base recording from taste sensilla of drosophila for analyzing responses of taste neurons

Um novo método para análise aprimorada da atividade dos neurônios gustativos do inseto

Insects, including drosophilid flies, are endowed with a sophisticated taste system that enables them to extract complex chemical information from their surroundings. This system allows them to discern the chemical composition of various substances, distinguishing between those that are nutritious and those that are harmful1,2.
At the core of this system are specialized structures known as taste hairs or sensilla, strategically located on various body parts. In drosophilid flies, these sensilla are located on the labellum, which is the major taste organ of the fly head1,2,3,4, as well as on the legs and wings1,2,5,6. The labellum is located at the tip of the proboscis and contains two lobes4,7,8. Each lobe is covered with 31 taste sensilla categorized as short, long, and intermediate4,7,8. These sensilla each house 2-4 taste neurons1,2,9,10. These taste neurons express members of at least four different gene families, namely the Gustatory receptor (Gr), Ionotropic receptor (Ir), Pickpocket (Ppk), and Transient receptor potential (Trp) genes1,2,11,12,13. This diversity of receptors and channels equips insects with the ability to recognize a wide array of chemical compounds, including both nonvolatile and volatile cues1,2,14.
For over 50 years, scientists have quantified the response of taste neurons and their receptors using a technique called tip recording3,4,6,8,13,15,16,17,18,19,20,21,22,23,24,25,26,27,28, 
29,30,31,32,33,34,35. However, this method has major limitations. First, neural activity can be measured only during contact with the stimulus, and not before or after contact. This limitation precludes the measurement of spontaneous spiking activity and prevents the measurement of OFF responses. Second, only tastants that are soluble in aqueous solutions can be tested.
These limitations can be overcome by a rarely used alternative electrophysiological technique called &#34;base recording.&#34; Here we describe this technique, which we have adapted from a method used by Marion-Poll and colleagues24, and show the crucial taste coding features that it can now conveniently measure14.

Autor em destaque: Avanços na quimiorrecepção - de receptores de odor de insetos a RNAs não codificantes

Gravação de base: uma técnica para analisar as respostas dos neurônios gustativos em Drosophila

Our lab investigates how animals detect and respond to odors, taste, and some pheromone. We've made significant strides in understanding the molecular basis of chemoreception. We've made several advances in the field, including the discovery of the first insect odor receptors, the discovery of the first insect taste receptors, and the elucidation of basic principles of the logic of odor and taste coding.This technique allows the measurement of test neuron activity before, during, and after the stimulus. Thus, it allows extensive analysis of OFF responses that occur after a taste stimulus. It also allows the measurement of taste neuron activity to the river of volatile compounds.The classical technique for taste electrophysiology was unable to measure taste responses to hydrophobic compounds such as long-chain pheromones that have very low solubility in water. However, with the help of this technique, we can overcome this limitation by separating the recording electrode from the glass stimulus capillary. Our research explores the functioning of the chemosensory systems in animals, focusing on their role in feeding and mating choices.We're currently examining non-coding RNAs and micro peptides in fruit flies'olfactory systems, and how mosquitoes and tsetse flies use these systems to find human hosts.

Gravação de base de Taste Sensilla de Drosophila para análise de respostas de neurônios gustativos

Para começar, coloque a lâmina contendo o labelo da drosófila no microscópio stage sob uma ampliação baixa. Usando um micromanipulador manual, insira o eletrodo de referência no olho oposto ao olho com o eletrodo de registro. Agora use um micro manipulador motorizado para focar a ponta do capilar de estímulo de vidro no final do campo de visão sob objetivas de baixa e alta ampliação.Em seguida, empregue um segundo micromanipulador motorizado sob baixa ampliação e aproxime o eletrodo de gravação do labelo. Sob alta ampliação, insira o eletrodo de gravação na base de um sensillum gustativo. Continue a inserção até que o som da atividade de disparo neuronal seja ouvido na saída de áudio do sistema IDAC.Assim que um sinal estável for detectado, pressione o botão iniciar gravação para iniciar a gravação do sinal usando o software associado ao sistema IDAC. Usando o manipulador motorizado, ajuste a ponta do capilar do vidro de estímulo para que cubra a ponta do sensillum gustativo. Para concluir o estímulo, retraia o capilar do estímulo de vidro do sensillum usando o manipulador motorizado.Os picos de Sensillum são categorizados em duas classes com base na amplitude. Picos maiores são atribuídos ao neurônio sensível amargo, enquanto picos menores são do neurônio responsivo ao açúcar. O neurônio sensível amargo no sensillum S5 respondeu ao odor de DEET, indicando ativação sensorial sem contato físico.Uma resposta on foi observada no sensillum I1 ao entrar em contato com um estímulo amargo, seguida por uma resposta off mais forte após o término do contato.

base-recording-from-taste-sensilla-drosophila-for-analyzing-responses

Research

JoVE Journal

Neuroscience

Base Recording from Taste Sensilla of Drosophila for Analyzing Responses of Taste Neurons

96 visualizações.  Yale University. Para começar, coloque a lâmina contendo o labelo da drosófila no microscópio stage sob uma ampliação baixa. Usando um micromanipulador manual, insira o eletrodo de referência no olho oposto ao olho com o eletrodo de registro. Agora use um micro manipulador motorizado para focar a ponta do capilar de estímulo de vidro no final do campo de visão sob objetivas de baixa e alta ampliação.Em seguida, empregue um segundo micromanipulador motorizado sob baixa ampliação e aproxime ...

Vídeo: Gravação de base de Taste Sensilla de Drosophila para análise de respostas de neurônios gustativos 

Base Recording: A Technique for Analyzing Responses of Taste Neurons in Drosophila

Insects taste the external world through taste hairs, or sensilla, that have pores at their tips. When a sensillum comes into contact with a potential food source, compounds from the food source enter through the pore and activate neurons within. For over 50 years, these responses have been recorded using a technique called tip recording. However, this method has major limitations, including the inability to measure neural activity before or after stimulus contact and the requirement for tastants to be soluble in aqueous solutions. We describe here a technique that we call base recording, which overcomes these limitations. Base recording allows the measurement of taste neuron activity before, during, and after the stimulus. Thus, it allows extensive analysis of OFF responses that occur after a taste stimulus. It can be used to study hydrophobic compounds such as long-chain pheromones that have very low solubility in water. In summary, base recording offers the advantages of single-sensillum electrophysiology as a means of measuring neuronal activity - high spatial and temporal resolution, without the need for genetic tools - and overcomes key limitations of the traditional tip recording technique.

A rarely used method of electrophysiological recording, base recording, allows analysis of features of taste coding that cannot be examined by conventional recording methods. Base recording also allows the analysis of taste responses to hydrophobic stimuli that cannot be studied using traditional electrophysiological methods.

Insects taste the external world through taste hairs, or sensilla, that have pores at their tips. When a sensillum comes into contact with a potential ...