Edda Thiels - US grants

DESCRIPTION (Investigator's abstract): Knowledge of the mechanisms that underlie activity-dependent neural plasticity is integral to understanding both normal and impaired memory function. The overall goal of the experiments proposed in this grant application is to study parameters critical for the induction and biochemical processes that underlie the maintenance and expression of long-term depression (LTD) of synaptic strength of glutamatergic synapses in the adult hippocampus in vivo. Research with formal models of learning and memory has shown that synaptic strength must have the capacity to both decrease and increase in a use-dependent manner for successful simulation of these cognitive processes. A common paradigm for inducing LTD is the delivery of prolonged low-frequency stimulation to the afferent pathway. However, LTD induced in such a fashion has been found to occur only in in vitro preparations using tissue from young animals. We recently have demonstrated that repeated paired-pulse stimulation of the commissural pathway reliably induces robust LTD of the commissural input to CA1 pyramidal cells in the adult hippocampus in vivo. Subsequent experiments have revealed that the induction of LTD by paired-pulse stimulation is dependent on N-methyl-D-aspartate (NMDA) receptor activation and, temporally overlapping, inhibitory input to the postsynaptic cell target mediated by activation of gamma-amino-butyric acid A (GABAA) receptors. If GABAergic inhibition is weak or absent during excitatory activation, then LTD fails to develop. The first aim of the present proposal is to test whether or not the degree of GABAergic inhibition during paired-pulse stimulation controls the effectiveness of a train of paired pulses to induce LTD in the adult hippocampus in vivo. The mechanisms that underlie the maintenance and expression of LTD induced by paired-pulse stimulation currently are unknown. Differential activation of protein kinases and phosphatases commonly is thought to play a critical role in regulating bidirectional activity-dependent synaptic plasticity. The second and the third aim of the present proposal therefore is to examine in the adult hippocampus in vivo changes in protein phosphatase and protein kinase activity, respectively, in association with LTD induced by paired-pulse stimulation. The proposed experiments involve a rare combination of electrophysiological, pharmacological, and biochemical techniques to gain insight into the mechanisms that underlie activity-dependent neural plasticity in the intact adult brain.

DESCRIPTION (provided by applicant): Stimuli that have been paired with reward can potentiate behavior aimed at obtaining the reward. This potentiating effect of Pavlovian conditioned cues on reward seeking, termed Pavlovian-instrumental transfer, generally is adaptive; however, it can become maladaptive when the reward is pursued in excess or induces damage, as is the case in drug addiction and obesity. The goal of the present proposal is to shed light on the neurobiological mechanisms underlying Pavlovian-instrumental transfer. We recently found that exposure to a reward-paired cue induces a transient increase in the activation of extracellular signal-regulated kinase (ERK) in the nucleus accumbens (NAc) of adult rats, and that this activation is required for the ability of the conditioned cue to potentiate reward seeking (Shiflett et al., 2008). These observations suggest an important role for NAc ERK in the mediation of the incentive-motivational properties of conditioned cues. The goal of the present proposal is to determine the receptors and cell types within the NAc involved in the cue-driven ERK activation (Specific Aim 1), the output targets of the NAc impacted by the cue-driven ERK activation (Specific Aim 2), and ERK-mediated intracellular events through which the cue-driven activation of this enzyme may facilitate reward seeking (Specific Aim 3). For each of these aims, our working hypotheses are as follows. The CS-driven ERK activation results from activation of D1 dopamine and NMDA glutamate receptors on NAc cells. The cell type that expresses CS-driven ERK activation are D1 receptor-expressing projection neurons that are part of direct, disinhibitory output pathways from the core and the shell of the NAc. CS-driven ERK activation within these projection neurons transiently raises their excitability, thereby facilitating output from these neurons onto their targets. The disinhibitory effect, in turn, promotes enhanced reward seeking. We will test these hypotheses using a powerful and innovative combination of behavioral, pharmacological, peroxidase- and double-flouresence immunohistochemical, retrograde tracing, and molecular techniques. The findings from the proposed work will advance our understanding of a fundamental aspect of behavior control by reward- predictive cues. The findings also will have implications for our understanding of the neural processes that contribute to costly behavioral aberrations, such as addiction and obesity, and the development of strategies for the treatment of these behavioral aberrations. PUBLIC HEALTH RELEVANCE: The proposed studies will uncover the neurobiological mechanisms through which conditioned cues potentiate reward-seeking behavior. The findings will advance our understanding of a basic kind of behavior control by environmental stimuli and have important implications for the development of strategies to treat drug addiction and obesity.

DESCRIPTION (provided by applicant): The dopamine transporter (DAT) plays a crucial role in the regulation of brain dopamine (DA) homeostasis. Through re-uptake of DA, DAT serves two important functions: the termination of synaptic transmission at dopaminergic terminals, and the replenishment of vesicular DA pools. In addition to uptake or direct transport, DAT can also function to release DA. This process, which is referred to as reverse transport or efflux, is the mechanism used by potent and highly addictive psychostimulants, such as amphetamine and its analogues, to increase extracellular DA levels in motivational and reward areas of the brain. It has long being recognized that DA neurons release DA through exocytotic and non-exocytotic processes. However, the exact mechanism by which physiological signals or psychostimulants, such as amphetamine, induce DA release through DAT still remains a complex and not completely understood area of research. Thus, examining the basic mechanism(s) that affect reverse transport through DAT is critical for both understanding fundamental aspects of DA regulation and clinical intervention in DA-related brain disorders associated with the therapeutic use and abuse of psychostimulants. The long-term goal of our research program is to identify and characterize signaling mechanisms that control DA release through DAT, and elucidate the molecular actions of psychostimulants. This application is based on our recent discovery that the beta upsilon subunit of G proteins (Gbetagamma) binds DAT and regulates transporter activity. This effect was demonstrated in cultured cells, brain synaptosomes, and in vivo. More importantly, activation of Gbetagamma promotes DAT-mediated DA efflux, whereas inhibition of Gbetagamma attenuates amphetamine-elicited DA efflux in cultured cells. Finally, activation of Gbetagamma enhances whereas inhibition of Gbetagamma reduces amphetamine-evoked locomotor activity in vivo. Based on these preliminary data, the central hypothesis of this proposal is that the interaction between DAT and beta upsilon subunits promotes DA release through DAT and is involved in the actions of amphetamine. In this proposal we will i) identify the Gbetagamma interaction site(s) in DAT and their role in transporter regulation, ii) test the hypothesis that Gbetagamma is involved in DAT-mediated DA efflux, and iii) test the hypothesis that Gbetagamma is involved in amphetamine's actions in vivo. The successful completion of the studies proposed here will provide a detailed characterization of the DAT-Gbetagamma interaction and a clear understanding of its contribution to DAT reverse transport. The fact that amphetamine induces DAT reversal suggests that DA can also be released through DAT under physiological conditions. Therefore, our proposed studies will define not only the role that Gbetagamma subunits play in the addictive properties of amphetamine, but also the contribution of Gbetagamma subunits to DA homeostasis as we grow our current understanding of the molecular details underlying physiological DAT reverse transport. The long-term goal of our research program is to identify novel therapeutic targets that can be used in the treatment of neuropsychiatric disorders, including drug addiction.