Methylphenidate (MPH) is the most commonly-prescribed medication for treating ADHD. Despite high prescription rates among kindergarten-aged children, MPH was not approved for use in children younger than nine, and research into its long-term consequences is lacking. Here, we examined the effects of early-life MPH exposure on anxiety-like behaviors in adulthood in normal rats and rats with dysfunctional central dopamine. On postnatal day (PD) 3, male and female rat pups were injected intracisternally with 6-OHDA or vehicle to generate normal and dopamine-deficient groups. In an initial pair of experiments, 6-OHDA (50, 100 and 150 µg/10µL infusion) was assessed for its ability to induce an ADHD-like phenotype. Subsequently, rats were lesioned with 6-OHDA (100 µg/infusion) or vehicle on PD3 and given MPH (0, 0.5, 2 or 5 mg/kg, i.p.) once daily for 10 days, starting on PD11. On PD60, anxiety-like behavior was assessed with light/dark box or social interaction tests. On PD65, all rats were tested on the elevated plus maze (EPM). Rats with neonatal 6-OHDA lesions exhibited anxiety-like behavior in the light/dark box test and on the EPM. However, there was a complex interaction between sex, lesion, and drug dose in the social interaction test. Pretreatment with 2 mg/kg MPH increased investigatory behaviors in non-lesioned females and decreased investigatory behaviors in lesioned females, suggesting that the long-term effects of early-life MPH in females depend on normal dopamine levels. Together, these experiments support the efficacy of preclinical ADHD models and diverse measures of anxiety-like behaviors when studying the effects of early-life MPH exposure.

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tDCS brain stimulation does not always work in the intended direction. It has been found to sometimes worsen behavior rather than improve it. A preliminary study shows that people high on sensation-seeking and lack of premeditation were prone to reverse effects of tDCS on performance on a Stop Signal Task. Both of these constructs are related to dopamine levels. Study 2 seeks to intentionally cause a reverse effect of tDCS by increasing participants’ dopamine levels via caffeine. There was not a significant interaction between tDCS and caffeine on errors on the Stop Signal Task in this study. However, other factors interacted with tDCS and caffeine including lack of premeditation. This two study package suggests the effects of tDCS are variable across individuals, with personality and neurochemistry both affecting behavioral outcomes of tDCS.

The neural mechanisms mediating one-trial and multi-trial behavioral sensitization during early ontogeny are poorly understood. The purpose of this thesis was to assess the importance of D2-like receptors for the induction of cocaine- and methamphetamine-induced one-trial and multi-trial behavioral sensitization during the middle and late preweanling period. In a series of four experiments, rats were injected with saline or the selective dopamine D2-like receptor antagonist raclopride 15 min prior to treatment with the indirect dopamine agonists cocaine or methamphetamine. Acute control groups received two injections of saline. The pretreatment regimens occurred on either PND 16 or PND 20 (one-trial behavioral sensitization) or PND 13-16 or PND 17-20 (multi-trial behavioral sensitization). On PND 17 or PND 21, rats were challenged with either cocaine or methamphetamine and sensitized responding was assessed. With only a single exception, both one -trial and multi-trial cocaine- and methamphetamine-induced sensitization was evident on PND 17 and PND 21. Importantly, the D2-like receptor antagonist raclopride did not prevent the induction of cocaine- or methamphetamine-induced one-trial behavioral sensitization. In regards to multi-trial behavioral sensitization, raclopride failed to inhibit cocaine -induced sensitized responding on PND 17 and PND 21. Interestingly, higher doses of raclopride (0.5 and 1 mg/kg) were able to prevent the induction of multi-trial methamphetamine-induced sensitization on PND 17. Therefore, D2-like receptor antagonism differentially affected methamphetamine -induced behavioral sensitization depending on whether a one-trial or multi-trial paradigm was employed. When considered together, these results suggest that the neural mechanisms underlying the methamphetamine -induced behavioral sensitization of preweanling rats differs depending on the type of experimental paradigm (one- vs multi-trial) being used. Other potential explanations (i.e., nonspecific antagonist effects, impact of contextual conditioning, etc.) for this interesting effect are presented in the Discussion.

Cathinones are psychostimulant compounds heavily implicated as drugs of abuse. They exert their physiological actions at the monoamine transporters, which are responsible for maintaining synaptic neurotransmitter homeostasis. Monoamine transporters produce currents during transport and have been shown to depolarize cell membranes and activate voltage-gated calcium channels in mammalian expression systems. This phenomenon is harnessed in an assay which measures these induced calcium transients, allowing for quantification of pharmacodynamic effects of compounds at monoamine transporters. It is unknown if this electrical coupling occurs in neurons, but the implications if it does are significant. In the current work, fluorescent resonance energy transfer studies of HEK cells expressing hDAT suggest that a subpopulation of monoamine transporters and calcium channels may be interacting directly. Additionally, this work presents calcium assay data comparing several novel methcathinone analogs. Of the compounds tested, a single α-methyl substituent at the α-carbon yields the greatest potency at hDAT. The implications of these results shed light on future psychostimulant studies and further define the physiological relationship of the components of a system used to study these compounds.

Reward has been shown to improve multiple forms of learning. However, many of these studies do not distinguish whether reward directly benefits learning or if learning is boosted by modulation of top-down factors such as attention and motivation. The work outlined in this dissertation explores the modulatory effects of reward and punishment without directly manipulating top-down factors such as attention or motivation. We achieved this goal by studying the effects of reward and punishment on incidental learning – a branch of procedural learning where learning occurs without intention and through repetition. Our results reveal that reward is able to bolster incidental learning during the performance and learning of an associative task, even when awareness of how to achieve the reward is minimized (Experiments 1 and 2). However, a similar benefit was not observed in an analogous set of experiments examining the effect of punishment on incidental learning (Experiments 3 and 4). A direct comparison between the effect of reward and punishment on incidental learning revealed a significant advantage for rewarded combinations over punishment. However, this advantage was only observed when high cognitive (associative) demands were emphasized (Experiment 6), as opposed to high motor demands (Experiment 5). Finally, we explored the role of dopamine in the effect of reward on incidental learning. Because dopamine neuron dynamics have been implicated in both reward processing and in various forms of learning, we hypothesized that patients with Parkinson's disease (PD), who experience an accelerated rate of death of dopamine neurons, would experience impaired learning from rewards compared to healthy older adults. Experiment 7 revealed a significant impairment in reward-related incidental learning for patients with Parkinson's disease relative to comparisons. The amount of levodopa medication taken by PD patients predicted the effect of reward, demonstrating a potential link between dopamine levels and the effect of reward on incidental learning. Together, this dissertation demonstrates that 1) reward improves incidental learning, 2) reward may be an exceptional form of feedback, as opposed to punishments, and 3) dopamine levels may potentially drive the effect of reward on incidental learning

Steroid hormones are known to have significant effects on a wide variety of biological processes. In particular, they serve as critical modulators of neural function and behavior and play critical roles in stress responses and neurologic disorders. Until recently the biological actions of steroid hormones were believed to operate primarily through activation of cognate nuclear hormone receptors or the allosteric modulation of ion channels (Majewskaet al., 1986). However, new signaling pathways involving G-protein coupled receptors (GPCRs) for steroid hormones have been recently identified in multiple different species, implicating steroid hormones in direct fast modulation of intracellular signaling and in turn behavior (Thomas et al., 2006, Gabor et al., 2015). In mammals G protein-coupled estrogen receptor 1 (GPER), also known as G protein-coupled receptor 30 (GPR30), is expressed throughout the body including in the nervous system and has been suggested to play a variety of roles in health and behavior (Prossnitz and Barton, 2011). Despite recent progress in this area from studies using rodent models, the mechanisms underlying "non-genomic” actions of steroids remain largely elusive. This gap in our understanding presents a significant scientific and clinical challenge to a comprehensive view of the role of steroid hormones in regulating both neural function, behavior and overall health of the organism. To understand the mechanisms for this unconventional steroid signaling we sought to use a simpler system to explore the functions of GPCR’s for steroid hormones. In 2005, Peter Evans’s group identified DopEcR, a unique GPCR in Drosophila melanogaster, which responds to ecdysone—the major steroid hormone in insects (Srivastava et al. 2005). This unconventional GPCR for steroid hormones is particularly interesting because it is a dual receptor that also responds to a structurally dissimilar compound, dopamine. DopEcR is preferentially expressed in the nervous system and has recently been implicated in modulating multiple behaviors including starvation-induced enhancement of sugar sensitivity (Inagaki et al., 2012), experience-dependent courtship suppression, habituation of the giant fiber pathway (Ishimoto et al., 2013) and ethanol-induced sedation (Petruccelli et al. 2016) in flies. DopEcR also plays a role in perception of sex pheromones in moths (Abrieux et al., 2013). More recently the mammalian GPCR for estrogen GPER has also been found to bind dopamine indicating that this unique attribute may be more prevalent among these novel GPCRs for steroids (Evans et al. 2013). Despite these previous findings, we still know little about how GPCRs for steroids modulate neurons at the cellular level and how they modulate behaviors. Therefore we sought to forge a more comprehensive understanding of the function of steroid signaling by characterizing DopEcR function in neuronal and behavioral modulation through GPCR’s. To characterize DopEcR’s function we looked at the consequences of DopEcR signaling at three levels: behavior, neuronal morphology and finally physiology. Because changes steroid hormones levels are often associated with environmental stressors we assayed the role of DopEcR in a stress related behavior: starvation-induced sleep suppression and hyperactivity. To look at DopEcR’s role in neuronal physiology we used bioluminescent calcium imaging to measure its effect on the stimulated calcium response in a brain structure critical for behavior. Finally we used principal clock neurons in the brain (PDF+ l-LNv neurons) as a model to examine DopEcR’s role in modulating plasticity and neuronal structure. In our present work described in Chapter 2, we found that the D1-like receptor, DopR1, modulates sleep and activity independent of starvation while DopEcR plays a role in mediating starvation-induced sleep suppression and enhanced activity. We found that knocking down EGFR in a DopEcR mutant background restored starvation induced changes in behavior, suggesting that DopEcR normally suppresses EGFR signaling to suppress sleep under starvation. In Chapter 4, we show that the nicotine-induced Ca2+-response was selectively enhanced in the medial lobes either in DopEcR mutant or in flies with DopEcR selectively knocked down within the MBs. Using a pharmacological approach, we show that the endogenous ligands of DopEcR mediated two different responses in the MBs: the steroid ligand ecdysone enhances activity in the calyx and cell body region, whereas monoaminergic ligand dopamine reduced activity in the medial lobes. In Chapter 5, we find that reducing DopEcR in PDF neurons results in reduced basal levels of bouton numbers. The reduction in bouton number is independent of cAMP signaling but instead relies on inhibition of EGFR signaling. Signifying that DopEcR may modulate EGFR associated signaling to make changes in the in the brain. These results demonstrate that DopEcR is able to modulate neuronal excitability, physical structure of neurons and the behavior of the organism. Interestingly it also indicates that DopEcR’s different ligands, dopamine and ecdysone, may have unique and spatially distinct effects on different brain structures or within the same structure. Overall, this study provides a solid foundation for understanding the roles and action mechanisms of GPCR-mediated steroid signaling in regulation of neural development, physiology and behavior.

Resurgence refers to the reappearance of an extinguished operant behavior when reinforcement for an alternative behavior is also discontinued. It is especially relevant to the reappearance of problem behavior because many behavioral interventions discontinue reinforcement for aberrant behavior while simultaneously reinforcing an appropriate response. Existing information about the neuropharmacology of resurgence is scarce, but suggests overlap between drug seeking observed in the resurgence model and drug seeking observed in the more widely studied reinstatement and renewal models. The aim of this dissertation was to explore additional neural systems relevant to reinstatement and renewal preparations within a resurgence paradigm to assess further overlap. The neuropharmacology of resurgence was examined in two studies via administration of two drugs that have proven effective in blocking drug seeking in reinstatement and renewal preparations. In two experiments, rats earned food pellets for pressing a target lever in Phase I. In Phase II, lever pressing no longer produced food, but food was delivered contingent on an alterative nose poke response. Finally in Phase III, neither response produced food deliveries. Prior to these Phase III sessions, separate groups of rats were injected with 0, 50, or 100 mcg/kg of the dopamine D-2 receptor antagonist raclopride in Experiment 1 or 0, 20, or 40 mcg/kg of alpha-2 agonist clonidine in Experiment 2. Both doses of raclopride were effective in blocking resurgence, but there was strong evidence that the higher dose did so via motor rather than motivational impairment. Furthermore, the lower dose significantly suppressed the alternative nose poke, which suggests motor impairment, as well. Only the higher dose of clonidine blocked resurgence, but did so with no evidence of motor impairment. Raclopride significantly impacted extinction of the alternative poke at both doses tested, whereas clonidine had no effect at either dose. The results of the present studies provide additional information about the neuropharmacology of resurgence, as well as additional evidence of overlap between resurgence, reinstatement, and renewal. The present results may also have implications regarding underlying neural mechanisms and for pharmacotherapies to attenuate relapse when alternative sources of reinforcement are thinned or discontinued.

The mesolimbic dopamine (DA) system has been implicated in providing the basis of pleasure, guiding the general mechanism of reinforcement as well as motivation. Support for these roles have grown from neurochemical research in the field of addiction. It is now well known that DA activity increases in the nucleus accumbens septi (NAcc) with exposure to addictive substances. Moreover, pharmacological manipulation of this system produces predictable changes in the administration of drugs of abuse, as well as natural reinforcers. This system is responsive to natural reinforcers and addiction may be the transference of routine mesolimbic function to environmental stimuli predictive of drug administration. The role of the NAcc in addiction specifically appears to be the facilitation of attention to drug-paired stimuli and addiction may be the behavioral manifestation of conditioned NAcc DA reactivity to the presence of drug-related stimuli. Although these findings have been reported in adults, few studies have focused on adolescence, the time when drug use/abuse begins. Adolescents may be particularly susceptible to addiction when considered in the light of this hypothesis. Recent research has revealed that the mesolimbic system of periadolescent animals is undergoing dramatic transition in functional tone. DA receptor and transporter levels are up regulated, synthesis rates are altered, and innervation from prefrontal cortex (PFC), involved in regulating tonic and phasic DA activity, is increasing. Consequently, during adolescence there is a dramatic change in tonic DA levels, variations in phasic responses to acute drug administration and alterations in how the system adapts to repeated drug exposure. The present study utilizes the procedures of conditioned place preference, Novelty preference and in vivo microdialysis to determine how this conditioning process changes during the period of adolescence. The results indicate that adolescents are different from adults not only on behavioral measures associated with drug abuse, but in their neurochemical responsiveness to alcohol, and that these differences are related to a general developmental aspect of adolescence that renders them susceptible to addiction.

No description available.