This is a competing renewal application for a U01 grant entitled """"""""Neurocircuitry Mapping and Genotyping Core"""""""";the application is submitted as a member of the NIAAA sponsored """"""""Integrative Neuroscience Initiative on Alcoholism (INIA)-West (G. Koob, PI). The application continues the focus of the current funding period on both research and core activities. Key core activities of the current funding period were a) the mastery of the use of the Weighted Gene Co-variance Network Analysis (WGCNA) for moderate to large sample sizes (lancu et al. 2010) and b) the development of a strategy for and implementation of quantitative RNAseq (Bottomly et al, 2011;Appendix A). With these tools in hand, we propose 1) to directly sequence the transcriptome ( ~ 25,000,000 75 bp reads/sample) in both replicate High Drinking in the Dark (HDID) mouse lines and in the HS/NPT control animals and 2) to sequence the transcriptome HDID animals that have completed the chronic intermittent ethanol (CIE) procedure with the appropriate control groups. The tissues needed for this analysis will be provided by the Crabbe U01. As the HDID and controls are derived from a 8- way inbred strain cross (Hitzemann et al. 1994), RNAseq is particularity useful, given that masking oligonucleotide array data is never optimal (see Walter et al. 2007,2009). N = 32/group;previous work (lancu et al. 2010) has illustrated that samples of this size are adequate for the proposed analyses. Samples are collected by laser capture micro-dissection (LCM);the regional priority for analysis will be the central nucleus of the amygdala (CeA) >the infralimbic cortex (IL) >the prelimbic cortex (PL). The occipital cortex (OC) will be used as a control region.
Aim 1 focuses on binge drinking whereas aim 2 focuses on how chronic ethanol exposure affects ethanol consumption in limited access 2-bottle choice paradigm. Our working hypothesis is that differences between co-expression networks and not the differential expression of individual genes have the greatest translational value (see e.g. Oti et al. 2008;Zhao et al. 2010).
In Aim 3, samples from ethanol exposed macaques (Grant U01- INlA-Stress) will be sequenced. Data from the CeA and cortical areas 25 and 32 will be compared to the results obtained in specific aims 1 and 2.
The purpose of the proposed research is to understand what genes are associated with animal models of excessive ethanol consumption. Detecting these genes and probably more importantly their associated gene networks may lead to new therapeutic targets for the treatment of alcoholism.
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