Efficacy of Administration Routes in Crayfish: Comparative Analysis of Intracoelomic and Intrapericardial Techniques Using Fluorescein Dye

Crayfish are emerging as model organisms for various disciplines. Moreover, decapod crustaceans also exhibit pain-like reactions and heightened anxiety when exposed to harmful stimuli, leading to short-term or persistent behavioral shifts. Awareness of decapod crustacean sentience and thus, suffering calls for refinement of current laboratory protocols. This study aims to enhance the standard methodology for injecting substances into crayfish by minimizing stress-inducing manipulation. We examined the impacts of various administration routes on the persistence of injected chemicals in marbled crayfish, its excretion, and animal survival. Fluorescein dye was used as a visual marker. It was administered via three alternative injection routes-intracoelomic (IC), intrapericardial administration through areola (IP-A), and intrapericardial administration through arthrodial membrane (IP-AM). Continuous video observations were made for a 4-h period under UV light, followed by intermittent observations at 12-h intervals over 48 h. The highest mortality (20%) was observed in IP-A administration. The IP-A method also provided the fastest systemic distribution of the dye in the body. Results indicated visibly higher urination frequency in IP-AM compared to IP-A. IC mirrored IP-AM outcomes without any observed mortality. We conclude that IC administration proved superior to intrapericardial methods, offering the least harmful but effective approach for crayfish injections.