Systemic and intra-accumbens microinjections of naltrexone interfere with tolerance to ethanol in rats

Rationale Evidence suggests a role for the opioid system in the control of ethanol reinforcement and drinking. Previous findings have shown that naltrexone, an opioid antagonist that decreases ethanol consumption in humans and experimental animals, reduces the acquisition of acute ethanol tolerance in rats. However, there are few data regarding the role of the opioid system in the acquisition of ethanol tolerance, particularly in brain areas involved in the rewarding actions of ethanol. Objectives This study investigates the effects of systemic and of intra-accumbens injections of naltrexone on the development of rapid tolerance to ethanol. Methods Wistar rats received intraperitoneal injections of naltrexone (0.1–3.0 mg/kg) or microinjections into the core or shell portions of the nucleus accumbens (5–20 μg) before ethanol (2.7 g/kg i.p.). The animals were tested for motor coordination on the tilting plane apparatus. Tolerance was assessed 24 h later by administering the same dose of ethanol to all animals and retesting them on the tilting plane. Results The second injection of ethanol resulted in less motor incoordination on Day 2, suggesting the development of rapid tolerance. Pretreatment with naltrexone, either i.p. (0.3 and 0.6 mg/kg) or intra-accumbens (5–20 μg), on Day 1, blocked the development of rapid tolerance to the motor-incoordinating effects of ethanol on Day 2 without affecting the motor performance of the animals on Day 1. Conclusions The results suggest that the opioid system may be involved in the development of ethanol tolerance, and that the nucleus accumbens may play a role in this phenomenon.     

Dopamine depletion augments endogenous opioid-induced locomotion in the nucleus accumbens using bothμ1 andδ opioid receptors

The aim of this study is to analyze further the opioid receptor subtypes involved in the augmentation of behavioral activity after dopamine depletion in the nucleus accumbens of rats. Initially, the opioid receptors involved in the augmentation of locomotion produced by endogenous opioids were evaluated by microinjection of kelatorphan, an inhibitor of proteolytic enzymes that inactivates enkephalin, with or without specific antagonists forμ ₁ orδ-opioid receptors, naloxonazine or naltrindole, respectively. Kelatorphan produced a dose-dependent increase in horizontal photocell counts and vertical movements. At all doses examined the behavioral response was augmented in rats sustaining accumbal dopamine lesions. The augmentation in dopamine-depleted rats was partially blocked by naloxonazine or naltrindole. Since the motor stimulant response to intra-accumbens microinjection of theδ-opioid agonist, [d-penicillamine^2,5]-enkephalin, was not augmented in a previous study, we tested the behavioral response to a new endogenousδ-opioid agonist, [d-Ala²] deltorphin I. The locomotor response to deltorphin was slightly augmented in dopamine-depleted rats. These data suggest that the augmentation in the motor response elicited by endogenous opioids after dopamine lesions in the nucleus accumbens involves bothμ ₁ andδ-opioid receptors.     

κ-opioid receptors are implicated in the increased potency of intra-accumbens nalmefene in ethanol-dependent rats

Previously, it was shown that ethanol-dependent animals display increased sensitivity to the general opioid receptor antagonist nalmefene compared to naltrexone. It was hypothesized that the dissociable effects of the two antagonists were attributable to a κ-opioid receptor mechanism. Nucleus accumbens dynorphin is upregulated following chronic ethanol exposure and such neuroadaptations could contribute to nalmefene's increased potency in ethanol-dependent animals. To test this hypothesis, male Wistar rats were trained to self-administer ethanol using an operant conditioning procedure. Animals were then implanted with bilateral intra-accumbens shell guide cannulae and assigned to either a chronic intermittent ethanol vapor-exposure condition (to induce dependence) or an air-exposed control group. Following a one-month exposure period, nalmefene, nor-binaltorphimine (nor-BNI; selective for κ-opioid receptors) or a combination of the selective opioid receptor antagonists CTOP and naltrindole (selective for the μ- and δ-opioid receptors, respectively) were site-specifically infused into the nucleus accumbens shell prior to ethanol self-administration sessions during acute withdrawal. Nalmefene and CTOP/naltrindole dose-dependently reduced ethanol self-administration in nondependent and dependent animals, whereas nor-BNI selectively attenuated ethanol self-administration in ethanol-dependent animals without affecting the self-administration of nondependent animals. Further analysis indentified that intra-accumbens shell nalmefene was more potent in ethanol-dependent animals and that the increased potency was attributable to a κ-opioid receptor mechanism. These data support the concept that dysregulation of DYN/κ-opioid receptor systems contributes to the excessive self-administration observed in dependent animals and suggest that pharmacotherapeutics for ethanol dependence that target κ-opioid receptors, in addition to μ- and δ-opioid receptors, are preferable to those that target μ- and δ-opioid receptor mechanisms alone.     

Role of protein kinase C epsilon (PKCɛ) in the reduction of ethanol reinforcement due to mGluR5 antagonism in the nucleus accumbens shell

Rationale  The type 5 metabotropic glutamate receptor (mGluR5) and the epsilon isoform of protein kinase C (PKCɛ) regulate ethanol intake, and we have previously demonstrated that mGluR5 receptor antagonism reduces ethanol consumption via a PKCɛ-dependent mechanism. Objectives  We explored the potential neuroanatomical substrates of regulation of ethanol reinforcement by this mGluR5-PKCɛ signaling pathway by infusing selective inhibitors of these proteins into the shell or core region of the nucleus accumbens (NAc). Methods  Male Wistar rats were trained to self-administer ethanol intravenously and received intra-NAc infusions of vehicle or the selective mGluR5 antagonist 3-((2-methyl-1,3-thiazol-4-yl)ethynyl)pyridine (MTEP) alone and in combination with a PKCɛ translocation inhibitor (ɛV1–2) or a scrambled control peptide (sɛV1–2). The effects of intra-NAc MTEP on food-reinforced responding and open-field locomotor activity were also determined. Results  MTEP (1 μg/μl) had no effect on ethanol or food reinforcement or locomotor activity when infused into either region. MTEP (3 μg/μl) reduced ethanol reinforcement when infused into the NAc shell but not the core, and this effect was reversed by ɛV1–2 (1 μg/μl) but not sɛV1–2 (1 μg/μl). In both regions, this concentration of MTEP did not alter food-reinforced responding or locomotor activity, and infusion of ɛV1–2 alone did not alter ethanol reinforcement. MTEP (10 μg/μl) reduced locomotor activity when infused into the shell; therefore, this concentration was not further tested on responding for ethanol or food. Conclusions  Blockade of mGluR5 receptors in the NAc shell reduces ethanol reinforcement via a PKCɛ-dependent mechanism.     

Involvement of μ- and δ-opioid receptors in the effects of systemic and locally perfused morphine on extracellular levels of dopamine, DOPAC and HVA in the nucleus accumbens of the halothane-anaesthetized rat

The effects of systemic and intra-accumbens infusion of morphine on the extracellular level of dopamine (DA) and its metabolites, DOPAC and HVA, were investigated in the nucleus accumbens (NAcc) of halothane-anaesthetized rats using in vivo microdialysis. Morphine in a dose of 1 or 5mg/kg i.v. produced a significant increase (60-100% of baseline levels) in the extracellular level of DA for at least 3 h. Morphine at 5, but not 1mg/kg, produced a small (10-15%) but significant reduction in the level of DOPAC when compared with saline in the first h following injection. Pretreatment with the preferential μ-opioid receptor antagonist naloxone in a dose of 1 or 3mg/kg i.p. significantly blocked the morphine-induced changes in the extracellular levels of DA and DOPAC. Pretreatment with the selective δ-opioid receptor antagonist, naltrindole, at 1mg/kg i.p. blocked only the morphine-induced decrease in DA metabolism. Furthermore, in the presence of naltrindole, systemic morphine induced a large and long-lasting increase in the level of DOPAC and HVA, which was significantly higher than in rats receiving combinations of saline/water + saline, saline/water + morphine and naltrindole + saline. When applied directly into the NAcc, morphine at concentrations of 125, 250 and 500 ng infused over 10min produced a dose-related increase in the extracellular level of accumbens DA with either no effect or a small reduction in the level of DOPAC and HVA. The effects of intra-accumbens morphine on DA levels were significantly blocked by pretreatment with i.p. naltrindole but not naloxone. These results indicate that, while systemic morphine probably increases DA via activation of μ-opioid receptors, local perfusion of morphine increases DA in the NAcc via activation of δ-opioid receptors located in the NAcc. Furthermore, under the conditions of the study, it appears that activation of μ- and δ-opioid receptors by morphine respectively increases and decreases DA metabolism. Received: 4 December 1995 / Accepted: 27 December 1996     

Effects of the local administration of selective μ-, δ-and κ-opioid receptor agonists on osteosarcoma-induced hyperalgesia

The stimulation of peripheral opioid receptors yields analgesic responses in a model of bone cancer-induced pain in mice. In order to know the type(s) of peripheral opiate receptors involved, the paw thermal withdrawal latencies were measured in C3H/HeJ mice bearing a tibial osteosarcoma, after administering selective agonists of μ-,δ-and κ-opiate receptors. The peritumoral administration of DAGO (0.6–6 μg) inhibited the osteosarcoma-induced hyperalgesia at doses ineffective in healthy animals, the highest one even increasing the withdrawal latencies over the control values. Naloxone-methiodide (2 mg/kg) and cyprodime (1 mg/kg), but not naltrindole (0.1 mg/kg) nor nor-binaltorphimine (10 mg/kg), antagonized DAGO-induced analgesic effects, these therefore probably being mediated through peripheral μ-opioid receptors. The peritumoral injection of DPDPE (100 μg) induced analgesia which was inhibited by naloxone-methiodide and naltrindole but not by nor-binaltorphimine. Cyprodime partially antagonized the analgesia induced by 100 μg of DPDPE, but did not modify the effect induced by 30 μg of this agonist—a dose that restores the hyperalgesic latencies up to the control values. The antihyperalgesic effect induced by the peritumoral administration of U-50,488H (1 μg) was antagonized by naloxone-methiodide and nor-binaltorphimine, but not by cyprodime nor naltrindole, thus suggesting the involvement of peripheral κ-opioid receptors. In conclusion, the stimulation of peripheral μ-, δ- and κ-opioid receptors is a pharmacological strategy useful for relieving this experimental type of bone cancer-induced pain, the greatest analgesic effect being achieved by stimulating peripheral μ-opioid receptors.     

Deep brain stimulation of the nucleus accumbens reduces ethanol consumption in rats

Recent studies have shown that deep brain stimulation (DBS) of the nucleus accumbens (NAcc) has an inhibitory effect on drug-seeking behaviors including reinstatement responding for cocaine. The objective of the present study was to expand on these findings by assessing the effects of DBS on behaviors related to alcohol consumption. The specific aim of this study was to determine whether DBS delivered to either the shell or core of the NAcc would reduce ETOH intake in rats using a two-bottle choice limited access procedure. Long Evans rats were induced to drink a 10% ethanol solution using a saccharin fading procedure. Bipolar electrodes were implanted bilaterally into either the core or shell of the NAcc. During testing animals received DBS 5 min prior to and during a 30-minute test session in which both ETOH and water bottles were accessible. Current was delivered at amplitudes ranging from 0 to 150 µA. ETOH consumption was significantly reduced from baseline levels at the 150 µA current for both shell and core electrode placements. A significant current effect was not found for water consumption for either site. These results provide evidence that DBS delivered either to the nucleus accumbens core or shell subregions can significantly reduce ethanol intake in the rat.     

Phase II trial of fenretinide (NSC 374551) in patients with recurrent small cell lung cancer

Background Alterations in retinoid signaling appear to be involved in the pathogenesis of small cell lung cancer (SCLC). Fenretinide [N-(4-hydroxyphenyl)retinamide], a synthetic retinoid, inhibits the growth of SCLC cells in vitro via the induction of apoptosis. Since these data suggested that SCLC is the adult solid tumor that is most susceptible to fenretinide, a trial to evaluate the clinical activity of fenretinide in patients with SCLC was considered the definitive test of its clinical potential in adult oncology. Methods Patients with progressive SCLC after one or two prior chemotherapy regimens and a performance status of 0–2 were eligible for the study. Patients with stable, treated brain metastases were eligible. Fenretinide 900 mg/m² twice daily was administered orally on days 1–7 of each 21-day cycle. Blood and saliva were collected pre-treatment and on day 7 of cycle 1 to measure fenretinide and retinol levels by high-pressure liquid chromatography (HPLC). Results Nineteen patients were enrolled. Fifteen patients had one prior chemotherapy regimen and four patients had two prior regimens. The median time from diagnosis to enrollment was 10 months. A median of two cycles of fenretinide was administered. There were no objective responses, but four of 17 evaluable patients (24%) had stable disease after 2–17 cycles. The median time to treatment failure was 5.7 weeks overall, while the four patients with stable disease demonstrated treatment failure at 11, 13, 19, and 52 weeks. Median survival was 25 weeks, with one patient alive 22 months after the start of treatment. The 1-year survival rate was 29%. Toxicity included mild, reversible visual changes (haziness, altered night vision), grade 1–3 nausea/vomiting, and grade 1–2 diarrhea. The mean day 7 plasma fenretinide level was 2.90 ± 1.66 μg/ml (7.40 ± 4.25 μM; n = 14). The mean pre-treatment and day 7 plasma retinol levels were 0.47 ± 0.16 μg/ml and 0.05 ± 0.07 μg/ml (n = 8), respectively. The mean day 7 salivary fenretinide level was 0.08 ± 0.18 μg/ml, with no correlation between salivary and plasma drug levels. Conclusions Fenretinide is well tolerated in patients with SCLC and stabilization of disease was noted in 24% of patients with this aggressive disease. However, after the first stage of enrollment, the response rate did not meet criteria to proceed with full trial accrual. Plasma concentrations of fenretinide that induce cytotoxicity in vitro in SCLC cell lines are clinically achievable, but there were no objective responses. Non-invasive drug monitoring using saliva underestimates systemic exposure.     

Anatomically dissociable effects of dopamine D1 receptor agonists on reward and relief of withdrawal in morphine-dependent rats

Rationale  Chronic opiate administration induces neuroadaptations within the nucleus accumbens (NAc) and ventral tegmental area (VTA) that can contribute to dependence. We have shown that morphine dependence shifts the behavioral consequences of D1 dopamine (DA) receptor signaling: systemic administration of a D1 receptor agonist is rewarding and blocks naloxone-precipitated withdrawal signs in morphine-dependent rats, but has minimal effects in nondependent rats. These data suggest that D1 receptors acquire the ability to regulate reward and withdrawal in morphine-dependent rats. The brain regions involved in these effects are not known. Objective  Studies were designed to test the hypothesis that the nucleus accumbens shell (NASh) and the ventral tegmental area (VTA) are important sites for mediating the behavioral effects of D1 receptor activation in morphine-dependent rats. Materials and methods  The effects of microinjecting the D1 receptor agonist SKF 82958 into the NASh or the VTA on place conditioning and somatic withdrawal signs were studied in morphine-dependent and nondependent rats. Results  Intra-NASh microinjection of SKF 82958 (1 μg/side) established conditioned place preferences in morphine-dependent but not nondependent rats, but had no effect on naloxone-induced place aversions or somatic withdrawal signs. Intra-VTA microinjection of SKF 82958 (2 μg) did not establish place preferences under any conditions, but blocked naloxone-induced place aversions without effects on somatic withdrawal signs. Conclusions  There is an anatomical dissociation between D1 receptor-mediated reward and relief of withdrawal in morphine-dependent rats. When combined, the individual effects of D1 receptor activation in the NASh and VTA on the affective signs of precipitated morphine withdrawal resemble those seen with systemic administration.     

Ethanol exposure differentially alters pro-enkephalin mRNA expression in regions of the mesocorticolimbic system

Rationale Opioid peptides have been suggested to play a major role in ethanol reinforcement mechanisms and alcohol drinking behaviour. However, in non-selected strains of rodents, it is not known whether opioid biosynthesis is a critical event in these processes.Objective The aim of this work was to study the effects of a high dose of ethanol (2.5 g/kg body weight) on pro-enkephalin (pro-enk) mRNA expression in brain regions of the mesocorticolimbic system for up to 24 h after drug administration.Materials and methods Male Wistar rats were administered with ethanol (2.5 g/kg body weight) or distilled water and were killed 30 min, 1, 2, 4, 8 or 24 h after treatment. Coronal brain sections (20 μ) were obtained and pro-enk mRNA expression was studied by in situ hybridization and densitometry.Results Acute ethanol administration induced a transient decrease and increase in pro-enk mRNA expression in the ventral tegmental area (33.2%) and prefrontal cortex (26.5%) 2 and 4 h after treatment, respectively. In contrast, ethanol induced prolonged increases in pro-enk mRNA expression in the core and shell regions of the nucleus accumbens, with different kinetics. Maximal effects were observed 2 h after ethanol exposure (core, 70.0%; shell, 60.0%).Conclusions Our results indicate that enkephalin expression in regions of the rat mesocorticolimbic system is differentially altered by acute ethanol treatment and suggest that enkephalins may play a key role in ethanol reinforcement mechanisms.     

100Hz电针通过激活伏隔核内κ阿片受体抑制可卡因CPP的表达

目的：探讨100Hz电针抑制大鼠可卡因条件位置偏爱（Conditioned Place Preference,CPP）表达的机制。方法：采用可卡因诱导大鼠CPP模型,观察（1）选择性κ阿片受体拮抗剂nor-BNI加100Hz电针对可卡因CPP表达的影响;（2）每次电针前给大鼠双侧伏隔核内注射nor-BNI,能否阻断电针对可卡因CPP的表达;（3）电针处理可卡因CPP大鼠伏核组织中κ阿片受体mRNA表达的变化。结果：（1）10μg／5μl nor—BNI侧脑室给药或0．3μg／1μl nor-BNI伏隔核内微注射预处理给药都能翻转100Hz电针对可卡因CPP的抑制作用;（2）100Hz电针能显著增加可卡因CPP大鼠伏隔核内κ阿片受体mRNA的表达。结论：100Hz电针通过激活伏隔核内κ阿片受体从而抑制可卡因CPP的表达。     

Role of dopamine D1-family receptors in dorsolateral striatum in context-induced reinstatement of heroin seeking in rats

Rationale  In humans, exposure to environmental contexts previously associated with heroin intake can provoke relapse to drug use. In rats, exposure to heroin-associated contexts after extinction of drug-reinforced responding in different contexts reinstates heroin seeking. This effect is attenuated by blockade of D₁-family receptors in lateral or medial accumbens shell, but not accumbens core. Objectives  In this study, we further characterized the role of striatal D₁-family receptors in context-induced reinstatement by assessing the effect of dorsolateral or dorsomedial injections of the D₁-family receptor antagonist SCH 23390 on this reinstatement. Materials and methods  Rats were trained to self-administer heroin (0.05–0.10 mg/kg per infusion) for 12 days; drug infusions were paired with a discrete tone–light cue. Subsequently, heroin-reinforced lever pressing was extinguished in the presence of the discrete cue in a nondrug context. During reinstatement tests under extinction conditions, the D₁-family receptor antagonist SCH 23390 (0.3–1.0 μg per side) was injected into the dorsolateral or dorsomedial striatum prior to exposure to heroin self-administration context or the nondrug (extinction) context. We then used a disconnection procedure to examine whether D₁-family receptors in the dorsolateral striatum and lateral accumbens shell jointly or independently support context-induced reinstatement. Results  Dorsolateral but not dorsomedial SCH 23390 injections attenuated context-induced reinstatement of heroin seeking. SCH 23390 injections into the dorsolateral striatum of one hemisphere and lateral accumbens shell of the other hemisphere were ineffective. Conclusions  Results indicate that dorsolateral striatum D₁-family dopamine receptors are critical for context-induced reinstatement of heroin seeking. Results also suggest that D₁-receptor-mediated dopamine transmission in the dorsolateral striatum and lateral accumbens shell independently support this reinstatement.     

Involvement of the opioid system in the effects induced by nicotine on anxiety-like behaviour in mice

Rationale Recent studies have revealed the participation of the endogenous opioid system in several behavioural responses induced by nicotine including antinociception, rewarding properties, and physical drug dependence. Objectives The present study was designed to examine the possible involvement of the various opioid receptors in the anxiolytic- and anxiogenic-like responses induced by nicotine in mice. Methods The acute administration of low (0.05) or high (0.8 mg/kg) doses of nicotine subcutaneously produced opposite effects in the elevated plus maze, i.e. anxiolytic- and anxiogenic-like responses, respectively. Animals were only exposed once to nicotine. The effects of the pretreatment with the μ-opioid receptor antagonist, β-funaltrexamine (5 mg/kg), the δ-opioid antagonist, naltrindole (2.5 mg/kg) and the κ-opioid antagonist, nor-binaltorphimine (2.5 mg/kg) intraperitoneally were evaluated on the anxiolytic- and anxiogenic-like responses induced by nicotine. Results β-funaltrexamine, but not nor-binaltorphimine or naltrindole, abolished nicotine-induced anxiolytic-like effects, suggesting an involvement of μ-opioid receptors in this behavioural response. On the other hand, naltrindole, but not nor-binaltorphimine or β-funaltrexamine, increased the anxiogenic-like responses of nicotine, suggesting an involvement of δ-receptors in this behavioural effect. Conclusions These results demonstrate that the endogenous opioid system is involved in the effects induced by nicotine on anxiety-like behaviour and provide new findings to further clarify the interaction between these two neurochemical systems.     

Effects of SR141716 and WIN 55,212-2 on tolerance to ethanol in rats using the acute and rapid procedures

Rationale Our previous findings have shown rapid cross-tolerance between ethanol and Δ⁹-tetrahydrocannabinol and that intraperitoneal (i.p.) injection of cannabinoid receptor type 1 (CB1R) antagonist SR141716 (SR) does not interfere with tolerance to either of these drugs in mice. Objectives This study investigates the effects of SR, alone or in combination with the CB receptor agonist WIN 55,212-2 (WIN), on the development of acute and rapid tolerance to the incoordinating effect of ethanol in rats. Materials and methods Male Wistar rats received SR, through i.p. (0.5–2.0 mg/kg) or intracerebroventricular (i.c.v.) injections (0.5–4.0 μg), alone or together with WIN (1.0 μg, i.c.v.), in combination with ethanol (2.7 g/kg, i.p.). Another group received WIN (1.0 μg, i.c.v.) in combination with ethanol (2.3 g/kg), and the rats were tested for motor coordination. Rapid tolerance was assessed 24 h later by administering ethanol to all animals and retesting them under the same dose regimen. Acute tolerance was evaluated for 75 min after ethanol (3.0 g/kg, i.p.) in animals treated with SR or WIN (i.c.v.). Results The reduced motor impairment on day 2 (i.e., rapid tolerance) was blocked by SR (i.p. and i.c.v.). WIN (1.0 μg, i.c.v.) facilitated rapid tolerance and also prevented the blockade of rapid tolerance by SR (1.0 μg, i.c.v.). In the acute tolerance procedure, SR did not affect the motor incoordination induced by ethanol. Conclusions The results suggest that the endocannabinoid system may contribute to the development of rapid tolerance to ethanol.     

Intracranial self-administration of MDMA into the ventral striatum of the rat: differential roles of the nucleus accumbens shell, core, and olfactory tubercle

Rationale Behavioral and anatomical data suggest that the ventral striatum, consisting of the nucleus accumbens and olfactory tubercle, is functionally heterogeneous. Cocaine and d-amphetamine appear to be more rewarding when administered into the medial olfactory tubercle or medial accumbens shell than into their lateral counterparts, including the accumbens core. Objectives We sought to determine whether rats self-administer the popular recreational drug (±)-3,4-methylenedioxymethamphetamine (MDMA) into ventrostriatal subregions and whether the medial olfactory tubercle and medial accumbens shell mediate MDMA’s positive reinforcing effects more effectively than their lateral counterparts. Results Rats receiving 30 mM MDMA into the medial olfactory tubercle, medial accumbens shell, or accumbens core, but not the lateral tubercle or lateral shell, showed higher self-administration rates than rats receiving vehicle. The medial shell supported more vigorous self-administration of MDMA at higher concentrations than the core or medial olfactory tubercle. In addition, intra-medial shell MDMA self-administration was disrupted by co-administration of the D1 or D2 receptor antagonists SCH 23390 (1–3 mM) or raclopride (3–10 mM). Conclusions Our data suggest that the ventral striatum is functionally heterogeneous. The medial accumbens shell appears to be more important than other ventrostriatal subregions in mediating the positive reinforcing effects of MDMA via both D1- and D2-type receptors. Together with previous data, our data also suggest that unidentified actions of MDMA interfere with the positive reinforcing effects of dopamine in the medial olfactory tubercle.     

Inflammatory processes enhance cAMP-mediated uterus relaxation in the pregnant rat: the role of TNF-α

The objective of this study was to assess the in vitro uterus relaxing potency of β₂-adrenergic receptor (β₂-AR) agonists in pregnant rats after in utero administration of the bacterial lipopolysaccharide, Escherichia coli endotoxin (LPS). The LPS (100 μg/kg) was injected into the uterine lumen on day 16 of pregnancy. The effects of β₂-AR agonist terbutaline was tested in vitro, in isolated uterine rings precontracted by electric field stimulation. Uterine β₂-AR densities were detected by radioligand binding assay, the activated G-protein levels were investigated by a radiolabelled GTP binding assay. Uterine cAMP accumulation and the serum tumor necrosis factor-α (TNF-α) levels were measured by enzyme immunoassay. The endotoxin-evoked preterm delivery occurred on day 21. Higher pD₂ values of terbutaline (p < 0.001) were detected in endotoxin-treated rats: 9.14 ± 0.36 vs. 7.71 ± 0.12 compared with sham-operated rats. The densities or the equilibrium dissociation constants of β₂-ARs were not different (p > 0.05) in LPS-treated vs. control animals. Serum TNF-α level rose threefold after LPS treatment, but this rise was abolished by thalidomide. In LPS + thalidomide-treated rats, the effect of terbutaline became similar to that in sham-operated controls. By the measurement of myometrial cAMP levels, we documented that the concentration–response curve of terbutaline on cAMP accumulation was shifted to the left in the LPS-treated rats, with a significant rise in the pD₂. We concluded that in the case of uterine inflammation, the in vitro uterus-relaxing potency of β₂-agonists enhances, which is possibly mediated by TNF-α and uterine cAMP levels and that may serve as a rationale for the use of β₂-AR agonists in the attenuation of preterm uterine contractions on an inflammatory basis.     

Intra-Accumbens trh prolongs maintenance of tolerance to hypothermic effect of ethanol in rats

Tolerance to the hypothermic effect of ethanol (Et–OH) developed in male Wistar rats treated daily with 5 g/kg of 20 per cent ET–OH v/v i.g. for 6 days. On Day 6 Et–OH treatment was discontinued and the animals were divided into two groups injected into the nucleus accumbens septi (NAS), either with pGlu-His-Pro-NH₂ (TRH) or saline. The peptide was injected via permanently implanted cannulae in doses of 10 or 30 m?g in 0.5 m?l on each side once daily. The effect of such treatment on the maintenance of Et–OH tolerance was assessed by measuring the temperature decrement in response to a challenge dose of Et–OH (5 g/kg i.p.) given on Day 10. It was found that in saline-treated rats the initial tolerance declined. TRH-injected groups exhibited constantly low response to hypothermic action of Et–OH. In a control experiment TRH was shown to have no effect on the body temperature after intra-accumbens application in Et–OH-naive rats.     

The role of vasopressin on the effect of U-50,488 to block the development of morphine tolerance and physical dependence

U-50,488, a selective κ-opioid receptor agonist, has been reported to inhibit the development of antinociceptive tolerance to morphine in mice, rats and guinea pigs, but the mechanism involved in this action remains unknown. Since U-50,488 has been reported to supress the plasma vasopressin level, we investigated the role of vasopressin with U-50,488 in the male Sprague Dawley rat in this study. Animals (230–270 g) were chronically treated with morphine (10 mg/kg, i.p.) twice a day for 6 days in order to induce tolerance to antinociceptive effect measured by tail-flick test. Withdrawl symptoms were precipitated by naloxone (10 mg/kg, i.p.) on day 7. U-50,488 (i.p.) or AVP (i.p. or i.c.v.) or U-50,488 and AVP was (were) coadministered with chronic morphine to investigate their effects on morphine tolerance and dependence. We found that coadministration of 8 mg/kg U-50,488 (i.p.) with morphine almost completely block morphine tolerance and partially block withdrawal symptoms. In contrast, coadministration of AVP (0.3 μg/kg, i.p., or 0.01 μg, i.c.v.) with morphine and U-50,488, the effects of U-50,488 to block morphine tolerance and dependence were reversed. In addition, treatment of AVP antagonist (dPTyr(Me)AVP, 0.5 μg/kg, i.p. or 0.5 μg, i.c.v.) has the similar effect as U-50,488 to block morphine tolerance. In summary, the effect of U-50,488 to block morphine tolerance and dependence may relate to its inhibitory effect on AVP release. Received: 20 February 1996 / Accepted: 14 October 1996     

Induction by mercury compounds of brain metallothionein in rats: Hg0 exposure induces long-lived brain metallothionein

Metallothionein (MT) is one of the stress proteins which can easily be induced by various kind of heavy metals. However, MT in the brain is difficult to induce because of blood-brain barrier impermeability to␣most heavy metals. In this paper, we have attempted to induce brain MT in rats by exposure to methylmercury (MeHg) or metallic mercury vapor, both of which are known to penetrate the blood-brain barrier and cause neurological damage. Rats treated with MeHg (40 μmol/kg per day × 5 days, p.o.) showed brain Hg levels as high as 18 μg/g with slight neurological signs 10␣days after final administration, but brain MT levels remained unchanged. However, rats exposed to Hg vapor for 7 days showed 7–8 μg Hg/g brain tissue 24 h after cessation of exposure. At that time brain MT levels were about twice the control levels. Although brain Hg levels fell gradually with a half-life of 26 days, MT levels induced by Hg exposure remained unchanged for >2␣weeks. Gel fractionation revealed that most Hg was in the brain cytosol fraction and thus bound to MT. Hybridization analysis showed that, despite a significant increase in MT-I and -II mRNA in brain, MT-III mRNA was less affected. Although significant Hg accumulation and MT induction were observed also in kidney and liver of Hg vapor-exposed rats, these decreased more quickly than in brain. The long-lived MT in brain might at least partly be accounted for by longer half-life of Hg accumulated there. The present results showed that exposure to Hg vapor might be a suitable procedure to provide an in vivo model with enhanced brain MT. Received: 25 June 1997 / Accepted: 4 November 1997     

Reduction in repeated ethanol-withdrawal-induced anxiety-like behavior by site-selective injections of 5-HT1A and 5-HT2C ligands

Rationale Anxiety-like behavior resulting from repeated withdrawals from chronic ethanol diets is counteracted by systemic administration of a 5-HT_2C receptor antagonist or a 5-HT_1A receptor partial agonist.Objectives This study investigated whether prior treatment with these agents into the amygdala, dorsal raphe nucleus, nucleus accumbens, or paraventricular nucleus during early withdrawals would ameliorate the social interaction deficits observed after a subsequent withdrawal.Methods Sprague–Dawley rats were exposed to three cycles of 5 days of forced ethanol diet (4.5%, w/v), with 2 days of control diet after the first and second cycles. Drugs were administered into one of four brain sites 4 h after removal of ethanol on the first and 2nd cycles but not the third. The social interaction test was performed 5 h after removal of ethanol on the third cycle. Drugs tested included SB-243213, a 5-HT_2C receptor inverse agonist; buspirone, a 5-HT_1A receptor partial agonist; and Ro 60 1075, a 5-HT_2C receptor agonist.Results Only SB-243213 (at 3 μg, but not at 1 and 0.3 μg) counteracted the social interaction deficits after injections into the amygdala, while buspirone (at 0.3 and 1 μg but not at 0.1 μg) reduced deficits only when given into the dorsal raphe nucleus. In contrast, the 5-HT_2C receptor agonist, Ro 60 1075, accentuated the behavioral deficit after two weekly injections into the amygdala.Conclusions These results are consistent with the involvement of 5-HT_2C receptors in the amygdala and 5-HT_1A autoreceptors in the dorsal raphe nucleus in repeated ethanol withdrawal-induced sensitization of anxiety-like behavior.For publication in Psychopharmacology.