Anxiolytic effects of N-(4,5-dihydro-5-oxo-1,2-dithiolo-[4,3,b]-pyrrole-6-yl)-N-methylformamide, a pyrroloformamide isolated from a marine Streptomyces sp., in adult zebrafish by the 5-HT system.

General anxiety disorders are among the most prevalent mental health problems worldwide. The emergence and development of anxiety disorders can be due to genetic (30-50%) or non-genetic (50-70%) factors. Despite medical progress, available pharmacotherapies are sometimes ineffective or can cause undesirable side effects. Thus, it becomes necessary to discover new safe and effective drugs against anxiety. This study evaluated the anxiolytic effect in adult zebrafish (Danio rerio) of a natural pyrroloformamide (PFD), N-(4,5-dihydro-5-oxo-1,2-dithiolo-[4,3,b]-pyrrole-6-yl)-N-methylformamide, isolated from a Streptomyces sp. bacterium strain recovered from the ascidian Eudistoma vannamei. The complete structure of PFD was determined by a detailed NMR analysis, including 1H-13C and 1H-15N-HBMC data. In addition, conformational and DFT computational studies also were performed. A group of fishes (n = 6) was treated orally with PFD (0.1, 0.5 and 1.0 mg/mL; 20 µL) and subjected to locomotor activity and light/dark tests, as well as, acute toxicity 96 h. The involvement of the GABAergic and serotonergic (5-HT) systems was investigated using flumazenil (a silent modulator of GABA receptor) and 5-HT1, 5-HT2A/2C and 5-HTR3A/3B receptors antagonists, known as pizotifen, granisetron and cyproheptadine, respectively. PFD was nontoxic, reduced locomotor activity and promoted the anxiolytic effect in zebrafish. Flumazenil did not inhibit the anxiolytic effect of the PFD via the GABAergic system. This effect was reduced by a pretreatment with pizotifen and granisetron, and was not reversed after treatment with cyproheptadine. Molecular docking and dynamics studies confirmed the interaction of PFD with the 5-HT receptor.Communicated by Ramaswamy H. Sarma.

Pyrroloformamide (PFD), isolated from the marine Streptomyces sp. associated ascidian Eudistoma vannamei, showed no toxicity in adult zebrafish but reduced its locomotor activity.The structural elucidation of PFD was determined by the analysis of 1D and 2D NMR and HRESIMS data.The density functional theory (DFT) study confirmed the existence of two conformers as determined by NMR spectra.The serotonergic system modulated the anxiolytic effect of PFD via the 5-HT receptor in adult zebrafish.Molecular docking and dynamics studies confirmed the interaction of PFD with the 5-HT receptor.

Combretum lanceolatum extract reverses anxiety and seizure behavior in adult zebrafish through GABAergic neurotransmission: an in vivo and in silico study.

Combretaceae are reported in the literature for presenting neuroprotective and anxiolytic effects in animal models. Combretum lanceolatum Pohl. has few scientific reports on its pharmacological effects. The aim of this study was to evaluate the anxiolytic and anticonvulsant effects of the ethanol extract from the leaves of C. lanceolatum Pohl. (EtFoCl) and its possible mechanism of GABAergic action in adult zebrafish. EtFoCl was subjected to determination of the total phenol concentration, identification of phytochemical flavonoids by HPLC and in vitro antioxidant activity test, open field test and 96-hour acute toxicity in zebrafish. Anxiolytic doses were tested for pentylenetetrazole-induced seizures in adult zebrafish. To study the mechanisms of action, molecular docking simulations were performed between the main phytochemicals and the GABAA receptor (anxiolytic activity) and carbonic anhydrase II (anticonvulsant). The non-toxic doses that caused motor impairment were assessed in acute and chronic anxiety using the light and dark test. EtFoCl had altered the animals' locomotion, presenting an effect similar to the anxiolytic and anticonvulsant. These effects were prevented with flumazenil (GABAA antagonist). The phytochemicals homoorientin and quercetin-3-O-galactoside coupling in a region close to that of the inhibitor diazepam (GABAA receptor). Regarding the anticonvulsant mechanism, Homoorientina and Isovitexina were identified as the most favorable for the complex form with the carbonic anhydrase enzyme. C. lanceolatum has pharmacological potential for the treatment of acute and chronic anxiety and seizures, which can be partially explained by an interaction with the GABAA receptor.Communicated by Ramaswamy H. Sarma.

Anxiolytic-like effect of natural product 2-hydroxy-3,4,6-trimethoxyacetophenone isolated from Croton anisodontus in adult zebrafish via serotonergic neuromodulation involvement of the 5-HT system.

Benzodiazepines are highly effective in combating anxiety; however, they have considerable adverse effects, so it is important to discover new safe anxiolytic agents. This study was designed to investigate the effect of the natural product 2-hydroxy-3,4,6-trimethoxyacetophenone (HTMCX) on anxiety and seizure behavior in adult zebrafish and its possible mechanisms of action. The acute toxicity of 96 h of HTMCX was analyzed, and the open and light/dark field tests (n = 6 animals/group) were used to assess the anxiety behavior of animals treated with HTMCX. In addition, the mechanisms of action were investigated with antagonists of the GABAA, 5-HT receptors, and molecular anchorage study. Pentylenetetrazole (PTZ) was used to induce seizure by immersion. As a result, acetophenone HTMCX (1, 3 and 10 mg/kg; v.o.) was non-toxic and affected locomotor activity. The higher doses (3 and 10 mg/kg; v.o.) produced signs of anxiolytic action in the light/dark test, and this effect was reversed by the pizotifen (antagonist 5HTR1 and 5HTR2A/2C), having the potential to form a complex with 5HTR1B. However, the anxiolytic effect of HTMCX has not been abolished by flumazenil (antagonist GABAA), cyproheptadine (antagonist 5HTR2A), and granisetron (antagonist 5HTR3A/3B). Therefore, HTMCX demonstrated an anxiolytic effect, suggesting that the 5HTR1 and 5HTR2C receptors may be involved in the pharmacological performance of this acetophenone in the central nervous system.

Anxiolytic-like effect of brominated compounds from the marine sponge Aplysina fulva on adult zebrafish (Danio rerio): Involvement of the GABAergic system.

Benzodiazepines are commonly used to treat disorders of the central nervous system, including anxiety. However, due to their adverse effects, there is a continuing interest in discovering new safe and effective drugs. Marine natural products have emerged as a prolific source of bioactive nitrogenated compounds. Aiming to discover new biologically active natural compounds, the marine sponge Aplysina fulva, a nitrogen-bearing heterocyst producer, was investigated. The main isolated compounds (4, 6, and 9) were evaluated on adult zebrafish (Danio rerio). A group of fishes (n = 6) was preliminarily subjected to acute toxicity, and open field tests using 0.1, 0.5, and 1.0 mg/mL (v. o.) of those compounds was performed. The anxiolytic effect was further investigated in the light/dark assay based on the locomotor response at zebrafish. Interactions through the GABAergic system were investigated using flumazenil, a silent modulator of GABA receptors. To improve the results, a study of molecular docking using the GABAA receptor also was performed. Based on the results, the bromotyrosine derivative compounds 4, 6, and 9 exhibited anxiolytic-like effects mediated by the GABAergic system.

Anxiolytic-like effect of chalcone N-{(4'-[(E)-3-(4-fluorophenyl)-1-(phenyl) prop-2-en-1-one]} acetamide on adult zebrafish (Danio rerio): Involvement of the GABAergic system.

Benzodiazepines are the standard drugs for the treatment of anxiety, but their undesirable side effects make it necessary to develop new anxiolytic drugs. The objective of this study was to evaluate the possible anxiolytic-simile effect of synthetic chalcone N-{(4'-[(E)-3-(4-fluorophenyl)-1-(phenyl) prop-2-en-1-one]} acetamide (PAAPFBA) on adult zebrafish (Danio rerio). PAAPFBA was synthesized with an 88.21% yield and its chemical structure was determined by 1H and 13C NMR. Initially, animals (n = 6/group) were treated (4 or 12 or 40 mg/kg, intraperitoneal) with PAAPFBA and were submitted to acute toxicity and open field tests. Then, other groups (n = 6/each) received PAAPFBA for the analysis of its effect on the Light & Dark Test. The participation of the GABAergic system was also assessed using the GABAA antagonist flumazenil. Molecular docking was performed using the GABAA receptor. The effect of PAAPFBA on anxiety induced by alcohol withdrawal was analyzed. PAAPFBA was non-toxic, reduced the locomotor activity, and showed an anxiolytic-like effect in both models. This effect was reduced by pre-treatment with the flumazenil. In agreement with in vivo studies, molecular docking indicated an interaction between chalcone and the GABAA receptor. The results suggest that PAAPFBA is an anxiolytic agent mediated via the GABAergic system.

Antinociceptive activity of ethanolic extract of Azadirachta indica A. Juss (Neem, Meliaceae) fruit through opioid, glutamatergic and acid-sensitive ion pathways in adult zebrafish (Danio rerio).

Neem fruit (Azadirachta indica A. Juss.) are popularly used to treat infections, diarrhea, fever, bronchitis, skin diseases, infected burns and hypertension. Although the antinociceptive and anti-inflammatory potential of A. indica has already been investigated in experimental models of pain and inflammation in mice, the current research is the first to report the evaluation of the capacity of A. indica fruit ethanolic extract (EtFrNeem) in acute pain attenuation using the adult zebrafish (Danio rerio) as an alternative model to the use in rodents. EtFrNeem was submitted to antioxidant action, preliminary chemical prospecting, FT-IR and determination of phenol and flavonoid content tests. Subsequently, EtFrNeem was tested for acute nociception and abdominal inflammation, locomotor activity, and acute toxicity in adult zebrafish. Possible neuromodulation mechanisms were also evaluated. EtFrNeem showed low antioxidant activity, but was shown to be rich in flavonoids. EtFrNeem showed no anti-inflammatory action, did not alter the locomotor system, and it was not toxic. However, EtFrNeem significantly reduced the nociceptive behavior induced by formalin, glutamate and acidic saline, when compared to the control group. These effects of EtFrNeem were significantly similar to those of morphine, used as a positive control. The antinociceptive effect of EtFrNeem was inhibited by naloxone, ketamine and amiloride. EtFrNeem has the pharmacological potential for acute pain treatment and this effect is modulated by the opioid system, NMDA receptors and ASICs channels. These results lead us to studies of isolation and characterization of EtFrNeem bioactive principles, using adult zebrafish as an experimental model.