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Fingerprinting of psychoactive drugs in zebrafish anxiety-like behaviors.

Maximino C, da Silva AW, Araújo J, Lima MG, Miranda V, Puty B, Benzecry R, Picanço-Diniz DL, Gouveia A, Oliveira KR, Herculano AM - PLoS ONE (2014)

Bottom Line: The observed phenotypes were clustered according to shared behavioral effects.Moreover, anxiolytic drugs all decreased, while anxiogenic drugs increased, serotonin turnover.These results underscore the power of behavioral profiling in adult zebrafish as an approach which combines throughput and physiological complexity in the pharmacological dissection of complex behaviors.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Neurociências e Comportamento "Frederico Guilherme Graeff", Departamento de Morfologia e Ciências Fisiológicas, Centro de Ciências Biológicas e da Saúde, Universidade do Estado do Pará, Marabá, Pará, Brazil.

ABSTRACT
A major hindrance for the development of psychiatric drugs is the prediction of how treatments can alter complex behaviors in assays which have good throughput and physiological complexity. Here we report the development of a medium-throughput screen for drugs which alter anxiety-like behavior in adult zebrafish. The observed phenotypes were clustered according to shared behavioral effects. This barcoding procedure revealed conserved functions of anxiolytic, anxiogenic and psychomotor stimulating drugs and predicted effects of poorly characterized compounds on anxiety. Moreover, anxiolytic drugs all decreased, while anxiogenic drugs increased, serotonin turnover. These results underscore the power of behavioral profiling in adult zebrafish as an approach which combines throughput and physiological complexity in the pharmacological dissection of complex behaviors.

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Related in: MedlinePlus

Drugs which cluster on the 'anxiolytic' group decrease 5-HT turnover in the brain.(A) Turnover rates, as measured by 5-HIAA:5-HT ratios, normalized to the values of vehicle-treated animals, for the following drugs: fluoxetine (FLX; chronic treatment with 10 mg/kg); chlordiazepoxide (CDZ; 0.02 mg/kg); clonazepam (CLZ; 0.05 mg/kg); diazepam (DZP; 1.25 mg/kg); buspirone (BUS; 50 mg/kg); ethanol (EtOH, 2.5%); dizocilpine (MK; 0.005 mg/kg); verapamil (VER; 5 mg/kg); WAY 100,635 (WAY; 0.03 mg/kg); and SB 224,289 (SB; 2.5 mg/kg). Asterisks mark statistically significant differences in relation to vehicle-treated animals (F10, 43 = 45.99, p<0.0001, one-way ANOVA followed by Dunnett's Multiple Comparison test). Bars represent mean (B) Correlation between turnover rates (Y-axis) and time spent in the white compartment (X-axis) for vehicle- and drug-treated animals (n = 4 for each point). Points represent means and error bars represent standard errors. A negative correlation is found between the decrease in serotonin turnover and the increase in time on white produced by a drug (r2 = 0.5688, p = 0.0073).
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pone-0103943-g004: Drugs which cluster on the 'anxiolytic' group decrease 5-HT turnover in the brain.(A) Turnover rates, as measured by 5-HIAA:5-HT ratios, normalized to the values of vehicle-treated animals, for the following drugs: fluoxetine (FLX; chronic treatment with 10 mg/kg); chlordiazepoxide (CDZ; 0.02 mg/kg); clonazepam (CLZ; 0.05 mg/kg); diazepam (DZP; 1.25 mg/kg); buspirone (BUS; 50 mg/kg); ethanol (EtOH, 2.5%); dizocilpine (MK; 0.005 mg/kg); verapamil (VER; 5 mg/kg); WAY 100,635 (WAY; 0.03 mg/kg); and SB 224,289 (SB; 2.5 mg/kg). Asterisks mark statistically significant differences in relation to vehicle-treated animals (F10, 43 = 45.99, p<0.0001, one-way ANOVA followed by Dunnett's Multiple Comparison test). Bars represent mean (B) Correlation between turnover rates (Y-axis) and time spent in the white compartment (X-axis) for vehicle- and drug-treated animals (n = 4 for each point). Points represent means and error bars represent standard errors. A negative correlation is found between the decrease in serotonin turnover and the increase in time on white produced by a drug (r2 = 0.5688, p = 0.0073).

Mentions: Cluster analysis revealed a high degree of predictive validity in the proposed assay (Figures 1–3). First, anxiolytic drugs with clinical efficacy (benzodiazepines, chronic [14 days] fluoxetine) cluster together with buspirone, and anxiogenic drugs (NMDA, 5-HTP) cluster with caffeine (Figure 1). Second, motor-stimulating drugs (which represent potential false positives in locomotor-based assays) form their own cluster (Figure 1). For example, a low dose of caffeine (Figure 3A), a low dose of ethanol (Figure 3B), and bupropion (Figure 3C) increased locomotion, without effects on ethological measures or time in the white compartment. Second, drugs with multiple targets (e.g., ethanol, caffeine) correlated with drugs in different cluster in a dose-dependent way (Figure 1); caffeine, for example, clustered with anxiogenic drugs at a higher dose, and with stimulant drugs at a lower dose (Figure 1). Third, anxiolytic/anxiogenic and locotomor stimulating effects closely followed those observed in mammals. Fourth, compounds which clustered on the “anxiolytic” effect (Figure 1) all reduced serotonin turnover, which was correlated with time spent on the white compartment in these groups (r2 = 0.5688, p = 0.007) (Figure 4). These analyses indicate that compounds with shared systems effects produce similar phenotypes which are conserved across vertebrates. While from a neuroanatomical and genomic point of view the serotonergic system diverges from that of mammals [45], these and other data strongly suggest that the function of the serotonergic system is conserved across vertebrates. It should also be observed that some behavioral components (time on white, thigmotaxis, latency to white and risk assessment) are more strongly affected by drug treatments (Figure 1), suggesting that those parameters have a stronger predictive value to pharmacological treatments. Interestingly, time on white and thigmotaxis cluster together, while latency to white and risk assessment fall together on another cluster. Erratic swimming and freezing, while affected by anxiogenic and anxiolytic drug treatments, show a weaker liability. These results are in accordance with those observed in the novel tank test [15], in which erratic swimming and freezing had weaker predictive power in relation to time in the upper half of the tank and latency to upper half.


Fingerprinting of psychoactive drugs in zebrafish anxiety-like behaviors.

Maximino C, da Silva AW, Araújo J, Lima MG, Miranda V, Puty B, Benzecry R, Picanço-Diniz DL, Gouveia A, Oliveira KR, Herculano AM - PLoS ONE (2014)

Drugs which cluster on the 'anxiolytic' group decrease 5-HT turnover in the brain.(A) Turnover rates, as measured by 5-HIAA:5-HT ratios, normalized to the values of vehicle-treated animals, for the following drugs: fluoxetine (FLX; chronic treatment with 10 mg/kg); chlordiazepoxide (CDZ; 0.02 mg/kg); clonazepam (CLZ; 0.05 mg/kg); diazepam (DZP; 1.25 mg/kg); buspirone (BUS; 50 mg/kg); ethanol (EtOH, 2.5%); dizocilpine (MK; 0.005 mg/kg); verapamil (VER; 5 mg/kg); WAY 100,635 (WAY; 0.03 mg/kg); and SB 224,289 (SB; 2.5 mg/kg). Asterisks mark statistically significant differences in relation to vehicle-treated animals (F10, 43 = 45.99, p<0.0001, one-way ANOVA followed by Dunnett's Multiple Comparison test). Bars represent mean (B) Correlation between turnover rates (Y-axis) and time spent in the white compartment (X-axis) for vehicle- and drug-treated animals (n = 4 for each point). Points represent means and error bars represent standard errors. A negative correlation is found between the decrease in serotonin turnover and the increase in time on white produced by a drug (r2 = 0.5688, p = 0.0073).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4117595&req=5

pone-0103943-g004: Drugs which cluster on the 'anxiolytic' group decrease 5-HT turnover in the brain.(A) Turnover rates, as measured by 5-HIAA:5-HT ratios, normalized to the values of vehicle-treated animals, for the following drugs: fluoxetine (FLX; chronic treatment with 10 mg/kg); chlordiazepoxide (CDZ; 0.02 mg/kg); clonazepam (CLZ; 0.05 mg/kg); diazepam (DZP; 1.25 mg/kg); buspirone (BUS; 50 mg/kg); ethanol (EtOH, 2.5%); dizocilpine (MK; 0.005 mg/kg); verapamil (VER; 5 mg/kg); WAY 100,635 (WAY; 0.03 mg/kg); and SB 224,289 (SB; 2.5 mg/kg). Asterisks mark statistically significant differences in relation to vehicle-treated animals (F10, 43 = 45.99, p<0.0001, one-way ANOVA followed by Dunnett's Multiple Comparison test). Bars represent mean (B) Correlation between turnover rates (Y-axis) and time spent in the white compartment (X-axis) for vehicle- and drug-treated animals (n = 4 for each point). Points represent means and error bars represent standard errors. A negative correlation is found between the decrease in serotonin turnover and the increase in time on white produced by a drug (r2 = 0.5688, p = 0.0073).
Mentions: Cluster analysis revealed a high degree of predictive validity in the proposed assay (Figures 1–3). First, anxiolytic drugs with clinical efficacy (benzodiazepines, chronic [14 days] fluoxetine) cluster together with buspirone, and anxiogenic drugs (NMDA, 5-HTP) cluster with caffeine (Figure 1). Second, motor-stimulating drugs (which represent potential false positives in locomotor-based assays) form their own cluster (Figure 1). For example, a low dose of caffeine (Figure 3A), a low dose of ethanol (Figure 3B), and bupropion (Figure 3C) increased locomotion, without effects on ethological measures or time in the white compartment. Second, drugs with multiple targets (e.g., ethanol, caffeine) correlated with drugs in different cluster in a dose-dependent way (Figure 1); caffeine, for example, clustered with anxiogenic drugs at a higher dose, and with stimulant drugs at a lower dose (Figure 1). Third, anxiolytic/anxiogenic and locotomor stimulating effects closely followed those observed in mammals. Fourth, compounds which clustered on the “anxiolytic” effect (Figure 1) all reduced serotonin turnover, which was correlated with time spent on the white compartment in these groups (r2 = 0.5688, p = 0.007) (Figure 4). These analyses indicate that compounds with shared systems effects produce similar phenotypes which are conserved across vertebrates. While from a neuroanatomical and genomic point of view the serotonergic system diverges from that of mammals [45], these and other data strongly suggest that the function of the serotonergic system is conserved across vertebrates. It should also be observed that some behavioral components (time on white, thigmotaxis, latency to white and risk assessment) are more strongly affected by drug treatments (Figure 1), suggesting that those parameters have a stronger predictive value to pharmacological treatments. Interestingly, time on white and thigmotaxis cluster together, while latency to white and risk assessment fall together on another cluster. Erratic swimming and freezing, while affected by anxiogenic and anxiolytic drug treatments, show a weaker liability. These results are in accordance with those observed in the novel tank test [15], in which erratic swimming and freezing had weaker predictive power in relation to time in the upper half of the tank and latency to upper half.

Bottom Line: The observed phenotypes were clustered according to shared behavioral effects.Moreover, anxiolytic drugs all decreased, while anxiogenic drugs increased, serotonin turnover.These results underscore the power of behavioral profiling in adult zebrafish as an approach which combines throughput and physiological complexity in the pharmacological dissection of complex behaviors.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Neurociências e Comportamento "Frederico Guilherme Graeff", Departamento de Morfologia e Ciências Fisiológicas, Centro de Ciências Biológicas e da Saúde, Universidade do Estado do Pará, Marabá, Pará, Brazil.

ABSTRACT
A major hindrance for the development of psychiatric drugs is the prediction of how treatments can alter complex behaviors in assays which have good throughput and physiological complexity. Here we report the development of a medium-throughput screen for drugs which alter anxiety-like behavior in adult zebrafish. The observed phenotypes were clustered according to shared behavioral effects. This barcoding procedure revealed conserved functions of anxiolytic, anxiogenic and psychomotor stimulating drugs and predicted effects of poorly characterized compounds on anxiety. Moreover, anxiolytic drugs all decreased, while anxiogenic drugs increased, serotonin turnover. These results underscore the power of behavioral profiling in adult zebrafish as an approach which combines throughput and physiological complexity in the pharmacological dissection of complex behaviors.

Show MeSH
Related in: MedlinePlus