Development of behavioural and physiological assays to assess discrimination of male and female odours in crayfish, Pacifastacus leniusculus

Many aquatic organisms use chemical signals to coordinate courtship. However, relatively few water-borne pheromones have been identified. A key obstacle hindering progress in the purification of crustacean pheromones has been the development of reliable bioassays. This study focuses on developing novel bioassays to guide the purification of sex pheromones in signal crayfish, Pacifastacus leniusculus. We aimed to elicit specific sexual behaviours in male crayfish in response to female urine or conditioned water released from a female dummy. A physiological assay, based on male heartbeat recordings, was developed to assess if physiological tests could provide quicker and more sensitive responses than the behavioural assay. Males exposed to female urine showed significantly increased levels of specific mounting behaviours in comparison to male urine or control water. However, other sexual behaviours such as seizing, turning and spermatophore deposition were not observed. The physiological assay demonstrated that a rapid change in the heart rate of male crayfish could be induced through exposure to odour from conspecific female crayfish. Our study indicates male crayfish can discriminate between male and female odours. Physiological measures provide a quick assay for sensitivity to a substance whilst behavioural assays indicate its functional significance.