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Title: Investigating the effects of a point mutation in the TRPC3 channel, cause of cerebellar ataxia in Moonwalker mice, on the Purkinje cells in mice
Author: Dulneva, Anna
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2012
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The Moonwalker (Mwk) mouse is a mouse model of cerebellar ataxia that harbours a point mutation in the Trpc3 gene. TRPC3 is a non-selective cation channel, most highly expressed in the Purkinje cells of the cerebellum. The gain-of-function mutation in the TRPC3 protein affects the development of Purkinje cell dendrites by reducing their branching, and also leads to abnormal motor coordination and cerebellar ataxia in Mwk mice at the age of 3 weeks. The aim of this thesis was to determine how the mutation in the TRPC3 channel results in the observed pathology. Proper function of the TRPC3 channel relies on its interaction with other proteins, hence we investigated binding partners of TRPC3. The study revealed PI synthase and CaMKIV as novel interaction partners of TRPC3. PI synthase is implicated in the upstream signalling events leading to TRPC3 activation, whereas CaMKIV is activated by Ca2+, possibly due to TRPC3 activation. We have identified alterations in phosphorylation of several key Ca2+ signalling proteins (CaMKII, CaMKIV, CREB and ERK), which indicates that there are changes in Ca2+ homeostasis in Mwk cerebella. Down-regulation of CaMKIV and up-regulation of CREB phosphorylation occurs as early as P21, which indicates that their abnormal activity could contribute to the Mwk phenotype. Microarray analysis comparing wild-type and Mwk Purkinje cells has revealed gene expression changes, which are likely due to abnormal Ca2+ signalling. Genes Ipo5, Opn3 and Sv2c are up-regulated at P11; Car2 and Stk17b are down-regulated at P14; and Cntn3 is up-regulated at P18 in Mwk Purkinje cells. High quality RNA from Purkinje cells was extracted using an optimised laser-capture microdissection method. Work on the Mwk mice points to the importance of TRPC3 activity for the proper development of Purkinje cell dendrites and depicts TRPC3 as a possible target for cerebellar ataxia treatment.
Supervisor: Davies, Kay Sponsor: Medical Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Life Sciences ; Biology ; Cell Biology (see also Plant sciences) ; Genetics (life sciences) ; Physiology and anatomy ; neuroscience ; brain ; cerebellum