Fig. 11

The D2 receptor antagonist sulpiride does not replicate or reverse the effects of AS1 in multiple choice assays. A Temperature choice assay (28.5 °C vs 37.5 °C) with various concentrations of sulpiride. All AS1-treated fish significantly preferred the 37.5 °C side, whereas fish treated only with sulpiride or vehicle solution significantly chose the 28.5 °C side. N = 28 larvae for 1% DMSO, 37 for 1 μM sulpiride, 41 for 5 μM sulpiride, 44 for 10 μM sulpiride, 30 for 30 μM sulpiride, 58 for 5 μM AS1, 49 for 1 μM sulpiride + 5 μM AS1, 52 for 5 μM sulpiride + 5 μM AS1, 54 for 10 μM sulpiride + 5 μM AS1, and 62 for 30 μM sulpiride + 5 μM AS1. B Swimming velocity of the fish in the experiment shown in A. AS1-treated fish have significantly lower velocities at 37.5 °C than at 28.5 °C at all concentrations of sulpiride tested apart from 1 μM. Additionally, AS1-treated fish have significantly lower swimming velocities at 37.5 °C than non-AS1 treated fish at all concentrations of sulpiride tested. * 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 AITC aversion assay with various concentrations of sulpiride. None of the sulpiride concentrations tested elicited a change in AITC aversion. N = 174 fish for DMSO, 36 fish for 1 μM sulpiride, 34 fish for 5 μM sulpiride, 177 fish for 10 μM sulpiride, and 33 fish for 30 μM sulpiride. D AITC aversion assay for various concentrations of sulpiride + 5 μM AS1, with DMSO control fish shown in C. At no concentration tested does sulpiride attenuate AS1-induced attraction to AITC. N = 169 fish for 5 μM AS1, 32 fish for 1 μM sulpiride + 5 μM AS1, 32 fish for 5 μM sulpiride + 5 μM AS1, 172 fish for 10 μM sulpiride + 5 μM AS1, and 33 fish for 30 μM sulpiride + 5 μM AS1. * represent the significant difference of experimental traces from the DMSO-treated control fish at each indicated time point. E The distance of each individual larva from the AITC-infused agarose at the final time point (t = 840 s) of the experiment shown in C and D. Whereas control and all sulpiride-only condition fish localize farther from the AITC source, all AS1-treated fish congregate closer to the AITC source. * presented directly over columns represent the significant difference between the DMSO-only control condition, and between other columns when indicated by lines. F Light/dark preference assay. No tested concentration of sulpiride was capable of reversing AS1-induced preference for the dark. N = 39 fish for 0 μM sulpiride, 40 for 10 μM sulpiride, 40 for 30 μM sulpiride, 77 for 0 μM sulpiride + 5 μM AS1, 79 for 10 μM sulpiride + 5 μM AS1, and 83 for 30 μM sulpiride + 5 μM AS1. */^#/⁺ p < 0.05, **/^##/⁺⁺ p < 0.01, ***/^###/⁺⁺⁺ p < 0.001. Two-way ANOVA with Tukey’s multiple comparisons test used in A–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