Pharmacometabolomic Signature of Ataxia SCA1 Mouse Model and Lithium Effects

<div><p>We have shown that lithium treatment improves motor coordination in a spinocerebellar ataxia type 1 (SCA1) disease mouse model (<i>Sca1^154Q/+</i>). To learn more about disease pathogenesis and molecular contributions to the neuroprotective effects of lithium, we investigated metabolomic profiles of cerebellar tissue and plasma from SCA1-model treated and untreated mice. Metabolomic analyses of wild-type and <i>Sca1^154Q/+</i> mice, with and without lithium treatment, were performed using gas chromatography time-of-flight mass spectrometry and BinBase mass spectral annotations. We detected 416 metabolites, of which 130 were identified. We observed specific metabolic perturbations in <i>Sca1^154Q/+</i> mice and major effects of lithium on metabolism, centrally and peripherally. Compared to wild-type, <i>Sca1^154Q/+</i> cerebella metabolic profile revealed changes in glucose, lipids, and metabolites of the tricarboxylic acid cycle and purines. Fewer metabolic differences were noted in <i>Sca1^154Q/+</i> mouse plasma versus wild-type. In both genotypes, the major lithium responses in cerebellum involved energy metabolism, purines, unsaturated free fatty acids, and aromatic and sulphur-containing amino acids. The largest metabolic difference with lithium was a 10-fold increase in ascorbate levels in wild-type cerebella (p<0.002), with lower threonate levels, a major ascorbate catabolite. In contrast, <i>Sca1^154Q/+</i> mice that received lithium showed no elevated cerebellar ascorbate levels. Our data emphasize that lithium regulates a variety of metabolic pathways, including purine, oxidative stress and energy production pathways. The purine metabolite level, reduced in the <i>Sca1^154Q/+</i> mice and restored upon lithium treatment, might relate to lithium neuroprotective properties.</p></div

Effect of lithium treatment on cerebellum metabolome.

<p>Metabolic network of wild-type and <i>Sca1^154Q/+</i> cerebellum phenotypes. <b>A.</b> Wild-type mice. <b>B.</b> SCA1 knock-in mice. Red nodes: Increased metabolite levels under Lithium treatment; blue nodes: decreased levels. Node shades indicate ANOVA significance levels, node size reflect differences in magnitude of regulation. Red lines: reactant pair relationships obtained from the KEGG reaction pair database. Yellow solid lines: chemical similarity >0.5 Tanimoto score (Tanimoto scores range between 0 to 1, where 1 reflects identical structures). Yellow broken lines: chemically closest structure at <0.5 Tanimoto scores. Green circles group significant compounds that changed only in the Wild-type genotype. Orange circles group significant compounds that changed in both genotypes.</p

Effect of Lithium treatment on blood plasma metabolic profile: Significantly different metabolites comparing Lithium treatment versus control conditions.

<p>Note: Bold indicates statistical significance. One-way ANOVA performed separately for wild-type and <i>Sca1^154Q/+</i> mice (see supplemental <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070610#pone-0070610-g002" target="_blank">Figure 2</a> for box-whisker plots). Abbreviation: Li, Lithium.</p

Box-and-whisker plots: genotype-dependent metabolites in cerebellum tissue with significant differences between lithium and controls (<i>p</i><0.05).

<p><b>A.</b> Box-whisker plots for selected significantly regulated metabolites. <b>B.</b> Box-whisker plots for significantly regulated metabolites of purine metabolism pathway. The whiskers encompass 1.5 of the interquartile range (IQR). Median value is indicated with a line in the box. Boxes are filled in color (dark grey: SCA1 knock-in; light grey: wild-type) when the samples are statistically different between the two lithium treatments. Abbreviations: Ctl, Control; KI, SCA1 knock-in; Li, Lithium; WT, Wild-type.</p

Genotype effect on metabolic profiles: Significantly different metabolites comparing wild-type versus <i>Sca1^154Q/+</i> mice under control conditions.

<p>Notes: Bold indicates statistical significance. One-way analysis of variance performed separately for cerebellum and blood plasma (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070610#pone-0070610-g002" target="_blank">Figure 2</a> and supplemental <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070610#pone-0070610-g001" target="_blank">Figure 1</a> for box-whisker plots). Abbreviation: NIST, National Institute of Standards and Technology.</p

Lithium treatment effect on blood plasma metabolome. Metabolic network of wild-type and <i>Sca1^154Q/+</i> plasma phenotypes.

<p><b>A.</b> Wild-type mice. <b>B.</b> SCA1 knock-in mice. Red nodes: Increased metabolite levels under Lithium treatment; blue nodes: decreased levels. Node shades indicate ANOVA significance levels, node size reflect differences in magnitude of regulation. Red lines: reactant pair relationships obtained from the KEGG reaction pair database. Yellow solid lines: chemical similarity >0.5 Tanimoto score (Tanimoto scores range between 0 to 1, where 1 reflects identical structures). Yellow broken lines: chemically closest structure at <0.5 Tanimoto scores.</p

Level of metabolites in SCA1 mice relative to wild-type non-treated mice: the treatment effect of lithium on impaired metabolites in SCA1 knock-in mice.

<p>Note: All values are relative to wild-type untreated animals. Abbreviations: Avg, Average; NIST, National Institute of Standards and Technology; SD, Standard deviation.</p

Effect of introducing the <i>Sca1^154Q/+</i> gene into the wild-type genetic background for plasma and cerebellum.

<p>Individual box-whisker plots for selected significantly regulated metabolites. The whiskers encompass 1.5 of the interquartile range (IQR). Median value is indicated with a line in the box. The confidence diamonds indicate average values when the two samples are statistically different (colored boxplots, red for blood and grey for brain). Abbreviations: KI, SCA1 knock-in; WT, Wild-type.</p

Supervised multivariate Partial Least Square separation of metabolic phenotypes.

<p><b>A.</b> Differences between plasma and cerebellum (vector v1) and between SCA1 and wild-type under control conditions (separated by vectors v3 and v4). <b>B.</b> Differences between Lithium treated and control conditions in cerebellum (vector v1) and between SCA1 and wild-type (separated by vectors v2 and v3). Abbreviations: Cereb, Cerebellum; Ctrl, Control; Li, Lithium; WT, Wild-type.</p

Overall study design with the number of tested animals.

<p>Overall study design with the number of tested animals.</p