Wouter Koek - US grants

DESCRIPTION (provided by applicant): Recreational use of gamma-hydroxybutyrate (GHB) is increasing dramatically. The pharmacological mechanisms by which GHB produces its abuse-related effects are poorly understood. GHB abuse is likely related to its subjective effects, and subjective effects of drugs in humans can often be predicted from drug discrimination experiments in animals. The discriminative stimulus effects of GHB are likely to be multidimensional and to involve several different receptor mechanisms. Some of these mechanisms may be unique to GHB (i.e., those involving specific GHB receptors), whereas others may be in common with other compounds (i.e., those involving GABAA and GABAB receptors). The studies proposed here will examine the involvement of these mechanisms in the discriminative stimulus effects of GHB under various conditions. Studies under Specific Aim I examine the ability of positive GABAA modulators and agonists at GABAA, GABAB and GHB receptors to substitute for GHB, and the ability of negative GABAA modulators and antagonists at GABAA, GABAB and GHB receptors to attenuate the discriminative stimulus effects of GHB. These substitution and antagonism studies are conducted under conditions involving different training doses of GHB, because the training dose often affects the pharmacological selectivity of the discrimination. Not only the training dose, but also the alterative training condition can affect the pharmacological selectivity, as shown by drug-drug discrimination studies. Studies under Specific Aim II attempt to increase the pharmacological selectivity of the discriminative stimulus effects of GHB by training to discriminate GHB not only from saline, but also from other drugs that share receptor mechanisms with GHB. It is hypothesized that 1) GABAA, GABAB, and GHB receptors each are involved in the discriminative stimulus of GHB, 2) when a low training dose is used the role of GABAA receptors is more prominent, and when a high training dose is used the discrimination will primarily involve GABAB and GHB receptors, and 3) when a high dose of GHB can be discriminated from a GABAB agonist, its discriminative stimulus effects will involve only GHB receptors. By identifying the role of specific receptors in abuse-related effects of GHB, future studies may be better able to develop specific, pharmacologically targeted therapies for GHB abuse.

[unreadable] DESCRIPTION (provided by applicant): Many people drink, but only some become alcoholic. Genetic factors likely play a role, but environmental factors such as stress may also be involved. Because of the possibility to manipulate both genetic and environmental factors, mice are the species of choice for the experimental study of gene-environment interactions. Alcohol and drug abuse are likely related to subjective effects, and such effects in humans can often be predicted from drug discrimination experiments in animals. Learning to discriminate a drug from its vehicle generally takes animals several months. Recently, a novel procedure was introduced in which mice acquire a drug discrimination in less than two weeks. The present application is aimed at exploiting this potentially useful drug discrimination methodology to examine genetic determinants of sensitivity to alcohol. Studies under Specific Aim I examine the sensitivity of different mouse strains to the discriminative stimulus effects of alcohol, and test the hypothesis that DBA/2J mice are more sensitive than C57BL/6J mice to these effects. For each of the strains, different groups of mice will be trained to discriminate alcohol at different doses, and training dose / discrimination accuracy data will be used to compare the sensitivity of the different strains. Studies under Specific Aim II begin to identify genetic factors that contribute to differences in the discriminative stimulus effects of alcohol, and test the hypothesis that mice with reduced or no expression of the serotonin transporter are less sensitive to the discriminative stimulus effects of alcohol. The approach will 1) exploit a procedure for rapidly assessing abuse-related effects of alcohol in mice, 2) provide new information about genetic determinants of sensitivity to alcohol, and 3) provide a comparative basis for future studies of developmental aspects of alcohol and drug abuse. Initiation of drug abuse occurs primarily during adolescence, and during this developmental period, drugs of abuse may have distinct effects, which could increase risk for abuse. In rodents, adolescence has been defined as spanning from postnatal days 28-42. Being able to train a discrimination within this two-week period is a prerequisite for studying the discriminative stimulus effects of drugs during adolescence in a rodent model. Thus, the rapid drug discrimination procedure in mice may be especially suitable to examine developmental aspects of abuse related drug effects. This future research will increase our understanding of risk factors for alcoholism and drug abuse and will help the development of effective approaches to prevention and treatment. [unreadable] [unreadable] [unreadable]

[unreadable] DESCRIPTION (provided by applicant): The abuse of prescription medications, which is a growing public health concern, has recently increased sharply, particularly in adolescents. Prescription opioid analgesics are now the third most widely abused illicit substances (after marijuana and amphetamine) in this age group. Adolescence appears to be a period of increased vulnerability to the addictive properties of drugs. Also, there is evidence that the effects of exposure to drugs of abuse such as nicotine during adolescence are long-lasting. Little is known, however, about the effects of opioids in adolescence. Most of what we know about opioid abuse and addiction has been learned from heroin addiction in adult individuals, and from research in adult animals. Information about abuse-related effects of opioids in adolescent animals is lacking partly because several of the procedures commonly used to measure these effects in rodents often require at least several weeks of training, whereas periadolescence in rodents lasts only two weeks. One possible approach is to use species with a longer adolescence, such as primates. However, a less expensive and more practical approach would be to modify lengthy procedures for use in adolescent rodents. The present application is aimed at using rapid procedures to study abuse-related effects of morphine in adolescent mice. Studies under Specific Aim I will examine stimulant-, rewarding-, and sedative effects of morphine in adolescent mice by measuring morphine-induced locomotion, conditioned place preference, and motor impairment, and will compare these effects with those observed in adult animals. Studies under Specific Aim II examine the effects of morphine exposure during adolescence on subsequent sensitivity to the stimulant-, rewarding-, and sedative effects of morphine during adolescence, in comparison with the consequences of such exposure in adults. Based on the results, doses of morphine will be selected that are equi-effective to alter subsequent opioid sensitivity in adolescents and adults. These doses with similar short-term effects in adolescents and adults will be used in studies under Specific Aim III to examine the effects of morphine exposure during adolescence on subsequent opioid sensitivity during adulthood, in comparison with effects of such exposure in adults. Together, these studies will answer the following questions: are adolescent animals 1) more sensitive to abuse-related effects of morphine, 2) more susceptible to the sensitizing effects of morphine exposure, and 3) more likely to show long-term consequences of such exposure? The information resulting from the present studies on age- and exposure dependent differences in opioid sensitivity in mice will provide a basis for future studies of gene-environment interactions such as the role of genetic vulnerability to environmental stress in the sensitivity to opioids. This research will increase our understanding of risk factors for abuse of prescription opioids and will help the development of approaches to prevent and treat this important public health issue. PUBLIC HEALTH RELEVANCE: The abuse of prescription opioid analgesics by adolescents is on the rise. This is particularly worrisome because little is known about the effects of opioids in adolescents, and because adolescence appears to be an especially vulnerable period during which drugs may have different, longer-lasting effects than during adulthood. The studies in this application use adolescent animal models to learn more about the risk factors for abuse of prescription opioid analgesics, which will help to develop approaches to prevent and treat this important public health issue. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Depression and related disorders are a major public health problem, compounded by the fact that at least half of patients are not effectively treated by currently available medications. Among the most commonly prescribed is the class of selective serotonin (5-HT) reuptake inhibitors (SSRIs), which act to inhibit 5-HT transporter (SERT) mediated 5-HT uptake. The increase in extracellular 5-HT that follows is thought to be critical for initiation of the cascade of downstream events needed for therapeutic effects. Although SERT is the major player regulating high-affinity 5-HT uptake, there is emerging evidence for an important role of organic cation transporter-3 (OCT3) and possibly the plasma membrane monoamine transporter (PMAT) in taking up 5-HT in brain. This raises the possibility that lack of therapeutic response following SERT blockade could be due to significant 5-HT uptake by OCT3 (and/or PMAT). Our studies using decynium-22 (D-22), a blocker of both OCT3 and PMAT, lend support to this idea. For example, D-22 augments the effect of an SSRI, fluvoxamine, to inhibit 5-HT uptake and to produce antidepressant-like effects in wildtype mice. Moreover, D-22 produces these effects also when given alone in mice that lack, or have reduced SERT expression. Thus, the antidepressant-like effect of D-22 appears to be most pronounced when SERT is either pharmacologically or genetically inactivated. We also found that OCT3 expression (but not PMAT) is increased in mice with a constitutive reduction of SERT, suggesting a compensatory role for OCT3. One important aspect of the proposed studies will be to examine the possibility that this also occurs after chronic treatment with SSRIs, which is known to reduce SERT expression. In addition to 5-HT, OCT3 (and PMAT) can transport norepinephrine (NE) and dopamine (DA), neurotransmitters also linked to the therapeutic action of current antidepressants. Taken together, the goals of the proposed studies are to (1) validate OCT3 (and/or PMAT) as the site where D-22 produces its antidepressant-like effect; (2) determine the relative importance of inhibition of 5-HT, NE and DA uptake in producing the antidepressant-like effect of D-22, and (3) examine the therapeutic potential of D-22 by studying its effect on biogenic amine uptake and antidepressant-like activity after its chronic administration. The results of these studies will help to establish OCT3 (and/or PMAT) as a novel target for the discovery of drugs with improved therapeutic potential, as well as provide a mechanism that can, at least in part, account for poor therapeutic response to current antidepressant drugs. PUBLIC HEALTH RELEVANCE: Although reports vary, it is estimated that major depression is unsuccessfully treated in more than half the patient population, underlining the urgent need to identify new targets for antidepressant medications. OCT3 is emerging as one such target. By validating this target, the experiments proposed here will lay the foundation for the discovery of novel antidepressants with marked therapeutic potential, especially in treatment resistant patients.

ABSTRACT Co-abuse of cocaine and alcohol is one of the most common, and dangerous drug pairings, as evidenced by their concurrent use being a major cause for emergency hospitalization. Thus, this drug combination is not only a serious health threat to the individual user, but a major public health burden. Currently, there are no effective treatments for addiction to cocaine and ethanol, underscoring the vital need to understand the mechanistic basis of this highly addictive drug pairing in order to discover new targets for therapeutic intervention. It is well-known that cocaine and ethanol each increase extracellular levels of dopamine (DA), serotonin (5-HT), and norepinephrine (NE), biogenic amine neurotransmitters that are strongly linked to the rewarding properties of drugs. Cocaine does this by inhibiting the high-affinity, low-capacity transporters for these neurotransmitters, DAT, SERT, and NET, respectively. However, the mechanisms by which ethanol does so are unclear. It is known that ethanol inhibits uptake of DA, 5-HT, and NE, however our published data, together with literature evidence, show this inhibition to be DAT-, SERT-, and NET-independent. Organic cation transporter 3 (OCT3) is a low-affinity, high-capacity transporter for DA, 5-HT, and NE, and is emerging as an important player in regulation of biogenic amine homeostasis. Interestingly, recent reports show that corticosterone, a blocker of OCT3, enhances cocaine-induced DA signaling and potentiates reinstatement of cocaine seeking via an OCT3-dependent mechanism. Moreover, we found that OCT3 expression is increased in mice lacking SERT (-/-), and that ethanol, and corticosterone, both inhibit 5-HT clearance in SERT-/- mice to a much greater extent than in their wild-type counterpart. Together, these findings raise the possibility that ethanol may interact with OCT3 to inhibit uptake of biogenic amines, thereby increasing the addictive properties of cocaine, and propagating the concurrent use of these drugs. To this end, the studies proposed in this exploratory R21 will test the overarching hypothesis that one mechanism by which ethanol increases extracellular DA, 5-HT and NE is by inhibition of their uptake via OCT3, and that this inhibition enhances the increase in biogenic amines produced by cocaine, which blocks their uptake via DAT, SERT and NET. Importantly, we will determine the OCT3-dependency of ethanol?s ability to enhance the rewarding properties of cocaine. We will use pharmacological and genetic approaches, combined with in vivo neurochemistry, and behavioral assays relevant for reward. Regardless of the outcome of these exploratory studies, results will fill fundamental knowledge gaps about the mechanism(s) through which ethanol inhibits uptake of biogenic amines and enhances rewarding effects of cocaine. Results from these studies will improve our understanding of mechanisms that make the abuse potential of concurrent alcohol and cocaine use so high. Importantly, these studies will form an essential platform on which to base larger scale studies probing novel molecular targets, putatively OCT3, for medications to treat abuse of alcohol and cocaine.