Behavioral severity across experimental groups—the top panel presents a heatmap illustrating the severity score for each experimental group, where colors represent the number of behavioral abnormalities observed (green = low, red = high). The bottom panel displays a behavioral presence matrix, indicating the presence (✔) or absence (-) of specific behavioral abnormalities across experimental groups. The severity score was calculated based on the total number of distinct behaviors exhibited within each group (n = 10 per group).

Behavioral severity across experimental groups with pairwise significance—box plot illustrating the distribution of behavioral severity scores across experimental groups. The y-axis represents the severity score, calculated as the total number of distinct behavioral abnormalities observed per group, while the x-axis denotes the different exposure conditions. Each box plot displays the median (horizontal line), interquartile range (box), and data variability (whiskers), with outliers represented as individual points. Statistical significance of pairwise comparisons between groups is indicated by horizontal brackets above the plot, with asterisks denoting significance levels (p < 0.01 (**), p < 0.0001 (****)).

Linear regression of mortality rate over time across experimental groups. The y-axis represents the mortality rate (%) recorded at four-time points (1, 7, 14, and 21 days), while the x-axis denotes the exposure duration. Colored markers (X) indicate observed mortality data for each experimental group while corresponding dashed lines represent the linear regression fit for each condition. Statistical significance was assessed using ANOVA and post hoc comparisons to evaluate differences in mortality trends between exposure groups.

Expression levels of BDNF across experimental groups. The box plot represents the relative expression levels of BDNF in zebrafish (n = six per group) exposed to different concentrations of DM, IMI, and their combinations. The y-axis indicates the normalized gene expression (GOI/GOR × 100), while the x-axis represents the experimental groups. Each box plot displays the mean (horizontal discontinued line), median (horizontal continued line), interquartile range (box), and data variability (whiskers), with individual data points shown. The control group serves as the baseline reference.

Expression levels of ntrk2a across experimental groups. Box plot illustrating the expression of ntrk2a (TrkB receptor isoform) in zebrafish following exposure to DM, IMI, and their mixtures. The y-axis represents normalized gene expression values (GOI/GOR × 100), while the x-axis corresponds to different experimental conditions. Statistical differences between groups were determined using ANOVA and Tukey post hoc tests (n = six).

Expression levels of ntrk2b across experimental groups. Box plot depicting ntrk2b expression levels, another TrkB receptor isoform, in zebrafish under pesticide exposure. The y-axis shows normalized gene expression values (GOI/GOR × 100), and the x-axis denotes the experimental groups. The plot provides insights into how pesticide exposure influences TrkB receptor expression, with statistical significance assessed using ANOVA and post hoc Tukey tests (n = six).

Expression levels of NGF across experimental groups—the relative expression levels of NGF in zebrafish exposed to different concentrations of DM, IMI, and their combinations (n = six per group). The y-axis indicates normalized gene expression (GOI/GOR × 100), while the x-axis represents the experimental groups. Each box plot displays the mean (horizontal discontinued line), median (horizontal continued line), interquartile range (box), and data variability (whiskers), with individual data points shown. Statistical analysis was performed using ANOVA followed by Tukey post hoc tests, with significance set at p < 0.05.

Expression levels of ntrk1 across experimental groups. Box plot illustrating the expression of ntrk1 (TrkA receptor) in zebrafish following exposure to different pesticide conditions (n = six per group). The y-axis represents normalized gene expression (GOI/GOR × 100), while the x-axis corresponds to the experimental groups. Statistical significance was determined using ANOVA and Tukey post hoc tests, with differences considered significant at p < 0.05. The data indicate alterations in ntrk1 expression in response to pesticide exposure, with potential implications for neurotrophic signaling.

Expression levels of ntf-3 across experimental groups. Box plot illustrating ntf-3 expression levels (n = six per group) in zebrafish exposed to DM, IMI, and their mixtures. Statistical significance was determined using ANOVA and Tukey post hoc tests.

Expression levels of ntf-4/5 across experimental groups. Box plot showing ntf-4/5 expression changes (n = six per group) across experimental conditions. Statistical comparisons were performed using ANOVA and Tukey post hoc tests.

Expression levels of ntrk3a (left) and ntrk3b (right) across experimental groups. Box plots displaying ntrk3a and ntrk3b expression levels (n = six per group). The data indicate significant downregulation in IMI and mixed-exposure groups (p < 0.0001).

Histological sections of key brain regions of control and DM-exposed fish. Representative cross-sections illustrating the structural organization of different brain regions in zebrafish from the control group and those exposed to increasing concentrations of deltamethrin (DM). Panels (1A–5A) correspond to the control group, while (1B–5B,1C–5C,1D–5D) depict sections from fish exposed to 0.25 µg L⁻¹, 0.5 µg L⁻¹, and 1 µg L⁻¹ DM, respectively. The images display the following brain regions: Olfactory bulbs (1A–1D), scale bar = 30 µm; Telencephalon (2A–2D), scale bar = 60 µm; Diencephalon and mesencephalon (3A–3D), scale bar = 100 µm; Rhombencephalon (4A–4D), scale bar = 100 µm; Medulla oblongata (5A–5D), scale bar = 60 µm. These histological sections highlight potential structural alterations associated with different pesticide exposures. Changes in tissue integrity, cellular arrangement, and neurotoxic effects are examined across experimental conditions.

Representative histological sections illustrating structural organization in various brain regions of zebrafish from the control group and those exposed to different concentrations of deltamethrin (DM) and imidacloprid (IMI) in combination. Panels (1A–5A) correspond to the control group, while (1B–5B,1C–5C,1D–5D) show sections from zebrafish exposed to 0.125 µg L⁻¹ DM + 6 µg L⁻¹ IMI, 0.25 µg L⁻¹ DM + 50 µg L⁻¹ IMI, and 0.5 µg L⁻¹ DM + 500 µg L⁻¹ IMI, respectively. The images display the following brain regions: Olfactory bulbs (1A–1D), scale bar = 30 µm; Telencephalon (2A–2D), scale bar = 60 µm; Diencephalon and mesencephalon (3A–3D), scale bar = 100 µm; Rhombencephalon (4A–4D), scale bar = 100 µm; Medulla oblongata (5A–5D), scale bar = 60 µm.