Pilot Projects Funded in 2015
for Funding Period January 1 - December 31, 2015
Role of oxidative homeostasis in risk for and treatment of ethanol withdrawal, and reward
PI: Kari Buck, Ph.D., Department of Behavioral Neuroscience, OHSU; Research Scientist, Portland VA Medical Center
Abstract: Although no animal model exactly duplicates alcoholism in humans, models for specific traits, including ethanol withdrawal (EW), are valuable genetic resources. Detection of quantitative trait loci (QTLs), chromosomal regions within which allele (gene) variation affects a complex trait, has been fundamental to identify genes that confer risk. Using preclinical models, we have identified QTLs with high certainty (p<10-5) with large effects on EW. Here we focus on one of these QTLs (referred to herein as Ew1) with proven effects on EW using acute and chronic models. We have finely mapped Ew1 to a small 0.4 Mb interval. This QTL is syntenic with a region of human 1q where multiple studies identify loci associated with alcohol dependence in humans. Interestingly, our preliminary data indicate that Ew1 may also affect ethanol conditioned place preference (CPP). Further, QTLs are detected in this region of mouse chromosome 1 for additional rewarding and aversive effects of ethanol. Thus, a hypothesis that has emerged is that the gene(s) underlying Ew1 may affect a variety of alcohol responses, including its rewarding and/or aversive actions, and that the human homolog of the underlying gene(s) may contribute to alcoholism risk, making the gene(s) a compelling translational target.
The overall objective is to identify the gene(s) underlying Ew1 effects, and to assess its role in ethanol rewarding/aversive effects. We have identified a high-quality quantitative trait gene (QTG) candidate (Ndufs2) crucially involved in oxidative homeostasis. We provide the first evidence that N-acetylcysteine (NAC), a powerful antioxidant (and already FDA approved), reduces EW. Results from our lab provide the first studies of mitochondrial respiratory chain (MRC) complex organization in mouse brain, and find remarkable genetic differences in complex I (within which NDUFS2 is a core subunit) containing supercomplex organization and activities. Together, our data implicate oxidative homeostasis as having an important role in genetic risk for EW. A growing body of evidence implicates naturally-occurring variation in redox homeostasis/stress in genetically influenced differences among relevant genetic models, disproportionately affecting tissues with high energy demand including brain. Thus, our results may direct future research in areas beyond the addictions. The proposed aims are a logical progression of this work and may be summarized: 1. Comparing Ew1 congenic and wild-type (WT) littermates, test for pleiotropic effects of Ew1 on CPP. 2. Using congenic and WT littermates, test NAC effects on withdrawal in dependent animals and use WGCNA on Ew1 congenic mice to establish a role for NAC in Ew1. 3. Using RNAi, rigorously test the role of Ndufs2 on EW.
Retrotransposon activity in the brain of an alcoholism non-human primate model
PI: Lucia Carbone, Ph.D., Department of Behavioral Neuroscience, OHSU
Abstract: One of the major health concerns linked to alcohol use disorder (AUD) is the increased risk of developing brain changes leading to impairment of learning, memory, and other cognitive deficits. However, the molecular mechanisms through which chronic alcohol use affects brain function and structure are still relatively unexplored. This project focuses on studying the effects of alcohol use on retrotransposons: genomic parasites making up >20% of mammalian genomes. Somatic retrotransposition (SR) has been recently described in the adult brain, challenging the dogma that neurons remain stable for an individual’s lifetime. Moreover, transcriptional activation retrotransposons has been observed in post-mortem brain of human alcoholics. While this evidence suggests that alcohol use can disrupt retrotransposon repression, the degree to which it can modify SR in the adult brain has not been studied. Our long-term goal is to determine to what extent and by which mechanism(s) chronic alcohol use affects SR in the brain. The goal of this pilot proposal is to show the feasibility of our approach and to provide initial evidence that alcohol use influences SR in the brain of a small cohort (n=8) of rhesus macaques self-administering alcohol or water. Rhesus is physiologically and genetically close to humans. Moreover, it minimizes confounding factors usually present and uncontrollable in the human population (e.g. diet, other drugs, social pressure) while providing access to tissues. In order to identify somatic insertions, we will isolate single-neurons using laser microdissection from the dentate gyrus of the hippocampus and apply retrotransposon capture sequencing (RC-seq). Somatic insertions will be validated by PCR and whole-genome sequencing. The dentate gyrus has been chosen because hippocampal dysfunction has been associated with alcohol use; moreover, it is where most adult neurogenesis occurs. This proposal has the potential to uncover a completely novel path through which alcohol use establishes structural and functional changes in the brain. An improved understanding of SR and potential association with the behavioral deficits observed in AUD, will allow for the development of targeted therapies.
Investigating the bidirectional relationship between alcohol and circadian gene expression
PI: Angela Ozburn, Ph.D., Department of Behavioral Neuroscience, OHSU; Research Biologist, Portland VA Medical Center
Abstract: Previous studies 1) support a role for circadian genes in regulating alcohol and drug reward, and 2) reveal that alcohol and other drugs of abuse alter circadian rhythms. However, there are no studies of the effects of alcohol intake on rythmic expression of circadian genes. Furthermore, no studies have examined whether changes in circadian gene expression are correlated across regions known to be important for circadian rhythms and alcohol intake. The proposed study plans to address this major gap in our knowledge and collect preliminary data that will be useful for guiding a future major grant application. One goal of this proposal is to determine the effects of chronic binge-like drinking on diurnal rhythms of circadian gene expression in the master circadian pacemaker (SCN), and in the reward- and stress-related brain region, the nucleus accumbens (NAc). Addicted individuals display disrupted rhythms, and chronic disruption or particular chronotypes may increase the risk for substance abuse and relapse. Therefore, it is likely that medications that “re-set” or entrain circadian rhythms might be therapeutic for the reducing alcohol consumption, and in fact studies in rats have suggested that this might be the case. It is unknown whether this compound ameliorates alcohol-induced changes in rhythmic expression of circadian genes. We will employ mice selectively bred to achieve high blood alcohol levels in the binge-drinking paradigm, drinking in the dark (HDID) to address this gap in our knowledge. Thus, the second goal of this proposal is to determine whether pharmacologically inhibiting CK1E/D, a key regulator of the period of the molecular clock, reduces binge-like alcohol drinking and alters circadian gene expression in the SCN and the NAc. Our overarching hypothesis is that chronic alcohol intake will reduce the period of rhythmic circadian gene expression in the NAc and that inhibiting CK1E/D will reduce alcohol intake and "re-set" or lengthen the molecular clock period. It is possible that these changes will be present in the SCN as well, which would test whether or not a connection exists between alcohol effects on the SCN and the NAc..
PARC Pilot Project Application Information
Each year, the Portland Alcohol Research Center (PARC) aims to fund two to four pilot research projects (up to $35K each). The PARC is especially interested in encouraging investigators new to alcohol research to submit applications. Pilot proposals that focus on the neuroadaptation to ethanol exposure and those with relevance to behavioral genomics are especially welcome.The call for pilot project applications generally is made each year on this website July 1st, with an application due date in late August, for funding to commence for selected projects the following January. Each proposal is reviewed by at least two members of our external scientific advisory board. Based on these reviews and their own reviews, Center Director Tamara Phillips and Scientific Director Robert Hitzemann make recommendations to the PARC Executive Committee for funding.