Drug dependence is a persistent problem throughout the world. Once addicted to a drug, many users have difficulty quitting use, even when they desire to stop using. This substance dependence is both psychological as well as neurophysiological. In 2008 the U.S. Department of Health classified 22 million Americans as having a significant degree of drug dependence. Additionally, the U.S. Office of National Drug Control Policy stated that the health care costs in 2004 for substance abuse was estimated to be comparable to cancer. There is therefore undoubtedly both an individual and societal need for therapeutic interventions in drug dependence.
While a great deal is known about the molecular action of these addictive substances, very little is understood about the underlying neurocircuitry of addiction. The process of addiction is a learned behavior and is not solely driven by homeostatic adaptations to the drug itself. Psychoactive drugs overrun the natural reward circuitry of the brain, forcing maladaptive learning of drug associated rewards which then become overvalued in comparison to natural rewards. This dissertation approaches the study of drug addiction using two separate techniques: one regionally specific to the nucleus accumbens, and one cell type specific to dopaminergic and GABAergic neurotransmitter systems of the central nervous system.
All drugs of abuse increase striatal dopamine release despite their widely varied mechanisms of action. The ventral striatum primarily consists of the Nucleus Accumbens (NAc), which has long been thought of as the origin of addiction-like behaviors. As such, the NAc has been proposed to be one of the primary reward centers of the brain. Utilizing viral delivery of short-hairpin RNA (shRNA) into the NAc, knockdown of specific subtypes of receptors can be achieved. In this manner, behavioral testing can investigate changes in rodent addiction-like behavior following region specific knock down of dopamine receptor isoforms.
The widespread dopaminergic connections leaving the ventral tegmental area (VTA) modulate the glutamatergic and GABAergic connections that instigate reward valuation and associative memory formation. An integrated view of these circuits is needed in order to better understand the progressive neurophysiological changes that are occurring in addiction. Bac transgenic techniques can be used to generate mouse lines with inducible Cre expression specifically within dopaminergic or GABAergic neuronal cell types. Utilizing these mouse lines, both neurotransmitter systems can be characterized in a manner never before possible.
In conclusion, this dissertation shows that techniques can now be employed to begin studying the neurocircuitry of addiction with regional or cell type specificity. The method for rapid screening of shRNA that is presented here will expedite research using transcriptional regulation. Our results on D2L knockdown of the dopamine receptor within the nucleus accumbens are an informative first pass at attempting to differentiate these near indistinguishable splice variants. The experimental mouse models generated by our bac recombineering will advance research into the role that these specific neurotransmitter systems are playing in addiction. Further understanding the brain regions and neurotransmitter systems associated with drug addiction can lead to potential pharmacotherapeutics.