Opiate withdrawal signs precipitated by naloxone following a single exposure to morphine: potentiation with a second morphine exposure

Recent studies in humans with no prior history of opiate abuse indicated that naloxone-precipitated signs of opiate withdrawal could be observed after a single exposure to morphine, and that the severity of withdrawal was enhanced following a second morphine exposure 24 h later. The current study was conducted to establish a paradigm in rodents that resembled these conditions described in humans. To that end, naloxone-precipitated (0.03–3.0 mg/kg) suppression of operant response rates and somatic signs of withdrawal following single or repeated treatments with morphine (5.0 mg/kg) were assessed in previously opiate-naive rats. In one group of rats, naloxone was administered 4 h after both the first and second morphine pretreatment, while in a separate group of rats naloxone was administered 4h after the second morphine pretreatment only. A single morphine pretreatment significantly increased naloxone’s potency to suppress operant response rates, and resulted in the precipitation by naloxone of certain somatic signs of withdrawal. The effects of naloxone on both dependent measures (operant response rates and somatic signs) were potentiated following a second morphine pretreatment, regardless of whether naloxone was administered following both morphine exposures or only following the second morphine exposure. Thus, repeated morphine administration appears to be the critical factor underlying the progressive increase in antagonist potency, whereas prior experience with naloxone is not a necessary factor. The results provide additional support for the hypothesis that the development of dependence on opiates is a progressive phenomenon that may begin with a single dosing. Received: 30 May 1996/Final version: 5 August 1996     

Conditioning processes contribute to severity of naloxone-precipitated withdrawal from acute opioid dependence

Rationale Single injections with morphine can induce a state of acute opioid dependence in humans and animals, typically measured as precipitated withdrawal when an antagonist such as naloxone is administered 4–24 h after morphine. Repeated treatment with morphine results in further increases in naloxone potency, and prior work has shown that this progressive shift in naloxone potency requires repeated naloxone experience under some but not all experimental conditions.Objective The current study sought to further characterize the experimental conditions that support naloxone experience-dependent and experience-independent potentiation of precipitated suppression of operant responding in morphine pretreated rats, and to assess more directly whether conditioning mechanisms may contribute to the former process.Methods Rats trained on an FR15 schedule for food received a total of five vehicle or morphine injections (5.6 mg/kg SC) at 4, 8, or 22 h prior to an operant session in which a cumulative dose-effect function for naloxone-induced suppression of responding was determined. Separate groups of animals at each interval between morphine and naloxone received cumulative naloxone dosing after all morphine pretreatments (NAL ALL DAYS) or after just the first and last morphine pretreatment (NAL FIRST/LAST). Additional groups of rats at the 4 h MOR–NAL interval received most of their naloxone cumulative dose-effect experience in either the home cage or in the operant context with levers retracted.Results Vehicle-pretreated (Morphine-Naive) rats showed little change in the naloxone dose-effect function even after five cumulative dose-effect determinations. With a single morphine pretreatment, naloxone potency was increased at 4 or 8 h post-morphine, but not at 22 h. With repeated morphine treatment, all MOR–NAL intervals resulted in significant shifts in naloxone potency across treatment days even when naloxone was administered only after the first and last morphine pretreatment. However, much greater shifts in naloxone potency were observed at 4-h and 8-h intervals when naloxone was administered on all treatment days. At the 22 h MOR–NAL interval, there was no further potentiation in naloxone potency with additional naloxone experience provided on the intermediate days. Finally, when the repeated naloxone experience occurred in the home cage at the 4-h interval, naloxone potency was identical to that seen after limited naloxone experience (NAL FIRST/LAST), and significantly less than naloxone potency in groups receiving repeated naloxone experience in the operant context.Conclusions The results suggest that conditioned withdrawal mechanisms may play a significant role in the initial development of opioid dependence.     

Repeated experience with naloxone facilitates acute morphine withdrawal: potential role for conditioning processes in acute opioid dependence

Single injections with morphine can induce a state of acute opioid dependence in humans and animals, typically measured as precipitated withdrawal when an antagonist such as naloxone is administered 4-24 h after morphine. Repeated treatment with morphine at 24-h intervals can result in a progressive shift in potency of naloxone to produce such acute withdrawal signs, including suppression of operant responding for food reward. The current study characterized fully both morphine and naloxone dose-effect functions in an effort to establish the relative contributions of repeated morphine vs. repeated naloxone (Nal) experience to these potency shifts. Rats trained on an FR15 schedule for food received four vehicle or morphine injections (0.56-5.6 mg/kg sc), spaced 24 h apart. Four hours after each morphine pretreatment (Repeat Nal), or 4 h after the fourth and final morphine pretreatment only (Single Nal), a cumulative dose-effect function for naloxone-induced suppression of responding was determined. Vehicle-pretreated (Morphine Naive) rats showed little change in the naloxone dose effect function even after four cumulative dose-effect determinations. By contrast, a progressive increase in naloxone potency was observed following successive pretreatments with morphine under Repeat Nal conditions, and the magnitude of naloxone potency shift was morphine dose dependent. At a morphine dose of 5.6 mg/kg, repeated naloxone experience in the presence of morphine was not an absolute requirement to produce an increase in naloxone potency across days, but repeated naloxone could potentiate the magnitude of the observed shift, indicating both experience-independent and experience-dependent processes at work. At lower doses of morphine (1.0 and 3.3 mg/kg) no shift in naloxone potency was observed across days of morphine treatment in the absence of repeated naloxone experience (Single Nal conditions), indicating an increasing contribution of naloxone experience-dependent processes as dose of morphine was decreased. It is argued that these experience-dependent processes in the progressive shift of naloxone potency observed in the current study may reflect an important role of conditioning in the early development of opioid dependence.     

Acute opiate withdrawal in rats undernourished during infancy: impact of the undernutrition method

Acute morphine withdrawal was assessed in adult rats following early postnatal undernutrition produced by two different methods (Large Litter procedure-20 pups/litter and Modified Slob procedure-rats cross-fostered on days 2, 4, and 6 to nonlactating dams for 24-hour periods). Response rates were first stabilized on a FR16 operant schedule. A single dose of morphine (20 mg/kg) was then administered, followed 4 h later by a single injection of naloxone (2.5 mg/kg). Males reared in large litters showed little behavioral disruption after morphine, suggesting either insensitivity to the opiate or the rapid development of tolerance. After naloxone. Modified Slob males displayed milder withdrawal than those in the well-nourished control or large litter groups. Thus the method of undernutrition influenced morphine's action and expression of withdrawal. A clear sex difference was also evident, females appearing to be generally less sensitive to the opiate- and naloxone-induced withdrawal than males. Body temperature underwent a characteristic elevation following morphine and a depression following naloxone across all groups, but undernutrition did not affect these responses. Hence, behavior proved to be the more sensitive measure for revealing differences in opiate dependence and withdrawal following early life undernutrition, under the test conditions employed.     

Rapid induction of dependence to morphine in rats

The rate of development of dependence to morphine was studied in female rats which were given increasing concentrations of morphine sulphate in their drinking fluid (5% sucrose solution). The occurrence of physical dependence was determined by the naloxone-precipitated withdrawal syndrome at various times during the 3-week experimental period. It was found that a significant degree of the withdrawal syndrome precipitated by naloxone was evident at 24 hr after starting administration of morphine; the syndrome reached its greatest intensity after the rats had received the opiate for 7 days. This study shows that dependence on morphine can be induced in rats by administration of the opiate in drinking fluid for a period shorter than 7 days.     

Relative potency of the opioid antagonists naloxone and 6-alpha-naloxol to precipitate withdrawal from acute morphine dependence varies with time post-antagonist

The current study compared the potency of naloxone versus 6-alpha-naloxol to precipitate opioid withdrawal under varying conditions of morphine pretreatment history using suppression of operant responding for food reward as the index of withdrawal. Male Wistar rats trained to respond on a lever for food reward received pretreatment with either Vehicle (Morphine-Naïve), a single subcutaneous (SC) injection of 5.6 mg/kg morphine (Single Morphine), or two morphine injections at 24 h intervals (Repeat Morphine), with varying doses of naloxone or 6-alpha-naloxol injected SC 4 h post-morphine and 5 min prior to the 30 min test session. When responding over the entire 30 min operant session was examined, naloxone was only 5-fold more potent than 6-alpha-naloxol in suppressing operant responding under Morphine Naïve conditions, but this increased to a 65-fold potency difference after Single or Repeat Morphine pretreatment. Examination of the relative potency of these antagonists in the Early Phase of operant testing (5-15 min post-antagonist) revealed an even greater 100-fold potency difference between naloxone and 6-alpha-naloxol, but in the Late Phase of testing (25-35 min post-antagonist), this had declined to a 9-fold potency difference, comparable to the relative potency of naloxone to 6-alpha-naloxol under Morphine-Naïve conditions. The results confirm a differential potency of naloxone to its reduced conjugate 6-alpha-naloxol in vivo, and extend the observation of this phenomenon to an acute (single) pretreatment with a low dose of morphine and an additional sign of opioid withdrawal to those previously used. However, the results also indicate that delay in onset of action of 6-alpha-naloxol at opioid receptors in the central nervous system may contribute significantly to its reduced potency relative to naloxone under certain morphine pretreatment conditions.     

Context- and cue-conditioned potentiation of acute morphine dependence and withdrawal

Single morphine injections induce a state of acute opioid dependence measured by an increase in naloxone potency to precipitate withdrawal. Repeated morphine exposure (daily/weekly intervals) results in further potentiation of naloxone potency, perhaps due to conditioning mechanisms. The current study tested the hypothesis that previously neutral stimuli could elicit a conditioned potentiation of the withdrawal response following acute bolus injections of morphine. Rats trained on an FR-15 schedule for food received five morphine injections (5.6 mg/kg) at daily intervals. Four hr after morphine injection on Conditioning Days (first 4 days), naloxone (1 mg/kg)-induced suppression of responding was paired either with operant context only, or with a tone/light conditioned stimulus (CS). On Test Day low dose naloxone (0.001-0.33 mg/kg) given 4-h post-morphine preceded the operant session. Rats exposed to naloxone repeatedly in the operant context without CS (Paired-Context Only) showed an increase in naloxone potency on Test Day relative to Unpaired Controls that received all morphine and naloxone in the home cage at a different time of day than operant testing. Rats exposed to the tone/light CS on Conditioning Days also showed a significant increase in naloxone potency relative to Unpaired Controls when the CS was represented on Test Day (Paired-CS), but not when the CS was omitted on the Test Day (Paired-CS/Test Context). Thus, conditioning processes appear to play a significant role in the early development of opioid dependence and withdrawal.     

Buprenorphine and a CRF1 antagonist block the acquisition of opiate withdrawal-induced conditioned place aversion in rats.

Conditioned place aversion in rats has face validity as a measure of the aversive stimulus effects of opiate withdrawal that reflects an important motivational component of opiate dependence. The purpose of the present study was to validate conditioned place aversion as sensitive to medications that will alleviate the aversive stimulus effects of opiate withdrawal in humans, and to extend this model to the exploration of the neuropharmacological basis of the motivational effects of opiate withdrawal. Male Sprague-Dawley rats were implanted with two subcutaneous morphine pellets and 5 days later began place conditioning training following subcutaneous administration of a low dose of naloxone. Animals were subjected to three pairings of a low dose of naloxone (15 microg/kg, s.c.) to one arm of a three-chambered place conditioning apparatus. Buprenorphine administered prior to each pairing dose-dependently blocked the place aversion produced by precipitated opiate withdrawal. A corticotropin-releasing factor-1 (CRF1) receptor antagonist (antalarmin) also reversed the place aversion produced by precipitated opiate withdrawal. Antalarmin did not produce a place preference or place aversion by itself in morphine-dependent rats. No effect was observed with pretreatment of the dopamine partial agonist terguride or the selective serotonin reuptake inhibitor fluoxetine. Also, chronic pretreatment with acamprosate (a glutamate receptor modulator used to prevent relapse in alcohol dependence) did not alter naloxone-induced place aversion. Buprenorphine by itself in dependent rats produced a mild place preference at low doses and a mild place aversion at higher doses. These results suggest that buprenorphine blocks the aversive stimulus effects of precipitated opiate withdrawal in rats and provides some validity for the use of place conditioning as a measure that is sensitive to potential opiate-dependence medications. In addition, these results suggest that CRF1 antagonists can block the aversive stimulus effects of opiate withdrawal and may be potential therapeutic targets for opiate dependence.     

The dopamine antagonist, alpha-flupenthixol, interferes with naloxone-induced place aversion learning, but not with acute opiate dependence in rats.

Pretreatment with the dopamine antagonist alpha-flupenthixol (alpha-flu) interfered with the establishment of a naloxone-induced place aversion in two experiments. In Experiment 1, the potential of pretreatment with alpha-flu to interfere with the establishment of a naloxone-induced place aversion was evaluated in rats administered morphine (Group MN) or saline (Group SN) 24 h prior to the naloxone conditioning trial. Naloxone-precipitated withdrawal from morphine administered 24 h prior to the trial produced a stronger place aversion than produced by naloxone alone. The neuroleptic, alpha-flu, attenuated the naloxone-induced place aversion, but did not selectively interfere with the place aversion produced by acute opiate dependence. Experiment 2 replicated demonstration of interference with naloxone-place aversion learning by neuroleptic pretreatment with the inclusion of saline controls. These results suggest that dopamine modulates either the aversive motivational properties of naloxone or learning, even in opiate na?ve rats.     

Attenuation of isobutylmethylxanthine-induced suppression of operant behavior by pretreatment of rats with clonidine

Administration of 3-isobutyl-1-methylxanthine (IBMX) to rats performing a FR30 operant for food reinforcement produces a dose-dependent suppression of behavior. Operant behavior suppressed by 5 mg IBMX/kg is attenuated by pretreatment, 30 min before the operant session, with the alpha 2 adrenergic agonist clonidine (5-30 micrograms/kg). Clonidine itself causes a dose-dependent reduction in FR30 responding prior to the administration of IBMX. However, doses of clonidine which also suppressed responding were not more effective than lower doses in attenuating the suppression of operant behavior caused by IBMX, perhaps due to postsynaptic or nonspecific actions of clonidine. Methylxanthines, alone or in combination with the opiate antagonist naloxone, produce signs of opiate withdrawal. This quasi-morphine withdrawal syndrome may be useful in studies of either the development or expression of opiate withdrawal. Since clonidine attenuates the rate-suppressant effect of IBMX, it is likely that a significant component of IBMX's behavioral effects are due to increases in NE neurotransmission. These results are similar to those obtained with true opiate withdrawal in rats, strengthening the idea that suppression of operant behavior by IBMX involves mechanisms in common with opiate withdrawal. It may be a useful way of objectively studying the expression of the withdrawal syndrome in the absence of opiates and/or a way of determining if a drug can selectively block withdrawal.     

Opiate antagonist--receptor interaction unchanged by acute or chronic opiate treatment.

The apparent pA2 of naloxone, a measure of the apparent receptor affinity for the antagonist, was estimated in naive rats, in rats after acute and various schedules of chronic morphine pretreatment, and in rats during abrupt withdrawal. The results were adjusted to compensate for the amount of pretreatment drug in the rat brains as, otherwise, such a drug could have inflated the apparent pA2 values (see appendix). After this adjustment, a tendency of the apparent pA2 values to be increased after chronic pretreatment disappeared. The apparent pA2 of naloxone in acutely pretreated rats was also unchanged from that in naive rats, while the results in abruptly withdrawn rats were ambiguous. Thus, no change in the affinity of the antagonist for opiate receptors was found and theories suggesting that a conformational change takes place in the opiate receptor during the development of tolerance/dependence could not be supported.     

Withdrawal from acute morphine dependence is accompanied by increased anxiety-like behavior in the elevated plus maze

Pretreatment with a single moderate dose of morphine (e.g. 5.6-10 mg/kg) 4-24 hr prior to challenge with an opioid antagonist such as naloxone results in reliable expression of behaviors that resemble aversive or emotional consequences of withdrawal from chronic opioid exposure, including suppression of operant responding, elevations in brain reward thresholds, and conditioned place aversion. Repeated daily or weekly treatment with these same morphine doses results in a progressive increase in naloxone potency to elicit these withdrawal signs. The current study sought to determine whether increased anxiety-like behavior during withdrawal from chronic opioid dependence is also seen after acute morphine exposure, and progresses with repeated intermittent treatment. Male Wistar rats were handled and injected with either vehicle or morphine for 4 consecutive days. Three injection regimens were employed: Morphine Naive (4 vehicle injections), Acute Morphine (3 vehicle injections, 4th injection 5.6 or 10 mg/kg morphine), or Repeat Morphine (all 4 injections with 5.6 or 10 mg/kg morphine). Acute pretreatment with 5.6 mg/kg or 10 mg/kg morphine resulted in time-dependent increases in exploration of the open arms of the plus maze in naloxone-naive rats when tested at 2, 4 or 8 hr after the final pretreatment injection, with the effects at the higher dose appearing later (4 hr) than after the lower dose (2 hr). This pattern of results, in combination with a separate study which confirmed a significant anxiolytic-like effect of a low dose of morphine (0.56 mg/kg) administered 15 min prior to test, suggested that low residual morphine levels remaining in plasma at 2-4 hr after 5.6 and 10 mg/kg morphine may be sufficient to elicit anxiolytic-like effects. Repeat treatment with either dose of morphine resulted in a further increase in the magnitude and duration of this anxiolytic-like effect. These effects had dissipated by 8 hr post-morphine, and therefore precipitation of withdrawal by one of several doses of naloxone (0.10-3.3 mg/kg) was assessed in separate cohorts of rats 8 hr after the final pretreatment under Morphine Naïve, Acute Morphine, or Repeat Morphine conditions. Naloxone resulted in a significant dose-dependent expression of anxiety-like behavior with no effects on general activity after Acute Morphine pretreatment at either 5.6 or 10 mg/kg morphine. A further significant shift in naloxone potency was observed after Repeat Morphine pretreatment at the 10 mg/kg but not the 5.6 mg/kg dose. Thus, anxiety-like behavior is a prominent feature of the negative emotional consequences of naloxone-precipitated withdrawal from acute opioid dependence.     

Dorsal and median raphe serotonergic system lesion does not alter the opiate withdrawal syndrome

Previous pharmacological studies have implicated serotonergic brain systems in opiate withdrawal. To test the hypothesis that serotonin (5-HT) has a critical role in the development of opiate withdrawal, we have employed a near-total brain 5-HT system lesion technique (90% depletion) using 5,7-dihydroxytryptamine combined with induction of opiate dependence by implantation of morphine pellets or by repeated injections of increasing doses of morphine. The effects of serotonergic neuron lesion were examined on spontaneous opiate withdrawal (changes in circadian locomotor activity) and naloxone-precipitated opiate withdrawal syndrome (the somatic aspect). The antiwithdrawal properties of clonidine, an alpha(2)-adrenoceptor agonist currently used for clinical treatment for the somatic signs of opiate withdrawal, were tested also in the lesioned rats. Our findings show that serotonergic lesions in morphine-dependent rats did not alter either the spontaneous or the naloxone-induced withdrawal syndrome (with exception of jumping behavior). Moreover, clonidine alleviated the naloxone-induced withdrawal syndrome in lesioned as well as in sham-operated morphine-dependent rats. These results demonstrate that 5-HT systems are not directly responsible for the development of the somatic opiate withdrawal syndrome in morphine-dependent rats.     

Chronic opiate treatment enhances both cocaine-reinforced and cocaine-seeking behaviors following opiate withdrawal

After chronic exposure to psychostimulants or opiates, self-administration or conditioned place preference with either class is increased (sensitized). Cross-sensitization of conditioned place preference, i.e., enhancement of psychostimulant-induced preferences after exposure to opiates, has also been described, but increases in cocaine self-administration after morphine pretreatment have not been reported. The present study evaluated effects of chronic morphine treatment on cocaine reinforcement. Opiate dependence was established in Wistar rats by administration of morphine as a constant infusion that was gradually increased to a dose of 50mg/kg per day over a 1-week period. Immediately after discontinuation of chronic morphine treatment, animals were allowed to acquire cocaine self-administration under a simple fixed-ratio schedule (FR-1), and were subsequently advanced to a progressive ratio schedule. Acquisition of cocaine self-administration under the FR-1 did not differ in saline- and morphine-pretreated animals. For cocaine self-administration under a progressive ratio schedule measured at 5 or more days after the onset of opiate withdrawal, chronic pretreatment with morphine increased the number of ratios completed, augmented final response requirements, and produced a more stable pattern of cocaine self-administration. Responding was also increased in morphine-pretreated animals during an initial extinction session. These results show that chronic opiate treatment can enhance both cocaine-reinforced and cocaine-seeking behaviors following opiate withdrawal. A similar effect may occur in human patients who discontinue methadone or other forms of replacement therapy for opiate abuse, and may contribute to relapse involving use of cocaine or other psychostimulants.     

Discrete cues paired with naloxone-precipitated withdrawal from acute morphine dependence elicit conditioned withdrawal responses

Acute bolus doses of morphine induce a state of acute opioid dependence as measured by naloxone-precipitated withdrawal. Repeated morphine and precipitated withdrawal experience further enhances naloxone-induced withdrawal severity, partly because of direct neuroadaptation to repeated morphine, and partly because of conditioned associations of context and withdrawal experience. To determine whether a discrete tone/light conditioned stimulus could elicit conditioned withdrawal responses in acute dependence, rats trained on a fixed-ratio-15 operant schedule for food reward received morphine (5.6 mg/kg) 4x at daily or weekly intervals, with each morphine injection followed at 4 h by naloxone (1.0 mg/kg) and an operant session. The conditioned stimulus was presented to a Paired group after each naloxone injection. Separate control groups experienced the conditioned stimulus either at a different time of the day or on a different day of the week than naloxone (Unpaired), received naloxone without any conditioned stimulus exposure [Paired-no conditioned stimulus (Paired-NO CS)] or received vehicle instead of naloxone before conditioned stimulus presentation (NaI-Naive). On the test day, all rats received vehicle before conditioned stimulus exposure. The conditioned stimulus alone reliably suppressed responding in Paired groups relative to control conditions with either daily or weekly intervals between conditioning sessions. The administration of morphine 4 h before conditioned stimulus exposure on the test day was not necessary to observe conditioned withdrawal. Thus, conditioned withdrawal is reliably established to discrete cues associated with naloxone-precipitated withdrawal from acute, infrequent (weekly) opioid exposure.     

Altered motivation and learning following opiate withdrawal: evidence for prolonged dysregulation of reward processing.

Opiate abuse has been associated with cognitive deficits in human addicts. To determine if prior opiate exposure alters the ability to learn, we trained animals in an instrumental learning task for a food reward. During a 2-week period after withdrawal, morphine-abstinent rats were significantly slower at learning an escalating fixed-ratio response for food reward compared to placebo-treated animals. When these same animals were trained in a conditioned suppression paradigm (two tone-shock pairings given in the operant box), the morphine-withdrawn animals showed greater retention by taking significantly longer to resume responding for food reward when the tone was presented. In a third experiment, morphine-abstinent rats withdrawn 2 or 5 weeks were tested for their ability to associate a highly palatable food reward with a specific environment using a place-conditioning paradigm. At 2 weeks postwithdrawal, morphine-abstinent rats did not show any significant place preference for a food they readily consumed, while placebo-treated rats readily learned to prefer the food-paired environment. At 5 weeks postwithdrawal, rats developed significantly less preference for food-associated cues, but more preference for morphine-associated cues, compared to placebo-treated animals. These data suggest that prior morphine exposure may have prolonged effects on the motivation for natural rewards, which in turn may compromise the ability of former addicts to overcome their addictions.     

Behavioral expression of opiate withdrawal is altered after prefrontocortical dopamine depletion in rats: monoaminergic correlates.

The objective of this study was to establish the effects of prefrontocortical dopamine depletion on opiate withdrawal and prefrontocortical neurochemical changes elicited by morphine dependence and withdrawal. The dopaminergic content was also measured in the nucleus accumbens during withdrawal, in order to detect reactive changes induced by prefrontocortical lesion. Withdrawal was induced by naloxone in morphine-dependent rats. Monoamine levels were analyzed post-mortem by high performance liquid cromatography. The results showed that chronic morphine dependence did not modify basal levels of monoamines in sham rats, revealing neuroadaptation of prefrontocortical dopamine, noradrenaline and serotonin systems to chronic morphine. The neuroadaptive phenomenon remained after prefrontocortical lesion (> 79% dopamine depletion). On the other hand, a strong increase of dopamine, noradrenaline, and serotonin contents in the medial prefrontal cortex of sham rats was detected during opiate withdrawal. However, in lesioned rats, the increase of prefrontocortical dopamine and serotonin content, but not that of noradrenaline, was much lower. In the nucleus accumbens, prefrontocortical lesion reactively enhanced the dopaminergic tone and, although opiate withdrawal reduced dopaminergic activity in both sham and lesioned rats, this reduction was less intense in the latter group. At a behavioral level, some symptoms of physical opiate withdrawal were exacerbated in lesioned rats (writhing, mastication, teeth-chattering, global score) and exploration was reduced. The findings hence indicate that: (i) prefrontocortical monoaminergic changes play a role in the behavioral expression of opiate withdrawal; (ii) the severity of some withdrawal signs are related to the dopaminergic and serotonergic tone of the medial prefrontal cortex rather than to the noradrenergic one, and (iii) an inverse relationship between mesocortical and mesolimbic dopaminergic systems exists.     

Long-term opiate effects on amphetamine-induced dopamine release in the nucleus accumbens core and conditioned place preference

Withdrawal following chronic exposure to opiates or other drugs of abuse, administered as frequent doses, or a chronic infusion can cause reductions in mesolimbic dopamine (DA) transmission. However, mesolimbic DA transmission can be enhanced by opiates or psychostimulants administered intermittently as a single daily injection. Both enhanced and attenuated responsiveness of the mesolimbic DA system may have important implications for substance abuse disorders. Previous studies have shown that procedures that use electrical stimulation or drug treatments to augment neurotransmitter release are more effective for demonstrating declines in mesolimbic DA transmission that persist for extended periods following opiate withdrawal. The present study evaluated the effects of pretreatment with noncontingent morphine on amphetamine-induced DA release in the nucleus accumbens core and conditioned place preference (CPP). Morphine pretreatment was administered as a constant infusion, which was gradually increased to a dose of 50 mg/kg/day over a 1-week period in Wistar rats. At 10 days after cessation of morphine pretreatment, baseline dialysate DA levels in the nucleus accumbens core were unchanged, but amphetamine-induced increases in DA were attenuated by greater than 50% in morphine-pretreated animals. Morphine pretreatment did not modify locomotor activity during conditioning sessions, expressed as absolute values or change in activity counts between saline and morphine injections. Place preference, conditioned by two morphine pairings at 10 and 11 days after the onset of opiate withdrawal, was enhanced by opiate pretreatment between 12 and 33 days after the onset of withdrawal. In conclusion, morphine pretreatment delivered as a constant infusion can have pronounced and long-lasting effects on DA release and CPP, which may have important implications for drug-seeking behavior and treatment of substance abuse disorders.     

Ketanserin and pirenperone attenuate acute morphine withdrawal in rats

The involvement of serotonin2 (5-HT2) receptors in the expression of opiate withdrawal was examined using a behavioral test for acute morphine dependence. The 5-HT2 antagonists, ketanserin and pirenperone, injected shortly before naloxone, attenuated the naloxone-induced suppression of an autoshaped lever-touch response in rats treated 4 h earlier with a moderate dose of morphine. A low dose of pirenperone was also effective in blocking withdrawal-induced hypothermia. These data support the hypothesis that 5-HT is involved in the expression of opiate withdrawal.     

Influence of omega-conotoxin on morphine analgesia and withdrawal syndrome in rats.

The effect of omega-conotoxin on opiate analgesia and withdrawal syndrome was investigated in rats. omega-Conotoxin given i.c.v. and i.p. caused weak analgesia in the tail-flick test. When the toxin (20 ng/rat) was given i.c.v. immediately before morphine (1.5 micrograms/rat i.c.v.) the resultant analgesic effect was additive. In contrast, the analgesia elicited by morphine (3 micrograms/rat i.c.v.) was greatly reduced after 24-h pretreatment with the toxin (20 ng/rat i.c.v.). The systemic administration of the toxin (10 micrograms/kg i.p.) did not affect morphine analgesia whether omega-conotoxin was coadministered with morphine (2.5 mg/kg i.p.) or was given 24 h before the opiate (5 mg/kg i.p.). omega-Conotoxin i.c.v. injected in morphine-dependent rats 15 min before naloxone challenge significantly attenuated the abstinence syndrome. On the contrary systemic administration of omega-conotoxin failed to suppress the morphine withdrawal syndrome. The present results suggest that omega-conotoxin affects both acute and chronic effects of morphine.