Suppression of GABA-induced abstinence behaviour in naive rats by morphine and bicuculline.

Intraperitoneal administration of n-dipropylacetate (DPA) to naive rats produced abstinence behaviour including shaking, digging, hunchback posture, piloerection and ptosis during 15 min and increased motor activity considerably. Treatment with a subconvulsive dose of the GABA antagonist bicuculline suppressed this DPA-induced abstinence behaviour, indicating that GABA was increased at receptor sites. Also morphine in a low dose of 1 mg/kg suppressed this behaviour, while administration of naloxone after morphine treatment could release the abstinence behaviour. Simultaneous treatment with morphine and naloxone or naloxone alone were without effect. The administration to DPA treated rats of doses higher than 1 mg/kg morphine resulted in a severe depression of motor activity. It is concluded that an increased availability of GABA at its receptor sites plays an important role in the behaviour observed after DPA administration. The experiments with morphine and naloxone suggest that morphine receptors are involved in DPA-induced abstinence behaviour.     

Co-administration of dextromethorphan during pregnancy and throughout lactation significantly decreases the adverse effects associated with chronic morphine administration in rat offspring.

In this study, we have focused our investigation of the facts whether co-administration of a NMDA antagonist dextromethorphan (DM) with morphine during pregnancy and throughout lactation could prevent the adverse effects associated with chronic morphine administration in rat offspring. Adult female Sprague-Dawley rats were randomly separated into four groups and were received subcutaneous injection of either saline, morphine, morphine + dextromethorphan or dextromethorphan twice a day and progressively increased 1 mg/kg at 7-day intervals from a beginning dose of 2 mg/kg for both morphine and dextromethorphan. The rats were mated between days 7 and 8. Administration of drugs was continued during pregnancy. After rat offspring were born, the doses of morphine or dextromethorphan injected into the maternal rats were increased by 1 mg/kg every two weeks till the offspring were 30 day old. The results showed that mortality of morphine group is much higher than control group. The offspring of morphine group weighed significantly less than control group on postnatal day 14 (p14), p30 or p60. The antinociceptive effect of morphine on p14 rats was reduced in the morphine group and indicated the development of morphine tolerance. The hippocampal NMDA receptor densities have been shown decreased on p14 rats. The precipitated withdrawal symptoms were assessed on p7 rats. Rats in morphine group showed greater frequency of abdominal stretch and wet dog shake in 2 hr than control group. On the other hand, co-administration of DM with morphine effectively prevented all these adverse effects of morphine to the offspring rats. DM co-administered with morphine also partially prevented the development of morphine tolerance in maternal rats. If this effect of dextromethorphan is applied to clinical pregnant patients with morphine addiction or chronic pain, it will have a great value for the benefit of their children.     

Central antinociceptive effects of lysine-vasopressin and an analogue

Vasopressin (VP) neurons project to extrahypothalamic sites involved in pain perception, including the substantia gelatinosa of the spinal cord as well as the trigeminal and vagus nerves. Previous studies have reported antinociceptive activity following intracerebroventricular (ICV) or subcutaneous (SC) VP injections (16–100 μg) on the tail-flick test while hyperalgesia has been observed in rats either genetically deficient in VP or treated with antisera to VP. The present study investigated whether nanogram (ng) doses of lysine-vasopressin (LVP) and a VP analogue with prolonged activity increased tail-flick latencies and flinch-jump thresholds following ICV or SC injections. LVP (150 and 500 ng, ICV) significantly increased tail-flick latencies while the analogue 1-deamino-(8-Lys-N^?-(Gly-Gly-Gly))-VP (500 ng, ICV) produced more powerful and prolonged analgesia. In contrast, latencies were not increased by SC injections of LVP (150–1500 ng). Further, flinch-jump thresholds were affected minimally by either ICV or SC LVP injections. These data suggest a role for VP in pain modulation and a central site of this action.     

Thermoregulation in hyperthyroid rats: mechanism underlying the lack of hypothermic response to morphine in hyperthyroid animals

Previous experiments in our laboratory demonstrated that morphine, at doses which produced pronounced hypothermia in normal rats, not only failed to decrease but instead increased body temperature in thyroxine-treated animals. The present studies were undertaken to further investigate these initial findings. In animals treated chronically with subcutaneous thyroxine, basal body temperatures were elevated and morphine induced only hyperthermia whether given subcutaneously (10 mg/kg) or centrally (30 micrograms) into the anterior hypothalamus. Basal oxygen consumption, which reflects metabolic heat production, was significantly elevated when compared to controls. In response to morphine, control animals showed decreased oxygen consumption while thyroxine-treated animals showed a slight increase. In both groups of animals, changes in core temperatures reflected changes in oxygen consumption. These results indicate that hyperthyroid animals fail to decrease body temperature in response to morphine because they are unable to decrease metabolic heat production. Morphine, acting at central hypothalamic sites, reduces heat production in normal animals, but in thyroxine-treated animals morphine cannot overcome the increased thermogenesis.     

Effect of negative air ions on morphine-induced changes in the latency of the tail-flick reflex

Adult male Long-Evans rats were exposed to negatively charged air ions at high concentrations (7 x 10(5)/cm3) for six days. Sham-exposed rats were treated identically except that the source of ions was not activated. At the end of the exposure, the latency of the tail-flick reflex was measured in each rat before and 30 and 60 min after an injection of morphine sulphate. The tail-flick reflex was initiated by thermal stimulation. Two heat settings were employed, the lower considered to impart a submaximal and the higher a maximal thermal stimulus. Three morphine doses were tested: 0.5, 1.0, and 1.5 mg/kg. Statistically significant differences between ion-exposed and sham-exposed rats were observed in tail-flick latencies 30 min after the administration of the two lower doses, but not after the highest dose of morphine sulphate. These differences were found at both intensities of thermal stimulation. Tail-flick latencies measured in each group prior to morphine injection were not affected by negative-ion exposure. The data indicate that exposure of rats to negative air ions tends to inhibit the action of morphine on the latency of the tail-flick reflex at morphine doses below 1.0 mg/kg.     

II. N-acetyl human β-endorphin-(1–31) alleviates the morphine withdrawal syndrome in rodents: A comparative study with clonidine

The potential effect of intracerebroventricular (icv) N-acetyl human β-endorphin-(1–31) on morphine dependence was examined in mice and rats. Animals were rendered tolerant-dependent by subcutaneous (sc) implantation of an oily suspension (10 ml/Kg mouse and 3 ml/Kg rat) containing 0.1 g/ml of morphine. After 72 h of chronic morphine, 1 mg/Kg sc naloxone precipitated in both species a withdrawal syndrome that was moderate in animals pretreated with the acetylated derivative of β-endorphin. Doses of 28 fmols/rat or 80 fmols/mouse N-acetyl human β-endorphin-(1–31) reduced the number of animals presenting the jumping behaviour, as well as the number of jumps recorded. Moreover, less than half of the rats presented the other withdrawal signs evaluated: squeak on touch, diarrhoea, chattering, chewing, ptosis and body shakes. This activity could be observed when N-acetyl human β-endorphin was injected 1 h to 24 h before naloxone; longer intervals resulted in a significant loss of this activity. The ₂ agonist clonidine given icv at pmol-nmol doses decreased the incidence of morphine withdrawal syndrome. Combinations of these two substances generally did not produce any further enhancement of the effects of clonidine and N-acetyl β-endorphin when used alone. Icv injections of the antagonist of ₂-adrenoceptors yohimbine prevented both clonidine and N-acetyl β-endorphin-(1–31) from reducing the jumping behaviour displayed by morphine-abstinent mice. It is suggested that N-acetyl β-endorphin produces this alleviation of the morphine withdrawal syndrome by improving the efficiency of ₂-mediated agonist effects after acting on a neural substrate that is distinct from the μ opioid receptor binding site.     

慢性吗啡耐受大鼠脑内孤啡肽生成与释放增加

本文彩放射免疫分析法测定了慢性吗啡耐受过程中大鼠脑室灌流液、中脑导水管周围灰质（ＰＡＧ）及杏仁核中孤啡肽（ＯＦＱ）免疫活性的动态变化。结果观察到：（１）大鼠连续５ｄ皮下注射递增剂量的盐酸吗中民慢性吗啡耐受,其脑室灌流中ＯＦＱ－ｉｒ随吗啡注射剂量和注射次数的增加逐渐上升,第５ｄ注射后较对照组升高了５２％;（２）皮下注射吗啡１ｄ、３ｄ、５ｄ的大鼠ＰＡＧ中ＯＦＱ－ｉｒ比对照组分别升高了１７％、４８％和８     

Analgesic effects of ethylketocyclazocine and morphine in rat and toad

We have previously found rat and toad (Bufo marinus) brain to contain inverse ratios of benzomorphan-preferring (kappa/sigma) and morphine-preferring (mu) opioid receptor types. The aim of the present study was to compare in vivo pharmacologic activity of a benzomorphan, ethylketocyclazocine (EKC) and morphine sulfate (MS) in rat and toad. Footshock intensity thresholds for eliciting locomotion were determined and dose-response curves for EKC and MS analgesia were obtained. Drugs were injected subcutaneously. In rats (high mu, low kappa in brain), both compounds produced analgesia and displayed similar sensitivity to naloxone antagonism. The analgesic effects of EKC and MS may, therefore, be mediated by a common receptor type (mu) in this pain test in rats. In toads (high kappa, low mu in brain), MS produced naloxone-reversible analgesia at doses 20-fold higher than were effective in rats. Toads did not display EKC analgesia at doses below those producing motor impairment. Moreover, 50-fold higher doses were required to produce such impairment in toads. Thirty minutes following subcutaneous injection of 3H-EKC, similar concentrations were found in rat and toad brain. Uptake into brain is probably not a factor in the behavioral resistance of toads to EKC.     

Morphine inhibits TRH-induced intestinal transit increases

The current study examined the effects of intraperitoneal (IP) and intracisternal (IC) administration of the opiate agonist, morphine, and an opioid, central beta-endorphin, on thyrotropin releasing hormone (TRH)-induced small intestinal transit increases. Anesthetized rats, 14-day and older, were studied to determine age-related differences. Results showed that in all age groups IP morphine (2 mg/kg) blocked TRH (15 μg)-induced increases in transit of a charcoal bolus. Morphine 1 μg and beta-endorphin 1 μg administered IC in 0.6 μl failed to block TRH (10 μg)-induced increases in intestinal transit in 14-day-old rats. However, both morphine and beta-endorphin 1 μg IC blocked TRH-induced increases in adult rats. Dose-response studies demonstrated that higher doses (> 1 μg) of morphine IC were required to block TRH-induced increases in preweaning rats.     

Effects of morphine on visual evoked responses recorded in five brain sites.

The effects of morphine on averaged evoked responses to visual stimulation were examined in specific brain structures relevant to pain, analgesia, tolerance and motor disturbances. Permanent electrodes (60 μ in diameter) were implanted stereotaxically in the central gray, mesencephalic reticular formation, caudate nucleus, parafasicular-centromedian complex and the lateral geniculate body as a control site. Visual evoked responses were obtained in unanesthetized, unrestrained rats prior to and following the administration of morphine in successive doses of 1, 5, 10 and 30 mg/kg and 1 mg/kg of naloxone (a morphine antagonist). The parafasicular-centromedian complex and the reticular formation exibited a progressive increase in response amplitude to increased dose of morphine. These effects were reversed by naloxone. In this study the parafasicular-centromedian complex was found to be the most sensitive structure to morphine, displaying the largest changes in response amplitude as a result of morphine administration.     

The effect of pentobarbital on the antinociceptive action of morphine in morphine-tolerant and non-tolerant rats

The interaction of sodium pentobarbital with morphine sulfate in both morphine-tolerant and non-tolerant rats was investigated using the tail-compression test for analgesia. Male Sprague-Dawley rats (300–350 g) were given pentobarbital (4, 8, or 16 mg/kg) 5 min before morphine (2, 4, 6, or 8 mg/kg). Control animals received two saline injections, or pentobarbital plus saline, or saline plus morphine. All injections were subcutaneous. Prior to the first injection, a baseline nociceptive threshold was determined for each rat by applying a modified micrometer to its tail and increasing the pressure until a squeak was elicited. Test readings were taken every half-hour for 2 hr beginning 30 min after the second injection. For the chronic studies, animals were first made tolerant to morphine by the administration of the narcotic twice a day for 3 days, increasing the dose from 10 to 50 mg/kg/injection. Identical testing procedures were then followed with these rats except that the test dose of morphine given on day 4 was in the range 8–128 mg/kg. It was found that Na pentobarbital, in the subanesthetic doses used, had neither antinociceptive nor hyperalgesic properties. Furthermore, the barbiturate had no effect on the antinociceptive action of morphine in either morphine-tolerant or non-tolerant rats.     

Restraint Stress Potentiated Morphine Sensitization: Involvement of Dopamine Receptors within the Nucleus Accumbens

Sensitization to psychostimulant drugs, as well as morphine, subjected to cross-sensitization with stress. The development of morphine sensitization is associated with enhancements in dopamine overflow in the Nucleus accumbens (NAc). This study aimed to examine the role of accumbal D1/D2-like dopamine receptors in restraint stress (RS) induced sensitization to morphine antinociceptive effects. Adult male Wistar rats weighing 220–250 g underwent stereotaxic surgery. Two stainless steel guide cannulae were bilaterally implanted, 1 mm above the NAc injection site. Different solutions of SCH-23390, as a D1-like receptor antagonist or sulpiride, as a D2-like receptor antagonist, were microinjected into the NAc five min before exposure to RS. Restraint stress lasted for 3 h, 10 min after RS termination; animals received a subcutaneous injection of morphine (1 mg/kg) for 3 consecutive days. The procedure was followed by a 5-day drug and/or stress-free period. After that, on the 9th day, the nociceptive response was evaluated by the tail-flick test. The results revealed that intra-NAc administration of D1/D2-like dopamine receptor antagonists, SCH-23390 or sulpiride, respectively, blocked morphine sensitization-induced by RS and morphine co-administration in rats for three consecutive days. This work provides new insight into the determinant role of accumbal dopamine receptors in morphine sensitization produced by RS-morphine co-administration.

     

Further studies on the understanding of Octodon degus natural resistance to morphine: a comparative study with the Wistar rat

1. Octodon degus shows higher levels of tolerance to morphine when compared with the Wistar rat. 2. In the formalin algesiometric test, this caviomorph is more resistant to pain (P less than 0.01) and to the analgesic effect of morphine (P less than 0.001). 3. CD50 and LD50 were significantly higher in Octodon degus as compared with Wistar rat. 4. Morphine caused in rat severe hypotension, while doses eight times higher in O. degus had a transient effect. 5. 3H-naloxone binding in adrenal glands of O. degus is higher than in other tissue samples assayed from the same animal or rats.     

Opioid effects on glucose and eicosanoid metabolism in isolated uterus of ovariectomized and non-ovariectomized restricted diet rats.

The effect of a 25-day restricted diet (50% of the normal food intake) on uterine glucose metabolism of ovariectomized (25 days) and non-ovariectomized rats, was studied. Underfeeding reduces (14)CO(2) production from U(14)C-glucose in intact animal. However, in spayed rats, results are the opposite. In intact rats receiving a low food intake, the effect of the addition to the KRB medium of various agonist opioids, was studied. Dinorphin A did not bring about any change. On the other hand, beta endorphin increased glucose metabolism. Also, the addition of Dago and Dadle increased (14)CO(2) production, while their corresponding specific blockers, beta-FNA and Naltrindole, reversed it. Ovariectomized rats subjected to food restriction are not affected by opioid agonists. In vitro morphine, like endogenous opioids, increased (14)CO(2) in intact restricted diet rats. Arachidonic acid metabolism in these rats show that underfeeding brings about a decrease in PGF(2 alpha) and PGE(2), but the addition of morphine does not alter this situation, for which eicosanoids metabolites are not related to the effect of morphine. The morphine effect was not altered by naloxone. The subcutaneous injection of morphine increased glucose metabolism in intact underfed animals, while naloxone reduced (14)CO(2) in spayed rats subjected to underfeeding. It can be concluded that uteri from ovariectomized rats receiving a restricted diet are influenced by a mechanism of upregulation related to endogenous opioids. These likely originate in other tissues, and so prevent us from seeing the morphine effect.     

Chronic morphine treatment increases the number, but decreases the affinity of [3H]-U69,593 binding sites in the rat heart.

In the present study, the effects of chronic morphine treatment on the binding properties of tritiated U69,593, a specific kappa-ligand, in the rat heart were determined. Adult rats were given morphine through osmotic pumps at a rate of 80 micrograms/hour supplemented by daily injection of morphine at increasing doses for 9 days. The increase in colonic temperature to morphine, used as an indicator of development of tolerance in the rat, was measured daily. It was shown that, on day 7 following morphine treatment, the rats developed tolerance to morphine as indicated by an attenuated hyperthermic response to morphine. The [3H]-U69,593 specific binding properties were determined by direct receptor binding assay. The binding sites increased gradually and reached a significantly higher level at day 10. Scatchard analysis showed that both Bmax and Kd increased at day 10 following morphine treatment, indicating an increase in number of sites and a reduction in affinity to the kappa-ligand. Acute morphine injection at a dose of 10 mg/kg did not cause any significant alteration of [3H]-U69,593 binding sites. Two days after withdrawal of morphine, the [3H]-U69,593 binding sites returned to the original level. The finding of the present study indicates that there is an up-regulation of number, but a reduction in affinity of kappa-binding sites following chronic morphine treatment.     

Morphone abstinence and quasi-abstinence effects after phosphodiesterase inhibitors and naloxone.

Naive or morphine-dependent rats received a single subcutaneous injection of a phosphodiesterase inhibitor; their behavioral responses were then recorded after a small subcutaneous dose of naloxone. In naive rats, the potent phosphidiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX) produced acutely a state in which a small dose of naloxone (0.03 to 1.0 mg/kg subcutaneously) precipitated a quasi-morphine abstinence syndrome that was difficult to distinguish from the true abstinence syndrome, precipitated by the same dose of naloxone in rats made dependent on morphine. IBMX also intensified the true morphine abstinence syndrome. The potency with which IBMX, theophylline, caffeine and RO 20–1724 exerted these effects corresponded with their potency as inhibitors of cyclic-3′, 5′-AMP phosphodiesterase in rat brain homogenate. These and previous findings indicate that: (i) morphine-abstinence effects express increased activity of a central cyclic AMP mechanism; and (ii) naloxone can potently stimulate behavior in animals not treated with any opiate drug.     

Methionine-enkephalin and beta-endorphin levels in spleen and thymus gland of morphine tolerant-dependent and abstinent rats

The effect of chronic administration of morphine and abrupt and naloxone-precipitated withdrawal on the levels of beta-endorphin and methionine-enkephalin in spleen, adrenals and thymus glands of Sprague-Dawley rats was determined. Rats were made tolerant to and dependent on morphine by subcutaneous implantation of 6 morphine pellets (75 mg morphine in each) during a 7-day period. The tolerant-dependent (with pellets intact) and abstinent (pellets removed 18 hours earlier) rats were sacrificed. In another group, rats with pellets intact were injected with naloxone and sacrificed 10 min later (precipitated abstinence). The weights of the tissues under any of the above treatments did not change nor did the levels of methionine-enkephalin and beta-endorphin in adrenals. The level of beta-endorphin was elevated in the spleen and thymus of morphine tolerant-dependent rats, while the levels of methionine-enkephalin in rats undergoing abrupt or naloxone-precipitated abstinence were significantly higher than in their respective placebo controls. The levels of methionine-enkephalin in the thymus gland of rats with placebo and morphine pellets left intact did not differ. It is concluded that in morphine tolerant-dependent rats the levels of beta-endorphin in spleen and thymus are elevated. During abrupt and naloxone-precipitated abstinence, the levels of methionine-enkephalin in the thymus gland are significantly elevated possibly due to an inhibition of their release. Since these opioid peptides have been implicated in immunomodulation, and alterations were seen in organs controlling immune function, the present results may be helpful in explaining altered immune function in morphine dependent and abstinent states.     

Dopamine antagonist binding: a significant decrease with morphine dependence in the rat striatum

(³H)-Spiroperidol specific binding was determined in striatal tissue of rats which received a single dose of, or made dependent on morphine. Acute morphine (30 mg/kg i.p.) did not alter (³H)-spiroperidol specific binding. However, morphine-dependent rats with two 50 mg pellets when withdrawn for 24 or 48 hours, significantly decreased the binding and increased K_d. Binding sites were reduced with a decrease in K_d in rats implanted with four-50 mg pellets or receiving high doses of morphine. These results indicate that binding characteristics of (³H)-spiroperidol depend on the relative dose of morphine used to induce dependence. Low dose dependence (2 pellets) results in a decrease in binding affinity while high dose dependence (4 pellets or chronic injection) results in an increase of (³H)-spiroperidol affinity in the presence of fewer binding sites.     

Effects of morphine or naloxone on kainic acid neurotoxicity.

Intraperitoneal or subcutaneous administration of kainic acid (KA) (5–15 mg/kg) to adult rats included a syndrome of wef wet dog shakes (WDS), convulsions and brain damage. Components of the syndrome were evoked in a dose-related manner with low doses inducing WDS only and progressively higher doses being associated with an increasing incidence of naloxone (4 mg/kg) 5 minutes prior to KA (12 mg/kg) resulted in a moderate reduction in the incidence of WDS, convulsions and brain damage. Administering morphine (5 or 10 mg/kg) 10 minutes prior to KA (7 mg/kg) markedly enhanced the neurotoxicity of KA as was evidenced in an increase in the incidence of convulsions and brain damage from 7% (KA alone) to 100% (morphine + KA). KA, a structural analog of the putative excitatory transmitter glutamate (Glu), is thought to exert its excitotoxic activity through Glu excitatory receptors. Additional studies are needed to elucidate the mechanism by which morphine and naloxone respectively enhance and suppress KA neurotoxicity and to clarify whether interaction of these agents at either opioid or Glu receptors plays a role in such phenomena.     

The pharmacology of endorphin modulation of chick distress vocalization

Intraventricular injections of beta-endorphin, gamma-endorphin and alpha-endorphin were demonstrated to reduce isolation-induced distress vocalization on 2-4 day old chicks in a dose response manner at doses as small as 12.5 picomoles (pmol). beta-Endorphin was more potent than the other peptides and morphine, while Met-enkephalin was without effect. However, the D-Ala2 substituted form of Met-enkephalin was as potent as morphine. None of the opioid peptides was effective when injected peripherally in doses of 400 pmol/g body weight. Extension of the interval between injection and behavioral observation from 4 minutes eliminated the ability of alpha- and gamma-endorphin to reduce the peeps. Specificity of the opioid effect was determined by testing intraventricular injections (200 pmol) of 9 other endogenously found peptides. Somatostatin, vasoactive intestinal peptide, and human pancreatic peptide reduced the vocalizations modestly, while alpha-MSH reliably increased them.