Additive effects of intra-accumbens infusion of the cAMP-specific phosphodiesterase inhibitor, rolipram and cocaine on brain stimulation reward.

Evidence from cocaine self-administration studies suggests that increasing the activity of cyclic AMP (cAMP) pathways within the nucleus accumbens may produce a reduction in cocaine's reinforcing effects. Rolipram may increase intra-cellular levels of cAMP by selectively inhibiting Type IV phosphodiesterases, enzymes that catalyze cAMP breakdown. The present study was undertaken to test the hypothesis that infusion of rolipram into the nucleus accumbens would decrease cocaine-induced enhancement of the sensitivity of brain stimulation reward (BSR) pathways. BSR thresholds were determined in rats after the systemic administration of cocaine (4 mg/kg IP) and the infusion of rolipram (0.2 microg/side) into the nucleus accumbens both alone and in combination. Thresholds also were determined after the systemic administration of rolipram alone and, as a positive control, for amphetamine (10 microg/side) infused into the nucleus accumbens. BSR thresholds were significantly lowered below baseline levels following d-amphetamine administration suggesting that cannulae were in place to allow perfusion of reward pathways. Compared to values for saline alone, thresholds were lower after the injection of cocaine (4 mg/kg IP) or the infusion of rolipram (0.2 microg/side) into the nucleus accumbens. Treatment with the combination of cocaine and intra-nucleus accumbens rolipram produced a greater lowering of the BSR threshold than did administration of either rolipram or cocaine alone. Systemic administration of rolipram (0.5 mg/kg IP) either blocked the effects of BSR or raised BSR thresholds and produced stimulation-induced head jerking in most of the test animals. These results suggest that infusion into the nucleus accumbens of rolipram, an agent that putatively elevates cAMP levels in this structure, can enhance the sensitivity of reward pathways to BSR and can augment cocaine's actions on these pathways.

Adenosine agonists CGS 21680 and NECA inhibit the initiation of cocaine self-administration.

Administration of the adenosine antagonist caffeine will facilitate the reinstatement of cocaine self-administration responding. This suggests that adenosine receptors may play a role in the motivational systems that regulate cocaine-seeking behaviors. If so then adenosine agonists may act to block cocaine self-administration. To test this hypothesis, the effects of the nonselective adenosine agonist NECA and of the A2A selective agonist, CGS 21680 on the self-administration of cocaine were determined. In these experiments, rats were allowed to obtain intravenous cocaine infusions (0.6 mg/kg/infusion) delivered under a Fixed Ratio 5 schedule. Treatment with either NECA or CGS 21680 in comparison to vehicle administration reduced the number of infusions received per session. This, primarily, was due to a marked increase in the latency for delivery of the first cocaine infusion. Responding after drug-induced delays tended to be at control levels. Adenosine agonists are known to have sedative effects and these actions might play a role in NECA and CGS 21680-induced increases in latencies for cocaine delivery. These results indicate that the administration of adenosine agonists may inhibit cocaine-seeking behaviors. The degree to which these actions are on motivational systems as opposed to involving less specific effects remains to be fully elucidated.

Cue-induced changes in basal local cerebral glucose utilization 13 days after morphine sensitization in the Fischer 344 rat: relevance for drug craving.

The present experiment tested the hypothesis that some persistent neural adaptation develops during the course of repeated sensitizing doses of morphine administered to rats. A sub-hypothesis was that this imprint would be of greater magnitude in the presence of morphine-conditioned cues. In order to test these hypotheses basal local cerebral metabolic rates for glucose (LCMR(glu)) were determined 13 days after the last of four 10-mg/kg doses of morphine administered in 36 h to Fischer 344 male rats. LCMR(glu) was determined using the 2-deoxy-D-[1-(14)C]glucose method (2-DG). Half of the rats, the conditioned group, were placed in the 2-DG chamber after each injection and half, the nonconditioned group, were placed in a neutral environment. A control group received only saline in lieu of morphine. All metabolic rates were determined in a nondrugged state. The major finding was large increases in metabolic rate throughout the forebrain in the sensitized rats. This was especially so in the conditioned group, 46 out of 93 areas examined had significant increases while in the nonconditioned group it was 25 out of 93. Both the core and shell of the nucleus accumbens showed significant elevations in metabolic rates in the presence of morphine cues but only the shell in the absence of the cues. There were no significant decreases in basal metabolic activity in any of the brain regions evaluated in either experimental group. The present finding suggests that changes in the brains of these morphine-sensitized rats may model the altered brain states responsible for drug craving in human drug addicts.

Fluoxetine blocks expression but not development of sensitization to morphine-induced oral stereotypy in rats.

RATIONALE: Repeated high doses of morphine in the rat cause stereotypic gnawing behavior that can be re-expressed by a low dose of morphine weeks and even months after the initial treatment. The determination of the role of serotonin in this sensitized morphine-induced behavior has both empirical and theoretical relevance. OBJECTIVES: To determine whether the serotonin-reuptake blocker fluoxetine will block the development and/or the expression of this opiate-induced stereotypy. METHODS: Rats were given four 10-mg/kg injections of morphine alone or with 5.0 mg/kg fluoxetine over a 36-h period. At weekly intervals for 6 weeks after the last of the sensitizing morphine doses, all rats were challenged with 4.0 mg/kg morphine. At week 2 and week 4, however, the morphine was co-administered with fluoxetine. RESULTS: Fluoxetine completely blocked the expression of the morphine-induced stereotypy; however, when the morphine/fluoxetine-treated rats were challenged with morphine alone, they expressed similar degrees of stereotypy as the rats that initially only received morphine. CONCLUSIONS: The results indicate that increasing synaptic serotonin will block the expression but not the development of sensitization to the oral stereotypic effects of repeated high doses of morphine. Also, despite the complete blocking of the morphine effect by fluoxetine during the sensitization phase, the presence of significant biting by these rats during the challenge with morphine alone argues that conditioning factors are not a necessary component for the morphine sensitization to develop.

The irreversible gamma-aminobutyric acid (GABA) transaminase inhibitor gamma-vinyl-GABA blocks cocaine self-administration in rats.

gamma-Vinyl gamma-aminobutyric acid (GABA) (GVG) is an irreversible inhibitor of GABA transaminase, the primary enzyme involved in GABA metabolism. Acute administration of GVG increases brain GABA levels and blocks cocaine-induced locomotor activity, cocaine-induced lowering of brain stimulation reward thresholds, and cocaine-induced conditioned place preference. To further evaluate the effects of GVG on cocaine-induced reward, we examined its effects on cocaine self-administration in male Wistar rats on fixed ratio 5 and progressive ratio schedules of reinforcement. Additionally, the effects of GVG on operant responding for a food reward were examined on the same two schedules to determine whether the effects of GVG were specific to cocaine reward or generalized to other types of reward. GVG dose dependently decreased responding for cocaine on both schedules of reinforcement, suggesting that GVG attenuated the reward value of the cocaine. Responding for food was also decreased by GVG, suggesting that the effects of increased GABA levels induced by GVG may have a general effect on central reward systems. Data from this and other studies indicate that GVG does not induce motor impairment, decrease spontaneous locomotor activity, or induce catalepsy. Taken together, these data suggest that increases in GABAergic activity induced by GVG have an attenuating effect on centrally mediated reward systems and that the GABA system may be a useful target in the development of new therapeutic strategies for cocaine addiction.

The type IV phosphodiesterase inhibitors, Ro 20-1724 and rolipram, block the initiation of cocaine self-administration.

The hypothesis that the selective activation of cyclic AMP (cAMP) signal transduction pathways will suppress the initiation of cocaine self-administration was examined in this investigation. To test this hypothesis, the effects of the administration of the cAMP-specific (type IV) phosphodiesterase inhibitors, rolipram and Ro 20-1724, on cocaine self-administration were determined. The effects of Ro 20-1724 treatment on operant responding for food also were examined. Both cocaine and food were delivered following a fixed-ratio 5 schedule. A significant increase in the latency for the delivery of the first cocaine infusion and a reduction in the number of infusions obtained per session were produced by treatment with either rolipram or Ro 20-1724. Similar effects on responding for food were seen with Ro 20-1724 administration. Responding after drug-induced delays tended to be at control levels. These results suggest that cAMP-specific phosphodiesterase inhibitors may inhibit the initiation of operant responding for either cocaine or food. However, the extent to which these actions involve specific effects on central motivational systems as opposed to other mechanisms remains to be determined.

Prefrontal cortex infusions of SCH 23390 cause immediate and delayed effects on ventral tegmental area stimulation reward.

A reward-relevant relationship between dopamine projection regions of the ventral tegmental area (VTA) was investigated through the use of brain stimulation reward (BSR) thresholds. Using a rate-free method, changes in VTA BSR thresholds were determined after intracranial injections of the dopamine D1 antagonist, SCH 23390 into the prefrontal cortex (PFC), or the nucleus accumbens (NAcc). Reward thresholds assessed immediately after the infusion of SCH 23390 into the NAcc (0.5 microgram/0.5 microliter/side) were significantly higher than those assessed just after saline infusions, indicating a drug-induced attenuation of the rewarding effects of the brain stimulation. The effects of this dose subsided when tested 24 h later. Conversely, intra-PFC infusions of SCH 23390 at the same dose (0.5 microgram/0.5 microliter/side) resulted in lowered BSR thresholds when rats were tested immediately after infusion. In addition, animals tested 24 h after receiving the lowest dose (0.125 microgram/0.5 microliter/side) demonstrated a robust delayed threshold-lowering effect. These immediate and delayed effects of the intra-PFC dopamine antagonist demonstrate a facilitation of VTA BSR and are consistent with the view that PFC dopamine serves a modulatory role over important reward elements within the NAcc. The deferred effects of intra-prefrontal cortex DA receptor blockade on brain stimulation reward thresholds may reflect adaptive responses of subcortical structures to changes in PFC dopamine neurotransmission. It has been suggested that neural adjustments of this type may underlie long term changes in central nervous system functioning brought about by disease, drug use or behavioral conditioning.

The synergistic effects of combining cocaine and heroin ("speedball") using a progressive-ratio schedule of drug reinforcement.

The relative reinforcing value of cocaine/heroin combination ("speedball") was compared in the rat using a progressive-ratio (PR) reinforcement schedule. The initial training for all rats was a combined dose of 18 microg/kg/inj of heroin (H) plus 300 microg/kg/inj of cocaine (C). Break points for the training dose and individual component doses were determined for half and double the training dose. Of the three doses of each treatment, only C yielded the expected monotonic increase in break point as a function of dose. Also, break points for C (300 and 600 microg/kg/inj) was greater than for the combination of C and H (18 H/300 C and 36 H/600 C microg/kg/inj), suggesting a greater reward value for C alone. The doses for these three drug treatments that produced saline level break points were then determined. At these lower doses, significant break points were obtained with the H/C combination at which the respective doses of H or C had break points identical to those of saline. These lower dose results indicate that the combination is clearly synergistic and that the discrepancy with doses at the opposite end of the dose response curve suggest that the PR schedule is vulnerable to drug-induced motor effects.

Acute and chronic fluoxetine treatment decreases the sensitivity of rats to rewarding brain stimulation.

The effects of fluoxetine on rewarding brain stimulation were determined in eight Wistar rats using a rate-independent discrete-trial threshold measure. Rats were implanted with bipolar, stainless steel electrodes either into the ventral tegmental area (VTA) or medial forebrain bundle (MFB). Acute administration of fluoxetine significantly raised the reward threshold (decreased sensitivity) at doses of 2.5, 5.0, 10.0, and 20.0 mg/kg, i.p., without altering latency of response. There were no significant differences between VTA and MFB groups. To determine the effects of chronic treatment, daily injections of 5.0 mg/kg fluoxetine were administered to rats for 21 days. Chronic treatment of fluoxetine continued to significantly elevate reward thresholds with no evidence of tolerance. The results of these experiments suggest that fluoxetine does not possess abuse potential and that serotonin produces an inhibitory effect on the mesolimbic dopaminergic reward system. Furthermore, these results suggest that the antidepressant effects of fluoxetine are not the direct result of excitation of brain reward systems, at least in the same manner as abused substances, for example, cocaine.

DAMGO and DPDPE facilitation of brain stimulation reward thresholds is blocked by the dopamine antagonist cis-flupenthixol.

The role of dopamine neurotransmission in opioid reward was investigated using a rate-independent measure for determining brain stimulation reward (BSR) thresholds. Intra-accumbens infusions of the mu- and delta-specific peptides, D-Ala2, N-Me-Phe4, Gly-ol5-Enkephalin and D-Pen2, D-Pen5-Enkephalin caused significant lowering of BSR thresholds. The dopamine D1/D2 antagonist, cis-flupenthixol, blocked these effects at a dose that did not significantly alter thresholds when given alone. These data suggest both mu- and delta-opioid potentiation of BSR is dopamine dependent.

Persistent increases in basal cerebral metabolic activity induced by morphine sensitization.

To characterize the underlying neuroanatomic substrate of morphine (MS) sensitization, changes in the local cerebral metabolic rate for glucose (LCMRglu) were examined in 95 brain regions of male F-344 rats using the 2-deoxy-D-[1-14C]glucose method. The results of these experiments demonstrate that MS-induced sensitization is manifested by increases in basal metabolic activity that last for at least 6 days. Although changes in basal metabolic rate were found to be more extensive in the presence of conditioned cues, the increases in LCMRglu in nonconditioned sensitized rats indicate a basic underlying pharmacologic effect of MS sensitization on basal brain activity. Regions in which MS sensitization had a lasting pharmacologic effect include the shell of the nucleus accumbens, the prelimbic area of the prefrontal cortex, and the dorsolateral prefrontal cortex. Interestingly, the core of the nucleus accumbens and regions of the caudate were found to have an increased LCMRglu only in the presence of conditioned cues, indicating conditioned brain activity without observable changes in behavior. The previous administration of an MS-sensitizing treatment was also found to alter the cerebral metabolic response to a subsequent acute MS challenge (0.5 mg/kg, subcutaneously), most notably in forebrain systems. The more widespread activation of brain structures in the basal state in the presence of conditioned cues suggests that these MS-sensitized rats may model an altered brain state related to craving in the abstinent opiate addict.

Gamma-vinyl GABA attenuates cocaine-induced lowering of brain stimulation reward thresholds.

Gamma-vinyl GABA (GVG, also referred to as vigabatrin), an irreversible inhibitor of GABA transaminase (GABA-T), raises levels of GABA in nerve terminals, inhibits striatal dopamine release, and attenuates cocaine-induced increases in extracellular dopamine in the striatum and nucleus accumbens. In order to determine the action of GVG on dopamine-mediated reward, we examined its effects on the threshold for rewarding brain stimulation in male F-344 rats. GVG dose-dependently raised brain stimulation reward (BSR) thresholds at doses of 200, 300, and 400 mg/kg without significant effects on motor performance as measured by response latencies. In order to determine if GVG had similar modulatory effects on cocaine-induced lowering of BSR thresholds, the effective doses of GVG were co-administered with 2.5 and 5.0 mg/kg cocaine, doses that significantly lower BSR thresholds. The 400 mg/kg dose of GVG significantly blocked the lowering of thresholds seen at each dose of cocaine. Cocaine in combination with 200 or 300 mg/kg GVG, doses of GVG that significantly raise BSR thresholds, resulted in thresholds not significantly different from those obtained with cocaine alone. These data demonstrate that, at the doses tested, GVG is more effective at modulating basal reward thresholds that at modulating thresholds lowered by cocaine, implying that as dopaminergic activity increases, GABAergic activity must also increase in order to exert its inhibitory influence on dopaminergic activity.

Involvement of delta- and mu-opioid receptors in the potentiation of brain-stimulation reward.

A rate-free method of determining brain-stimulation reward thresholds was used to identify the rewarding effects of the delta-opioid receptor and mu-opioid receptor agonist peptides, [D-Pen2, D-Pen5]enkephalin (DPDPE) and [D-Ala2-MePhe4-Gly(o1)5]enkephalin (DAMGO). The nucleus accumbens-delivered opioid receptor agonists produced marked lowering of the threshold for ventral tegmental area brain-stimulation reward. No change in baseline thresholds was seen after peripheral administration of the nonpeptide delta-opioid receptor antagonist, naltrindole. However, an unexpected finding was that naltrindole blocked the threshold-lowering effects of both DPDPE and DAMGO. These data demonstrate nucleus accumbens activation of delta- and mu-opioid receptors and ventral tegmental area brain-stimulation reward share common brain substrates. In addition, the interference of both delta- and mu-opioid receptor mediated reward by naltrindole may have implications for therapeutic use.

Acute stress causes mucin release from rat colon: role of corticotropin releasing factor and mast cells.

We determined the effects of immobilization stress on rat colonic mucus release and mast cell degranulation and examined whether corticotropin releasing factor (CRF) was involved in these responses. After 30-min immobilization, rats were killed, colonic mucosal explants were cultured, and levels of rat mast cell protease II (RMCP II) and prostaglandin E2 (PGE2) were measured. Mucin release from explants was assayed by incorporation of [3H]glucosamine into colonic mucin and by histological evaluation of goblet cell depletion. Stress caused significant increases of colonic RMCP II, PGE2, and mucin release and fecal pellet output and caused an approximately 10-fold increase in colonic mucosal levels of cyclooxygenase-2 (COX-2) mRNA. These stress-associated changes were reproduced by intravenous or intracerebral injection of CRF in conscious, nonstressed rats. Pretreatment of rats with the CRF antagonist alpha-helical-CRF9-41, hexamethonium, atropine, or bretylium, or the mast cell stabilizer lodoxamide inhibited stress-induced release of RMCP II, PGE2, and mucin, whereas indomethacin prevented mucin release but not mast cell degranulation. Hexamethonium and CP-96,345, a substance P antagonist, inhibited fecal pellet output caused by stress. We conclude that CRF released during immobilization stress increases colonic transit via a neuronal pathway and stimulates colonic mucin secretion via activation of neurons and mast cells.

Low-dose apomorphine attenuates morphine-induced enhancement of brain stimulation reward.

Thresholds for brain stimulation reward (BSR) delivered to the medial forebrain bundle-lateral hypothalamus were determined by means of a rate free psychophysical method. Lower doses of apomorphine (0.5 to 0.2 mg/kg) produced modest elevations in BSR thresholds. A 0.4 mg/kg dose of apomorphine resulted in emergence of stereotypic behaviors and the loss of stimulus control. Morphine's BSR threshold lowering effects were significantly blocked by the concurrent administration of a 0.1 mg/kg dose of apomorphine. These results support the hypothesis that presynaptic dopamine neurons are involved in the mediation of morphine's reinforcing effects and that dopamine autoreceptor agonists may be of some use in the pharmacotherapy of opiate abuse.

Naloxone alters the local metabolic rate for glucose in discrete brain regions associated with opiate withdrawal.

The current 2-deoxy-D-[1-14C]glucose investigation was performed to test the hypothesis that endogenous opioids influence basal synaptic activity within discrete brain regions. To examine this hypothesis, the effects of naloxone (1.0 mg/kg s.c.) on local cerebral metabolic rate for glucose (LCMRglu) in 84 brain regions were compared to saline controls. The specificity of naloxone's effects for opioid receptors was assessed by the coadministration of the opiate agonist morphine in a separate group. In naloxone-treated rats, there was a significant decrease in LCMRglu in the locus coeruleus (LC) and an increase in the central nucleus of the amygdala (CAMY), supporting a tonic influence of endogenous opioids on these regions. These metabolic changes were reversed by coadministered morphine, indicating that naloxone's metabolic actions are specific for opioid receptors. Based on the role of the LC and CAMY in opiate withdrawal, the present results suggest a subthreshold naloxone precipitated withdrawal from endogenous opioids. Although morphine administered alone significantly reduced LCMRglu in 16 brain regions, these did not include the LC or the CAMY. These results identify brain regions in which synaptic activity is under tonic modulation by endogenous opioids.

Reexpression of morphine-induced oral stereotypy six months after last morphine sensitizing dose.

We describe three experiments, two of which were designed to determine the duration of the sensitization to morphine-induced oral stereotypy caused by three high doses of morphine (MS) administered 12 h apart (10, 20, and 20 mg/kg). In one of these experiments, we also administered an additional 40 mg/kg. The third experiment was designed to determine the role of mode of drug administration in the development of MS-induced sensitization; drug was delivered by an implanted minipump. By the third high dose, all rats manifested marked repetitive gnawing and biting behavior. The stereotypy was reexpressed up to over 300 days in repeatedly challenged animals and up to at least 180 days in animals receiving only one MS challenge. The development of sensitization was not altered by the mode of drug administration.

Chronic escapable footshock causes a reduced response to morphine in rats as assessed by local cerebral metabolic rates.

The 2-deoxy-D-[14C]glucose (2-DG) method was used to examine the effects of morphine sulfate (MS) on local cerebral metabolic rates for glucose (LCMRglu) in male F-344 rats required to turn a wheel manipulandum in order to escape from nociceptive footshock. This nociceptive stimulus was identical with that utilized in a previous 2-DG study from this laboratory [15] except that animals were exposed to 15 daily 30 min sessions of footshock prior to the 2-DG testing day rather than a single footshock exposure. This allows a direct comparison of the effects of morphine in chronic and acute pain. Unlike the acute footshock study, morphine in chronic footshock rats did not have a significant effect compared with chronic footshock alone in any of the 73 measured brain structures, including limbic and midline thalamic structures previously shown to be important in morphine-induced analgesia during acute pain [15]. Whereas 93% of measured cerebral structures showed decreases in LCMRglu following morphine administration in the acute footshock rats, morphine given to chronic footshock rats caused decreases in only 56% of the structures as compared with chronic footshock plus saline. It is hypothesized that these differential effects of morphine are due in part to a habituation to the chronic stressor such that chronic footshock rats are less stressed than acute footshock rats. Additionally, it is suggested that chronic exposure to pain produces a constant elevation of opioid peptides leading to opioid receptor downregulation and consequently morphine tolerance. These results demonstrate that, even in the presence of the same nociceptive stimulus, morphine can have widely disparate effects on brain metabolism if there are differences in the pain history of the animal.

The effect of MK-801 and SCH23390 on the expression and sensitization of morphine-induced oral stereotypy.

Repeated high doses of morphine sulfate, administered in a 24-36 h period, stimulates the expression of oral stereotypy in rats. Sensitization to this effect of morphine is demonstrated by the reexpression of the stereotypy by the administration of 4.0 mg/kg of morphine one week following the original exposure. To investigate the role of N-methyl-D-aspartic acid (NMDA) and D1 dopamine (DA) receptors in the acute expression and sensitization of morphine-induced oral stereotypy, rats were administered four injections of morphine (10.0 mg/kg) one injection every 12 h and observed for the expression of stereotypic behaviors following pretreatment with selective antagonists. Pretreatment with the NMDA antagonist, MK-801 (0.7 mg/kg), before each of the four morphine injections antagonized both the initial expression of oral stereotypy and the development of sensitization. In contrast, the DA D1 receptor antagonist SCH23390 (40.0 micrograms/kg) administered during the four high-dose treatments with morphine antagonized the initial expression of oral stereotypy and not the development of sensitization. These findings implicate glutamate's action at the NMDA receptor in both the acute expression of morphine-induced oral stereotypy, and the development of sensitization of this morphine effect, whereas DA D1 receptors may only be involved in the acute expression of the stereotypy.