Research ArticleNEUROSCIENCE

Mass spectrometry imaging identifies abnormally elevated brain l-DOPA levels and extrastriatal monoaminergic dysregulation in l-DOPA–induced dyskinesia

See allHide authors and affiliations

Science Advances  06 Jan 2021:
Vol. 7, no. 2, eabe5948
DOI: 10.1126/sciadv.abe5948
  • Fig. 1 Tissue distribution of DA in basal ganglia structures and regional quantitation of DA.

    DA distribution in basal ganglia structures in (A) control, MPTP, non-LID, and LID tissue sections, scaled to 0 to 100% of maximum intensity and (B) to 0 to 20% of maximum intensity. Lateral resolution, 80 μm; all images are log-transformed and RMS-normalized; scale bars, 4 mm. Coronal level, −4 mm ac. (C) Regional quantitation of DA; y axis shows log2AUC of DA. n = 6 for Ctrl, non-LID, and LID; n = 5 for MPTP. A two-way ANOVA followed by a Tukey’s multiple comparisons test was performed. *P < 0.05; **P < 0.01; ***P < 0.001. Amyg, amygdala; Cd, caudate; Ctrl, control; GPe, external globus pallidus; GPi, internal globus pallidus; Hip, hippocampus; Hy, hypothalamus; PrG, precentral gyrus; Put, putamen; StT, nucleus of stria terminalis; Thal, thalamus.

  • Fig. 2 Regional alterations of metabolites in non-LID versus LID.

    (A) Nissl-stained macaque brain tissue section at −4 mm ac with annotated brain regions. (B) Regional heat maps color-coded according to log2FC between non-LID and LID. Blue indicates elevated levels in non-LID, red indicates higher levels in LID, and white indicates no change. (C) Regional quantitation of metabolites in non-LID and LID. Log2-transformed AUC values for each sample are shown. Blue, non-LID; red, LID. Crossbar and error bars show means and SDs. A Shapiro-Wilk normality test was used to determine normal distribution of samples; a Student’s t test was used when passing normality test; otherwise, a Mann-Whitney U test was used. *P < 0.05; **P < 0.01; ***P < 0.001. For all metabolites in all regions, n = 6, except n = 5 for PoG in LID group, n = 3 for PoG in non-LID group, and n = 5 for Hip in non-LID. Statistical results are summarized in table S2. ac, anterior commissure; ACgG, anterior cingulate gyrus; Clau, claustrum; Ent, entorhinal area; Ins, insula; ITG, inferior temporal gyrus; MTG, middle temporal gyrus; PoG, postcentral gyrus; STG, superior temporal gyrus.

  • Fig. 3 MALDI-MS images of neurotransmitters and metabolites in non-LID and LID.

    (A) Nissl-stained macaque brain tissue section at −4 mm ac with annotated brain regions. (B) Catecholaminergic metabolic pathway. All MALDI-MS images are RMS-normalized and log-transformed intensities. l-DOPA, 3-OMD, 3-MT, HVA, and DOPAC are scaled to 0 to 100% of maximum intensity, and DA and NE are scaled to 0 to 10% of maximum intensity. (C) GABA, scaled to 0 to 100% of maximum intensity. (D) 5-HT metabolic pathway: 5-HT scaled to 0 to 15%, 5-HIAL scaled to 0 to 20%, and 5-HIAA scaled to 0 to 50% of maximum intensity. For all images: left section, non-LID; right section, LID; scale bar, 20 mm; lateral resolution, 150 μm. Enzymes involved are annotated by the arrows. COMT, catechol-O-methyl transferase; MAO, monoamine oxidase; AADC, aromatic l-amino acid decarboxylase; DβH, dopamine-β-hydroxylase; AD, aldehyde dehydrogenase.

  • Fig. 4 Correlation of l-DOPA to 3-OMD or DA in non-LID and LID.

    (A) LR of the change in 3-OMD content in response to l-DOPA. (B) LR of the change in DA content in response to l-DOPA. (A and B) Blue, non-LID; red, LID. A significant linear relationship is indicated with *P < 0.05, **P < 0.005, and ***P < 0.001. For all linear models, n = 6 in the LID group, and Cd, Hy, Put, and StT in the non-LID group. For the remaining non-LID regions, n = 5 in Amyg and Thal, n = 4 in Clau, Ctx, GPe, and GPi, n = 3 in Hip in both correlations, n = 4 in Ent in l-DOPA to 3-OMD, and n = 3 in Ent in l-DOPA to DA. (C) PCA model computed from results of LR models in (B); blue, non-LID; red, LID; purple, loadings. (D) PCA model computed from results of LR analysis for change in DA in response to increased l-DOPA in LID only. Regions are color-coded according to hierarchical clustering and grouping structures that respond similarly to changes in l-DOPA content. P-intercept, P value for intercept; P-slope, P value for slope; R-squared, R2; Residual SE, SE of residuals.

  • Fig. 5 DA metabolism in striatum.

    (A) DOPAC/DA ratio (MAO metabolism of DA) and (B) 3-MT/DA ratio (COMT metabolism of DA) in Cd and Put in non-LID (blue) and LID (red). Bars show means, and error bars show SDs (n = 6 for both groups in both regions). A Mann-Whitney U test was used; results are summarized in table S4. *P < 0.05.

  • Fig. 6 Comparison of 5-HT, DA, and NE localization in basal ganglia and hippocampus.

    (A) Nissl-stained section showing basal ganglia structures at −6 mm ac and MALDI-MS images of 5-HT (scaled to 0 to 25%), DA (scaled 0 to 30%), and NE (scaled to 0 to 40%) in non-LID and LID. Lateral resolution, 50 μm. Scale bars, 5 mm. White arrows indicate DA and NE patches that colocalize. (B) Nissl-stained section of Hip at −4 mm ac and MALDI-MS images of 5-HT, DA, and NE (all scaled to 0 to 20%) in control, non-LID, and LID macaque brain. Lateral resolution, 80 μm. Scale bars, 2 mm. For (A) and (B), images are RMS-normalized and log-transformed, and overlays show DA in green and 5-HT in magenta. CA, Cornu ammonis; CAml, molecular layer of the Hip; CArd, stratum radiatum; DGgc, granular layer of the dentate gyrus; DGml, molecular layer of the dentate gyrus; HDG, hilus of the dentate gyrus; so, stratum oriens; pcl, pyramidal cell layer; pre, presubiculum; sub, subiculum; TCd, tail of caudate nucleus.

  • Fig. 7 Mapping of DA throughout cortical areas and layers.

    (A) Nissl-stained coronal tissue section at −4 mm ac illustrating the location of the analyzed cortical area. (B) MS images of DA, 5-HT, and NE in PrG and (C) TG at −4 mm ac. Color intensity scale is 0 to 50% for DA and 5-HT and 0 to 30% for NE. Lateral resolution, 50 μm; scale bar, 1 mm; images are RMS-normalized. The arrows in (B) and (C) show the orientation of the sample pointing toward the outer layers of the cortex. (D) Nissl-stained cortex section overlaid with DA distribution illustrating the three layers of the cortex that were analyzed. (E) Relative quantitation of DA in outer, middle, and deep cortical layers at −4 mm ac in the PrG, Ins, and TG. Bars show means, and error bars show SDs; blue, non-LID; red, LID. Statistics were performed using a two-way ANOVA and a post hoc Sidak’s multiple comparisons test (n = 6 for both groups in all cortical layers). *P < 0.05; **P < 0.01; ***P < 0.001. Two-way ANOVA results are summarized in table S5.

Supplementary Materials

Stay Connected to Science Advances

Navigate This Article