The presence of morphine-like and codeinelike substances was demonstrated in the pedal ganglia, hemolymph, and mantle tissues of the mollusc Mytilus edulis. The pharmacological activities of the endogenous morphine-like material resemble those of authentic morphine. Both substances were found to counteract, in a dose-dependent manner, the stimulatory effect of tumor necrosis factor ax or interleukin la on human monocytes and Mytilus immunocytes, when added simultaneously to the incubation medium. The immunosuppressive effect of this opiate material expresses itself in a lowering of chemotactic activity, cellular velocity, and adherence. Codeine mimics the activity of authentic morphine, but only at much higher concentrations. Specific high-affinity receptor sites (jx3) for morphine have been identified on human monocytes and Mytilus immunocytes. In Mytilus recovering from experimentally induced stress, the return of "alerted" immunocytes to a more inactive state appears to be due to a significant rise in the content of morphine-like material in the pedal ganglia and hemolymph at this time. Thus, morphine may have a role in calming or terminating the state of immune alertness.The occurrence throughout the animal kingdom of messenger molecules identical with, or closely related to, those known in mammals has been amply documented by biochemical identification, functional analysis, and the demonstration of specific receptor sites (1-6). This applies in particular to classical neurotransmitters, neuropeptides, and cytokines. The demonstration by Spector and coworkers (9-11) and others (7, 8) added to the sample in a 6:10 (vol/vol) ratio (HCl/sample) to a final strength of 10%]. The samples were then heated at 100°C for 30 min, cooled, and centrifuged at 10,000 x g for 20 min. The resulting precipitate was discarded and the supernatant was extracted with 5 vol of 10% (vol/vol) 1-butanol in chloroform. The supernatant was adjusted to pH 8.8-9.0. The organic phase was back-extracted into 2 vol of 0.01 M HCl. Morphine recovery was 50-70%. The aqueous phase was evaporated to dryness and dissolved in 10 ml of phosphate-buffered saline (PBS, pH 7.4) and then the pH was adjusted to 8.5-9.0. Samples were passed through a Sep-Pak C18 cartridge. Morphine and codeine were eluted from the cartridge with 7 ml of 0.1 M pyridine in acetic acid (pH 6.2) containing 2.5% (vol/vol) 1-propanol. The eluate was evaporated to dryness and dissolved in 250 ,ul of 1 mM HCl. An aliquot of 200 ,ul was injected onto a C18 reverse-phase HPLC column (LiChrosorb, 0.4 x 25 cm, Merck). The samples were eluted with 0.1 M pyridine in acetic acid (pH 6.2) followed by a linear gradient of 0-25% 1-propanol/0.1 M pyridine in acetic acid at a flow rate of 1.5 ml/min. Four 1-min fractions were collected during morphine and codeine elution as determined by known standards. These fractions were then evaporated, dissolved, and assayed for morphine and codeine by a sensitive radioimmunoassay (RIA) described by . Polyclonal antibodies generated against 3...
