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1. Opioid (mu, delta, kappa) and cannabinoid (CB1, CB2) receptors are coupled mainly to Gi/Go GTP-binding proteins. The goal of the present study was to determine whether different subtypes of opioid and cannabinoid receptors, when coexpressed in the same cell, share a common reservoir, or utilize different pools, of G proteins. 2. The stimulation of [35S]GTPgammaS binding by selective opioid and cannabinoid agonists was tested in transiently transfected COS-7 cells, as well as in neuroblastoma cell lines. In COS-7 cells, cotransfection of mu- and delta-opioid receptors led to stimulation of [35S]GTPgammaS binding by either mu-selective (DAMGO) or delta-selective (DPDPE) agonists. The combined effect of the two agonists was similar to the effect of either DAMGO or DPDPE alone, suggesting the activation of a common G-protein reservoir by the two receptor subtypes. 3. The same phenomenon was observed when COS-7 cells were cotransfected with CB1 cannabinoid receptors and either mu- or delta-opioid receptors. 4. On the other hand, in N18TG2 neuroblastoma cells, which endogenously coexpress CB1 and delta-opioid receptors, as well as in SK-N-SH neuroblastoma cells, which coexpress mu- and delta-opioid receptors, the combined effects of the various agonists (the selective cannabinoid DALN and the selective opioids DPDPE and DAMGO) were additive, implying the activation of different pools of G proteins by each receptor subtype. 5. These results suggest a fundamental difference between native and artificially transfected cells regarding the compartmentalization of receptors and GTP-binding proteins.

1. The present study reexamines a previous notion on opioid stimulation of cyclic GMP (cGMP) formation and the retraction of the original findings. 2. The effect of opioid agonists on cGMP accumulation in two cell lines of neuronal origin was measured. The proportion of cGMP stimulation in NG108-15 neuroblastoma x glioma hybrid cells resembled the proportion of [Ca2+]in elevation by opioids in this culture. The failure of opioids to stimulate cGMP formation in SK-N-SH human neuroblastoma coincided with the lack of cGMP stimulation by other Ca2+ mobilizing agents in these cells. The nitric oxide donor nitroprusside elevated cGMP in both cell lines. 3. The implication of the opioid-Ca(2+)-NO-cGMP cellular pathway for opioid activity in vivo is discussed.

1. Aggregating fetal rat brain cells express a significant amount of proenkephalin A (PENK) mRNA, a selective radioimmunoassay shows that this mRNA is also translated into enkephalins. 2. Depolarization with potassium chloride (KCl) or veratridine increases the expression of PENK mRNA in a time-dependent fashion, with a maximal increase of sixfold. It is interesting, however, that depolarization of the same cultures with KCl has no effect on the expression of prodynorphin mRNA. 3. An increase in PENK mRNA levels has been also observed in cultures treated with 8-Br-cAMP, phorbol 12-myristate-13-acetate (TPA), or dexamethasone. 4. However, incubation of the cultures with the opioid agonist etorphine or the antagonist naltrexone did not alter PENK gene expression, suggesting that there is not feedback control of opioids on PENK biosynthesis in these cells. 5. The increase in PENK mRNA in depolarized and in TPA-dexamethasone-, or 8-Br-cAMP-treated cultures was not accompanied by a significant increase in the amount of free immunoreactive met-enkephalin. Fetal brain cell cultures are therefore a useful neuronal model system for studying the mechanism that regulated the expression of PENK mRNA.

The dopamine-stimulated adenylate cyclase activity was studied both in vivo and in vitro in the central nervous system of the bivalve mollusc Mytilus edulis. Dopamine, epinine, and apomorphine stimulated the enzyme system. Fluphenazine, haloperidol, chlorpromaxine, and to a lesser extent BOL inhibited the dopamine-stimulated adenylate cyclase. Etorphine, beta-endorphine, DALA, and methionine enkephalin depressed cyclic AMP levels. This phenomena was naloxone reversible. In addition, the opioids inhibited the stimulation of adenylate cyclase by dopamine. This phenomena was also naloxone reversible. The study demonstrates an interaction among dopamine, the opioids, and cyclic AMP.

1. This report further demonstrates that etorphine influences presynaptic dopamine release, which in turn centrally modulates peripheral cilioinhibition. 2. In older animals cilioinhibition has become enhanced due to a lack of responsiveness to endogenous opioids which results in greater dopamine release, causing a higher level of cilioinhibition as demonstrated by challenging the visceral ganglia with etorphine or destroying the dopaminergic component with 6-hydroxydopamine. 3. Only the central cilioinhibitory, not the peripheral inhibitory response, mechanism appears to be altered in older animals. Thus, the alteration appears in the central integrative mechanisms involved with regulating ciliary activity. 4. The KCl-stimulated release of dopamine is unaltered in both young and old organisms, whereas the opiate inhibition of the KCl-stimulated release of dopamine is reduced in older organisms. Thus, the aging-associated alteration is associated with a specific process. 5. The reduction of opioid influence and the resulting enhanced cilioinhibitory activity may make the organisms more susceptible to environmental stress.

A commercial neuroleptanalgesic acepromazine-etorphine combination administered intramuscularly to four horses produced a severe tachycardia and an increase in muscular tone, together with hypoxaemia, hypercapnia, metabolic acidosis associated with an increase in the packed cell volume and hyperglycaemia. No electrolyte changes were found. After reversal of the action of etorphine with diprenorphine, there was a prolonged decrease in the calcium and phosphorus serum concentrations and decreases in the packed cell volume and the total protein serum concentration. In a second experiment on the same four horses, glyceryl guaiacolate (10 g/100 kg body weight intravenously) was given as soon as the horses were anaesthetized with acepromazine-etorphine. The muscular rigidity disappeared and the tachycardia was less evident. There was a more pronounced hypoxaemia but the changes in the other parameters were similar to those in the first experiment. It was concluded that the neuroleptanalgesic-glyceryl guaiacolate combination is not a safe anaesthetic procedure in horses.

Opioid receptors and enkephalinergic neurons in the central nervous system of Mytilus edulis have been reported. Also known is that the lateral epithelium of the gill is innervated by serotonergic, cilioexcitatory neurons and dopaminergic, cilioinhibitory neurons. The aim of the present report is to look for an effect of opioid agonists on the nervous control of the lateral cilia. Dopamine applied to the cerebral ganglion inhibited the activity of lateral cilia in the gill. This effect was blocked by the application of several opioids to the visceral ganglion. The block was reversed by the application of naloxone to the visceral ganglion. Dopamine applied to the visceral ganglion also inhibited lateral ciliary activity as shown earlier. Opioids applied to the visceral ganglion partially blocked this effect but this was overcome by higher concentrations of dopamine. Preparations with low endogenous rates of ciliary beating were stimulated by the application of opioids to the visceral ganglion. Naloxone blocked this effect. Preparations with high endogenous rates of ciliary beating were inhibited by the application of naloxone to the visceral ganglion. Electrical stimulation of the cerebrovisceral connective produced excitatory and inhibitory effects depending on the rate of stimulation. Morphine applied to the visceral ganglion diminished the cilioinhibitory effects and enhanced the cilioexcitatory effects of electrical stimulation. Morphine applied to the gill had no effect on the cilioinhibitory action of dopamine applied to the visceral ganglion. There was no observable effect of opioids applied to the gill and no alteration in the cilioinhibitory effect of dopamine or the cilioexcitatory effect of serotonin applied directly to the gill in the presence of opioids. Specific opioid binding sites were found in the visceral ganglion but were not found in gill, palp, mantle, or visceral mass tissue. A dopamine-stimulated adenylate cyclase activity was again found in the visceral ganglion and the gill. Etorphine reduced the dopamine stimulation of cyclase in the ganglion but not in the gill. It is postulated that a cilioinhibitory, dopaminergic mechanism includes nerves running from the cerebral ganglion to the gill with synaptic transmission in the visceral ganglion that can be modulated by opioids.

As Mytilus edulis (Bivalvia) ages, the number of stereospecific opiate binding sites in the visceral ganglia significantly decreases. The number of high-affinity etorphine sites decreased by 30%; the number of naloxone sites decreased by 23%. The affinity of the ligands did not vary with age. In addition, the augmentation by opioids of dopamine levels in the visceral ganglia was less in older animals and was accompanied by a reduced sensitivity of the ganglia to etorphine and DAMA (D-Ala2,Met5-enkephalin-amide).

The effects of dopamine and octopamine on adenylate cyclase activity were studied on the head homogenate of adult Culex pipiens mosquitoes in vitro. Both dopamine and octopamine were shown to increase the cyclic AMP content in the homogenate. The antagonist haloperidol blocked the production of cyclic AMP induced from dopamine but had no effect on the production of cyclic AMP induced by octopamine at the concentrations tested. The opiate agonist etorphine was ineffective at reducing cyclic AMP levels induced by either dopamine or octopamine at the concentrations tested.

Extensive conformational calculations were performed on the potent opiate analgesics etorphine, PET, R30490 and etonitazene to determine all of their many low-energy conformations. The results were used to characterize four possible models for binding of a simple pharmacophore, comprising two phenyl rings plus a protonated nitrogen, to opiate analgesic receptors. These four models may define the necessary three-dimensional features leading to particular opiate actions. The model favoured for mu receptor activity can accommodate a protonated nitrogen, an aromatic ring (which may be substituted with an electronegative group) and a second lipophilic group. These structural features must be presented in a precise three-dimensional arrangement. It appears likely that a hydrophilic substituent in a certain region of the analgesic pharmacophore may also interact with the receptor as a secondary binding group.