The aim of the present study was to commence a characterisation of some of the basic pharmacological properties of venom from the soldierfish (Gymnapistes marmoratus). Soldierfish venom was prepared by extraction into 10% glycerol and centrifugation to remove insoluble material. Protein content was determined and venom concentrations were expressed as μg venom protein. Soldierfish venom (0.5 15 μg/ml) produced concentration-dependent contractile responses in guinea-pig isolated ileum (GPI) and longitudinal smooth muscle (LSM) preparations. The muscarinic receptor antagonist atropine (10 nM) significantly inhibited responses of LSM to soldierfish venom (2.5 μg/ml). Responses to soldierfish venom (4 5 μg/ml) in GPI were not significantly affected by the ganglion-blocking drug mecamylamine (10μM) or by incubation with blood cholinesterase. The cyclooxygenase inhibitor indomethacin (2 μM) significantly inhibited responses to soldierfish venom (2.5 μg/ml) in LSM. Neither the thromboxane A2/prostaglandin H2 receptor antagonist GR32191B (1 μM) nor the leukotriene receptor antagonist S B205312 (10 nM) significantly affected responses to soldierfish venom (5μg/ml) in GPI. Responses to soldierfish venom (2.5 5 μg/ml) were not significantly inhibited by the histamine receptor antagonist mepyramine (0.5 μM), the angiotensin-converting enzyme inhibitor captopril (2μM) or the neurokinin-1 receptor antagonist CP-99,994 (0.1 μM) in LSM. The angiotensin AT1 receptor antagonist EXP3174 (0.1 μM) also tailed to inhibit significantly the responses to soldierfish venom (5 μg/ml) in GPI. A fluorometric assay for the detection of 5-hydroxytryptamine (5-HT) and related compounds indicated a level in soldierfish venom of 1.60 ± 0.01 ng of 5-HT-like substance per μg venom protein. Soldierfish venom (0.5-10 μg/ml) produced concentration- dependent contractile responses in rat isolated stomach fundus strips, and these responses (2.5 μg/ml) were significantly inhibited by the 5-HT1/5- HT2 receptor antagonist methysergide (0.1 μM). These results suggest that soldierfish venom may stimulate the release of acetylcholine to act at muscarinic receptors on guinea-pig gastrointestinal smooth muscle. The venom also appears to be causing the release of cyclooxygenase products, such as prostaglandins, and contains 5-HT, or a 5-HT-like substance, that acts directly at 5-HT receptors.