Fig. 12

The D2 receptor agonist sumanirole maleate does not replicate or reverse the effects of AS1 in multiple assays. A Temperature choice assay (28.5 °C vs 37.5 °C) with various concentrations of sumanirole. All AS1-treated fish significantly preferred the 37.5 °C side regardless of sumanirole concentration, whereas fish treated only with sumanirole or vehicle solution significantly chose the 28.5 °C side. N = 44 fish for DMSO, 41 for 1 μM sumanirole, 59 for 5 μM sumanirole, 44 for 10 μM sumanirole, 48 for 30 μM sumanirole, 28 for 5 μM AS1, 46 for 1 μM sumanirole + 5 μM AS1, 50 for 5 μM sumanirole + 5 μM AS1, 37 for 10 μM sumanirole + 5 μM AS1, and 53 for 30 μM sumanirole + 5 μM AS1. B Velocity data for fish in the experiment shown in A. AS1 treatment significantly increases swimming velocity in the 28.5 °C zone and decreases swimming velocity in the 37.5 °C zone, regardless of the concentration of sumanirole co-applied to the zebrafish. Sumanirole alone has no effect upon swimming velocity in either zone. * denotes significant differences in swimming velocities between the 28.5 and 37.5 °C zones for the same group of fish, + denotes significant difference from the 0 μM AS1 37.5 °C swimming velocity, and # denotes significant difference from the 0 μM AS1 28.5 °C swimming velocity. C, D AITC aversion assay. Sumanirole alone does not affect aversion to this noxious chemical stimulus, but at higher concentrations partially attenuates AS1-induced attraction to AITC. N = 68 for 0 μM sumanirole, 36 for 1 μM sumanirole, 36 for 5 μM sumanirole, 61 for 10 μM sumanirole, 63 for 30 μM sumanirole, 64 for 0 μM sumanirole + 2.5 μM AS1, 34 for 1 μM sumanirole + 2.5 μM AS1, 34 for 5 μM sumanirole + 2.5 μM AS1, 61 for 10 μM sumanirole + 2.5 μM AS1, and 63 for 30 μM sumanirole + 2.5 μM AS1. * represent the significant difference of experimental traces from the DMSO-treated control fish at each indicated time point. # represent the significant difference of experimental traces compared to AS1-only control fish at the indicated time points. E Distance of each individual larva from the AITC stimulus at the final time point (840 s) from the experiment shown in C, D. * presented directly over columns represent the significant difference between the DMSO-only control condition, and between other columns when indicated by lines. F Phototaxis assay. AS1-treated fish significantly chose the dark half of the arena regardless of what concentration of sumanirole was co-applied, whereas vehicle and sumanirole-only treated fish significantly chose the light half of the arena. N = 23 fish for 0 μM sumanirole, 37 for 1 μM sumanirole, 41 for 5 μM sumanirole, 40 for 10 μM sumanirole, 40 for 0 μM sumanirole + 2.5 μM AS1, 46 for 1 μM sumanirole + 2.5 μM AS1, 43 for 5 μM sumanirole + 2.5 μM AS1, and 40 for 10 μM sumanirole + 2.5 μM AS1. */^#/⁺ p < 0.05, **/^##/⁺⁺ p < 0.01, ***/^###/⁺⁺⁺ p < 0.001. Two-way ANOVA with Sidak’s multiple comparisons test used in A, B. Two-way ANOVA with Tukey’s multiple comparisons test used in C, D, F. One-way ANOVA with Tukey’s multiple comparisons test used in E. For all temperature and light/dark choice experiments, a one-sample t test was performed with a hypothetical mean of 50% to determine if fish were significantly choosing one side of the arena over the other