Fear and anxiety have proven to be essential during the evolutionary process. However, the mechanisms involved in recognizing and categorizing threat probability (i.e. low to high) to elicit the appropriate defensive behavior has yet to be determined. In this study we investigated the cerebellar contribution in evoking appropriate defensive escape behavior using a purely cerebellar, neurodegenerative mouse model for spinocerebellar ataxia type 6 (SCA6) which is caused by an expanded CAG repeat in exon 47 of the P/Q type calcium channel α1A subunit. These mice overexpress the carboxy-terminus of the P/Q type calcium channel containing an expanded 27 CAG repeat specifically in cerebellar Purkinje cells (CT-longQ27PC). We found that our CT-longQ27PC mice exhibit anxiolytic behavior in the open field, elevated plus maze and light/dark place preference tests which could be recovered with more threatening conditions such as brighter lighting, meowing sounds and an ultrasound repellent. Their innate fear to find safety in the Barnes maze and visual cliff tests was also diminished with subsequent trials which could be partially recovered with an ultrasound repellent in the Barnes maze. However, under higher threat conditions such as in the light/dark place preference with ultrasound repellent and in the looming tests, CT-longQ27PC mice responded with higher defensive escape behaviors as controls. Moreover, CT-longQ27PC mice displayed increased levels of CT-labeled aggregates compared to controls. Together these data suggest that cerebellar degeneration by overexpression of CT-longQ27PC is sufficient to impair defensive escape responses in those mice.
Increasing evidence suggests that astrocytes play an important role in the progression of Parkinson's disease (PD). Previous studies on our parkin knockout mouse demonstrated a higher accumulation of damaged mitochondria in astrocytes than in surrounding dopaminergic (DA) neurons, suggesting that Parkin plays a crucial role regarding their interaction during PD pathogenesis. In the current study, we examined primary mesencephalic astrocytes and neurons in a direct co‐culture system and discovered that the parkin deletion causes an impaired differentiation of mesencephalic neurons. This effect required the parkin mutation in astrocytes as well as in neurons. In Valinomycin‐treated parkin‐deficient astrocytes, ubiquitination of Mitofusin 2 was abolished, whereas there was no significant degradation of the outer mitochondrial membrane protein Tom70. This result may explain the accumulation of damaged mitochondria in parkin‐deficient astrocytes. We examined differential gene expression in the substantia nigra region of our parkin‐KO mouse by RNA sequencing and identified an upregulation of the endoplasmic reticulum (ER) Ca2+‐binding protein reticulocalbin 1 (RCN1) expression, which was validated using qPCR. Immunostaining of the SN brain region revealed RCN1 expression mainly in astrocytes. Our subcellular fractionation of brain extract has shown that RCN1 is located in the ER and in mitochondria‐associated membranes (MAM). Moreover, a loss of Parkin function reduced ATP‐stimulated calcium‐release in ER mesencephalic astrocytes that could be attenuated by siRNA‐mediated RCN1 knockdown. Our results indicate that RCN1 plays an important role in ER‐associated calcium dyshomeostasis caused by the loss of Parkin function in mesencephalic astrocytes, thereby highlighting the relevance of astrocyte function in PD pathomechanisms.
Background: Parkinson’s disease is a neurodegenerative disease caused by the loss of dopaminergic neurons in the substantia nigra pars compacta. Among the first identified causes of autosomal recessive Parkinson’s disease were mutations in the parkin gene. Independently, we and other groups have developed various parkin knockout mice, and none displayed dopaminergic degeneration in the substantia nigra. Interestingly, dopaminergic degeneration in the substantia nigra has been reported in a parkin knockout line (exon 3 deletion) carrying an additional mutation (D257A) in the mitochondrial DNA polymerase (polg) gene (mutator). The mutator mice show accelerated mutation rates in mitochondrial DNA resulting in a premature-aging phenotype. Methods: To verify this finding, we crossed our parkin-deficient mice with the mutator mice, and characterized phenotypic changes of the parkin/mutator double mutant mice up to one year of age. We examined their locomotion and motor coordination behaviors by using the open field, the rotarod, and the pole test, subsequently investigating their nigrostriatal axis by counting TH-positive cells in every tenth section throughout the entire substantia nigra pas compacta and their termini in the striatum. Results: The double mutants did not display additional deficiencies in locomotion in our behavior tests. We could also not detect dopaminergic neurodegeneration in the substantia nigra pars compacta of aged double mutants measured by levels of tyrosine hydroxylase positive neurons in the substantia nigra pars compacta as well as in striatal terminals. Conclusion: Our results do not support the hypothesis that the polgD257A mutation contributes to the age-related vulnerability of dopaminergic neurons in parkin-deficient mice. Keywords: Parkin, neurodegeneration, polgD257A, mutator, substantia nigra
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