(A) Establishment of a Repetition Index (RI) uncovers quantifiable Tetrahydrocannabinol (THC)-induced behavioral stereotypy in adult zebrafish that differs from swimming patterns in the low concentration ethanol vehicle (B). (C) A schematic trajectory with inner and outer zones and formula to calculate the RI. (D) Repetitive patterns are extracted as cycle sets from the raw tracking and summed to give a RI value. (E) THC (1 μM) elicited a significantly greater RI than the control condition with EtOH (0.0015%) (****p < 0.0001), calculated as a mean ± SEM, (F) and caused a significant reduction in velocity during exposure (**p = 0.0077). Controls EtOH (n = 38), 40 nM THC (n = 6), 1 μM THC (n = 23) and 2 μM THC (n = 12). Time point t and the whole recording time interval T. Values without a letter in common are statistically different to the control condition (p < 0.05, Kruskal–Wallis and Dunn's multiple comparisons test).

(A) Co-administration of Tetrahydrocannabinol (THC) and N-methyl-D-aspartate (NMDA) affects the THC-induced behavioral stereotypy in adult zebrafish. (B) At 100 μM NMDA with 1 μM THC the THC-induced Repetition Index (RI), calculated as a mean ± SEM, was not statistically different from the controls (EtOH) (ns p > 0.9999). (C) Nor did NMDA and THC co-administration affect the swimming velocity during exposure. Controls EtOH (0.0015%) (n = 19), 1 μM THC (n = 23), 100 μM NMDA (n = 6), 20 μM NMDA + 1 μM THC (n = 4), 30 μM NMDA + 1 μM THC (n = 4), 40 μM NMDA + 1 μM THC (n = 4) and 100 μM NMDA + 1 μM THC (n = 8). Mean RI values of 20 mM NMDA (n = 4), 30 mM NMDA (n = 4) and 40 mM NMDA (n = 4) were similar to 100 μM NMDA. Values without a letter in common are statistically different to the control condition (p < 0.05, Kruskal–Wallis and Dunn's multiple comparisons). Figure 2A was created with BioRender.com.

Co-administration of Tetrahydrocannabinol (THC) and GABA_A receptor antagonist pentylenetetrazol (PTZ) affects the THC-induced behavioral stereotypy in adult zebrafish. (A) At 1.5 mM PTZ with 1 μM THC the THC-induced Repetition Index (RI), calculated as a mean ± SEM, was not statistically different from the controls (EtOH) (ns p = 0.1578). (B) 1.5 mM PTZ and 1 mM PTZ with 1 μM THC caused significant increases in swimming velocity compared to controls (EtOH) (*p < 0.05). Controls EtOH (0.0015%) (n = 19), 1 μM THC (n = 23), 1.5 mM PTZ (n = 12), 0.2 mM PTZ + 1 μM THC (n = 6), 1 mM PTZ + 1 μM THC (n = 5), 1.5 mM PTZ + 1 μM THC (n = 12) and 2 mM PTZ + 1 μM THC (n = 6). Mean RI values of 0.2 mM PTZ (n = 6) and 2 mM PTZ (n = 6), were similar to 1.5 mM PTZ. Values without a letter in common are statistically different to the control condition (p < 0.05, Kruskal–Wallis and Dunn's multiple comparisons test).

Co-administration of Tetrahydrocannabinol (THC) and the inverse CB₁R agonist AM251 did not significantly reduce the THC-induced behavioral stereotypy in adult zebrafish. (A) At 1.8 μM AM251 with 1 μM THC the THC-induced Repetition Index (RI), calculated as a mean ± SEM, was statistically different from the controls (DMSO) (*p = 0.0279). (B) Nor did AM251 and THC co-administration affect the swimming velocity during exposure. Controls (1% DMSO, n = 6), 1 μM THC (n = 23), 1.8 μM AM251 (n = 6) and 1.8 μM AM251 + 1 μM THC (n = 12). Values without a letter in common are statistically different to the control condition (p < 0.05, One-way ANOVA and Dunnett's multiple comparisons tests for (A) and Kruskal–Wallis and Dunn's multiple comparisons test for (B)).

Co-administration of Tetrahydrocannabinol (THC) and the inverse CB2R agonist AM630 significantly reduced the THC-induced behavioral stereotypy in adult zebrafish. (A) At 3.5 μM AM630 with 1 μM THC the THC-induced Repetition Index (RI), calculated as a mean ± SEM, was not statistically different from the controls (DMSO) (ns p > 0.9999). (B) AM630 and THC co-administration did not affect the swimming velocity during exposure. Controls (DMSO 1%, n = 6), 1 μM THC (n = 23), 3.5 μM AM630 (n = 5) and 3.5 μM AM630 + 1 μM THC (n = 6). Values without a letter in common are statistically different to the control condition (p < 0.05, Kruskal–Wallis and Dunn's multiple comparisons test).

Co-administration of Tetrahydrocannabinol (THC) and the atypical antipsychotic sulpiride significantly reduced the THC-induced behavioral stereotypy in adult zebrafish. (A) At 10 μM sulpiride with 1 μM THC and 100 μM sulpiride with 1 μM THC, the THC-induced Repetition Index (RI), calculated as a mean ± SEM, was not statistically different from the controls (EtOH) (ns p > 0.05). (B) None of the co-administrations significantly altered the swimming velocity during exposure (ns p > 0.05). Controls (EtOH) (0.0015% (n = 19)), 1 μM THC (n = 23), 10 μM sulpiride (n = 6), 100 μM sulpiride (n = 8) 10 μM sulpiride + 1 μM THC (n = 8) and 100 μM sulpiride + 1 μM THC (n = 8). Values without a letter in common are statistically different to the control condition (p < 0.05, Kruskal–Wallis and Dunn's multiple comparisons test).

Potential mechanisms underlying the associations between Cannabis consumption and risks of psychosis/schizophrenia-like symptoms, with the current study’s pharmacological approaches. Δ9-tetrahydrocannabinol (THC) reduces N-methyl-D-aspartate (NMDA) release via NMDA receptor antagonism. THC also inhibits γ‐aminobutyric acid (GABA) release from interneurons in the prefrontal cortex, potentiating dopamine release. Both NMDA receptor hypoactivity and heightened dopaminergic signaling are hallmarks of schizophrenia. Pentylenetetrazol (PTZ).