Tolerance to morphine-induced analgesia in mice: Magnetic fields function as environmental specific cues and reduce tolerance development

Mice receiving daily injection of morphine (10 mg/kg) developed tolerance to morphine-induced analgesia, such that after 5–7 days of treatment their thermal response (paw licking) latencies in the hot plate test were indistinguishable from those of control animals. Exposure to a rotating magnetic field for thirty minutes before the daily morphine administrations significantly reduced the development of tolerance. These magnetic exposure also significantly increased over 7–10 days the basal nociceptive thresholds and paw licking response latencies of saline treated mice. Control and sham exposed mice that were fully tolerant to the analgesic effects of morphine failed to show any tolerance to morphine-induced analgesia when exposed to the magnetic stimuli prior to injection. Likewise, the partial tolerance to morphine shown by mice exposed to the rotating magnetic field pre-injection environmental cues was eliminated when control or sham pre-injection cues lacking the magnetic stimuli were provided. In all cases tolerance to morphine-induced analgesia was evident in the subsequent re-test with the original cues. These results indicate that magnetic field exposure can reduce the development of tolerance to the analgesic effects of morphine. They also show that magnetic stimuli function as significant environmental cues for the development of tolerance to morphine-induced analgesia. This suggests that magnetic stimuli affect both the associative (classical conditioning) and non-associative (physiological, pharmacological) mechanisms involved in the development of opiate tolerance.     

Exposure to time varying magnetic fields associated with magnetic resonance imaging reduces fentanyl-induced analgesia in mice

The effects of exposure to clinical magnetic resonance imaging (MRI) on analgesia induced by the mu opiate agonist, fentanyl, was examined in mice. During the dark period, adult male mice were exposed for 23.2 min to the time-varying (0.6 T/sec) magnetic field (TVMF) component of the MRI procedure. Following this exposure, the analgesic potency of fentanyl citrate (0.1 mg/kg) was determined at 5, 10, 15, and 30 min post-injection, using a thermal test stimulus (hot-plate 50 degrees C). Exposure to the magnetic-field gradients attenuated the fentanyl-induced analgesia in a manner comparable to that previously observed with morphine. These results indicate that the time-varying magnetic fields associated with MRI have significant inhibitory effects on the analgesic effects of specific mu-opiate-directed ligands.     

Dissociation of tolerance to the analgesic and hypothermic effects of morphine by using thyrotropin releasing hormone

The effects of thyrotropin releasing hormone (TRH) on tolerance to the analgesic and hypothermic effects of morphine were determined in male Swiss Webster mice. The tolerance to morphine was induced by SC implantation of a morphine pellet containing 75 mg morphine free base for 3 days. Subcutaneous injections of TRH (4 mg/kg) twice a day inhibited tolerance to the analgesic effect of morphine, as evidenced by a greater degree of analgesia in TRH treated mice as compared with similarly treated vehicle injected controls. The same treatment, however, failed to modify tolerance to the hypothermic effect of morphine. These effects were produced with alterations in brain or plasma levels of morphine. It is concluded that tolerance to the two pharmacological effects of morphine may involve separate mechanism.     

Environmental and intraperitoneal ethanol influences morphine antinociceptive effect in mice

The ability of acute environmental or intraperitoneal (i.p.) ethanol to influence morphine antinociceptive effect was studied in mice. In order to induce tolerance to morphine analgesia, mice received daily injections of 10 mg/Kg morphine over a period of 10 days. Mice were divided into three groups: i.p. ethanol (E), environmental ethanol (E*), and control saline (M). During the induction of tolerance these groups were treated identically except on days 1 and 11. On these days, 10 minutes prior to morphine injection, mice received either i.p. ethanol (1g/Kg), environmental ethanol (a bottle of 10% ethanol placed next to the animals cage during the experiments), or an equivalent volume of saline. Analgesia was assessed using a standard hot plate protocol and dose-response cumulative curves for morphine analgesia were obtained on days 1 and 11. On day 1, both the i.p. and environmental administration of ethanol showed similar morphine-potentiation effects [Mean Effective Dose: ED50 (M1)=4.5 mg/kg; ED50 (E1)=2.4 mg/kg; ED50 (E*1)=2.1 mg/kg]. On day 11, control group mice showed a reduction of morphine analgesia at test [ED50 (M11)=14.1 mg/kg]. Mice receiving i.p. and environmental ethanol again showed a leftward shift in dose-response cumulative curves for morphine antinociception with respect to controls [ED50 (E11)=9.1 mg/kg; ED50 (E*11)=4.7 mg/kg]. I.p. ethanol administration at non-antinociceptive doses enhances the morphine antinociception effect similarly in tolerant and non-tolerant (naive) mice. The presence of environmental ethanol can also induce a similar pattern of increase in morphine antinociception effect.     

Comparison of the effects of specific and nonspecific inhibition of nitric oxide synthase on morphine analgesia,tolerance and dependence in mice

The effects of L-Canavanine, a selective inducible nitric oxide synthase (NOS) inhibitor and N(G)-nitro-L-arginine methyl ester (L-NAME), a nonselective NOS inhibitor, on pain threshold and morphine induced analgesia, tolerance and dependence in mice were investigated and compared. Morphine was administered by subcutaneous implantation of a pellet containing 40 mg free base and withdrawal was precipitated by intraperitoneal (i.p.) injection of naloxone (2 mg/kg). L-Canavanine (200 mg/kg, i.p.) did not affect the pain threshold, morphine-induced analgesia and the induction and expression phases of morphine tolerance and dependence. L-NAME (20 mg/kg, i.p.) significantly (p < 0.05) enhanced the pain threshold, potentiated morphine-induced analgesia and attenuated the expression phase of morphine dependence which has been characterized by withdrawal signs and body weight loss, but did not modify the induction phase of morphine tolerance and dependence. It is concluded that constitutive NOS isoforms which were inhibited by L-NAME may be involved specifically in the mechanisms of morphine induced analgesia, tolerance and dependence.     

Cancer promotion in a mouse-skin model by a 60-Hz magnetic field: II. Tumor development and immune response.

This paper describes preliminary findings on the influence of 60-Hz (2-mT) magnetic fields on tumor promotion and co-promotion in the skins of mice. The effect of magnetic fields on natural killer (NK) cell activity in spleen and blood was also examined. Groups of 32 juvenile female mice were exposed to the magnetic field as described in part I. The dorsal skin of all animals was treated with a subthreshold dose of the carcinogen 7,12-dimethyl-benz(a)anthracene (DMBA). One week after the treatment, two groups were sham exposed (group A) or field exposed at 2 mT (group B) 6 h/day for 21 weeks, to test whether the field would act as a tumor promoter. No tumors developed in these two groups of mice. To test whether the magnetic field would modify tumor development by directly affecting tumor growth or by suppressing immune surveillance, two additional groups of mice were treated weekly with the tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) and then either sham exposed (group C) or field exposed (group D). The time to appearance of tumors was shorter (but not statistically so) in the group exposed to magnetic fields and TPA. Some differences in NK cell activity and spleen size were observed between the sham- and field-exposed groups.     

Opiate Modulation of Striatal Dopamine and Hippocampal Norepinephrine Release Following Morphine Withdrawal

When opiates are abruptly withdrawn after chronic treatment, increases in hippocampal noradre-nergic function are observed which are accompanied by decreases in striatal dopamine release. The latter effects have to shown to persist for several weeks following the onset of opiate withdrawal. We examined the long-term effects of opiate withdrawal on 4-aminopyridine and potassium stimulated release of striatal dopamine and hippocampal norepinephrine. Tissue samples were obtained either from rats that had been exposed to opiate withdrawal following a seven day morphine infusion or sham treated control subjects. At 48 hours after the onset of withdrawal (cessation of morphine infusions), slices were loaded with [³H] neurotransmitter, washed extensively, and exposed to different drug treatments. 4-aminopyridine induced concentration related increases in striatal dopamine release, which was 36% calcium independent. Similar values for fractional release of striatal dopamine were obtained in morphine withdrawn and control subjects, for both potassium and 4-aminopyridine induced release. In addition, thresholds for 4-aminopyridine or potassium induced release of striatal dopamine did not differ between control and morphine withdrawn subjects. Treatment with 1.0 M morphine sulfate potentiated potassium evoked release of norepinephrine to an equal extent in both morphine withdrawn and sham treated hippocampal tissue. Exposure to a threshold concentration of potassium (8.0 mM), stimulated increased release of hippocampal norepinephrine in a significantly greater fraction of tissue samples obtained from morphine withdrawn animals. Although these results do not support changes in striatal dopamine release following opiate withdrawal, opiate mechanisms appear to be important determinants of in vitro hippocampal norepinephrine release.     

NMDA receptor antagonism disrupts the development of morphine analgesic tolerance in male, but not female C57BL/6J mice

Multiple studies demonstrate that coadministration of N-methyl-D-aspartate (NMDA) receptor antagonists with the opioid agonist morphine attenuates the development of analgesic tolerance. Sex differences in the effects of noncompetitive, but not competitive NMDA receptor antagonists on acute morphine analgesia, have been reported in mice, yet the role of sex in modulation of morphine tolerance by NMDA receptor antagonists has yet to be addressed. Therefore, we tested whether there is a sex difference in the effect of NMDA receptor antagonists on the development of morphine analgesic tolerance in C57BL/6J mice. Acutely, at a dose required to affect morphine tolerance in male mice, the noncompetitive NMDA receptor antagonist dizocilpine (MK-801) prolonged morphine analgesia similarly in both sexes in the hot plate and tail withdrawal assays. In the hot plate assay, coadministration of MK-801 or the competitive antagonist 3-(2-carboxpiperazin-4-yl)propyl-1-phosphanoic acid (CPP) with morphine attenuated the development of tolerance in male mice, while having no effect in females. Like normal and sham females, ovariectomized mice were similarly insensitive to the attenuation of morphine tolerance by MK-801 in the hot plate assay. Surprisingly, in the tail withdrawal assay, MK-801 facilitated the development of morphine-induced hyperalgesia and tolerance in males but not females. The results demonstrate that male mice are more sensitive to modulation of nociception and morphine analgesia after repeated coadministration of NMDA receptor antagonists. Furthermore, the underlying mechanisms are likely to be different from those mediating the sex difference in the modulation of acute morphine analgesia that has previously been reported.     

Exposure to a hypogeomagnetic field or to oscillating magnetic fields similarly reduce stress-induced analgesia in C57 male mice

Previous studies have shown that exposure to altered magnetic fields alters analgesic responses in a variety of species, including humans. Here we examined whether deprivation of the normally occurring geomagnetic field also affects stress-induced analgesia, by measuring the nociceptive responses of C57 male mice that were restraint-stressed in a hypogeomagnetic environment (inside a mu-metal box). Stress-induced analgesia was significantly suppressed in a manner comparable to that observed in mice that were either exposed to altered oscillating magnetic fields or treated with the prototypic opiate antagonist naloxone. These results represent the first piece of evidence that a period in a hypogeomagnetic environment inhibits stress-induced analgesia.     

Footshock induced analgesia in mice: its reversal by naloxone and cross tolerance with morphine

Using the abdominal constriction response as the criterion for analgesia, mice tested immediately after a period of footshock showed a significant analgesic response compared with non-footshocked controls. Footshock induced analgesia could not be elicited in mice that had been made tolerant to morphine or in mice that had been pretreated with the narcotic antagonist naloxone. It is concluded that footshock induced analgesia in the mouse is due to the release of endogenous opioid peptides.     

ACUTE ETHANOL EXPOSURE DECREASES THE ANALGESIC POTENCY OF MORPHINE IN MICE

Chronic (7 days), forced ethanol drinking can decrease the analgesic potency of opioid agonists in mice. In the present study, the effect of short-term ethanol treatment was examined using forced ethanol access and ethanol injection protocols. Mice were given forced access to 1, 3 or 7% (v/v) ethanol for 24 hr and then tested for s.c. morphine analgesia using the tailflick assay. Controls had access to water. Another group of mice was injected i.p. with 2.5 g/kg ethanol or water 4 times over a 21 hr period and tested 3 hr after the final injection for morphine analgesia. Other mice were injected once i.p. with 1, 2 or 3 g/kg ethanol or water and tested 24 hr later using the tailflick. In the forced access study, ethanol dose-dependently decreased morphine's analgesic potency with the highest dose (7%) producing a 1.6-fold shift in the ED₅₀. This decrease in morphine potency was similar to that found in a related study using 7% ethanol for 7 days (1.8-fold shift). Repeated ethanol injections significantly reduced the analgesic potency of morphine (1.9-fold shift), whereas, a single injection of 1, 2 or 3 g/kg ethanol did not alter the potency of morphine. Control studies indicated that neither 24 hr water nor food deprivation affected morphine potency. Overall, these data show that sustained exposure to ethanol over a 24 hr period will dose-dependently decrease morphine's analgesic potency. © 1998 Elsevier Science Inc.     

Assessment of temporal changes in pulmonary edema with NMR imaging

Nuclear magnetic resonance imaging (NMRI) parameters [longitudinal relaxation time (T1), transverse relaxation time (T2), and signal intensity] acquired at a magnetic field of 2.35 T were validated with a study of nine different phantom gel solutions. This technique was then applied to study 13 anesthetized supine cats, among which 10 had lung edema induced by oleic acid (0.075 ml/kg); the result was compared with postmortem analyses of lung water. Three animals (series A) were imaged until the edema was first visualized in NMRI, usually 15-20 min after oleic acid infusion. Another seven animals (series B) were imaged over 4-5 h. As lung water increased, so did the signal intensity. When edema first appeared, T1, T2, and the volume of the edematous region within the slice in the upper lobes showed no gravity-dependent differences; this was confirmed by postmortem measurements (series A) of lung water. With time, gravity-dependent regions displayed greater volumes of edematous regions and greater T1 values (P less than 0.01), suggesting a continued accumulation of lung water. In comparison, nondependent regions displayed constant volumes of edematous region and lesser T1 values (P less than 0.01), suggesting an increased protein concentration but no change in lung water. This study suggests the potential applicability of NMRI parameters in the assessment of pulmonary edema.     

Temporal Variation in Drug Interaction Between Lithium and Morphine‐Induced Analgesia

The administration‐time‐dependent aspects of the drug interaction between lithium and morphine‐induced analgesia were studied using the mouse hot‐plate test at six different times of day, each scheduled at 4 h intervals. Lithium treatment alone, in doses of 1 to 10 mmol/kg administered intraperitoneally (i.p.) did not significantly alter test latencies compared to the corresponding clock‐time in saline‐injected controls. Basal pain sensitivity and morphine‐induced antinociceptive activity displayed significant circadian rhythms as assessed by the hot‐plate response latencies, with higher values occurring during the nocturnal activity than during the daytime rest span. Acute administration of lithium, in a dose of 3 mmol/kg, 30 min prior to morphine dosing did not influence morphine‐induced analgesia compared to all the clock‐time test‐matched morphine groups, except the 9 HALO (Hours After Lights On) one. There was a prominent potentiation of the morphine‐induced antinociception at this biological time during combined drug treatment. The latter finding demonstrates that administration‐time‐dependent differences in drug‐drug interactions need to be considered in both experimental designs and clinical settings.     

EFFECTS OF MICROWAVES AND ELF MAGNETIC FIELD ON THE PHAGOCYTIC ACTIVITY OF VARIOUSLY TREATED RAT MACROPHAGES

The purpose of this study was to investigate the effects of 9450-MHz microwaves and extremely low frequency magnetic fields (ELFMF) on the phagocytic activity of rat macrophages in control rats and those treated with vitamins C and E. In the microwave group, 24 albino Wistar rats were exposed to microwaves (2.65 mW/cm², specific absorption rate [SAR]: 1.80 W/kg) for 1 h/day for 21 days. Thirty-two albino Wistar rats were divided into four groups (one control, three experimental) (n = 8). The rats in the first exposure group were only exposed to microwaves for 1 h per day for 21 days. In addition to exposure with microwaves as in the first experimental group, vitamins E and C (150 mg/kg/day) were injected intraperitoneally into the rats in the second and third exposure groups, respectively. In the magnetic field exposure group, 26 albino Wistar rats were divided into two groups: the sham (n = 12) and exposed groups (n = 14). The rats in the experimental group were exposed to ELFMF (50 Hz, 0.75 mT) for 3 h/day for 3 weeks. After completing the exposure period, the rats were sacrificed under ketalar anesthesia. The viability of isolated alveolar macrophages of rats in the microwave and ELF groups was determined and compared to sham groups. The results were analyzed with the Mann–Whitney U test. In the microwave group, the phagocytic activity in the experimental groups was found to be higher than the sham groups. However, with phagocytic activity in rats treated with both microwaves and vitamins, only the vitamin C group was significant (p < 0.05). In the magnetic field group, the phagocytic activity of rats exposed to ELFMF was lower than that of the sham group, but the results were not significant (p > 0.05). Rectal temperatures of microwaveexposed groups were found to be significantly higher compared to the control group (p < 0.05).     

Longitudinal study of Pex1-G844D NMRI mouse model: A robust pre-clinical model for mild Zellweger spectrum disorder

Zellweger spectrum disorders (ZSD) are inborn errors of metabolism caused by mutations in PEX genes that lead to peroxisomal biogenesis disorder (PBD). No validated treatment is able to modify the dismal progression of the disease. ZSD mouse models used to develop therapeutic approaches are limited by poor survival and breeding restrictions. To overcome these limitations, we backcrossed the hypomorphic Pex1 p.G844D allele to NMRI background. NMRI mouse breeding restored an autosomal recessive Mendelian inheritance pattern and delivered twice larger litters. Mice were longitudinally phenotyped up to 6 months of age to make this model suitable for therapeutic interventions. ZSD mice exhibited growth retardation and relative hepatomegaly associated to progressive hepatocyte hypertrophy. Biochemical studies associated with RNA sequencing deciphered ZSD liver glycogen metabolism alterations. Affected fibroblasts displayed classical immunofluorescence pattern and biochemical alterations associated with PBD. Plasma and liver showed very long-chain fatty acids, specific oxysterols and C₂₇ bile acids intermediates elevation in ZSD mice along with a specific urine organic acid profile. With ageing, C₂₆ fatty acid and phytanic acid levels tended to normalize in ZSD mice, as described in patients reaching adulthood. In conclusion, our mouse model recapitulates a mild ZSD phenotype and is suitable for liver-targeted therapies evaluation.     

Quantitative comparison of ohmefentanyl isomers induced conditioning place preference in mice

Differences of analgesia and withdrawal response among ohmefentanyl stereoisomers have been studied. In the present study, Quantitative comparison of reinforcing effects of ohmefentanyl stereoisomers and morphine was performed by using a conditioned place preference design in mice. Results showed that morphine and ohmefentanyl stereoisomers were able to increase significantly the time spent in the drug-paired side with respect to vehicle treated animals. A good linear correlation between doses of drugs and number of mice with place preference was found within a given dose range. On the basis of the dose-response curve analysis, ohmefentanyl stereoisomers displayed a significant difference in place preference ED50. The addictive index (analgesic ED50/place preference ED50) was used to assess the addictive potential of drugs. It was demonstrated that the addictive potential of ohmefentanyl stereoisomers did not exhibit a large difference as addictive index. Among these stereoisomers, the addictive potential of compound F9208 was markedly lower than that of morphine.     

Reversal of the antinociceptive effects of centrally-administered morphine by the benzodiazepine receptor antagonist Ro 15-1788

The effects of the benzodiazepine receptor antagonist, Ro 15-1788, were examined on analgesia induced by morphine after central (intracerebroventricular, i.c.v., or intrathecal, i.t.) and systemic administration. Analgesia was assessed in squirrel monkeys trained to respond under an electric shock tiltration procedure and in mice using the radiant heat tail-flick test. Central and systemic administration of morphine produced antinociceptive effects that were antagonized by 0.1 mg/kg of naloxone in both species. Ro 15-1788 antagonized the effects of morphine after central (i.c.v. or i.t.) administration but did not alter the effects of morphine given by the systemic route. This novel interaction suggests that Ro 15-1788 may be useful in pharmacologically separating neural substrates subserving opiate analgesia.     

Dietary-induced hypertrophic--hyperplastic obesity in mice

Metabolically intact NMRI mice and genetically obese NZO mice were fed ad lib. either a high-carbohydrate diet (standard) or a high-fat diet for a period of about 11 (NMRI mice) or 38 (NZO mice) wk. In both strains of mice, body weight increased more in the groups fed the high-fat diet. However, caloric intake by NMRI mice fed the high-fat diet was less than that of the controls. In NMRI mice fed the high-fat diet, epididymal and subcutaneous fat cell volumes increased; when these mice were fed the standard diet, only epididymal fat cell volume increased. Epididymal and subcutaneous fat cell numbers increased only in the group fed the high-fat diet. In NMRI mice fed either diet, the postprandial blood glucose was lower in older animals, but plasma insulin remained unchanged. The glucose tolerance deteriorated insignificantly. In NZO mice fed either diet, epididymal fat cell volumes and fat cell numbers increased. In this strain of mice the postprandial blood glucose and plasma insulin exhibited the strain-specific pattern, independent of the diet. In older animals fed either diet the glucose tolerance decreased.     

Central administration of neuropeptide FF and related peptides attenuate systemic morphine analgesia in mice

Neuropeptide FF (NPFF) belongs to an opioid-modulating peptide family. NPFF has been reported to play important roles in the control of pain and analgesia through interactions with the opioid system. However, very few studies examined the effect of supraspinal NPFF system on analgesia induced by opiates administered at the peripheral level. In the present study, intracerebroventricular (i.c.v.) injection of NPFF (1, 3 and 10 nmol) dose-dependently inhibited systemic morphine (0.12 mg, i.p.) analgesia in the mouse tail flick test. Similarly, i.c.v. administration of dNPA and NPVF, two agonists highly selective for NPFF(2) and NPFF(1) receptors, respectively, decreased analgesia induced by i.p. morphine in mice. Furthermore, these anti-opioid activities of NPFF and related peptides were blocked by pretreatment with the NPFF receptors selective antagonist RF9 (10 nmol, i.c.v.). These results demonstrate that activation of central NPFF(1) and NPFF(2) receptors has the similar anti-opioid actions on the antinociceptive effect of systemic morphine.     

Enterostatin (VPDPR) has anti-analgesic and anti-amnesic activities

Enterostatin (VPDPR), an anorexigenic peptide derived from the amino terminus of procolipase, significantly inhibited analgesia induced by the mu-opioid agonist morphine (5 mg/kg, s.c.) after i.c.v. administration to mice at a dose of 100 nmol. On the other hand, VPDPR (approximately 200 nmol, i.c.v.) did not attenuate analgesia induced by the kappa-opioid agonist D-Phe-D-Phe-D-Nle-D-Arg-NH2 (100 microg/mouse, i.c.v.) or delta-opioid agonist DTLET (4 nmol/mouse, i.c.v.). VPDPR (100 nmol, i.c.v.) significantly improved amnesia induced by scopolamine (0.2 mg/kg, i.p.) in mice. However, VPDPR did not enhance memory in normal mice at the same dose.