The primary focus of this research is to develop a better understanding of the pharmacological mechanisms underlying the behavioral effects of cocaine that lead to its abuse, and the consequences of that abuse. This better understanding will advance basic knowledge of the pharmacology of cocaine, and drug abuse. In addition, there is a large unmet medical need for cocaine addiction treatments. Recent (1999 to 2002) annual estimates of the number of individuals using cocaine range from 2 to 3.2 million in the United States alone. This research will ultimately lead to the discovery of new treatment modalities for cocaine abuse. Effective treatments for drug abuse will ultimately have a positive public health impact in curtailing drug abuse and the transmission of HIV infection. ? ? Cocaine and many other drugs, including analogues of benztropine (BZT), bind with high affinity to the dopamine transporter (DAT), which is thought to be the substrate through which cocaine acts to produce its behavioral effects that lead to its abuse. However, the BZT analogues are distinct from cocaine in their behavioral activity. Because of these pharmacological differences, and the potential of BZT analogues to serve as medical treatments for cocaine abuse, we have extensively studied compounds from this chemical group.? ? Previous studies of BZT analogues have found them to inhibit dopamine uptake like cocaine, but with less effectiveness than cocaine in producing behavioral effects related to drug abuse. As many of the BZT analogues are also muscarinic antagonists, studies have assessed whether nonselective muscarinic antagonists decrease the effects of cocaine because . Previous studies of interactions of muscarinic antagonists and cocaine effect were conducted with nonselective muscarinic antagonists. Because the BZT analogues show preferential affinity for the M1 muscarinic receptor subtype, we examined interactions of cocaine and the preferential M1 antagonists, telenzepine (TZP) and trihexyphenidyl (TXP). Dose-dependent increases in extracellular levels of dopamine in the NAc shell and core, and the prefrontal cortex, were produced by cocaine, but not by the preferential M1 antagonists, TZP and TZP. When administered with cocaine, however, both M1 antagonists dose-dependently increased the effects of cocaine on dopamine in the NAc shell, and these effects were selective in that they were not obtained in the NAc core or in the prefrontal cortex. Telenzepine also increased locomotor activity, although the effect was small compared to that of cocaine. The locomotor stimulant effects of trihexyphenidyl on the other hand, approached those of cocaine. Telenzepine attenuated whereas trihexyphenidyl enhanced the locomotor stimulant effects of cocaine, with neither drug facilitating cocaine-induced stereotypy. In contrast, both M1 antagonists enhanced the subjective effects of cocaine dose-effect curve. The most important implication of the results is that the muscarinic antagonist effects of BZT analogues do not account for their differences from cocaine, suggesting that these drugs may interact with the DAT in a novel manner.? ? To investigate possible mechanisms for the differences between cocaine and BZT analogues, we compared the subjective effects of different inhibitors with their molecular mode of interaction at the DAT. We determined how different inhibitors affected accessibility of the sulfhydryl-reactive reagent 2-(trimethylammonium) ethyl-methanethiosulfonate to an inserted cysteine (I159C), which is accessible when the extracellular transporter gate is open but inaccessible when it is closed. The data indicated that cocaine analogs bind an open conformation, whereas BZT analogues (and analogues based on the sigma antagonist, rimcazole) bind a closed conformation. Next, we investigated the changes in inhibition potency of 3Hdopamine uptake of the compounds at a mutant DAT (Y335A) characterized by a global change in the conformational equilibrium. We observed a close relationship between the decrease in potencies of inhibitors at this mutant and cocaine-like responding in rats trained to discriminate cocaine from saline injections. Our data suggest that chemically different DAT inhibitors stabilize distinct transporter conformations and that this in turn affects the cocaine-like subjective effects of these compounds in vivo.? ? Cocaine causes biochemical changes in the brain areas involved in learning and memory, including hippocampus and cortex, whose role in drug reinforcement is now being actively investigated. Thus, we studied molecular events in the hippocampus and frontal cortex of rats that exhibited a conditioned place preference (CPP) through treatment with cocaine exclusively in a unique environment. After cocaine conditioning, gene expression in the hippocampi and frontal cortices of conditioned rats, was compared to that in cocaine treated but not conditioned, and saline-treated controls. Our study revealed that 214 transcripts were differentially regulated in the hippocampi of cocaine-paired rats. These include genes that play roles in protein phosphorylation, RNA processing and protein synthesis, ubiquitin-dependent protein degradation and cytoskeleton organization. In contrast, 39 genes were differently expressed in the frontal cortex. Our data support the possibility that molecular changes in the hippocampus might participate in the formation and maintenance of memory patterns induced by cocaine in the brain. Differences in the transcriptional responses in the hippocampus and cortex suggest the primary importance of the hippocampus for recent memory processing associated with cocaine-induced CPP.? ? Drugs of abuse including cocaine, are often taken in combinations, including in combination with drugs that are not ordinarily abused alone. Drugs that act as histamine antagonists are typically safe when administered alone, but can be abused in combination with other drugs of abuse. The pattern of activation of dopamine neurotransmission in the nucleus accumbens of rats produced by H1 histamine antagonists which have behavioral effects like those of psychostimulant drugs was examined. Diphenhydramine and (+)-chlorpheniramine were compared with triprolidine, a potent and selective H1 antagonist and ())-chlorpheniramine which is less active than its enantiomer at H1 receptors. Affinities of the drugs to DA, serotonin, and norepinephrine transporters at H1 receptors and potencies for DA uptake inhibition in striatal synaptosomes were determined to assess mechanisms by which the compounds increased DA levels. Intravenous diphenhydramine (1.03.0 mg/kg) (+)- and ())-chlorpheniramine (1.05.6 mg/kg) but not triprolidine (1.03.0 mg/kg) elicited a cocaine-like pattern of stimulation of DA transmission with larger effects in the NAc shell than core. The absence of stereospecific effects with chlorpheniramine enantiomers along with the lack of an effect with triprolidine suggest that the effects on DA transmission were not related to H1 receptor antagonism. Although in vivo potencies were not directly related to DA transporter affinities, it is hypothesized that actions at that site modulated by other actions, possibly those at the serotonin transporter, are primarily responsible for the neurochemical actions of the drugs on DA neurotransmission and might underlie the occasional misuse of these medications.
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