Danny Winder - US grants

The goal of this research project is to further our understandingof the neuronal effects of ethanol (EtOH) that contribute toacute intoxication. Ethanol potently inhibits the function of N- methyl-D-aspartate type glutamate receptors (NMDARs). In most neuronal preparations examined to date, this inhibition is selective with respect to other glutamate receptors. Glutamate is the major excitatory neurotransmitter in the mammalian CNS, and activation of NMDARs by this neurotransmitter has been implicated in a number of CNS functions including motor control and information storage. There is a wealth of evidence indicating that EtOH inhibition of NMDAR function contributes to aspects of acute intoxication, including cognitive impairment and sedation. The mechanism of EtOH action on NMDARs is not fully understood. In addition, there is little definitive information in the literature about the molecular properties of NMDARs that confer EtOH sensitivity. The studies proposed in this application will address these issues by testing two hypotheses. The first aim will test the hypothesis that EtOH inhibition of NMDAR function will be altered in neurons isolated from selected brain regions of mice lacking the epsilon1 or epsilon2 subunits or in which the t-terminal region of these subunits has been deleted. This will be tested by examining EtOH inhibition of receptors in whole-cell recordings from CNS neurons acutely isolated or grown in cell culture. Neurons from the neocortex and cerebellar cortex of wild-type and mutant mice will be examined. It is expected that EtOH will more potently inhibit NMDARs in neocortical neurons from wild-type mice relative to epsilon2 knockout and c-terminal truncated animals. The epsilon1 knockout and c-terminal truncated mice should show a loss of developmental changes in EtOH sensitivity of NMDARs in neocortical neurons. Ethanol sensitivity of NMDA receptors is likely to be enhanced in cerebellar granule cells from epsilon1 mutant mice relative to wild-type mice. NMDAR-mediated synaptic transmission in the CA1 region of hippocampal brain slices will also be examined in the wild-type and epsilon1 mutant mice. It is predicted that EtOH will produce greater inhibition of transmission in wild-type than in epsilon1 knockout and c-terminal truncated mice. The second aim will test the hypothesis that key amino acid residues in the pore-loop and third membrane spanning (TMIII) domains of the NMDAR1 subunit confer EtOH sensitivity on the NMDAR. This will be examined by whole cell electrophysiological experiments in HEK 293 cells expressing recombinant receptors containing mutant or wild-type NMDAR1 subunits. Ethanol sensitivity of wild-type and receptors with single-point mutations in the pore-loop and TMIII regions will be determined. Thorough examination of biophysical and pharmacological properties of mutant receptors will be carried out to determine if mutations specifically affect EtOH sensitivity. The proposed experiments will add to ow knowledge of the molecular basis of EtOH effects on NMDARs. It is hoped that the outcome of these experiments will provide a basis for the development of treatments that can counteract some of the damaging neural effects of EtOH.

DESCRIPTION (Provided by applicant): Glutamatergic transmission plays a critical role in physiological and pathophysiological behavior, from learning and memory to drug addiction. A fundamental property of many glutamatergic synapses is that they undergo long-term potentiation (LTP). As a lasting enhancement of transmission in response to transient stimuli, LTP is an attractive candidate cellular mechanism underlying some aspects of learning, memory and drug addiction. Neuromodulatory neurotransmitters play key roles in regulating both these behaviors and glutamatergic synaptic transmission. Indeed, "metaplastic" processes initiated by prior synaptic activity or neuromodulators regulate LTP. However, the mechanisms underlying this "metaplasticity" are unknown. Neuromodulators activate three classes of receptors, those coupled to Gs, Gq and Gi families of heterotrimeric G-proteins. A large body of evidence suggests that Gs-coupled receptors influence a variety of aspects of synaptic transmission, cellular excitability and LTP. This regulation occurs in large part through the recruitment of the cAMP and p42-44 MAP kinase (ERK) signaling cascades in hippocampal neurons. In contrast, relatively little is known of the role of Gi-linked receptors in LTP. However, Gi-linked receptors have been demonstrated to play key roles in learning and memory, as well as drug addiction. Further, like Gs-coupled receptors, Gi-linked receptors can regulate the cAMP and ERK signaling cascades, though in more complex ways. In non-native systems Gi-linked receptors activate ERK and differentially regulate cAMP levels. While it is known that in CAl Gi-linked receptors regulate synaptic transmission and cellular excitability, it is unclear whether native Gi-linked receptors exhibit the complex regulation of cAMP levels and ERK activation seen in the non-native systems. Finally, Gs-coupled receptors are unlikely to be activated independently in vivo, but instead will likely be co-activated with Gi-linked receptors by endogenous ligands. In this application, the roles of Gi-linked receptors in LTP will be addressed using a combination of biochemical, genetic and electrophysiological approaches in mouse hippocampal slices. The coupling of Gi-linked receptors to the cAMP and ERK signaling cascades will be examined in area CAl of mouse hippocampus using a combination of biochemical, electrophysiological and imaging approaches. In parallel, Gi-linked receptor regulation of LTP, particularly those forms that require the activation of these cascades, will also be examined.

Description (provided by applicant): Parkinson's disease (PD) is a neurodegenerative disorder in which a loss of dopamine leads to disordered signaling in the basal ganglia. While therapeutic interventions with dopamine mimetics or precursors are useful in early stages of the disease, they lose effectiveness in later stages. We hypothesize that the loss of I P effectiveness of pharmacotherapies is due to a reduction of dendritic spines on medium spiny neurons in striatum, observed both in PD as well as in animals experimentally depleted of dopamine. Spine decreases in dopamine depletion models are paralleled by increases in the number of "perforated" synapses, and by an, increase in excitatory transmission, suggesting that dopainine depletion may promote dysregulated synaptic plasticity. However, to date little is known of the signal transduction mechanisms through which dopamine regulates these processes. We hypothesize that the increased excitatory transmission initiated by dopamine depletion in the striatum leads to increased PP2B activity. This increased activity is exacerbated by a lack of control of DARPP-3 2 via the DI receptor activation, leading to PP2B and PP 1 mediated suppression of synaptic plasticity and retraction of dendritic spines. We will test this idea directly by taking both genetic and pharmacological approaches to determine the role that PP2B plays in striatal synaptic plasticity, as well as determining to what degree PP2B activity is regulated by-dopamine depletion. Using MPTP injections in mice, we will determine whether genetic inhibition of PP2B in medium spiny neurons decreases dopamine-depletion induced reductions in dendritic spine density.

[unreadable] DESCRIPTION (provided by applicant): Epidemiological evidence suggests that there is an inverse correlation between the age of first exposure to a drug of abuse and the likelihood of developing dependence, suggesting that periadolescence is a time of high vulnerability to drugs of abuse compared with adulthood. One key difference between periadolescents and adults is in their respective responses to stressors. Periadolescence is associated with a period of increased risk-taking, which is accompanied by hyporesponsiveness to many stressors compared to adults. In contrast, periadolescent animals have higher basal levels of circulating corticosterones, and under certain contexts appear more anxious than adults. Stress and anxiety play critical roles in drug abuse, thus an understanding of the basic mechanisms underlying these age-specific differences in anxiety-related behaviors will lead to more specific and age-appropriate therapeutic interventions both for drug abuse and anxiety disorders in general. The ventral noradrenergic bundle (VNAB) emanating from the nucleus of the solitary tract plays a key role in stress/anxiety responses. Lesions of the VNAB or manipulations of noradrenergic transmission within the major target of this projection, the bed nucleus of the stria terminalis (BNST), abolish stress-induced reinstatement of cocaine intake and conditioned place aversion to morphine withdrawal. The BNST is an assembly of sub-nuclei within the extended amygdala with rich interconnections between stress and reward centers. Stimulation of the anterolateral regions of the BNST evokes firing of dopaminergic neurons of the VTA, as well as increases PVN output, increasing peripheral corticosterone levels. Thus altering the level of output of the BNST would be predicted to have a dramatic impact on behavioral responses to drugs of abuse. We have identified two adrenergic receptors that play opposing roles in regulating inputs to the BNST. Here we will test the hypothesis that this regulation is altered in the periadolescent compared to the adult BNST. We will also determine whether psychostimulants, which indirectly target these receptors through altering endogenous catecholamine levels, produce similar effects on transmission. Finally, we will assess the impact of alterations in the relative levels of the receptors on both regulation of transmission and basal anxiety levels by screening recombinant inbred strains of mice (BxDs) with varying levels of receptor expression. [unreadable] [unreadable]

[unreadable] DESCRIPTION (provided by applicant): Alcohol addiction is a complicated disorder thought to involve an intersection of aspects of the individual, the environment, and alcohol itself. Stress/anxiety-related behaviors are an example of an interaction between the environment and the individual that may play a critical role in alcohol addiction. The bed nucleus of the stria terminalis (BNST) is a brain region thought to play a critical role, both in regulating stress and anxiety responses, and in mediating interactions between the stress and reward circuitries. A growing body of evidence suggests that this region may play a key role in aspects of stress-induced alcohol intake. [unreadable] [unreadable] We have found that NMDA receptor dependent synaptic plasticity can be evoked at glutamatergic [unreadable] synapses in this region, and that this plasticity, and NMDA receptor function itself, is regulated by acute [unreadable] ethanol administration. In this competing renewal, we aim to determine the mechanisms underlying the in vitro acute effects of ethanol in this brain region. In addition, we wish to understand what effects more behaviorally relevant chronic administration of ethanol to the whole animal has on BNST function. [unreadable] [unreadable] Finally, evidence suggests that 1) withdrawal from chronic administration of alcohol is associated with an increase in extracellular corticotrophin releasing factor (CRF) levels in the BNST, 2) alterations in CRF signaling alter alcohol drinking behavior, 3) CRF in the BNST is required for stress-induced reinstatement of psychostimulants, and 4) CRF regulates NMDA receptor function, and NMDA receptor-dependent synaptic plasticity in other brain regions. Thus, we will assess the effects of CRF receptor activation on glutamatergic synaptic transmission in the BNST. In total, the successful completion of these studies will advance our understanding of the mechanisms underlying the interactions of alcohol and anxiety centers, and set the stage for a screen for novel pharmaceutical and behavioral approaches to ameliorate stress/anxiety induced alcohol abuse. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Addiction is a tremendous health and financial burden on our society. A large literature indicates that dopaminergic transmission in the brain plays a key role in the reward system that mediates many behavioral responses to drugs of abuse. The ventral tegmental area (VTA) is a critical dopaminergic cell nucleus that is activated by multiple drugs of abuse, and is an integral component of the reward system. In addition though, a more dispersed group of dopamine neurons rostral and caudal to the VTA exists within the periaqueductal grey (PAG) referred to as the A10dc group of neurons. Little is known of the function of these neurons;however, recent studies have indicated that these cells play important roles in pain processing and in reward sensation. Moreover, these dopaminergic neurons have a discrete projection into the forebrain, predominantly into a region of the brain referred to as the extended amygdala. A large literature indicates that 1) the extended amygdala plays an important role in long-term modifications of behavior produced by drugs of abuse, and 2) dopaminergic transmission in the extended amygdala is key to some reward and anxiety responses. The advent of new genetic reporter strategies has allowed for the generation of new mouse reagents that allow investigators to target neurons for study that heretofore could not be isolated for examination. We propose to characterize and utilize two lines of mice already in our mouse colony that express enhanced green fluourescent protein under the direction of either the tyrosine hydroxylase promotor, or as a fusion construct with the endogenous dopamine transporter, to allow the a priori identification of A10dc neurons for electrophysiological analysis. Addiction poses an enormous health and financial burden on our society. Currently, our understanding of the brain circuitries involved in addiction is far from complete. The successful completion of these proposed studies will result in important new information about neurons that may be involved in addiction, potentially creating new targets for therapeutics development.

? DESCRIPTION (provided by applicant): Alcoholism is a tremendous health and financial burden on our society. A growing literature indicates that the extended amygdala plays a key role in stress-reward interactions that may mediate key behavioral responses to chronic alcohol exposure. We demonstrated that chronic intermittent alcohol exposure up regulates CRF receptor dependent augmentation of glutamate release, as well as postsynaptic NMDA receptor function at extended amygdala glutamate synapses. We propose here to test a two-hit model, whereby affective disruptions produced by chronic alcohol exposure and withdrawal depend upon the coordinated pre- and postsynaptic regulation of extended amygdala glutamate synapses. We propose a series of experiments to map out mechanisms by which CRF receptor signaling regulates glutamate synapses, and to explore the alterations in NMDA receptor function that occur with alcohol exposure and withdrawal. We further propose to explore the impact of disruption of these processes on alcohol-abstinence induced depressive behaviors.

DESCRIPTION (provided by applicant): Addiction is a tremendous health and financial burden on our society. A growing literature indicates that norepinephrine in the brain plays a key role in stress-reward interactions that may mediate key behavioral responses to drugs of abuse. A previously unappreciated group of noradrenergic neurons in the field of addiction, cells that project through the ventral noradrenergic bundle (VNAB), are thought to supply the key norepinephrine. The primary target of the VNAB in the brain is a group of nuclei referred to as the extended amygdala. In the previous funding period, we identified actions of each of the major classes of noradrenergic receptors on excitatory synaptic transmission in the bed nucleus of the stria terminalis, a major component of the extended amygdala. Here, we propose experiments to assess molecular mechanisms involved in these actions, and to begin to place these actions in the context of microcircuitry within the extended amygdala. Further, we propose to examine whether overlapping populations of neurons in the extended amygdala are activated by drugs of abuse and stressors, and whether this activation is regulated by noradrenergic ligands. In total, the proposed work will begin to define specific mechanisms likely to play key roles in stress-induced reinstatement of drug seeking behavior, thus providing new potential opportunities for therapeutic development. PUBLIC HEALTH RELEVANCE: Addiction poses an enormous health and financial burden on our society. Currently, our understanding of the brain circuitries involved in addiction is far from complete. The successful completion of these proposed studies will result in important new information about neurons that may be involved in addiction, potentially creating new targets for therapeutics development.

DESCRIPTION (provided by applicant): Depression is a major health problem. Traditional pharmacological treatments have targeted the catecholamine systems with selective reuptake inhibitors, but these have the disadvantages that a) they take extended treatment to achieve efficacy, b) they often require continued dosing, and c) a significant proportion of the clinical population is treatment resistant. The glutamate neurotransmitter system has gained increased attention in the last several years with the advent of the use of the drug ketamine as an antidepressant. Ketamine, which is an antagonist of the NMDA class of glutamate receptors, is advantageous in that it is both rapidly acting and produces persistent effects after a single dose. Animal model studies suggest that ketamine works by producing homeostatic plasticity at glutamate synapses, however the key circuits modified to produce antidepressant actions are not clear. In this proposal we test the hypothesis that NMDA receptors in a region of the brain known as the bed nucleus of the stria terminalis play a key role in the antidepressant actions of ketamine. Moreover, we propose that this occurs via an interaction with the corticotropin receptor factor (CRF) system in this region.

This proposal is for the renewal of a graduate training program at Vanderbilt University that is structured to support the early phases of neuroscience predoctoral education and training. In support of the overall NIH mission, the overarching objective of the program is to provide an exceptional training environment for the next generation of neuroscientists, and is built on the foundation of a strong training faculty with exceptional records of scholarship, research support and graduate mentoring. The heart of this mission is expressed in the academic and research goals of the program, which are to provide our students with a strong didactic foundation in the neurosciences through our core curriculum offerings, and to provide them with the opportunity to carry out state-of- the-art neuroscience research in the laboratories of a group of highly successful and committed mentors. In addition, the program has strong emphases on professional development, diversity, quantitative literacy and rigorous science, with the objective of building the requisite skills needed for success in graduate school and beyond, and of training an inclusive cadre of future independent investigators in neuroscience research. The Neuroscience Graduate Program at Vanderbilt is an interdisciplinary program that encompasses 5 different colleges and schools and 18 departments. Students can enter the program either directly or via three umbrella ?feeder? programs (IGP/MSTP/CPB). Traditional and emerging areas of research strength in the program include: addiction, attention, brain evolution, cell signaling, cognitive neuroscience, circadian rhythms and sleep, CNS drug development, development, developmental disabilities, molecular genetics, neurodegeneration and neurotoxicity, neuroimaging, plasticity, psychiatric illness, sensory and multisensory systems, synaptic transmission, and vision. The program is currently home to 86 trainees and 76 training faculty. The proposal requests an increase in support from 8 to 10 slots and provides the rationale and justification for this request.

Project Summary/Abstract Sleep disorders constitute a major public health problem in the United States, and abnormalities in arousal often occur in concert with other neurological or neuropsychiatric diseases. The locus coeruleus (NE), the major source of norepinephrine (NE) in the brain, promotes arousal, attention, and wakefulness, but the neural substrates that transduce these actions of the LC have not been fully identified. One intriguing candidate is an understudied population of wake-promoting dopamine (DA) neurons in the ventral periaqueductal gray (vPAG). The LC projects to the vPAG, and our preliminary data indicate that NE provides excitatory drive onto DA neurons in this brain region via ?1-adrenergic receptors (?1ARs), suggesting that NE-DA interactions in the vPAG may promote arousal. The goal of this proposal is to delineate this LC-vPAG circuit. In Aim 1, we will use tract tracing and immunohistochemistry at the electron microscopic level to map the connections between the LC and vPAG as well as specific localization of ?1ARs. In Aim 2, we will use ex vivo slice electrophysiology to determine the cellular mechanisms underlying the ability of NE to enhance excitatory drive onto vPAG DA neurons. In Aim 3, we will use DREADD chemogenetics to assess the behavioral consequences of inhibiting or stimulating various nodes of the LC-vPAG arousal circuit. These studies will define a novel arousal circuit and identify candidate neuroanatomical and molecular targets for the treatment of both primary and disease- associated deficits.

Alcohol use disorders (AUDs) are disabling and prevalent conditions that affect almost one-third of Americans. In addition to the devastating effects of AUDs, secondary effects include increased risk for suicide, depression, and substance use disorders. While initial treatments exist, unfortunately, relapse is common, making long- term recovery difficult. Given the high rates of relapse, interventions that seek to prevent relapse have high potential impact. Animal models of addiction have substantially informed our understanding of the stages of addiction? binge/intoxication, withdrawal/negative affect, and preoccupation/anticipation?and their underlying pathophysiology. Chronic alcohol exposure causes neuroadaptive brain changes in an attempt to maintain homeostasis. During early abstinence, hyperactive stress systems result in negative affect, anxiety, and depression which are thought to lead to relapse through negative reinforcement. An emerging theme from animal models is the involvement of the extended amygdala, including the bed nucleus of the stria terminalis (BNST), in withdrawal and reinstatement. While animal models of addiction are heavily studied under the assumption of their utility in translation to humans, the role of the BNST in human alcohol abstinence remains unknown. A major barrier to this work has been technological limitations in neuroimaging of the BNST because of its small size. We have recently overcome this challenge and have characterized the BNST neural circuitry in humans and have developed novel methods to examine BNST function. Our published findings show vast consistencies in BNST circuitry across species, providing a necessary foundation for translational studies. Our pilot studies show that the BNST is specifically engaged in mildly stressful situations where an upcoming threat is unpredictable. To investigate the role of the BNST in human addiction, we will study adults with moderate- severe AUDs who have been abstinent for one month, a critical time for relapse. The study will focus on three specific aims: (1) Determine whether there are intrinsic differences in BNST function and BNST circuit connectivity during early abstinence; (2) Determine whether there are task-based differences in BNST function and BNST circuit connectivity during early abstinence; (3) Determine the relationship between BNST function/connectivity, stress response (cortisol and skin conductance), and anxiety/depression symptoms in early abstinence. Based on findings from animal models, we predict that during early abstinence, the BNST will show hyperactivity and altered connectivity both ?at-rest? (intrinsic) and in response to a mildly stressful task. Furthermore, we predict that variation in stress response will mediate the relationship between BNST function/connectivity and anxiety and mood symptoms. The successful completion of this study will fill a critical knowledge gap, assessing the predictive validity of animal models championed by much of the neurobiology community that have heavily implicated the extended amygdala and BNST in alcohol use disorders. Elucidating these neural mechanisms is a crucial step in identifying novel targets for the prevention and treatment of alcohol use disorders.

Project Summary Addiction is a tremendous health and financial burden on our society. A growing literature indicates that norepinephrine in the brain plays a key role in stress-reward interactions that may mediate key behavioral responses to drugs of abuse. A previously unappreciated group of noradrenergic neurons in the field of addiction, cells that project through the ventral noradrenergic bundle (VNAB), are thought to supply the key norepinephrine. The primary target of the VNAB in the brain is a group of nuclei referred to as the extended amygdala. In the previous funding periods, we identified actions of each of the major classes of noradrenergic receptors on excitatory synaptic transmission in the bed nucleus of the stria terminalis, a major component of the extended amygdala. Moreover, we identified a novel mechanism whereby the noradrenergic system interacts with the corticotropin releasing factor receptor signaling system to drive recruitment of specific populations of VTA projecting neurons. We also identified novel actions of alpha2-adrenergic receptors in regulation of excitatory drive into the BNST. Adrenergic ligands have been identified as potential prophylactic therapeutic candidates in treating addiction. While results in human studies have been encouraging, their overall success in improving outcomes has been modest. We propose that this is in part due to the many disparate actions that the receptors regulated by these ligands regulate, and that if we could develop a more specific understanding of the regulated actions that were key to drug-seeking, we could fine tune treatment strategies to increase effectiveness. Here, we propose experiments to delineate specific pathways through which catecholamine- CRF signaling interactions regulate stress-induced cocaine seeking, and alpha2-adrenergic receptor-induced suppression of stress-induced reinstatement.

Project Summary Addiction is a tremendous health and financial burden on our society. A growing literature indicates that norepinephrine in the brain plays a key role in stress-reward interactions that may mediate key behavioral responses to drugs of abuse. A previously unappreciated group of noradrenergic neurons in the field of addiction, cells that project through the ventral noradrenergic bundle (VNAB), are thought to supply the key norepinephrine. The primary target of the VNAB in the brain is a group of nuclei referred to as the extended amygdala. In the previous funding periods, we identified actions of each of the major classes of noradrenergic receptors on excitatory synaptic transmission in the bed nucleus of the stria terminalis, a major component of the extended amygdala. Moreover, we identified a novel mechanism whereby the noradrenergic system interacts with the corticotropin releasing factor receptor signaling system to drive recruitment of specific populations of VTA projecting neurons. We also identified novel actions of alpha2-adrenergic receptors in regulation of excitatory drive into the BNST. Adrenergic ligands have been identified as potential prophylactic therapeutic candidates in treating addiction. While results in human studies have been encouraging, their overall success in improving outcomes has been modest. We propose that this is in part due to the many disparate actions that the receptors regulated by these ligands regulate, and that if we could develop a more specific understanding of the regulated actions that were key to drug-seeking, we could fine tune treatment strategies to increase effectiveness. Here, we propose experiments to delineate specific pathways through which catecholamine- CRF signaling interactions regulate stress-induced cocaine seeking, and alpha2-adrenergic receptor-induced suppression of stress-induced reinstatement.

? DESCRIPTION (provided by applicant): This project focuses on the role of signal transduction mechanisms in hippocampal neuronal plasticity and memory formation. In the most recent Project Period we have been exploring the role of the TET Dioxygenase family of signal transduction cascades in hippocampal neuronal plasticity and learning, focusing on the mechanisms through which this pathway controls memory-associated gene transcription. We initiated our studies in this area about 10 years ago by determining that epigenetic mechanisms including histone acetylation and DNA Methyltransferase activity are necessary for NMDA receptor-dependent LTP in area CA1. We then transitioned to studies in the behaving animal and discovered that epigenetic mechanisms are activated in the hippocampus with contextual associative conditioning, and that alterations in epigenetic mechanisms are necessary for fear conditioning and for spatial learning. Studies from a wide variety of laboratories have now shown that epigenetic signaling is involved in many forms of synaptic plasticity and learning, in essentially every species that has so far been examined including humans. Moreover, in the last Project Period, we as well as the Tsai lab discovered that TET1 Oxygenase is a controller of active DNA demethylation in the CNS, and that this pathway regulates memory capacity in the behaving animal. Given the clear importance of understanding the roles and regulation of TET Oxygenase-dependent epigenetic mechanisms in neural plasticity and learning, for the next Project Period we propose to continue our investigations into the genomic, epigenomic, and functional targets of TET Oxygenase regulation in the hippocampus. We will pursue the following three Specific Aims: 1: To test the hypothesis that TET1 drives active demethylation at memory-associated genes. 2: To test the prediction that site-specific gene demethylation is sufficient to alter gene expression and regulate memory formation. 3: To test the specific hypothesis that TET1 regulates memory capacity via controlling homeostatic synaptic plasticity and cell-wide excitability in hippocampal pyramidal neurons. By focusing on these important new targets for the TET Oxygenase pathway in hippocampus, we will continue to formulate a comprehensive model of epigenetic regulation of transcription-dependent molecular mechanisms in neuronal plasticity and memory.