funding and awards
Molecular Neurobiology of Human Drug Abuse [R01DA015446]
Opiate addiction is now a national epidemic marked by an increased incidence of abuse and overdose of heroin and elevated misuse of prescription opioids which also bears a high incidence of overdose death. Despite this growing opiate problem there remains a lack of knowledge about the molecular neuropathology of this human disorder. A fundamental core of our reverse translational research efforts has thus been to fill critical gaps of knowledge by direct investigation of the brains of human heroin abusers. Such efforts have identified the transcription factor ets-like kinase 1 (ELK1) as a central candidate in human heroin abusers. ELK1 disturbance was downstream of the mu opioid receptor (the pharmacological target of heroin metabolites and prescription opiates) and mitogen-activated protein kinase (MAPK) pathway and it was found by in silico analysis of microarray data to target the promoters of a large percentage of down-regulated genes in the nucleus accumbens (NAc) of heroin abusers. Surprisingly, very limited knowledge exist about ELK1 in relation to drug abuse, but this transcription factor has been implicated in other fields in cellular differentiation and synaptic plasticity that are highly relevant to the pathophysiology of addiction disorders. We hypothesize that heroin abuse leads to dysregulated ELK1-mediated transcriptional activity of target genes involved in the reorganization of striatal synapses and that modulate drug-seeking behaviors. We propose: (1) to determine ELK1-mediated transcriptional regulation in neurons within striatal and mesocorticolimbic brain regions of human heroin abusers by conducting ELK1 ChIP in combination with high throughput sequencing (ChIP-seq) on the nucleus accumbens, dorsal striatum and medial orbitofrontal cortex of heroin abusers. This data will also be integrated with transcriptome analysis to determine the relationship to gene expression. (2) To identify the epigenetic landscape across the ELK1 gene that contributes to its dysregulation in heroin abusers. (3) To investigate the causal role of ELK1 in structural plasticity and heroin seeking behavior by use of animal models. The multidisciplinary approach will expand significant insights about novel targets for treatment interventions and the dataset accrued by this body of work will be a unique and valuable resource to the field given the current lack of such human brain data.
Cannabidol as treatment intervention for opiate relapse
Opioid abuse is a significant global public health problem. Of the over million opiate-dependent subjects today, only less than a quarter of such individuals receive treatment. Using a strategy of indirectly regulating neural systems to modulate opioid-related behavior, our preclinical rodent studies demonstrated that cannabidiol (CBD), a nonpsychoactive component of cannabis, specifically inhibits cue-induced heroin-seeking behavior. The fact that drug craving is generally triggered by exposure to conditioned cues suggests that CBD might be an effective treatment for heroin craving, specially given its protracted impact on behavior. It is the goal of this project to (1) determine the safety and basic pharmacokinetic characteristics of cannabidol when administered concomitantly with opiate in humans and (2) characterize the acute and short-term effects of cannabidol administration on cue-induced craving in drug-abstinent heroin-dependent subjects using a random double blind design.
Molecular Neurobiology of Drug Addiction/Project 4 [P01DA008227]
The objective of this project is to elucidate transcriptional and epigenetic mechanisms underlying glutamatergic dysregulation in mesocorticolimbic circuits in heroin abuse, a chronic relapsing disorder. Aberrant glutamatergic transmission in the nucleus accumbens (NAc) and medial prefrontal cortex (mPFC) is central to drug-seeking behavior and addiction vulnerability. Our research focuses on studies of the human brain for which limited molecular insights are available. Preliminary data from microarray analysis already revealed distinct impairments of NAc glutamate-related gene expression in heroin abusers. The majority of these genes are upregulated, indicating that transcriptional change may be predominantly activation. Bioinformatic prediction of potential transcriptional regulators of these gene alterations identified DNA-binding factors SP1 and CREB1 as most likely regulators. SP1 and CREB1 are known to interact with various molecules involved in histone modification, such as histone acetyltransferases (e.g., Ncoa1, Ncoa3) that were also affected in the heroin population, which is mechanistically consistent with the observed predominance of upregulated glutamatergic genes. We will validate these findings using a comprehensive view of the genome by RNA-seq next-generation sequencing (Program Core) in the NAc of another subset of heroin abusers and control subjects, and will also examine the transcriptome in the mPFC, which provides glutamatergic innervation to the NAc. Specific glutamatergic disturbances will be assessed within discrete striatal output pathways using laser capture microdissection. To interrogate the molecular mechanisms by which glutamatergic aberrations are maintained, we will perform parallel studies in the rat heroin self-administration model in which viral-mediated gene manipulation strategies will be used to evaluate causal relationship of gene targets to heroin-seeking and intake. ChIP-Seq will be conducted (Program Core) to map genome-wide binding of a select number of target transcription factors, chromatin modifiers and histone marks in the NAc and mPFC. Overall, the novel research in this Project, which is fully integrative with the Core and other Projects of the PPG, will advance molecular knowledge about the pathophysiology of addiction disorders.
Neurodevelopmental Effects of Cannabis and its Epigenetic Regulation [R01DA030359]
Emerging lines of evidence suggest that interactions between genes and the environment play a critical role in individual vulnerability to psychiatric disorders including drug addiction. The epigenome is influenced by environment and thus is a highly relevant biological candidate to maintain persistent aberrant neuronal processing as a result of developmental drug exposure. The developing brain may be particularly sensitive to epigenetic influences, given the dynamic neuroplasticity characteristic of this period. Marijuana (Cannabis sativa) is the illicit drug most commonly used by pregnant women and teenagers. Our studies of human fetuses with maternal cannabis use revealed selective alterations of striatal preproenkephalin (PENK) and dopamine receptor D2 gene expression (predominantly enriched in striatopallidal neurons), but not prodynorphin or dopamine D1 receptors (enriched in striatonigral neurons). Similar gene expression patterns were detected in the ventral striatum of rats exposed to -tetrahydrocannabinol (THC), the psychoactive component of cannabis, prenatally as well as during adolescence. Importantly, PENK and D2 gene expression impairments persisted into adulthood following either prenatal or adolescent THC exposure and the animals exhibited increased heroin self-administration and inhibitory control deficit, phenotypes predictive of drug addiction vulnerability. Epigenetic modifications are capable of maintaining modified gene expression states that can persist throughout development and it is an intriguing possibility that such mechanisms would underlie the long-term effects of cannabis exposure. In this project, we propose to study chromatin modification at specific regulatory regions of the PENK and D2 genes in the ventral striatum of adult rats with developmental THC exposure. The fact that PENK and D2 genes are preferentially expressed in subpopulations of stratial neurons that constitutes the striatopallidal pathway suggests a potential THC-sensitivity of genes aligned to this circuit. We set out to discover novel striatopallidal genes that are affected by developmental THC exposure and explore the epigenetic mechanisms that regulate their expression. Furthermore, we will investigate the causal relationship between THC-sensitive striatopallidal genes and addiction-related behaviors by use of gene manipulation in the ventral striatum. Overall, these studies will provide important mechanistic insights into the long-term impact of developmental THC exposure that would enable the development of targeted treatment interventions for addiction vulnerability and other related psychiatric disorders associated with developmental insults.
Multigenerational Epigenetic Effects of Cannabis Exposure [R01DA03360]
Marijuana (Cannabis sativa) is the most commonly abused illicit drug by teens and young adults of childbearing age in the USA with significant social and public health implications. While research efforts have begun to characterize the behavioral and neurobiological consequences of direct repeated exposure to cannabis, the possible impact across multiple generations is not known. Preliminary data obtained in our lab suggests that adult rats from parents with adolescent exposure to -tetrahydrocannabinol (THC; primary psychoactive component of cannabis) exhibit reduced motivation and a depression-like phenotype, as well as anxiety-related behaviors that develop in response to their tendency for increased heroin intake. In addition, both male and female F1 offspring show decreased striatal expression of genes that encode several components of the dopamine, opioid and glutamate neurotransmitter systems. Female F1 offspring tended to have more significant disturbances, emphasizing the importance of gender that could potentially be relevant to sex differences seen in psychiatric disorders. The goal of this proposal is to characterize the whole population of mRNAs in the nucleus accumbens (NAc) of F1 offspring from parents with a history of THC exposure by transcriptome sequencing, as well as to study whole genome DNA methylation and chromatin modifications to identify the epigenetic milieu underlying the cross-generational disturbances in this mesolimbic brain region. The molecular studies will be combined with behavioral tests to assess reward-, depression- and anxiety-like behaviors in F1 males and females. We will address the causality between gene expression impairments and specific behavioral phenotypes using in vivo gene manipulations in the NAc. In order to determine whether there is persistent transgenerational transmission, phenotypic and epigenetic marks shown to be impaired in the F1 generation will be followed into the F2 progeny. Overall, these studies will provide fundamental mechanistic insights into the cross-generational impact of cannabis exposure that could guide the development of therapeutic interventions for the spectrum of behavioral phenotypes associated with vulnerability to addiction and psychiatric disorders.