The experiments reported in this thesis studied the contributions of opioid and NMDA receptors to predictive fear learning, as measured by freezing in the rat. The first series of experiments (Chapter 2) used a within-subject one-trial blocking design to study whether opioid receptors mediate a direct action of predictive error on Pavlovian association formation. Systemic administrations of the opioid receptor antagonist naloxone or intra-vlPAG administrations of the selective μ-opioid receptor antagonist CTAP prior to Stage II training prevented one-trial blocking. These results show for the first time that opioid receptors mediate the direct actions of predictive error on Pavlovian association formation. The second series of experiments (Chapter 3) then studied temporal-difference prediction errors during Pavlovian fear conditioning. In Stage I rats received CSA ?? shock pairings. In Stage II they received CSA/CSB ?? shock pairings that blocked learning to CSB. In Stage III, a serial overlapping compound, CSB → CSA, was followed by shock. The change in intra-trial durations supported fear learning to CSB but reduced fear of CSA, revealing the selective operation of temporal-difference prediction errors. This bi-directional change in responding was prevented by systemic NMDA receptor antagonism prior to Stage III training. In contrast opioid receptor antagonism differentially affected the learning taking place during Stage III, enhancing learning to CSB while impairing the loss of fear to CSA. The final series of experiments (Chapter 4) then examined potential neuroanatomical loci for the systemic effects reported in Chapter 3. It was observed that intra-BLA infusion of ifenprodil, an antagonist of NMDA receptors containing the NR2B subunit, prevented all learning during Stage III, whereas intra-vlPAG infusion of the μ-opioid receptor antagonist CTAP facilitated learning to CSB but impaired learning to CSA. These results are consistent with the suggestion that opioid receptors in the vlPAG provide an important contribution to learning. Importantly, this contribution of the vlPAG is over and above its role in producing the freezing conditioned response. Furthermore, the findings of this thesis identify complementary but dissociable roles for amygdala NMDA receptors and vlPAG μ-opioid receptors in predictive fear learning.

Methadone is currently the most widely used agent to manage opioid dependence, but clinical experience has highlighted some limitations with its use. In particular, a relatively high proportion of patients complain of breakthrough withdrawal symptoms ( non - holding ) at apparently adequate methadone doses. Levo - alpha - acetylmethadol ( LAAM ) is a long acting opioid that is likely to benefit methadone non - holders ; however, relatively little is known about its pharmacology at steady state. The primary aim of this thesis was to evaluate LAAM as an alternative maintenance pharmacotherapy to methadone for the treatment of non - holders ; subsidiary aims were to elucidate the pharmacodynamics and pharmacokinetics of LAAM and its active metabolites ( nor - and dinor - LAAM ), and to examine the in vitro activity of LAAM, nor - and dinor - LAAM. Sixteen methadone maintenance patients ( non - holders = 8 ) were recruited to participate in a randomised, crossover trial of LAAM and methadone. At steady state there were two testing sessions ( 24 h for methadone and 48 h for LAAM ) that featured the concurrent measurement of plasma drug concentrations and both subjective and physiological indices of opioid effect. Cognitive and psychomotor functions were also assessed once during each inter - dosing interval study. Ten age - and gender - matched controls were also tested. The peak magnitude of methadone ' s and LAAM ' s effects were similar. Compared to methadone, LAAM was associated with more stable and less severe withdrawal and mood disturbance. The general pattern of symptom complaints and cognitive function was similar for both drugs. Severity of mood disturbance and withdrawal was similar in holders on methadone and LAAM, but was greater in non - holders when they were taking methadone than LAAM. In comparison to plasma ( R ) - ( - ) methadone, plasma nor - and dinor - LAAM concentrations fluctuated little over the dosing interval. Furthermore, nor - and dinor - LAAM were both more potent in the guinea - pig ileum bioassay, and had greater affinity for mu opioid receptors in receptor binding studies, than LAAM. In conclusion, LAAM converted methadone non - holders into LAAM holders. It is proposed that it is the relatively flat plasma concentration - time profile for nor - and dinor - LAAM that confer stability of opioid effect, minimising withdrawal. Therefore, LAAM may have a role in selected patients, whose response to methadone is suboptimal. / Thesis (Ph.D.)--School of Medical Sciences, 2006.

Graduation date: 1999

Graduation date: 1996

The methadone maintenance treatment population suffers from high rates of comorbid psychiatric and substance use disorders. Despite a more than 40-year treatment history, not all patients are satisfied with methadone treatment and more than half of the patients complain of significant inter-dose withdrawal at least some of the time. The objectives of this research were to investigate the pharmacological response to methadone under the influence of comorbid major depressive disorder and smoking; and to identify factors other than physical withdrawal symptoms that can differentiate patients based on their complaints of dissatisfaction with treatment. In Study 1, seven depressed methadone maintenance patients experienced more opioid withdrawal symptomatology over a 24-hour methadone-dosing interval than 10 nondepressed methadone patients. Depression severity was significantly correlated with trough opioid withdrawal severity. This suggests that depression or depressive symptoms are related to reported opioid withdrawal. In Study 2, many factors other than physical opioid withdrawal symptoms were able to differentiate patients who were satisfied with treatment (holders, n=25), partially satisfied with treatment (partial holders, n=35), and not satisfied with treatment(nonholders, n=30). Results suggested that these patient satisfaction groups cluster differently depending on physical opioid withdrawal, mood, psychological distress, and personality. Nonholders experienced more physical withdrawal symptoms, craving for opioids, and negative drug effects. Holders had less psychological distress and experienced less negative mood states than the other groups. Partial holders had less agreeable personalities compared to patients in the other groups. In Study 3, opioid and nicotine withdrawal symptoms and effects were measured in 40 methadone-maintained patients who were current smokers during trough and peak methadone effects, both pre and post-nicotine administration. Cigarette smoking enhanced opioid withdrawal suppression during the peak methadone condition, methadone attenuated nicotine withdrawal, and methadone and nicotine shared many of the same main effects, suggesting that smoking and methadone effects may be inseparable dimensions. In summary, the results of these studies suggest that in addition to physical symptoms, mood related factors are important to opioid withdrawal perception and that the mood factors and drug interactions can impact on a patient’s perception of satisfaction with methadone treatment.

The methadone maintenance treatment population suffers from high rates of comorbid psychiatric and substance use disorders. Despite a more than 40-year treatment history, not all patients are satisfied with methadone treatment and more than half of the patients complain of significant inter-dose withdrawal at least some of the time. The objectives of this research were to investigate the pharmacological response to methadone under the influence of comorbid major depressive disorder and smoking; and to identify factors other than physical withdrawal symptoms that can differentiate patients based on their complaints of dissatisfaction with treatment. In Study 1, seven depressed methadone maintenance patients experienced more opioid withdrawal symptomatology over a 24-hour methadone-dosing interval than 10 nondepressed methadone patients. Depression severity was significantly correlated with trough opioid withdrawal severity. This suggests that depression or depressive symptoms are related to reported opioid withdrawal. In Study 2, many factors other than physical opioid withdrawal symptoms were able to differentiate patients who were satisfied with treatment (holders, n=25), partially satisfied with treatment (partial holders, n=35), and not satisfied with treatment(nonholders, n=30). Results suggested that these patient satisfaction groups cluster differently depending on physical opioid withdrawal, mood, psychological distress, and personality. Nonholders experienced more physical withdrawal symptoms, craving for opioids, and negative drug effects. Holders had less psychological distress and experienced less negative mood states than the other groups. Partial holders had less agreeable personalities compared to patients in the other groups. In Study 3, opioid and nicotine withdrawal symptoms and effects were measured in 40 methadone-maintained patients who were current smokers during trough and peak methadone effects, both pre and post-nicotine administration. Cigarette smoking enhanced opioid withdrawal suppression during the peak methadone condition, methadone attenuated nicotine withdrawal, and methadone and nicotine shared many of the same main effects, suggesting that smoking and methadone effects may be inseparable dimensions. In summary, the results of these studies suggest that in addition to physical symptoms, mood related factors are important to opioid withdrawal perception and that the mood factors and drug interactions can impact on a patient’s perception of satisfaction with methadone treatment.

Adolescence in humans is a vulnerable period for illicit drug use, and teenage onset of drug use is associated with long-term addiction. Adolescent sensitivity to drug reinforcement, relapse, and withdrawal has not been explored thoroughly in animal models, especially considering opiate drugs such as morphine and heroin. The present series of studies profiles adolescent sensitivity to opiates using adolescent and adult male rats to test for age differences in opiate self-administration, reinstatement, withdrawal signs, locomotor sensitization, and even brain activation during drug-seeking. To test for acute sensitivity to the reinforcing effects of morphine or heroin, we compared patterns of self-administration by adolescent vs. adult male rats on various schedules of reinforcement, drug doses, and daily access conditions. Using fixed ratio schedules and short daily access, adolescents self-administered less morphine than adults, an effect commonly interpreted as higher drug sensitivity. In contrast, escalation of morphine intake under long access conditions was similar across ages, as was heroin intake using fixed or progressive ratio schedules of reinforcement. To test for enduring effects of opiates, we compared opiate-seeking in the absence of the drug in tests of extinction responding and cue-induced reinstatement. Regardless of the acute effects of morphine or heroin, all adolescent treatment groups showed attenuated opiate-seeking compared to adults. Next we considered behavioral correlates of reinforcement, drug withdrawal and locomotor sensitization, during and after escalating doses of experimenter-administered heroin. Consistent with attenuated opiate-seeking, adolescents exhibited attenuated somatic and locomotor signs of withdrawal compared with adults, although locomotor sensitization was similar across ages. Finally, the medial prefrontal cortex (mPFC) is a brain region heavily implicated in drug reinforcement, so we used tissue levels of Fos-like immunoreactivity to compare activation of this region by heroin-seeking. Indeed mPFC activation was absent in rats that self-administered heroin as adolescents, but robust in adults. Together these behavioral and neuroanatomical results surprisingly suggest that adolescent male rats are less sensitive than adults to some acute and enduring effects of opiates, and may predict better response profiles among younger human addicts. Through future studies, adolescent rats may provide a new model to help identify treatments for drug abuse.

Thesis (Ph.D.) in Nursing--University of Maine, 2009. / Includes vita. Includes bibliographical references (leaves 134-152).

(-) Codeine, (-) morphine, and their semi-synthetic derivatives play an integral role in medicinal analgesia. Due to a complex list of undesirable side effects, their effective use is often complicated and troublesome giving cause for the investigation of novel semi-synthetic analogs for efficacy and side-effect profile. It was envisioned that new and interesting codeine analogs could be synthesized via an opening of a hindered 7,8-[alpha]-epoxide. Classically, hindered epoxides are formed via halohydrin formation and subsequent closure. Interestingly, the 7,8-epoxide formed via bromohydrin closure was resistant to reaction with small nucleophiles, such as oxygen and hydride, but reactive towards large and nucleophilic atoms, such as sulfur and bromide. It was discovered that the epoxide was in fact the less hindered 7,8-[Beta] epoxide via x-ray analysis of various compounds. This hinted at an unexpected rearrangement which most likely occurred during the bromohydrin formation due to the severe steric interactions present in the core structure of codeine. Due to the reversibility of bromonium ion formation, a highly hindered double bond can produce the opposite configuration of what is expected when subjected to aqueous brominating conditions. Many popular alkaloids, including codeine and galanthamine, are biosynthetically formed via a spirocyclic dienone intermediate. In nature these intermediates are formed via an enzymatically driven phenolic oxidation; however in the lab this reaction has proven difficult to reproduce. In a previous Magnus publication, (±) codeine and (-) galanthamine, were synthesized via a common spirocyclic cross-conjugated dienone intermediate similar to the intermediate found in nature. Most importantly, this intermediate was formed without a phenolic oxidation. Instead, a para-alkylation of an appropriately substituted phenol efficiently created the key intermediate. Expanding on this phenolic alkylation strategy, various biaryl systems were built in order to investigate the scope and limitations of this reaction. Multiple para- alkylations proved successful while ortho- alkylations unveiled an interesting rearrangement which occurs during the reaction. Lastly, it was determined that a 7-membered ring could not be set using a phenolic alkylation strategy. / text

Neuropathic pain is a prevalent disease with no effective, safe treatments and limited knowledge on the mechanisms involved. One target for neuropathic pain treatment may be the blockade of Dynorphin A (Dyn A). Dyn A is a unique endogenous ligand that possesses well-known neuroinhibitory effects via opioid receptors and neuroexcitatory effects that are mediated through the bradykinin 2 receptors (B2Rs). Extensive SAR was carried out to develop a ligand for the blockade of the excitatory actions of Dyn A at the B2R. A lead ligand was able to block Dyn A-induced hyperalgesia in naïve animals and was effective in a neuropathic pain model. However, the ligand was susceptible to enzymatic degradation. In an effort to increase the stability, modifications of the ligand using non-natural amino acids were performed. Analogues substituted at or near the N-terminus with a D-isomer retained binding at the receptor as well as provided a large increase in stability. These ligands were also found to be non-toxic in a cell toxicity assay. Dyn A has been found to not activate the classical signaling of the B2R, PI hydrolysis or Ca²⁺ mobilization. In an effort to determine Dyn A's signaling, a study was done examining up-regulation of phosphorylated proteins. It was found that Dyn A did not activate; pERK, 7 PKC isoforms or PKA. A well known B2R antagonist, HOE140, was found to have low affinity at rat and guinea pig brain B2Rs but high affinity in the guinea pig ileum. Further examination revealed that this discrepancy in binding may arise from a different isoform of the B2R that has not been previously examined. To date, we have discovered Dyn A analogues that have high affinity for the B2R, are very stable, and have low toxicity. The signaling pathway is still not fully understood, but further studies are underway. Also, there is evidence that the B2R in which the analogues are interacting at may be a different form than what has previously been described. Targeting this different isoform of the B2R with our current stable ligands may provide beneficial therapeutics for the treatment of neuropathic pain without the cardiovascular liabilities.