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(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).
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High doses of the Cannabis constituent Δ9-tetrahydrocannabinol (THC) increase the risk of psychosis in humans. Highly accessible animal models are needed to address underlying mechanisms. Using zebrafish with a conserved endocannabinoid system, this study investigates the acute effects of THC on adult zebrafish behavior and the mechanisms involved....
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... adult zebrafish. To determine the behavioral effects of THC, adult EK-WT zebrafish were individually immersed in 40 nM, 1 μM or 2 μM THC for 20 min and compared to control zebrafish exposed to the ethanol vehicle (0.00006%, 0.0015%, 0.003%) ( Fig. 2A). No significant difference was found between the ethanol control concentrations (Supplementary Fig. S1) and therefore they were grouped together in Fig. 1E,F. However, in THC-treated individuals, we noted an abnormal behavioral pattern that had the characteristic of repetitive circling (Fig. ...Context 2
... of THC, adult EK-WT zebrafish were individually immersed in 40 nM, 1 μM or 2 μM THC for 20 min and compared to control zebrafish exposed to the ethanol vehicle (0.00006%, 0.0015%, 0.003%) ( Fig. 2A). No significant difference was found between the ethanol control concentrations (Supplementary Fig. S1) and therefore they were grouped together in Fig. 1E,F. However, in THC-treated individuals, we noted an abnormal behavioral pattern that had the characteristic of repetitive circling (Fig. ...Context 3
... to the ethanol vehicle (0.00006%, 0.0015%, 0.003%) ( Fig. 2A). No significant difference was found between the ethanol control concentrations (Supplementary Fig. S1) and therefore they were grouped together in Fig. 1E,F. However, in THC-treated individuals, we noted an abnormal behavioral pattern that had the characteristic of repetitive circling (Fig. ...Context 4
... order to measure this behavioral abnormality, we developed a computational method to quantify the repetition index (RI) (Fig. 1C and see "Methods"). The mean RI, the standard error of the mean (SEM) and the range for the control condition and the THC conditions were plotted in Fig. 1E. No difference was observed between males and females. THC evoked prominent circling behavior in 69.6% (n = 23) of fish at 1 μM, with a RI significantly higher than the ethanol ...Context 5
... order to measure this behavioral abnormality, we developed a computational method to quantify the repetition index (RI) (Fig. 1C and see "Methods"). The mean RI, the standard error of the mean (SEM) and the range for the control condition and the THC conditions were plotted in Fig. 1E. No difference was observed between males and females. THC evoked prominent circling behavior in 69.6% (n = 23) of fish at 1 μM, with a RI significantly higher than the ethanol controls (****p < 0.0001, Kruskal-Wallis; ****p < 0.0001, Dunn's multiple comparisons test) (Fig. 1E). The tracks of one fish at 40 nM THC were categorized as ...Context 6
... range for the control condition and the THC conditions were plotted in Fig. 1E. No difference was observed between males and females. THC evoked prominent circling behavior in 69.6% (n = 23) of fish at 1 μM, with a RI significantly higher than the ethanol controls (****p < 0.0001, Kruskal-Wallis; ****p < 0.0001, Dunn's multiple comparisons test) (Fig. 1E). The tracks of one fish at 40 nM THC were categorized as circular swimming but the mean RI was not significantly different from the controls (ns p > 0.9999, Dunn's multiple comparisons test). 2 μM THC elicited strong circling in 25% (n = 12) of the fish, but this was not sufficient to cause a significantly higher mean RI compared to ...Context 7
... was no significant difference in mean velocity before or during exposure in the ethanol control, 40 nM THC or 2 μM THC conditions. At the concentration with the highest mean RI, 1 μM THC, velocity was significantly reduced during drug immersion (**p = 0.0019, Kruskal-Wallis; **p = 0.0077, Dunn's multiple comparisons test) (Fig. 1F). Taken together, 1 μM THC evoked the strongest circling behavior while simultaneously dampening overall velocity. Such dampening of velocity may be related to or independent of the circling behavior. The results from 1 μM THC administration in EK-WT fish (n = 23) and corresponding ethanol (0.0015%) controls (n = 19) were used in the ...Context 8
... (NMDA) attenuates THC-induced behavioral stereotypy. Based on the results from the THC dosage tests (Fig. 1E,F), 1 μM THC was selected for the following experiments to examine if agonism of the NMDAR by NMDA could attenuate the THC behavioral stereotypy. 1 μM THC was coadministered with 20 μM, 30 μM, 40 μM, and 100 μM NMDA and compared to controls with ethanol (0.0015%) or NMDA at 20 μM, 30 μM, 40 μM or 100 μM. The mean RI, the SEM and the ...Context 9
... RI was significantly higher than the controls (DMSO) (*p = 0.0380, One-way ANOVA; *p = 0.0279, Dunnett's multiple comparisons test) and 58.3% (n = 12) of the fish exhibited stereotyped circling (Fig. 4A). 1.8 μM AM251 alone also elicited repetitive behavior detected by the algorithm but this was not visually analogous to the clear THC-circling (Fig. 1A). Neither the vehicle DMSO (1%), nor co-treatment with 1.8 μM AM251 significantly altered velocity (ns p = 0.5969, Kruskal-Wallis) (Fig. 4B). Data from 7 fish was excluded due to failed tracking, leaving data from 47 ...Context 10
... a new analytical method that we have developed, this study demonstrated that 1 μM THC administration in adult zebrafish triggered a shift from typical navigational locomotor patterns to a repetitive circling behavior, which was ameliorated by the antipsychotic sulpiride ( Figs. 1 and 6). This behavioral phenotype appears analogous to THC's effect in rats 24 and the effect of NMDAR antagonists in zebrafish models of psychosis 16,17,38 . ...Context 11
... raw x and y co-ordinates from the inner zone (Fig. 1C) of the Ethovision tracking were used to calculate an unbiased Repetition Index (RI) using the following algorithm in Python and ...Context 12
... the temporal and spatial threshold values as follows: www.nature.com/scientificreports/ Optimization: To detect and extract repetitions in the movement trajectory M (Fig. 1), the algorithm was optimized with a sliding window size of 500 time points (θ time-window ) (2:45 min), a minimum repetitive behavior threshold of standard deviation = 0.01 (<θ std-change ), and a repetitive interval threshold of 700 time points (θ repetition-time ) (3:51 min). The thresholds were determined by trialing different ...Context 13
... of 500 time points (θ time-window ) (2:45 min), a minimum repetitive behavior threshold of standard deviation = 0.01 (<θ std-change ), and a repetitive interval threshold of 700 time points (θ repetition-time ) (3:51 min). The thresholds were determined by trialing different values and a) visually comparing how well the extracted cycle sets (e.g, Fig. 1D) captured the repetitive movements and b) how well the RI value reflected the repetitive behavior (i.e., a higher value for strong circling and a lower value for random swimming trajectories). Anonymized tracking images (e.g., Fig. 1A,B) were independently sorted into high and low repetitive behavior by two researchers. If ...Context 14
... were determined by trialing different values and a) visually comparing how well the extracted cycle sets (e.g, Fig. 1D) captured the repetitive movements and b) how well the RI value reflected the repetitive behavior (i.e., a higher value for strong circling and a lower value for random swimming trajectories). Anonymized tracking images (e.g., Fig. 1A,B) were independently sorted into high and low repetitive behavior by two researchers. If discrepancies between the manual sorting and the corresponding RI values occurred, the algorithm's threshold values were ...Context 15
... 10% of the total distance across the x and y axes, near the edges of the tank, were designated as thigmotaxis margins (Fig. 1C). Detected movement in this region was removed. (2) The standard deviation between the x and y co-ordinates within the specified window was calculated (3) and if below 0.01 (<θ std-change ), the trajectory was considered repetitive and set to 1, i.e. the fish returns to the same co-ordinates during the time frame. If the standard ...Context 16
... was considered repetitive and set to 1, i.e. the fish returns to the same co-ordinates during the time frame. If the standard deviation was above the threshold, the trajectory was considered arbitrary and set to 0. (4) Next, if the event set to 1 had a duration longer than 700 time points (>θ repetition-time ), it was extracted as a cycle set (Fig. 1D), (5) while shorter events < θ repetition-time , were excluded ( Supplementary Fig. S5). Thus, the standard deviation between the x and y co-ordinates within the specified window had to be close to 0 and the minimum duration of the behavior 3:51 min in order for the algorithm to extract the behavior. (6) The durations of all cycle sets ...Context 17
... the behavior 3:51 min in order for the algorithm to extract the behavior. (6) The durations of all cycle sets were summed, divided by the total time interval T and normalized into RI values ranging between 0 to 1. Higher RI values signify intensified and prolonged repetitions in swimming trajectory, such as the circling or eight-shaped patterns (Fig. 1A), whereas values nearer 0 indicate more random movement (Fig. ...Context 18
... behavior. (6) The durations of all cycle sets were summed, divided by the total time interval T and normalized into RI values ranging between 0 to 1. Higher RI values signify intensified and prolonged repetitions in swimming trajectory, such as the circling or eight-shaped patterns (Fig. 1A), whereas values nearer 0 indicate more random movement (Fig. ...Similar publications
Hypofunctioning of NMDA receptors, and the resulting shift in the balance between excitation and inhibition, is considered a key process in the pathophysiology of schizophrenia. One important manifestation of this phenomenon is changes in neural oscillations, those above 30 Hz (i.e., gamma-band oscillations), in particular. Although both preclinica...
Citations
... AM630 can block the effect of phytocannabinoid THC [79] and CB 2 receptor agonist JWH133 [80]. Few studies have examined the effects of AM630 using zebrafish models, however Dahlén et al. [81] found AM630 to eliminate the effects of THC-induced behavioural stereotypy while having no significant effect on zebrafish behaviour . ...
... All zebrafish were administered treatments via in-water immersion according to Dahlén et al. [81]. All treatment concentrations were determined from previous studies [14,81] or pilot testing. ...
... All zebrafish were administered treatments via in-water immersion according to Dahlén et al. [81]. All treatment concentrations were determined from previous studies [14,81] or pilot testing. On testing days, zebrafish were transferred by netting into a 3-liter habitat tank and were habituated in the testing room for a minimum of 15 min prior to treatment administration. ...
Terpenes are the most extensive and varied group of naturally occurring compounds mostly found in plants,
including cannabis, and have an array of potential therapeutic benefits for pathological conditions. The endocannabinoid system can potently modulate anxiety in humans, rodents, and zebrafish. The ‘entourage effect’
suggests terpenes may target cannabinoid CB1 and CB2 receptors, among others, but this requires further
investigation. In this study we first tested for anxiety-altering effects of the predominant ‘Super-Class’ terpenes,
bisabolol (0.001%, 0.0015%, and 0.002%) and terpinolene (TPL; 0.01%, 0.05%, and 0.1%), in zebrafish with the
open field test. Bisabolol did not have an effect on zebrafish behaviour or locomotion. However, TPL caused a
significant increase in time spent in the inner zone and decrease in time spent in the outer zone of the arena
indicating an anxiolytic (anxiety decreasing) effect. Next, we assessed whether CB1 and CB2 receptor antagonists,
rimonabant and AM630 (6-Iodopravadoline) respectively, could eliminate or reduce the anxiolytic effects of TPL
(0.1%) and β-caryophyllene (BCP; 4%), another super-class terpene previously shown to be anxiolytic in
zebrafish. Rimonabant and AM630 were administered prior to terpene exposure and compared to controls and
fish exposed to only the terpenes. AM630, but not rimonabant, eliminated the anxiolytic effects of both BCP and
TPL. AM630 modulated locomotion on its own, which was potentiated by terpenes. These findings suggest the
behavioural effects of TPL and BCP on zebrafish anxiety-like behaviour are mediated by a selective preference for
CB2 receptor sites. Furthermore, the CB2 pathways mediating the anxiolytic response are likely different from
those altering locomotion.
... In humans, PCP induces analgesia, ataxia, euphoria, anxiety, hallucinations, and psychosis (Kyzar et al., 2012). Amphetamine Benvenutti et al., 2021; Cocaine -Patiño et al., 2008;Pisera-Fuster et al., 2019) Nicotine Kyzar et al., 2012) Ketamine Benvenutti et al., 2021;Swain et al., 2004) Carty et al., 2019;Jensen et al., 2018;Nazario et al., 2015) THC Carty et al., 2019;Dahlén et al., 2021) Current Trends in Natural Sciences Vol. 12, Issue 23, pp. 266-273, 2023https://doi.org/10.47068/ctns.2023 ...
Polymeric materials became of crucial interest in the research area over the recent years due to worldwide production increase of these materials not always being followed by knowledge of their possible detrimental effects. Therefore, in the present study we analyzed the different effects of polymeric materials at various exposure times. Our results focused on some general effects in terms of anxiety and swimming performance over 120h of exposure and multiple repeated doses of both Polyethylene and Polypropylene microplastics. Polyethylene showed an anxiogenic potential, while Polypropylene exhibited an anxiolytic potential at the first two doses. In terms of swimming performance, an increase of specific parameters was observed throughout the repeated doses in the case of Polyethylene, whereas in the case of Polypropylene they decrease. These results were compared with those from the specialized existing literature on the effects of some of the most common hallucinogenic and psychoactive substances. Our results highlight that the mechanism of action of the aforementioned materials is different and show toxic effects from the first dose. Polyethylene has a similar outcome to the one in the case of some substances, especially hallucinogenic, such as LSD, mescaline, MDMA, phencyclidine, and nicotine. On the other hand, polypropylene has a similar effect to psychoactive drugs, cannabidiol, tetrahydrocannabinol, but also MDMA at certain doses. Bases on these observations, further studies should be conducted on the similarities of the detrimental effects.
... GO terms associated with ∆ 9 -THC exposure were searched using the g:profiler tool [35]. Ensembl gene data were used to map medaka genes of interest to their human orthologs in order to take full advantage of the many functional and non-inferred electronic annotations based on the human genome [36]. Using the online g-profiler ortholog search tool, we converted the Ensembl medaka genome ID into a human ID. ...
With the legalization of marijuana smoking in several states of the United States and many other countries for medicinal and recreational use, the possibility of its release into the environment cannot be overruled. Currently, the environmental levels of marijuana metabolites are not monitored on a regular basis, and their stability in the environment is not well understood. Laboratory studies have linked delta 9-tetrahydrocannabinol (Δ9-THC) exposure with behavioral abnormalities in some fish species; however, their effects on endocrine organs are less understood. To understand the effects of THC on the brain and gonads, we exposed adult medaka (Oryzias latipes, Hd-rR strain, both male and female) to 50 ug/L THC for 21 days spanning their complete spermatogenic and oogenic cycles. We examined transcriptional responses of the brain and gonads (testis and ovary) to Δ9-THC, particularly molecular pathways associated with behavioral and reproductive functions. The Δ9-THC effects were more profound in males than females. The Δ9-THC-induced differential expression pattern of genes in the brain of the male fish suggested pathways to neurodegenerative diseases and pathways to reproductive impairment in the testis. The present results provide insights into endocrine disruption in aquatic organisms due to environmental cannabinoid compounds.
... 7 THC dysregulates the brain striatum and is linked to social withdrawal, cognitive deficits and may activate a schizophrenic endophenotype in humans, according to Norris et al (2019). Increasing the action of CB-2 receptors, and lowering GABA stimulation to the forebrain, both lowered THC-induced stereotypies in the fish (Dahlen et al 2021). ...
... Studies have shown mice lacking CB-2 enzymes and schizophrenic humans, show reduced reflexive "startle responses" due to genetic deficits in the brain circuit which unconsciously monitors the atmosphere (Dahlen et al 2021). CNS over-excitation may be changed through CB-1 and CB-2 receptor balance, as the midbrain dopaminergic system contains many cannabinoid receptors (Dahlen et al 2021). ...
... Studies have shown mice lacking CB-2 enzymes and schizophrenic humans, show reduced reflexive "startle responses" due to genetic deficits in the brain circuit which unconsciously monitors the atmosphere (Dahlen et al 2021). CNS over-excitation may be changed through CB-1 and CB-2 receptor balance, as the midbrain dopaminergic system contains many cannabinoid receptors (Dahlen et al 2021). CBD works on both CB-1 and CB-2 receptors through two different pathways, it tends to also improve overall cognition in those with mental illnesses (Mouhamed et al 2018). ...
States in the US began to legalize marijuana nearly a decade ago. This review provides a short story of the research which led to the substitution of cannabinoids for opioids. Through the collection of 2941 peer-reviewed sources, three themes emerged: Replacing opioids, psychosis and cannabinoids, and the question of CBD, THC or both for the treatment of pain. With a certain percentage of the population being genetically predisposed to a CB-2 deficient response, they are at high risk of neuropsychiatric reactions to THC products. Many states have approved the sale of marijuana products which are seven times the safe dose for a normal person. More oversight and research is needed for both the safety of individuals who are self-medicating, and the education of medical prescribing professionals.
... Hyperlocomotion and stereotyped behaviors are thought to evaluate positive psychotic symptoms in mice (Eichler et al., 1980;Angermeyer and Matschinger, 2004;Morrens et al., 2006;van den Buuse, 2010;Forrest et al., 2014;Schubart et al., 2014;Compton et al., 2015;Svoboda et al., 2015;Kaufmann et al., 2018;Ma and Guest, 2018;Dahlén et al., 2021). ...
Deep brain stimulation (DBS) is a clinical intervention for the treatment of movement disorders. It has also been applied to the treatment of psychiatric disorders such as depression, anorexia nervosa, obsessive-compulsive disorder, and schizophrenia. Psychiatric disorders including schizophrenia, bipolar disorder, and major depression can lead to psychosis, which can cause patients to lose touch with reality. The ventral tegmental area (VTA), located near the midline of the midbrain, is an important region involved in psychosis. However, the clinical application of electrical stimulation of the VTA to treat psychotic diseases has been limited, and related mechanisms have not been thoroughly studied. In the present study, hyperlocomotion and stereotyped behaviors of the mice were employed to mimic and evaluate the positive-psychotic-like behaviors. We attempted to treat positive psychotic-like behaviors by electrically stimulating the VTA in mice and exploring the neural mechanisms behind behavioral effects. Local field potential recording and in vivo fiber photometry to observe the behavioral effects and changes in neural activities caused by DBS in the VTA of mice. Optogenetic techniques were used to verify the neural mechanisms underlying the behavioral effects induced by DBS. Our results showed that electrical stimulation of the VTA activates local gamma-aminobutyric acid (GABA) neurons, and dopamine (DA) neurons, reduces hyperlocomotion, and relieves stereotyped behaviors induced by MK-801 (dizocilpine) injection. The results of optogenetic manipulation showed that the activation of the VTA GABA neurons, but not DA neurons, is involved in the alleviation of hyperlocomotion and stereotyped behaviors. We visualized changes in the activity of specific types in specific brain areas induced by DBS, and explored the neural mechanism of DBS in alleviating positive psychotic-like behaviors. This preclinical study not only proposes new technical means of exploring the mechanism of DBS, but also provides experimental justification for the clinical treatment of psychotic diseases by electrical stimulation of the VTA.
... For centuries, the cannabis plant (Cannabis sativa and Cannabis indica) has been used as a source of food, fiber, and medicine [1][2][3][4]. In recent decades, scientific interest in cannabis has increased considerably, as its bioactive compounds have shown promising potential in the treatment of numerous musculoskeletal and neurological diseases in humans [5,6]. ...
... In human medicine, cannabinoids are already considered to be antiemetic, antispastic, analgesic, and appetite-stimulating compounds [2,5]. Their therapeutic effects have also been examined in a series of syndromes, including multiple sclerosis [15], Dravet syndrome [16,17], epilepsy [18], fibromyalgia [19], anxiety [20], schizophrenia [21], chronic pain [22], and cancer [23,24]. ...
... Cannabinoid use is still subject to uncertainty over aspects such as the dosing and side-effect profiles, and there is an overall lack of knowledge of their underlying mechanism of action; clinicians are therefore often reluctant to prescribe cannabis [25,28]. However, cannabis shows a lower potential to cause dependence (8.9%) than do other common substances of abuse, such as cocaine (20.9%), alcohol (22.7%), and nicotine (67.5%) [29]; moreover, it has recently been suggested that susceptibility to psychosis-like symptoms varies between cannabis consumers, as it involves a complex interplay between environmental factors and genetic predispositions [2]. Cannabinoids aside, it is also worth noting that potentially synergistic effects of phytocannabinoids and terpenoids have been reported in the treatment of pain, inflammation, depression, and anxiety [12]. ...
For centuries, the cannabis plant has been used as a source of food, fiber, and medicine. Recently, scientific interest in cannabis has increased considerably, as its bioactive compounds have shown promising potential in the treatment of numerous musculoskeletal and neurological diseases in humans. However, the mechanisms that underlie its possible effects on neurodevelopment and nervous-system functioning remain poorly understood and need to be further investigated. Although the bulk of research on cannabis and cannabinoids is based on in vitro or rodent models, the zebrafish has now emerged as a powerful in vivo model for drug-screening studies and translational research. We here review the available literature on the use of cannabis/cannabinoids in zebrafish, and particularly in zebrafish models of neurological disorders. A critical analysis suggests that zebrafish could serve as an experimental tool for testing the bioactivity of cannabinoids, and they could thus provide important insights into the safety and efficacy of different cannabis-extract-based products. The review showed that zebrafish exhibit similar behaviors to rodents following cannabinoid exposure. The authors stress the importance of analyzing the full spectrum of naturally occurring cannabinoids, rather than just the main ones, THC and CBD, and they offer some pointers on performing behavioral analysis in zebrafish.
Cannabinoids are active substances present in plants of the Cannabis genus. Both the Food and Drug Administration (FDA) and European Medicines Agency (EMA) have approved several medicinal products containing natural cannabinoids or their synthetic derivatives for the treatment of drug-resistant epilepsy, nausea and vomiting associated with cancer chemotherapy, anorexia in AIDS patients, and the alleviation of symptoms in patients with multiple sclerosis. In fact, cannabinoids constitute a broad group of molecules with a possible therapeutic potential that could be used in the management of much more diseases than mentioned above; therefore, multiple preclinical and clinical studies on cannabinoids have been carried out in recent years. Danio rerio (zebrafish) is an animal model that has gained more attention lately due to its numerous advantages, including easy and fast reproduction, the significant similarity of the zebrafish genome to the human one, simplicity of genetic modifications, and body transparency during the early stages of development. A number of studies have confirmed the usefulness of this model in toxicological research, experiments related to the impact of early life exposure to xenobiotics, modeling various diseases, and screening tests to detect active substances with promising biological activity. The present paper focuses on the current knowledge of the endocannabinoid system in the zebrafish model, and it summarizes the results and observations from studies investigating the pharmacological effects of natural and synthetic cannabinoids that were carried out in Danio rerio. The presented data support the notion that the zebrafish model is a suitable animal model for use in cannabinoid research.
