Opioid enhancement of evoked [Met5]enkephalin release requires activation of cholinergic receptors: possible involvement of intracellular calcium (original) (raw)
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Proceedings of the National Academy of Sciences, 1991
This laboratory has previously demonstrated that there is an opiate receptor-mediated, concentrationdependent modulation of the electrically stimulated release of enkephalin from the guinea pig myenteric plexus. Low doses of opioids (nanomolar) enhance release, whereas higher concentrations (10-100 nM) inhibit release. We now demonstrate that the in vivo i.p. administration of the islet-activating protein from pertussis toxin (PTX; 50 jtg/500 g of body weight) markedly diminishes the potency of jA, 6, or K-selective opioids to inhibit the evoked release of enkephalin. In contrast, PTX is without effect on the enhancement of enkephalin release observed after treatment with nanomolar concentrations of the above opioids. Conversely, pretreatment with cholera toxin (CTX; 0.01 nM for 3 hr in vitro) has no effect on the , 6, or K opioid inhibition of evoked enkephalin release but abolishes the ability of nanomolar concentrations of these agonists to enhance stimulated enkephalin release. These data indicate that different classes of guanine nucleotide-binding proteins (G proteins) appear to mediate the opioid enhancement or inhibition of stimulated enkephalin release. Furthermore, they suggest that a PTX-sensitive G protein (G1 or Go) and a CTX-sensitive G protein (Gs) are integral components of the mechanism that mediates opioid inhibition and opioid enhancement, respectively, of evoked enkephalin release. To our knowledge, this report represents the first demonstration that Gs-coupled opiate receptors (in addition to those that are coupled to G.) can modulate transmitter release.
Neuroscience Letters, 1992
The possible opioid control through ~,/t and x receptors of the spinal release of Met-enkephalin-like material (MELM) was investigated in halothane-anaesthetized rats. The intrathecal perfusion of the 6 agonist DTLET (10 pM) or the/1 agonist DAGO (10 #M) resulted in a marked inhibition of MELM release, which could be prevented by the selective antagonists naltrindole and naloxone, respectively. Although the ~c agonist U 50488 H (10/~M) was inactive per se, it completely suppressed the inhibitory effect of DAGO, without affecting that of DTLET. As the selective x antagonist norbinaltorphimine blocked the action of U 50488 H, it can be concluded that x receptors modulate the ,u-(but not the 6-) mediated feed back control of spinal enkephalinergic neurones.
European Journal of Pharmacology, 1992
The release of substance P (SP) from spinal dorsal horn slices is partially inhibited by micromolar concentrations of selective 6-opioid receptor agonists. In the present study, we have examined the effect of nanomolar concentrations of [D-PenZ,D -' Pen5]enkephalin (DPDPE, 6-opioid receptor agonist) and low micromolar of concentrations morphine on K+-evoked SP release from rat trigeminal nucleus caudalis (TNC) slices. DPDPE and morphine inhibited SP release with an apparent maximal effect at 3 nM and at 3/xM, respectively. DPDPE and morphine produced U-shaped concentration-response curves that were completely autoinhibited at 100 nM DPDPE and 1 /zM morphine. The inhibition of SP release produced by 3 nM DPDPE and 3 /zM morphine was blocked by the opioid receptor antagonists naloxone (30 nM; non-selective) and ICI 174,864 (0.3/xM; &selective) but not by nor-binaitorphimine (3 nM n-BNI; K-selective), naloxonazine (1 nM; ~cselective) or /3-funaltrexamine (20 nM /3-FNA; ~-selective). These findings indicate that 6-opioid receptor-mediated inhibition of SP release from TNC can be achieved by nanomolar concentrations of selective 6-opioid receptor agonists. Activation of ~-opioid receptors by morphine might be involved in the residual analgesia observed after/Zl-opioid receptor blockade and in the analgesia produced by high doses of morphine.
European Journal of Pharmacology, 1992
The release of substance P (SP) from spinal dorsal horn slices is partially inhibited by micromolar concentrations of selective 6-opioid receptor agonists. In the present study, we have examined the effect of nanomolar concentrations of [D-PenZ,D-' Pen5]enkephalin (DPDPE, 6-opioid receptor agonist) and low micromolar of concentrations morphine on K+-evoked SP release from rat trigeminal nucleus caudalis (TNC) slices. DPDPE and morphine inhibited SP release with an apparent maximal effect at 3 nM and at 3/xM, respectively. DPDPE and morphine produced U-shaped concentration-response curves that were completely autoinhibited at 100 nM DPDPE and 1 /zM morphine. The inhibition of SP release produced by 3 nM DPDPE and 3 /zM morphine was blocked by the opioid receptor antagonists naloxone (30 nM; non-selective) and ICI 174,864 (0.3/xM; &selective) but not by nor-binaitorphimine (3 nM n-BNI; K-selective), naloxonazine (1 nM; ~cselective) or /3-funaltrexamine (20 nM /3-FNA; ~-selective). These findings indicate that 6-opioid receptor-mediated inhibition of SP release from TNC can be achieved by nanomolar concentrations of selective 6-opioid receptor agonists. Activation of ~-opioid receptors by morphine might be involved in the residual analgesia observed after/Zl-opioid receptor blockade and in the analgesia produced by high doses of morphine.
Feedback inhibition of met-enkephalin release from the rat spinal cord in vivo
Synapse, 1992
The possible existence of a feedback control by endogenous opioids of the spinal release of met-enkephalin-like material was assessed in vivo, in halothane-anesthetized rats whose intrathecal space was continuously perfused with an artificial cerebrospinal fluid supplemented with various opioid-related drugs. Both the intrathecal perfusion of the p agonist D-Ala2-D-MePhe4-Gly-o15-enkephalin (DAGO) (10 pM) and the a agonist Tyr-D-Thr-Gly-Phe-Leu-Thr (DTLET) (10 pM) produced a significant inhibition of the spinal outflow of met-enkephalin-like material. The effect of DAGO, but not that of DTLET, could be prevented by naloxone (10 pM), and, conversely, the effect of DLTET, but not that of DAGO, was no longer observed in the presence of naltrindole (10 pM). Therefore naloxone and naltrindole acted as potent and selective p and a antagonists, respectively, when perfused at 10 pM in the intrathecal space of halothane-anesthetized rats. As expected from the lack of a tonic opioid control of spinal enkephalinergic neurones, neither naloxone nor naltrindole alone affected the spontaneous outflow of metenkephalin-like material. However, naltrindole, but not naloxone, markedly increased the spinal overflow of met-enkephalin-like material due to intrathecal administration of either porcine calcitonin (10 p M) or the peptidase inhibitors thiorphan (10 pM) plus bestatin (20 pM). These data suggest that a, but not p, receptors are involved in aphasic opioid inhibitory control of the release of met-enkephalin-like material in the rat spinal cord. This control would be functional only when the concentration of extracellular endogenous opioid(s) reaches a critical level, as probably occurring in pain-suffering subjects. Opioid antagonists acting selectively on the receptors involved in this feedback control might constitute a new class of analgesic drugs. o 1992 WiIey-Liss, Inc.
Proceedings of the National Academy of Sciences, 1987
Experiments were performed in order to determine whether the state of tolerance to and dependence upon opiates is associated with changes in one or more of the characteristics of the electrically induced release of methionine enkephalin from enteric ganglia. Acute morphine pretreatment substantially reduces the magnitude of the evoked release of this peptide from opiate-naive ilea. However, the rate of the evoked release of enkephalin from morphine-pretreated, tolerant/dependent preparations is indistinguishable from that observed for untreated, naive ilea. Paradoxically, 15 min after acute in vitro withdrawal of morphine from such preparations, the presence of morphine appears to be a prerequisite for the manifestation of electrically evoked release of methionine enkephalin. The evoked release of this peptide from ilea 60 min after withdrawal is no longer dependent upon morphine. Moreover, the magnitude of the increase in the rate of enkephalin release from these preparations is almost double that observed for opiate-naive ilea. These data indicate that the manifestation of opiate tolerance/dependence for the release of methionine enkephalin from enteric ganglia comprises several adaptive processes, the consequences of which can be observed at different stages of withdrawal.
Neurochemical Research, 1996
Opiates and opioid peptides carry out their regulatory effects mainly by inhibiting neuronal activity. At the cellular level, opioids block voltage-dependent calcium channels, activate potassium channels and inhibit adenylate cyclase, thus reducing neurotransmitter release. An increasing body of evidence indicates an additional opposite, stimulatory activity of opioids. The present review summarizes the potentiating effects of opioids on transmitter release and the possible cellular events underlying this potentiation: elevation of cytosolic calcium level (by either activating Ca 2. influx or mobilizing intracellular stores), blockage of K + channels and stimulation of adenylate cyclase. Biochemical, pharmacological and molecular biology studies suggest several molecular mechanisms of the bimodal activity of opioids, including the coupling of opioid receptors to various GTP-binding proteins, the involvement of different subunits of these proteins, and the activation of several intracellular signal transduction pathways. Among the many experimental preparations used to study the bimodal opioid activity, the SK-N-SH neuroblastoma cell line is presented here as a suitable model for studying the complete chain of events leading from binding to receptors down to regulation of transmitter release, and for elucidating the molecular mechanism involved in the stimulatory effects of opioid agonists.
Opioid inhibition of synaptic transmission in the guinea-pig myenteric plexus
British Journal of Pharmacology, 1985
Intracellular recordings were made from neurones in the myenteric plexus of the guinea-pig ileum. Presynaptic nerves were excited by a focal stimulating electrode on an interganglionic strand. 2 Fast excitatory postsynaptic potentials (e.p.s.ps) were depressed in amplitude by morphine and [Met5]enkephalin in the concentration range of I nM-I gM. Nicotinic depolarizations evoked by exogenously applied acetylcholine (ACh) were not affected by these opioids. 3 Hyperpolarization of the presynaptic fibres probably contributed to the depression of the fast e.p.s.p. because fast e.p.s.ps evoked by low stimulus voltages were more depressed than those evoked by high stimulus voltages and fast e.p.s.ps resulting from activation of a single presynaptic fibre were blocked in a non-graded manner. 4 Opioids depressed the slow e.p.s.p. in those neurones in which they did not change the resting membrane potential. 5 The slow e.p.s.p. was increased in amplitude in those neurones hyperpolarized by opioids. Depolarizations resulting from application of barium, substance P or ACh were also enhanced by opioids. Equivalent circuit models in which opioids increase, and substance P or ACh decrease, the same potassium conductance could account for this enhancement. 6 The actions of opioids were prevented or reversed by naloxone (1 nM-l JM). 7 It is concluded that morphine and enkephalin inhibit the release of ACh and a non-cholinergic transmitter from fibres of the myenteric plexus, and that this may involve a hyperpolarization of presynaptic fibres. Additionally, opioids can interact postsynaptically with other substances which affect membrane potassium conductances.
Opioid receptors in the spinal cord produce strong analgesia, but the mechanisms controlling their activation by endogenous opioids remain unclear. We have previously shown in spinal cord slices that peptidases preclude μ-opioid receptor (MOR) internalization by opioids. Our present goals were to investigate whether enkephalin-induced analgesia is also precluded by peptidases, and whether it is mediated by MORs or δ-opioid receptors (DORs). Tail-flick analgesia and MOR internalization were measured in rats injected intrathecally with Leu-enkephalin and peptidase inhibitors. Without peptidase inhibitors, Leu-enkephalin produced neither analgesia nor MOR internalization at doses up to 100 nmol, whereas with peptidase inhibitors it produced analgesia at 0.3 nmol and MOR internalization at 1 nmol. Leu-enkephalin was ten times more potent to produce analgesia than to produce MOR internalization, suggesting that DORs were involved. Selective MOR or DOR antagonists completely blocked the analgesia elicited by 0.3 nmol Leu-enkephalin (a dose that produced little MOR internalization), indicating that it involved these two receptors, possibly by an additive or synergistic interaction. The selective MOR agonist endomorphin-2 produced analgesia even in the presence of a DOR antagonist, but at doses substantially higher than Leu-enkephalin. Unlike Leu-enkephalin, endomorphin-2 had the same potencies to induce analgesia and MOR internalization. We concluded that low doses of enkephalins produce analgesia by activating both MORs and DORs. Analgesia can also be produced exclusively by MORs at higher agonist doses. Since peptidases prevent the activation of spinal opioid receptors by enkephalins, the coincident release of opioids and endogenous peptidase inhibitors may be required for analgesia.