ZFIN Figure: Esancy et al., 2023, Fig. 6

Fig. 6

The 5HT2B, sigma-1, and melanocortin-4 receptors do not appear to mediate the effects of AS1. A Thermal preference assay (28.5 °C vs 37.5 °C) with the 5HT2B agonist BW723C86. All AS1-treated fish significantly prefer the 37.5 °C half of the arena, regardless of whether BW723C86 was co-applied. All conditions that did not receive AS1 significantly chose the 28.5 °C side. At no concentration did BW723C86 influence AS1-induced attraction towards noxious heat. N = 46 fish for vehicle-treated controls, 43 for 1 μM BW723C86, 53 for 5 μM BW723C86, 53 for 10 μM BW723C86, 56 for 5 μM AS1, 50 for 1 μM BW723C86 + 5 μM AS1, 52 for 5 μM BW723C86 + 5 μM AS1, and 59 for 10 μM BW723C86 + 5 μM AS1. B Thermal preference assay (28.5 °C vs 37.5 °C) with the 5HT2B antagonist LY266097. All AS1-treated fish significantly prefer the 37.5 °C half of the arena, regardless of whether LY266097 was co-applied. All conditions that did not receive AS1 significantly chose the 28.5 °C side. At no concentration did LY266097 influence AS1-induced attraction towards noxious heat. N = 58 larvae for the DMSO condition, 57 for 1 μM LY266097, 50 for 5 μM LY266097, 50 for 10 μM LY266097, 59 for 5 μM AS1, 61 for 1 μM LY266097 + 5 μM AS1, 59 for the 5 μM LY266097 + 5 μM AS1, and 56 for the 10 μM LY266097 + 5 μM AS1. C Thermal preference assay (28.5 °C vs 37.5 °C) with the Sigma-1 receptor agonist PRE-084. All AS1-treated fish significantly prefer the 37.5 °C half of the arena, regardless of whether PRE-084 was co-applied. All conditions that did not receive AS1 significantly chose the 28.5 °C side. At no concentration did PRE-084 influence AS1-induced attraction towards noxious heat, despite potentiating preference for the 28.5 °C zone. N = 90 fish for the DMSO condition, 27 for 1 μM PRE-084, 31 for 5 μM PRE-084, 25 for 10 μM PRE-084, 37 for 5 μM AS1, 40 for 1 μM PRE-084 + 5 μM AS1, 43 for 5 μM PRE-084 + 5 μM AS1, and 46 for 10 μM PRE-084 + 5 μM AS1. D Thermal preference assay (28.5 °C vs 37.5 °C) with the Sigma-1 receptor antagonist BD1063. All AS1-treated fish significantly prefer the 37.5 °C half of the arena, regardless of whether BD1063 was co-applied. All conditions that did not receive AS1 significantly chose the 28.5 °C side. At no concentration did BD1063 influence AS1-induced attraction towards noxious heat. N = 30 for the DMSO condition, 26 for 1 μM BD1063, 30 for 5 μM BD1063, 28 for 10 μM BD1063, 27 for 5 μM AS1, 30 for 1 μM BD1063 + 5 μM AS1, 29 for 5 μM BD1063 + 5 μM AS1, and 28 for 10 μM BD1063 + 5 μM AS1. E Thermal preference assay (28.5 °C vs 37.5 °C) with the sigma-1 receptor agonist cutamesine (SA 4503). All AS1-treated fish significantly preferred the 37.5 °C half of the arena, regardless of whether cutamesine was co-applied. All conditions that did not receive AS1 significantly chose the 28.5 °C side. At no concentration did cutamesine have any effect upon temperature preference when applied alone, nor did it significantly reduce AS1-induced attraction to noxious heat. N = 30 fish for the DMSO condition, 29 for 1 μM cutamesine, 25 for 5 μM cutamesine, 54 for 10 μM cutamesine, 52 for 5 μM AS1, 49 for 1 μM cutamesine + 5 μM AS1, 52 for 5 μM + 5 μM AS1, and 58 for 10 μM cutamesine + 5 μM AS1. F Thermal preference assay (28.5 °C vs 37.5 °C) with the melanocortin 4 receptor antagonist ML00253764. All AS1-treated fish significantly prefer the 37.5 °C half of the arena, regardless of whether ML00253764 was co-applied. All conditions that did not receive AS1 significantly chose the 28.5 °C side. At no concentration did ML00253764 reduce AS1-induced attraction towards noxious heat, even though one concentration (5 μM) did slightly potentiate preference towards the 28.5 °C zone. N = 30 fish for the DMSO condition, 31 for 1 μM ML00253764, 28 for 5 μM ML00253764, 31 for 10 μM ML00253764, 31 for 5 μM AS1, 32 for 1 μM ML00253764 + 5 μM AS1, 32 for 5 μM ML00253764 + 5 μM AS1, and 31 for 10 μM ML00253764 + 5 μM AS1. * p < 0.05, ** p < 0.01, *** p < 0.001. Two-way ANOVA with Sidak’s multiple comparisons test used for A; two-way ANOVA with Tukey’s multiple comparisons test used for B–F. To determine if fish were significantly choosing one side of the arena over the other, a one-sample t test was performed with a hypothetical mean of 50%

Expression Data

Expression Detail

Antibody Labeling

Phenotype Data

Phenotype Detail

Acknowledgments

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