Análise da exposição a antibióticos e da modulação do sistema purinérgico em parâmetros comportamentais, bioquímicos e em um modelo de nocicepção em Peixe-zebra

Abstract

Since their discovery, antibiotics have become essential for health in humans and other species. Studies demonstrated the interaction of the use of antibiotics with well-being, defense, and other physiological aspects, but also with the damage that these compounds can cause in species such as fish, which do not even have a defined standard model of pain. Zebrafish has been a widely used model in translational biomedical research. This study evaluated the effect of antibiotics and modulation of the purinergic system on behavioral and biochemical parameters as well as in a zebrafish nociception model. The effects of 96h- exposure to Oxytetracycline (OTC), at concentrations of 2, 10, 20, and 100 mg/L, were evaluated in adults and larvae, as well as the effects of Florfenicol (FF) in larvae, at concentrations of 0.001, 0.005, 2 and 10 mg/L. Our results demonstrated that OTC induced an anxiety-like behavior in adults and reduced cortisol levels. To confirm the anxiety-like behavior, the fish were exposed to Clonazepam (0.006mg/L) for 10 min. Clonazepam reversed anxiety caused by OTC in the novel tank test and social interaction parameters, but no reversed cortisol levels. In larvae with 4 dpf, we demonstrated that exposure to OTC + FF (10mg/L - 10mg/L) for 96 h increased the mobile time and the number of entries in the aquarium center, accompanied by a reduction in the turn angle. We also observed a reduction in the antibiotic-induced optomotor response. In addition to the findings related to antibiotic exposure, we also studied a model of acetic acid (AA)-induced pain in zebrafish larvae. Our results demonstrated a reduction in distance covered, acceleration, and moving time for one hour after pain induction by AA in larvae with 5 dpf. To observe the possible prevention of pain, we used 0.05mg/L paracetamol (PAR) and 0.005mg/L Ibuprofen (IBP) one hour before exposing the larvae to AA. Our results demonstrated that PAR prevented AA-induced behavioral changes in all parameters, but IBP did not prevent larval pain. Surprisingly, dimethylsulfoxide (DMSO) at a concentration of 0.3% prevented AA-induced behavioral changes. PROB, an inhibitor of Pannexin 1 (PANX-1) prevented the changes in distance and movement observed at both AA concentrations tested. However, A740003, a P2X7 receptor antagonist, did not prevent behavioral changes in the AA-induced pain model. We also investigated the preventive role of ecto-5’-nucleotidase adenosine 5'-(α,β-methylene) diphosphate (AMPCP) and adenosine deaminase erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) inhibitors in the pain model induced by AA. Treatment with AMPCP or EHNA, followed by exposure to AA, did not prevent behavioral changes in the pain model in any of the parameters tested. At 0.050% AA, optomotor behavior was compromised, but AMPCP or EHNA did not prevent it. After AA-induced pain, adenosine monophosphate (AMP) hydrolysis increased in zebrafish larvae. However, exposure to AMPCP or EHNA did not prevent changes in AMP hydrolysis. This study reinforces the importance of zebrafish as an animal model for toxicological studies and the investigation of new analgesic pathways.