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Nitric oxide (NO) signalling is at the forefront of intense research interest because its many effects remain controversial and seemingly contradictory. This paper examines its role as a potential mediator of pain and tolerance. Within this context discussion covers endogenous morphine, documenting its ability to be made in animal tissues, including nervous tissue, and in diverse animal phyla. Supporting morphine as an endogenous signalling molecule is the presence of the newly cloned mu3 opiate receptor subtype found in animal (including human) immune, vascular and neural tissues, which is coupled to NO release. Importantly, this mu opiate receptor subtype is morphine-selective and opioid peptide-insensitive, further highlighting the presence of morphinergic signalling coupled to NO release. These findings provide novel insights into pain and tolerance as morphinergic signalling exhibits many similarities with NO actions. Taken together, a select morphinergic signalling system utilising NO opens the gate for the development of novel pharmaceuticals and/or the use of old pharmaceuticals in new ways.

The parasitic worm Ascaris suum contains the opiate alkaloid morphine as determined by HPLC coupled to electrochemical detection and by gas chromatography/mass spectrometry. The level of this material is 1168 ± 278 ng/g worm wet weight. Furthermore, Ascaris maintained for 5 days contained a significant amount of morphine, as did their medium, demonstrating their ability to synthesize the opiate alkaloid. To determine whether the morphine was active, we exposed human monocytes to the material, and they immediately released nitric oxide in a naloxone-reversible manner. The anatomic distribution of morphine immunoreactivity reveals that the material is in the subcuticle layers and in the animals’ nerve chords. Furthermore, as determined by RT-PCR, Ascaris does not express the transcript of the neuronal μ receptor. Failure to demonstrate the expression of this opioid receptor, as well as the morphine-like tissue localization in Ascaris, suggests that the endogenous morphine is intended for secretion into the microenvironment.

Samples of mosquitoes in the Culex pipiens L. complex from Memphis, Tenn., were collected from June to November 1985 and examined in regard to allozyme frequencies and ratios of the two arms of the phallosome of the male genitalia (DV/D). The dominant allozymes of hexokinase (HkA) and 6-phosphogluconate dehydrogenase (PgdF) significantly increased in frequency during this period as did the mean DV/D ratio. An analysis of gene frequencies by species group designated by DV/D ratios revealed no significant differences among Cx. pipiens, Cx. quinquefasciatus Say, and intermediates. The lack of association of gene frequencies with the taxa determined by the DV/D ratio indicated that although allozyme frequencies were correlated temporally with the DV/D ratio in the population, they were not associated with subspecies. These results are consistent with previous work that has shown latitudinal association and thermal stability differences in the major allozymes of these enzymes in the Cx. pipiens complex.

The tissue distribution, course of secretion, and sex differences of morphine were delineated in Ascaris suum. Nitric oxide (NO) release in various tissues in response to morphine and its metabolite morphine-6-glucuronide (M6G) were also examined. Ascaris suum of both sexes along with their incubation fluid were analyzed for morphine concentrations by high-performance liquid chromatography (HPLC) over a 5-day period. Various tissues were also dissected for HPLC and NO analysis. Morphine was found to be most prevalent in the muscle tissue, and there is significantly more morphine in females than males, probably because of the large amounts present in the female uterus. Morphine (10(-9) M) and M6G (10(-9) M) stimulated the release of NO from muscles. Naloxone (10(-7) M) and N-nitro-L-arginine methyl ester (10(-6) M) blocked (P < 0.005) morphine-stimulated NO release from A. suum muscle tissue. D-Phe-Cys-Tyr-D-Trp-Om-Thr-Pen-Thr-NH2 (CTOP) (10(-7) M) did not block morphine's NO release. However, naloxone could not block M6G-stimulated NO release by muscles, whereas CTOP (10(-7) M) blocked its release. These findings were in seeming contradiction to our earlier inability to isolate a mu opiate receptor messenger RNA by reverse transcriptase-polymerase chain reaction using a human mu primer. This suggests that a novel mu opiate receptor was possibly present and selective toward M6G.

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