2004 ACVP Student Poster Award Abstract

EFFECTS OF ANESTHETICS ON NEUTROPHIL FUNCTION IN FISH
D.M. Herolt1a+, D. Pali_2a, C.B. Andreasen3a, and J.A. Roth4a
1Veterinary Summer Scholar Research Program, Department of 2Natural Resource Ecology and Management, 3Veterinary Pathology, 4Veterinary Microbiology and Preventive Medicine, aCollege of Veterinary Medicine, Iowa State University, Ames, IA

Anesthetic efficacy and two parameters of neutrophil function, oxidative burst and degranulation of primary granules, were compared among three anesthetics in the fathead minnow (Pimephales promelas, Rafinesque 1820): tricaine methanesulfonate (MS-222), clove oil (EUG) and metomidate (MTMD).

Adult fish were maintained in a recirculation system. To test the effects of anesthetics on neutrophil function, fish were divided into six treatment groups, a baseline level control group, and a positive control group exposed to the same handling and crowding stress, but without use of anesthetic. Fish were assayed for neutrophil function at 0h (before treatment), 24h, 72h and 7 days. Reduction of cytochrome C was used to detect superoxide generation and measure oxidative burst. The degranulation of primary granules was measured as exocytosis of myeloperoxidase (MPO) using 3, 3’, 5, 5’-tetramethylbenzidine as a substrate.

The optimum anesthetic concentration was established as: MS-222 75 mg L-1, EUG 30 mg L-1 and MTMD 4 mg L-1. The cytochrome C assay did not reveal significant changes in superoxide generation. Degranulation of primary granules was decreased to 75% of baseline (100%) level activity 24h post treatment with MS-222, but was not significantly affected when MTMD and EUG were used. In all experiments, fish from the positive control group showed a significant (P < 0.001) decrease in degranulation (60-75% of baseline level) after 24h.

We conclude that MS-222 did not, while metomidate and eugenol did, prevent a decrease in degranulation. Exocytosis of MPO proved to be a viable assay for measuring effects of stress on neutrophil function in fish.

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