FACS: A novel method to study neurons altered during cocaine sensitization

Yale University, New Haven CT

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Abstract

DESCRIPTION (provided by applicant): Cocaine exerts a high toll on our society. These studies will help us to better understand how the environment that a former cocaine user associates with drug can cause relapse to cocaine use. This understanding should lead to better treatments for cocaine addiction. Many former cocaine abusers cite environmental cues (such as seeing cocaine paraphernalia) as a direct cause of their relapse to cocaine use. Our long-term goal is to understand how the environment modulates cocaine-related behaviors such as self-administration and reinstatement. To develop the techniques necessary for this, we begin with the simpler model of context-specific locomotor sensitization. We hypothesize that the learned associations between cocaine and the drug administration environment are mediated by changes in a minority of sparsely distributed neurons called "neuronal ensembles". These neuronal ensembles are selected by the cocaine-paired environmental stimuli. This idea is supported by the finding that selective lesions of cocaine-activated neuronal ensembles in the nucleus accumbens of transgenic c-fos-lacZ rats strongly attenuate the expression of context-dependent sensitization to cocaine (Koya et al., SFN 2007). The study proposed here attempts to identify unique neuroadaptations in these selectively activated neuronal ensembles;these are different from the more commonly examined neuroadaptations found in homogenates of brain tissue. Bruce Hope's lab has a special tool for identifying these activated neuronal ensembles: a c-fos-lacZ transgenic rat. Because all c-fos-positive cells in this animal express beta-galactosidase, we can separate the few activated cells from the majority of surrounding minimally activated cells. This allows us to study the activated cells specifically, instead of studying a homogenate of the relevant brain area, which is likely to obscure the molecular changes seen in only the activated cells. We will use Fluorescence Activated Cell Sorting to separate the population of highly activated cells from the majority of relatively non-activated neighboring cells. We will then use quantitative real time PCR to characterize the different molecular changes occurring in these cells as a result of context-dependent cocaine exposure.