Hypothermia-induced dystonia and abnormal cerebellar activity in a mouse model with a single disease-mutation in the sodium-potassium pump

<div><p>Mutations in the neuron-specific α₃ isoform of the Na⁺/K⁺-ATPase are found in patients suffering from Rapid onset Dystonia Parkinsonism and Alternating Hemiplegia of Childhood, two closely related movement disorders. We show that mice harboring a heterozygous hot spot disease mutation, D801Y (α₃^+/D801Y), suffer abrupt hypothermia-induced dystonia identified by electromyographic recordings. Single-neuron <i>in vivo</i> recordings in awake α₃^+/D801Y mice revealed irregular firing of Purkinje cells and their synaptic targets, the deep cerebellar nuclei neurons, which was further exacerbated during dystonia and evolved into abnormal high-frequency burst-like firing. Biophysically, we show that the D-to-Y mutation abolished pump-mediated Na⁺/K⁺ exchange, but allowed the pumps to bind Na⁺ and become phosphorylated. These findings implicate aberrant cerebellar activity in α₃ isoform-related dystonia and add to the functional understanding of the scarce and severe mutations in the α₃ isoform Na⁺/K⁺-ATPase.</p></div

Twenty-seven genes relevant to lens placode formation and lens morphogenesis show differential expression in Pax6^−/− E9.5 mutated lens placodes.

<p>(A) A list of 27 genes includes a combination of well-characterized genes in lens biology and selected differentially expressed genes in Pax6 null (<i>Sey</i>) cortex <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054507#pone.0054507-Holm1" target="_blank">[45]</a>. The differentially expressed genes in Pax6^−/− E9.5 wild type and mutated lens placodes were identified using the Illumina Mouse6 bead microarrays as described elsewhere <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054507#pone.0054507-Huang1" target="_blank">[96]</a>. Twenty-four of the 27 genes were differentially expressed in at least 50% of experiments. (B) Relative expression levels of <i>Fat4</i>, <i>Trpm3</i>, <i>Pax6</i>, <i>Has2</i>, <i>Efnb2</i>, and <i>Nav1</i> in wild type (WT, black bars) and Pax6^−/− (open bars) lens placode and mutated ectoderm were determined using qRT-PCR as described in Methods.</p

Down-regulation of Kif1b and Snca in Sye/Sey forebrains.

<p>In situ hybridization analysis of E14 wild type (WT) and Pax6^Sey/Sey mouse embryos. Ganglionic eminence, GE.</p

Pax6 regulates expression of Snca.

<p>(A) Identification of Pax6-binding region by ChIP-Chip in lens chromatin and corresponding luciferase reporter constructs for transfection assays. (B) Pax6 regulates <i>Snca</i> promoter/distal region in cultured cells. Transient transfections were performed in P19 embryonic carcinoma and in αTN4-1 lens cell as described in Methods. (C) Prediction of Pax6 binding sites with novel Pax6 DNA binding motifs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054507#pone.0054507-Xie1" target="_blank">[48]</a>. (D) EMSA validation of Pax6 binding to the probes identified by motif 1-1, 1-2 and 3-3. PD/HD, recombinant Pax6 protein containing both Pax6 paired domain (PD) and homeodomain (HD). P6CON, DNA-binding concensus for Pax6 paired domain.</p