large-scale extraction of crude sex pheromones from 6-day-old virgin c. elegans hermaphrodites

<meta charset="utf8"></head><body>Extração e quantificação de respostas voláteis de feromônios sexuais em C. elegans

The interplay between chemical communication and reproductive success is a fundamental principle across the animal kingdom1,2,3,4,5,6,7,8,9,10. Sex pheromones trigger a wide array of sexually dimorphic behaviors essential for locating mates, coordinating the steps involved in finding and attracting a partner, and ultimately promoting the propagation of a species11,12,13,14,15,16,17. Significant progress has been made in understanding pheromone signaling, but the molecular mechanisms, neural circuits, and computational processes governing these interactions often remain incompletely defined18,19,20,21,22,23,24,25,26.
The nematode Caenorhabditis elegans provides a powerful model for dissecting these questions. Notably, C. elegans exhibits an unusual reproductive strategy-hermaphrodites can self-fertilize but also outcross with males27,28,29,30,31,32,33. This flexibility requires a robust communication system to signal reproductive status. C. elegans is known for its well-characterized water-soluble pheromones, the ascarosides, which play varied roles in development, behavior, and social interactions. Recent discoveries have unveiled a distinct class of volatile sex pheromones employed by nematodes. These pheromones are specifically produced by sexually mature C. elegans and C. remanei virgin females and sperm-depleted hermaphrodites, serving as an attractant for adult males29,34,35. This attractant exhibits remarkable sexual dimorphism in its production and perception. The female somatic gonad governs the synthesis of this volatile sex pheromone, and production dynamically reflects reproductive status, ceasing upon mating and resuming several hours later29,34.
Understanding nematode sex pheromone communication provides insights into the evolution of chemical communication systems, the interplay between reproductive state and behavior, and the mechanisms underlying sexually dimorphic neural processing24,26,36,37,38,39. Studies implicate the amphid neuron AWA in males as critical for pheromone detection, with the G-protein-coupled receptor SRD-1 playing a key role in pheromone detection in males24. C. elegans is well-suited for studying animal chemical communication, especially sex pheromone signaling, due to its reliance on the olfactory system for mate-searching. While much is known about ascaroside signaling, the volatile sex pheromone system offers unique opportunities for comparison25,26,36,40,41,42,43,44,45,46,47,48,49,50, 
51,52,53,54,55,56,57. Moreover, C. elegans is a powerful genetic model organism due to its fully sequenced genome, clearly defined cellular lineage, and well-characterized olfactory mutants.
However, the complete neural circuitry involved in processing this pheromone, the computations that translate its perception into targeted mate-searching behaviors, and its biosynthesis regulation remain to be fully elucidated. Further investigations into these processes are crucial for understanding the diverse mechanisms governing animal communication and reproductive behaviors. The identification of key genes involved in pheromone synthesis, secretion, and perception promises to unveil novel molecular players in animal communication.&#160;The assays&#160;described here provide a basis to address these&#160;questions.

<meta charset="utf8"></head><body>Autor em destaque: examinando a influência volátil do feromônio sexual no comportamento masculino de C. elegans

Extração de feromônios sexuais voláteis e ensaio de quimioatração em Caenorhabditis elegans

Our research examines how the volatile sex pheromone affects male C.elegans behavior using chemotaxis assay. It aims to identify the molecular mechanism of pheromone production and the neuronal mechanism of it is perception. Individual worm to worm variability in response to pheromone, coupled with the influence of environmental factors such as temperature, humidity, and light, can introduce significant noise into the data.Moreover, the complex nature of worm behaviors complicates data interpretation. Addressing those challenges requires careful experimental design. The protocol is straight forward and highly reproducible.It is designed to optimize pheromone yet while minimizing time and resource consumption. The chemotaxis assay provides quantitative data on male attraction, enabling statistical analysis and comparison. This protocol provides a completed picture of pheromone communication.

Extração em larga escala de feromônios sexuais brutos de hermafroditas C. elegans virgens de 6 dias

large-scale-extraction-crude-sex-pheromones-from-6-day-old-virgin-c

Research

JoVE Journal

Behavior

Large-Scale Extraction of Crude Sex Pheromones from 6-Day-Old Virgin C. elegans Hermaphrodites

148 visualizações. Depois de sincronizar os vermes Caenorhabditis Elegans adultos saudáveis, lave-os cinco vezes com tampão M9. Para sincronizar worms no estágio L1, gire-os no buffer M9 por 12 a 15 horas para interromper o desenvolvimento. Observe os vermes L1 presos em uma placa de cultura NGM de 10 centímetros semeada com bactérias OP50.Para minimizar a presença de machos em uma placa hermafrodita, verifique a placa dois dias após a liberação do verme e remov...