, its protein distribution in the CNS has not been fully addressed. Here, we described the regional and cellular ADAM10 protein expression in C57BL/6 mice examined by immunofluorescence 1) throughout the adult mouse brain, cerebellum and spinal cord in vivo and 2) in different cell types as neurons, astrocytes, oligodendrocytes and microglia in vitro. We observed ADAM10 expression through the whole CNS, with a strong expression in the hippocampus, in the hypothalamus and in the cerebral and piriform cortex in the brain, in the Purkinje and in granular cell layers in the cerebellum and in the spinal cord to a lower extent. In vivo, ADAM10 protein expression was mainly found in neurons and in some oligodendroglial cell populations. However, in primary cultures we observed ADAM10 expression in neurons, oligodendrocytes, astrocytes and microglia. Interestingly, ADAM10 was not only found in the membrane but also in cytoplasmic vesicles and in the nucleus of primary cultured cells. Overall, this work highlights a wide distribution of ADAM10 throughout the CNS. The nuclear localization of ADAM10, probably due to its intracellular domain, emphasizes its role in cell signalling in physiological and pathological conditions. Further investigations are required to better elucidate the role of ADAM10 in glial cells.
Topics: Animals; ADAM10 Protein; Mice, Inbred C57BL; Neurons; Mice; Membrane Proteins; Central Nervous System; Spinal Cord; Amyloid Precursor Protein Secretases; Astrocytes; Microglia; Cells, Cultured; Oligodendroglia; Male; Brain; Cerebellum
PubMed: 38548249
DOI: 10.1016/j.brainres.2024.148888 -
ELife Mar 2024The cell-type-specific expression of ligand/receptor and cell-adhesion molecules is a fundamental mechanism through which neurons regulate connectivity. Here, we...
The cell-type-specific expression of ligand/receptor and cell-adhesion molecules is a fundamental mechanism through which neurons regulate connectivity. Here, we determine a functional relevance of the long-established mutually exclusive expression of the receptor tyrosine kinase Kit and the trans-membrane protein Kit Ligand by discrete populations of neurons in the mammalian brain. Kit is enriched in molecular layer interneurons (MLIs) of the cerebellar cortex (i.e., stellate and basket cells), while cerebellar Kit Ligand is selectively expressed by a target of their inhibition, Purkinje cells (PCs). By in vivo genetic manipulation spanning embryonic development through adulthood, we demonstrate that PC Kit Ligand and MLI Kit are required for, and capable of driving changes in, the inhibition of PCs. Collectively, these works in mice demonstrate that the Kit Ligand/Kit receptor dyad sustains mammalian central synapse function and suggest a rationale for the affiliation of Kit mutation with neurodevelopmental disorders.
Topics: Mice; Animals; Purkinje Cells; Stem Cell Factor; Cerebellum; Cerebellar Cortex; Interneurons; Receptor Protein-Tyrosine Kinases; Mammals
PubMed: 38536959
DOI: 10.7554/eLife.89792 -
Journal of Environmental Sciences... Aug 2024Tetrabromobisphenol A (TBBPA) is a widely used brominated flame retardant. There is evidence showing that TBBPA can exert thyroid disrupting effects in mammals, but...
Tetrabromobisphenol A (TBBPA) is a widely used brominated flame retardant. There is evidence showing that TBBPA can exert thyroid disrupting effects in mammals, but different results were also reported, along with inconsistent reports regarding its neurotoxicity. Here, we investigated thyroid disrupting effects and neurotoxicity of TBBPA (5, 50, 500 µg/(kg·day)) to male mice following maternal and direct exposure through drinking water, with the anti-thyroid drug propylthiouracil (PTU) as the positive control. On postnatal day (PND) 15, we expectedly observed severe thyroid compensatory hyperplasia and cerebellar developmental retardation in PTU-treated pups. The highest dose of TBBPA also caused thyroid histological alteration but had no effects on cerebellar development in terms of Purkinje cell morphology and the thickness of the internal granular layer and the molecular layer of the cerebellum. During puberty and adulthood, the thyroid morphological alterations became more pronounced in the TBBPA-treated animals, accompanied by decreased serum thyroid hormone levels. Furthermore, the 50 and 500 µg/(kg·day) TBBPA groups showed a significant decrease in the serum level of serotonin, a neurotransmitter associated with anxiety behaviors. Correspondingly, the highest dose group displayed anxiety-like behaviors in the elevated plus-maze test on PND 35, but this neurobehavioral alteration disappeared on PND 56. Moreover, no changes in neurobehavioral parameters tested were found in TBBPA-treated animals at puberty and adulthood. Altogether, all observations show that TBBPA can exert thyroid disrupting effects but has little overt impact on brain development and neurobehaviors in mice, suggesting that thyroid disruption does not necessarily cause overtly adverse neurodevelopmental outcomes.
Topics: Mice; Animals; Male; Thyroid Gland; Polybrominated Biphenyls; Brain; Flame Retardants; Mammals
PubMed: 38527875
DOI: 10.1016/j.jes.2023.10.028