Naloxone produces hypothermia in rats pretreated with beta-endorphin and morphine

The effects of naloxone (an antagonist of opioids at opiate receptors) on the thermoregulatory responses were assessed in three groups of naive (saline-treated), morphine-tolerant (24 hr after a dose of 100 mg/kg (s.c.) of morphine-oil suspension or three doses of 100 mg/kg (i.p.) of morphine and beta-endorphin-tolerant (24 hr after an intraventricular dose of 100 μg of beta-endorphin) rats at an ambient temperature of 22°C. Both morphine and beta-endorphin produced hypothermia, catatonia and sedation in naive rats. The hypothermia was due to decreased metabolic rate and cutaneous vasodilation. However, both in morphine-tolerant and beta-endorphin-tolerant rats, morphine and betaendorphin each produced hyperthermia rather than hypothermia. There were no changes in behavior. The hyperthermia was due to increased metabolism. Furthermore, naloxone administration produced a dose-dependent hypothermia and abstinence syndrome in both morphine-tolerant and beta-endorphin-tolerant rats, but not in naive rats. The hypothermia in response to naloxone in opioid-tolerant rats was brought about by both decreased metabolism and cutaneous vasodilatation. The data indicate that pretreatment of rats with opioids alters the thermoregulatory effects of naloxone.     

An opioid pancreatic peptide produces ileal muscle inhibition and naloxone-reversible analgesia.

The opioid activity of immunoreactive beta-endorphin-like peptide extracted from pork pancreas duplicates the effects of morphine and synthetic beta-endorphin when measured by inhibition of isolated guinea pig ileal muscle response to electro-stimulation in vitro and by morphine-like analgesia following intravenous injection in the mouse. These responses are reversed by the opiate antagonist naloxone, indicating that a potent opioid mu receptor binding ligand is present in pancreatic extract. These findings imply a pancreatic source of plasma immunoreactive beta-endorphin that may explain a number of physiological and behavioral effects generally attributed to hypophyseal beta-endorphin alone.     

Opiate antagonism reduces placentophagia and pup cleaning by parturient rats

Since endogenous opiate mechanisms are activated during parturition, the present study examined in rats the effects of opiate antagonism on maternal care during and shortly after parturition. Endogenous opiate mechanisms were blocked in late pregnant rats by (1) naltrexone pellet implants (Experiment 1); (2) acute naloxone injections of 10 mg/kg (Experiment 2) or 0.1 mg/kg (Experiment 7); or (3) induction of opiate tolerance (Experiment 3). All methods resulted in a significant decrease in placentophagia and/or in cleaning pups of umbilical cords and birth fluids (Experiment 6). Other aspects of maternal care appeared relatively unaffected and 24 hr pup survival rats were lowered only by induction of morphine tolerance (probably via its effects on the young). In nonpregnant females, naloxone produced a small but significant decrease in placentophagia (Experiment 4) whereas morphine-tolerant nonpregnant females consumed placentas as readily as controls (Experiment 5). Thus the inhibition of placentophagia produced by opiate antagonism may be specific to conditions associated with parturition. These findings suggest that endogenous opiates support placenta eating and pup cleaning during and immediately after birth. Mediation may be via opiate effects on ingestive behavior, and/or via a reduction in the stress of parturition which otherwise can interfere with the female's ability to perform these tasks.     

Arterial catecholamine levels in morphine-treated rats subjected to sympathetic nerve stimulation.

1. The effect of acute or chronic morphine treatment on the changes in arterial noradrenaline and adrenaline levels in response to sympathetic nerve stimulation was studied in rats. 2. Rats which had been chronically treated with morphine in their drinking fluid for 21 days were shown to be morphine-tolerant, as revealed by the tail-immersion test for analgesia. 3. It was found that animals given either acute or chronic morphine treatment had similar basal concentrations of arterial catecholamines to their controls. 4. Sympathetic nerve stimulation produced significant increases in arterial noradrenaline and adrenaline levels in both the control and morphine-treated animals. However, the degree of arterial noradrenaline elevation was significantly less in morphine-tolerant animals. 5. This phenomenon was not observed in acutely morphine-treated rats or at 2 weeks following opiate withdrawal in animals which had been treated previously with morphine for 3 weeks. 6. The findings suggest that chronic morphine treatment in rats not only leads to opiate tolerance but also reduces catecholamine release in response to sympathetic nerve stimulation.     

Dependence liability of enkephalin in the myenteric plexus of the guinea pig.

The endogenous ligands of opiate receptors, methionine-eukephalin and leucine-enkephalin, were tested for their ability to maintain or to reinduce dependence in myenteric plexus-longitudinal muscle strips taken from tolerant/dependent guinea pigs. Both peptides fully substituted for the function morphine exerts in this preparation. It is concluded that the enkephalins posses a high degree of dependence liability.     

Substance P and opioid interaction on stimulated and non-stimulated guinea pig ileum

In the past, substance P (SP) has been suggested to be both an opiate agonist and an antagonist. It therefore seemed appropriate to examine potential interactions of SP and opioids on guinea pig ileum. On non-stimulated ileal strips SP caused a dose responsive increase in contraction. Pretreatment of the tissue with morphine (3, 30, 300, 3000 nM), enkephalin (1.42, 14.2, 142, 1420 nM), naloxone (5 nM), or atropine (0.144 μM) did not significantly alter the spasmogenic effect of SP. On stimulated guinea pig ileum, whereas morphine and enkephalin inhibited the electrically induced twitch, SP administration resulted in contraction of the tissue. Additionally, neither strongly effective nor sub-threshold doses of SP antagonized the effects of the narcotics. These data are discussed in terms of separate receptors mediating the effects of the opiates and SP on guinea pig ileum.     

Modification of rat thermal responses to morphine by alpha-FMH suggests a role for neural histamine in morphine tolerance

In rats, morphine may either raise or lower body temperature depending on the dose. A morphine dose of 50 mg/kg, i.p., consistently produced a nearly maximal hypothermic response in non tolerant rats, whereas this dosage induced an elevation of body temperature in tolerant rats. In rats pretreated with alpha-fluoromethylhistidine (alpha-FMH), an irreversible inhibitor of histidine decarboxylase which induces a reduction in brain histamine synthesis, this morphine dose of 50 mg/kg, i.p. produced an elevation of rectal temperature resembling that observed in morphine-tolerant rats. To confirm the suggestion that hyperthermic effects of the higher dose of morphine in morphine-tolerant rats or in alpha-FMH-pretreated rats could be related to a possible involvement of mediators of fever, e.g. prostaglandins, animals were pretreated with acetylsalicylic acid (aspirin, Bayer) 30 mg/kg, i.p., 60 min before morphine. Results showed that acetylsalicylic acid prevented the hyperthermic response of morphine, resulting in a fall in body temperature. Since morphine releases histamine and alpha-FMH inhibits histamine synthesis, our data demonstrating that an inhibitor of prostaglandin-synthetase showed efficacy only in animals responding with fever to the higher dose of the opiate, suggests a physiological antagonism between histamine and prostaglandins on mechanisms underlying hyper/hypothermic responses to morphine.     

Abatement of morphine-induced slowing in gastrointestinal transit by Dai-kenchu-to, a traditional Japanese herbal medicine

As a way of alleviating severe constipation in cancer patients taking morphine to relieve pain, effects of Dai-kenchu-to (DKT), a traditional Japanese herbal medicine (Kampo medicine), on gastrointestinal transit in mice or on the isolated guinea pig ileum were studied in special reference to morphine. Without altering the anti-nociceptive effect of morphine, DKT was significantly effective against morphine-induced disorder of gastrointestinal transit in mice as assessed by the charcoal meal test for the intestine and measurement of transit time for the colon tract. The results of in vitro studies with guinea pig ileum suggest that abatement of morphine-induced disorder of transit by DKT is caused by both moderate contraction of morphine-treated longitudinal muscle and relaxation of morphine-induced tonic contraction of circular muscle.     

Effects of capsaicin on vascular smooth muscle

Acute administration of capsaicin in vitro produced either vascular smooth muscle contraction (cat middle cerebral artery) or smooth muscle relaxation (guinea pig carotid artery and thoracic aorta). Prior in vivo treatment with capsaicin abolished the relaxation response of guinea pig vessels to acute capsaicin. Instead a contractile response was seen after chronic capsaicin treatment, suggesting that the relaxation response produced by capsaicin is due to release of a vasodilator substance. Substance P caused relaxation in both cat cerebral arteries and the guinea pig thoracic aorta, an effect which was abolished or reduced by endothelial damage. However, responses to acute capsaicin were not altered by endothelial damage, suggesting that substance P does not mediate the relaxation response to acute capsaicin administration. Exposure to capsaicin in vitro did not affect the neurogenic vasodilator response of cat cerebral arteries and did not alter substance P levels. Therefore, it was concluded that the acute effect of capsaicin is composed of two components. A contractile response is most likely due to direct effects on vascular smooth muscle, while a relaxation response is attributed to release of an as yet unidentified bioactive substance distinct from substance P.     

眼镜蛇神经毒素对大鼠脑内脑啡肽含量及豚鼠回肠和小鼠输精管收缩的影响

连续注射1wk眼镜蛇神经毒素后,用放射免疫测定法测定对照组及神经毒素组大鼠5个脑区亮啡肽样免疫反应物质及甲硫啡肽样免疫反应物质的含量。神经毒素组两种脑啡肽样物质在下丘脑、中脑+丘脑两个脑区的含量变化均出现明显升高(p<0.05),海马及后脑的含量均无显著变化。吗啡在很低浓度对低频电刺激豚鼠回肠及小鼠输精管收缩有明显抑制作用。神经毒素的存在与否并不影响吗啡的效应。结果提示,神经毒素可能并不直接与吗啡受体产生相互作用,但其镇痛机理可能涉及中枢内源性阿片肽能系统。     

Anti-morphine anti-idiotypic antibodies. Opiate receptor binding and isolated tissue responses

Anti-idiotypic antibodies which recognize the opiate receptor were generated in guinea pigs following immunization against purified rabbit anti-morphine antibodies. The anti-idiotypic antibodies produced a concentration-dependent inhibition of [3H]naloxone binding to opiate receptors in a membranous mouse brain preparation. Saturation analysis indicated that the antibodies produced a non-competitive inhibition of naloxone binding. The ability of the antibodies to interact with biological systems was investigated in in vitro systems. In both the isolated guinea pig ileal longitudinal muscle and mouse vas deferens, the antibodies produced a concentration-dependent, opiate agonist-like action. The anti-morphine anti-idiotypic antibodies appear to interact specifically with the opiate receptor and may serve as useful tools in characterization of this receptor system.     

The opiate-like action of tilidine is mediated by metabolites

Summary The analgesic effect of tilidine in rats is completely antagonized by the narcotic antagonist naloxone. Radioreceptor assays revealed, however, that the main metabolites of tilidine, nortilidine and bisnortilidine, rather than tilidine exhibit affinity to opiate receptors. These findings were confirmed in studies using the electrically stimulated guinea pig ileum and the mouse vas deferens. Chronic tilidine administration to rats caused a considerable degree of physical dependence, which was expected from the ability of the intact animal to metabolize tilidine. In the isolated ileum from chronically morphinized guinea pigs, both nortilidine and bisnortilidine fully substituted for morphine in preventing induction of withdrawal, indicating dependence liability of these metabolites.     

Effect of kappa-opioid receptor agonists on morphine analgesia in morphine-naive and morphine-tolerant rats

The effect of i.p. administration of kappa-opioid receptor agonists, bremazocine, tifluadom and U-50,488H on morphine (8 mg/kg i.p.)-induced analgesia in morphine-naive and morphine tolerant male Sprague-Dawley rats was determined using the tail-flick test. The tolerance to morphine in the rats was induced by s.c., implantation of six morphine pellets during a 7-day period. Implantation of morphine pellets resulted in the development of tolerance as evidenced by the decrease in the analgesic response to morphine when compared to placebo pellets implanted rats. Bremazocine (0.3, 1.0 and 3.0 mg/kg) and U-50,488H (16 mg/kg) antagonized morphine-induced analgesia in morphine-naive rats while tifluadom (8 and 16 mg/kg) potentiated the effect. In morphine-tolerant rats, bremazocine (3 mg/kg) and U-50,488H (16 mg/kg) potentiated morphine-induced analgesia. Tifluadom at any of the doses had no effect on morphine-induced analgesia in morphine-tolerant rats. These results provide evidence that different kappa-opioid agonists modify morphine-induced analgesia differentially in morphine-naive and morphine-tolerant rats.     

Metabolism of dynorphin A(1-13)

Dynorphin A(1-13), a tridecapeptide of the endogenous opioid peptides, has modest effects in reducing mild opiate withdrawal in humans. Previous studies revealed that dynorphin also potentiates the analgesic effect of morphine in morphine-tolerant rats and mice. The therapeutic potential of dynorphin A(1-13) is limited due to extensive metabolism by human metabolic enzymes resulting in an in vivo half-life of less than one minute. Chemical modifications of dynorphin A(1-13), such as N-methylation of Tyr1 and amidation of the C-terminus have been shown to be effective in protecting against the proteolytic enzymes in human plasma. This article is a general review of the metabolism of dynorphin A(1-13) in human plasma and CSF.     

Changes in morphine-induced activation of cerebral Na(+),K(+)-ATPase during morphine tolerance: biochemical and behavioral consequences

There is ample evidence of the biological changes produced by the sustained activation of opioid receptors. We evaluated the adaptive changes of cerebral Na(+),K(+)-ATPase in response to the sustained administration of morphine (minipumps, 45mg/kg/day, 6 days) in CD-1 mice and the functional role of these changes in opioid antinociception. The antinociceptive effect of morphine as determined with tail-flick tests was reduced in morphine-tolerant mice. There were no significant changes in the density of high-affinity Na(+),K(+)-ATPase α subunits labeled with [(3)H]ouabain in forebrain membranes from morphine-tolerant compared to those of morphine-naive animals. Western blot analysis showed that there were no significant differences between groups in the changes in relative abundance of α(1) and α(3) subunits of Na(+),K(+)-ATPase in the spinal cord or forebrain. However, the morphine-induced stimulation of Na(+),K(+)-ATPase activity was significantly lower in brain synaptosomes from morphine-tolerant mice (EC(50)=1.79±0.10μM) than in synaptosomes from morphine-naive mice (EC(50)=0.69±0.12μM). Furthermore, adaptive alterations in the time-course of basal Na(+),K(+)-ATPase activity were observed after sustained morphine treatment, with a change from a bi-exponential decay model (morphine-naive mice) to a mono-exponential model (morphine-tolerant mice). In behavioral studies the antinociceptive effects of morphine (s.c.) in the tail-flick test were dose-dependently antagonized by ouabain (1 and 10ng/mouse, i.c.v.) in morphine-naive mice, but not in morphine-tolerant mice. These findings suggest that during morphine tolerance, adaptive cellular changes take place in cerebral Na(+),K(+)-ATPase activity which are of functional relevance for morphine-induced antinociception.     

Effects of morphine and naloxone on separation distress and approach attachment: Evidence for opiate mediation of social affect

In order to determine the relationship between endorphins and social attachment, the effects of morphine (an opiate agonist) and naloxone (an opiate antagonist) on various indices of attachment in guinea pigs were studied. In infants, crying or separation-induced distress vocalizations were significantly decreased by single injections of low morphine doses (0.25, .050 and 0.75 mg/kg) in a dose-dependent manner. Naloxone (1 mg/kg) reliably increased separation distress vocalizations in both juvenile and adult guinea pigs. Therefore, similar to opiate withdrawal symptoms, separation distress appeared to be alleviated by morphine and potentiated by naloxone. As for approach attachment, offspring/maternal proximity-maintenance time was significantly decreased by morphine (1.0, 2.5 and 5.0 mg/kg), suggesting that opiates may be capable of replacing a function normally subserved by endorphins in reinforcing attachments. These data support the hypothesis that an endorphin-based addiction-like process may underlie the maintenance of social attachments, and that separation distress may reflect a state of endogenous “endorphin withdrawal”.     

Mechanism of morphine block of electrical activity in ganglia of Auerbach''s plexus

A study was made of the effect of morphine on electrical activity within single ganglia of Auerbach's plexus of guinea pig longitudinal muscle-myenteric plexus as monitored by means of external electrodes. Morphine produces a concentration dependent block of single spike activity. This effect is competitively antagonized by naloxone. The ED₅₀ for morphine effect is about 7 × 10⁻⁷ M. Naloxone and dextrorphan have no effect on electrical activity. Acetylcholine in the concentration range of 10⁻⁷−10⁻⁵ M augments electrical activity of ganglia. Morphine has little if any effect on the enhanced stimulation produced by acetylcholine thus indicating that the drug does not act directly upon the ganglion. Our results suggest that a specific opiate receptor is present on preganglionic nerve terminals and that morphine and other opiates block ganglionic transmission by inhibiting the release of preganglionic acetylcholine.     

Effects of methylnaltrexone on guinea pig gastrointestinal motility

The purpose of the present study was to compare the effects of methylnaltrexone (MNTX), a peripherally acting μ opioid receptor (μOR) antagonist, on gastrointestinal (GI) motility in naïve vs. opiate chronically treated guinea pigs in vitro and in vivo. We have used the electrically stimulated muscle twitch contractions of longitudinal muscle-myenteric plexus (LMMP) preparations and total GI transit as measure of GI motility. In LMMP preparations of naïve guinea pigs, MNTX (1–30 μM) induced a significant, dose–response reduction of morphine-induced inhibition of electrically stimulated muscle twitch contractions, with an IC₅₀ of 9.4 10^?8M. By contrast, MNTX abolished the inhibitory effect of acute morphine at any concentrations tested (1–30 μM) in the guinea pigs chronically treated with opiates. In vivo, MNTX (10–50 mg s.c.) did not affect GI transit in naïve guinea pigs when administered acutely or for five consecutive days, but reversed the GI transit delay induced by chronic morphine treatment. These findings show that MNTX is effective in reversing opiate-induced inhibition of GI motility acting at peripheral μ opioid receptors, but does not exert a pharmacologic effect on GI transit in the absence of opiate stimulation.     

A novel combination of opiates and endothelin antagonists to manage pain without any tolerance development

Several neurotransmitter mechanisms have been proposed as playing a role in the development of morphine tolerance. We provide evidence for the first time that endothelin antagonists can restore morphine analgesia in morphine-tolerant rats and prevent the development of tolerance to morphine. Studies were carried out in rats and mice treated with implanted placebo or implanted morphine pellet. The maximal tail-flick latency in morphine pellet + vehicle-treated rats (7.54 seconds) was significantly lower when compared with placebo pellet + vehicle-treated rats (10 seconds), indicating that tolerance developed to the analgesic effect of morphine. BQ123 potentiated tail-flick latency by 30.0% in placebo-tolerant rats and 94.5% in morphine-tolerant rats compared with respective controls. BMS182874 potentiated tail-flick latency by 30.2% in placebo-tolerant rats and 66.7% in morphine-tolerant rats. The enhanced analgesic effect of morphine after treatment with endothelin antagonists could be blocked by naloxone, indicating an opiate-mediated effect; but naloxone binding to brain membranes was not affected by BQ123. Guanosine triphosphate binding was stimulated by morphine and endothelin-1 in non-tolerant mice and not in morphine-tolerant mice; however, guanosine triphosphate binding was stimulated by BQ123 in morphine-tolerant mice and was unaffected in non-tolerant mice. These results suggest that uncoupling of G-protein occurs in morphine tolerance and endothelin antagonist restores the coupling of G-protein to its receptors. A combination use of endothelin antagonist and opiates could provide a novel approach in improving analgesia and eliminating tolerance.     

Opiate regulation of maternal behavior in the rat

The effects of the opiate agonist morphine, the opiate antagonist naloxone and the weak opiate nonanalgesic dextrophan on the expression of maternal behavior were investigated in a series of three experiments. In the first experiment treatment of rats with morphine (5 mg/kg, subcutaneously) after ovariectomy and hysterectomy on day 17 of gestation resulted in a disruption in the onset and quality of maternal responsiveness in the homecage and in a T-maze test. The duration of morphine's acute disruptive action was 2–4 hours. In the second experiment concurrent treatment of morphine-injected rats with naloxone prevented the disruptive effects of morphine in both the homecage and T-maze tests. The effects of morphine did not appear to result from a severe alteration in activity levels as measured in an open-field test, although morphine did increase activity slightly by the fifth day of treatment. In the third experiment treatment of rats after ovariectomy plus hysterectomy on day 17 of gestation with dextrophan failed to disrupt maternal behavior. These results indicate that morphine disrupts maternal behavior through an opiate receptor mechanism, and suggests to us that endogenous opiates may mediate the expression of maternal behavior under certain physiological conditions.