Effects of anesthetic compounds on responses of earthworms to electrostimulation

Earthworms play an important role in biomedical research, and some surgical procedures require anesthesia. Anesthetic treatments used so far usually induce convulsive body movements connected with extrusion of coelomocyte-containing coelomic fluid that may affect experimental results. Extensive movements connected with the expulsion of coelomic fluid are exploited by immunologists as a method of harvesting immunocompetent coelomocytes from worms subjected to mild electrostimulation (4.5V). The aim of the investigations was to find anesthetic drugs without unintentional coelomocyte depletion. Experiments were performed on adult specimens of Dendrobaena veneta, the coelomocytes of which consist of amoebocytes and riboflavin-storing eleocytes. Earthworm mobility was filmed and extrusion of coelomocytes was quantified by detection of eleocyte-derived riboflavin in immersion fluid. Treatments included earthworms (1) immersed either in physiological saline (controls) or in a solution of one of the tested anesthetic drugs; (2) electrostimulated immediately after anesthesia, and (3) electrostimulated a second time after a 1-hour recovery period. The well-established fish and amphibian anesthetic agent MS-222 induced coelomocyte expulsion. In contrast, solutions of the mammalian local anesthetic drug, prilocaine hydrochloride (0.25-0.5%, 5-10 min) caused temporal earthworm immobilization followed by recovery, thus showing utility as an efficient earthworm anesthetic.