Depression-like behavior in the forced swim test.

<p>Boxplots show the results of the depression-like behavior in the forced swim test (FST). Young naive (3 month) α-GAL A deficient (Fabry KO) and wildtype (WT) male mice were investigated. No difference was found between the genotypes. Fabry KO: young (3 months; 10 male). WT: young (3 months; 10 male).</p

Anxiety-like behavior in the elevated plus maze test.

<p>Boxplots show the results of anxiety-like behavior in the elevated plus maze test (EPM). Young (3 months) and old (≥18 months) α-GAL A deficient (Fabry KO) and wildtype littermate (WT) male mice were investigated in the naïve state and after i.pl. complete Freund`s adjuvant (CFA) injection into the right hind paw. A) Time spent in the open arms was not different between naïve genotypes and age-groups, except for young WT mice compared to young Fabry KO mice (p<0.05). I.pl. CFA injection led to a decrease of time spent in the open arms in young Fabry KO mice compared to young naïve Fabry KO mice (p<0.05). After CFA injection time spent in open arms was also lower in young Fabry KO mice than in young WT mice (p<0.05). In old mice CFA i.pl. reduced time spent in open arms in Fabry KO (p<0.01) and WT mice (p<0.05). B) There was no intergroup difference in the number of entries into the open arms in naïve and CFA treated mice except for old CFA treated Fabry KO mice, which entered less often than old naïve Fabry KO mice (p<0.05). C) No difference was found in young age-groups for time spent in closed arms. Old CFA treated mice of both genotypes spent more time in the closed arms compared to old naive mice (Fabry KO: p<0.01; WT: p<0.001). D). There was no difference in entries into the closed arms between age-groups and genotypes in the naïve state. After CFA injection young Fabry KO mice more often entered the closed arms than young WT mice (p<0.05). Also, old CFA treated Fabry KO mice had more entries into the closed arm than old naïve mice (p<0.05). Fabry KO: young (3 months; naïve: 10 male; CFA: 6 male), old (≥18 months; naïve: 10 male; CFA: 6 male). WT: young (3 months; naïve: 10 male; CFA: 6 male), old (≥18 months; naïve: 10 male; CFA: 6 male). *p<0.05, **p<0.01.</p

Experimental design.

<p>Timelines show the test sequences at baseline and after i.pl. injection of complete Freund`s adjuvant (CFA) into the right hind paw. Anxiety- and depression-like behavior was investigated in young (3 months) and old (≥18 months) α-GAL A deficient (Fabry KO) and wildtype littermate (WT) male mice in the naïve state on three consecutive days of the first test week and learning behavior on five days of the second test week (A). Anxiety-like behavior was additionally tested one and 48 hours after CFA injection when pain maximum was assumed [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0180601#pone.0180601.ref022" target="_blank">22</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0180601#pone.0180601.ref023" target="_blank">23</a>] (B). Abbreviations: EPM = elevated plus maze; FST = forced swim test; LDB = light-dark box; MWM = morris water maze; OF = open field test.</p

Anxiety-like behavior in the light-dark box test.

<p>Boxplots show the results of anxiety-like behavior in the light-dark box test (LDB). Young (3 months) and old (≥18 months) old α-GAL A deficient (Fabry KO) and wildtype littermate (WT) male mice were investigated in the naïve state and after i.pl. complete Freund`s adjuvant (CFA) injection into the right hind paw. There was no intergroup difference between genotypes, age-groups, and treatment for the time spent in the light box (A), entries into the light box (B), time spent in the dark box (C) and entries into the dark box (D) except for the comparison of young naïve WT mice with young CFA treated WT mice (p<0.05 each). Fabry KO: young (3 months; naïve: 10 male; CFA: 6 male), old (≥18 months; naïve: 10 male; CFA: 6 male). WT: young (3 months; naïve: 10 male; CFA: 6 male), old (≥18 months; naïve: 10 male; CFA: 6 male).</p

Anxiety-like behavior in the open field test.

<p>Boxplots show the results of the anxiety-like behavior in the open field test (OF). Young (3 months) and old (≥18 months) old α-GAL A deficient (Fabry KO) and wildtype littermate (WT) mice were investigated in the naïve state and after i.pl. complete Freund`s adjuvant (CFA) injection into the right hind paw. No difference between genotypes, age- and treatment groups was found in the time spent in the center zone (A), except for young WT mice compared to young Fabry KO mice in the naïve state (p<0.05). Total distance travelled (B) was not different between genotypes, age- and treatment groups except for young Fabry KO mice in the naïve state compared with old Fabry KO mice (p<0.01). Fabry KO: young (3 months; naïve: 10 male; CFA: 6 male), old (≥18 months; naïve: 10 male, CFA: 6 male). WT: young (3 months; naïve: 10 male; CFA: 6 male), old (≥18 months;naïve: 10 male; CFA: 6 male). *p<0.05, **p<0.01.</p

Learning behavior in the Morris water maze test.

<p>Bar graphs show the results of the learning behavior in the Morris water maze test (MWM). Naive young (3 months) and old (≥18 months) α-GAL deficient (Fabry KO) and wildtype littermate (WT) male mice were investigated. Test duration on training days displayed not relevant differences between genotypes and age-groups and continuously decreased from training day one to four. Fabry KO: young (3 months; 10 male), old (≥18 months; 10 male). WT: young (3 months; 10 male), old (≥18 months; 10 male). *p<0.05, **p<0.01.</p

Cytokine gene expression before and after CCI.

<p>The box- and whisker plots illustrate the relative gene expression of pro- and anti-inflammatory cytokines in the ipsilateral sciatic nerve (A), and ipsilateral lumbar spinal cord (B) 28 days after CCI. A) CCI leads to an increase in TNF and IL-1ß gene expression in the ipsilateral sciatic nerve. The increase in IL-1ß gene expression is higher in IL-4 ko mice compared to wt mice. IL-10 gene expression increases only in IL-4 ko mice. B) CCI leads to an increase in TNF, IL-1ß, IL-10, and IL-13 gene expression only in IL-4 ko mice; this increase is higher compared to wt mice after CCI (TNF: p = 0.002; IL-1ß: p<0.001; IL-10: p = 0.011; IL-13: p = 0.026). Asterisk: *p<0.05, **p<0.01, ***p<0.001.</p

Analgesic effect of morphine in IL-4 ko and wt mice.

<p>Paw withdrawal latencies to thermal stimuli and withdrawal thresholds to von Frey hairs in wt and IL-4 ko mice after CCI (A, B) and additional morphine treatment (C, D). CCI leads to thermal (A) and mechanical (B) hypersensitivity lasting up to day 28 after CCI. Mice received morphine i.p. (abbreviated as M in the graphs) at day seven post-surgery. Only IL-4 ko mice showed elevation of mechanical withdrawal thresholds and prolongation thermal withdrawal latencies at 2 h after morphine i.p. (A, B). Asterisk: *p<0.05, **p<0.01, ***p<0.001.</p

Boxplots show the intraepidermal nerve fiber density (IENFD) in affected versus unaffected skin of patients with postherpetic neuralgia (PHN).

<p>In affected skin IENFD is lower than in corresponding (contralateral) unaffected skin (*p<0.05). The horizontal black line in the box marks the median value.</p

Behavioral tests in naïve IL-4 ko and wt mice.

<p>The bars illustrate the results of the behavioral tests in naïve wt and IL-4 ko mice. A) IL-4 ko mice do not differ from wt mice in withdrawal latencies to heat. B) IL-4 ko mice have reduced paw withdrawal thresholds to mechanical stimulation with von-Frey filaments (***p<0.001). IL-4 ko mice do not differ from wt mice in paw withdrawal time to acetone (C), and in withdrawal latencies of the hind limb upon pressure to the gastrocnemius muscle (D).</p