Name: Configuração de Tanque Experimental e Configuração de Gravação de Vídeo para Testes Comportamentais em Zebrafish Adulto
Uploaded: 2023-11-03T12:40:00
Description: Watch this Scientific Journal Video about Experimental Tank Configuration and Video Recording Setup for Behavioral Testing in Adult Zebrafish at JoVE.com

experimental tank configuration and video recording setup for behavioral testing in adult zebrafish

Bateria de Testes Comportamentais para Avaliação de Efeitos Neurotóxicos em Peixe-Zebra Adulto

The zebrafish (Danio rerio) is a popular model vertebrate species for ecotoxicology, drug discovery, and safety pharmacology studies. Its low cost, well-established molecular genetic tools, and conservation of key physiological processes involved in the morphogenesis and maintenance of the nervous system make zebrafish an ideal animal model for neuroscience research, including neurobehavioral toxicology1,2. The main endpoint for evaluating the neurotoxicity of a chemical was, until recently, the presence of neuropathological effects. Lately, however, behavioral endpoints have been incorporated into neurotoxicological screening protocols, and these functional outcomes are now commonly used to identify and determine the potential neurotoxicity of chemicals3,4. Moreover, behavioral endpoints are highly relevant from an ecological point of view, as even a very mild behavioral change in fish could endanger the survival of the animal in natural conditions5.
One of the most used behavioral assays in adult zebrafish research is the novel tank test (NTT), which measures anxiety-like behavior6,7. In this assay, fish are exposed to novelty (fish are placed in an unfamiliar tank), a mild aversive stimulus and their behavioral responses are observed. NTT is used to assess basal locomotor activity, geotaxis, freezing, and erratic movements of fish, principally. Erratic8 is characterized by abrupt changes of direction (zigzagging) and repeated episodes of accelerations (darting). It is an alarm reaction and is usually observed before or after freezing episodes. Freezing behavior corresponds to a complete cessation of the fish&#39;s movements (except for opercular and ocular movements) while on the bottom of the tank, as distinguished from immobility caused by sedation, which causes hypolocomotion, akinesia, and sinking8. Freezing is usually related to a high state of stress and anxiety and is also part of submissive behavior. Complex behaviors are excellent indicators of the state of anxiety of animals. NTT has been shown to be sensitive to pharmacological and genetic manipulation9, making it a valuable tool for studying the neural basis of anxiety and related disorders.
Zebrafish are a highly social species, so we can measure a wide range of social behaviors. The shoaling test (ST) and the social preference test (SPT) are the most used assays to assess social behavior10. The ST measures the tendency of fish to group together11 by quantifying their spatial behavior and movement patterns. ST is useful for studying group dynamics, leadership, social learning, and understanding the social behavior of many fish species12. The SPT in adult zebrafish was adapted from Crawley&#39;s preference for social novelty test for mice13&#160;and quickly became a popular behavioral assay for the study of social interaction in this model species14. These two tests have also been adapted for use in drug screening assays and have shown promise for identifying novel compounds that modulate social behavior15,16.
In general, behavioral assays in adult zebrafish are powerful tools that can provide valuable information on the behavior mechanisms or the neurophenotypes of active compounds and abused drugs17. This protocol details how to implement these behavioral tools7 with basic material resources and how to apply them in toxicity assays to characterize the effects of a wide range of neuroactive compounds. In addition, we will see that the same tests can be applied to assess the neurobehavioral effects of acute exposure to a neuroactive compound (methamphetamine) but also to characterize these effects after chronic exposure to environmental concentrations of a pesticide (glyphosate).

Autor em destaque: Avaliação de mudanças comportamentais ambientais e induzidas por drogas em peixes-zebra adultos 

Avaliação da Neurotoxicidade em Danio rerio Adulto utilizando uma Bateria de Testes Comportamentais em um Único Tanque

Our main research aims to identify the behavioral effects induced by environmental pollutants and drugs in fish. Also trying to understand the mechanisms leading to these effects. A social and anxiety-like behavior cannot be conveniently addressed using embryos.We conduct studies targeting these behaviors in the adult Zebrafish model. This protocol details how to implement behavioral tools with basic material resources to facilitate the reproducibility of tests and harmonize the results obtained within the scientific community. The assays presented in this protocol have proven to be highly reproducible and sensitive to genetic and pharmacological manipulations, making them variable and noninvasive tools fostering the neural and molecular mechanisms underlying behavior.Once a behavioral effort has been identified, the most challenging part is to determine the specific modes of action leading to the observative efforts. Starting by the molecular initiating event where predictions can provide some clues on the molecular initiating event, we are still far from obtaining accurate predictions. We are currently interested in the determining the phase validity of the chemical models of Parkinson's disease developed in embryonic and adult Zebrafish by using MPTP and rotenone with the same, the kinematic of the gait-like movement or during an explosive movement like the acoustic evoked escape response, will be evaluated.

Watch this Scientific Journal Video about Experimental Tank Configuration and Video Recording Setup for Behavioral Testing in Adult Zebrafish at JoVE.com

Configuração de Tanque Experimental e Configuração de Gravação de Vídeo para Testes Comportamentais em Zebrafish Adulto

Para começar, monte os tanques experimentais quadrados, câmeras e computadores. Para ensaios de comportamento individual de peixes-zebra. Encha o tanque com sete litros de água de peixe bem oxigenada a 28 graus Celsius.Ajuste a posição do tanque na frente da câmera para evitar imagens distorcidas. Usando a luz de fundo LED, verifique a configuração da iluminação e ligue a câmera. Em seguida, abra o gerenciador de câmera para verificar a disponibilidade da câmera jiggy no computador.Inicie o software de controle da câmera e abra a opção Câmera. Em seguida, selecione Modo Monocromático e ajuste o tamanho da imagem. Abra as propriedades da câmera e, em Câmera, defina o relógio de pixels como máximo e a taxa de quadros para 30 quadros por segundo.Em seguida, ajuste a exposição. Agora, em Imagem, defina o ganho como zero e ajuste os níveis de preto. Na guia Tamanho, ajuste as dimensões da janela para a região que requer gravação.Uma vez feito, feche as propriedades da câmera. Em seguida, crie uma pasta geral para a sessão de experimento para salvar as configurações e os vídeos da câmera. Em seguida, clique em Arquivo, Salvar parâmetros e em arquivo para salvar as configurações.Depois de configurar todas as câmeras, abra gravar sequência de vídeo e selecione Criar para salvar como um novo arquivo de vídeo. Selecione quadros máximos e insira 10, 800 na caixa de quadros. Em seguida, selecione Calcular taxa de quadros.Introduza os peixes um a um na parte inferior dos tanques experimentais para testes comportamentais e clique em Gravar para iniciar a gravação. Para o ensaio de cardume, introduzir o cardume no fundo do tanque experimental. Para o teste de preferência social, introduzir os peixes um a um no fundo dos diferentes tanques experimentais.Quando uma caixa de bate-papo com um número máximo de quadros atingidos aparecer na tela, clique em Aceitar. Em seguida, selecione Fechar para concluir a gravação. Por fim, retire os peixes registrados para testes comportamentais do tanque experimental e separe-os dos peixes ingênuos.