| In Chapter I, target-based pharmacology (TBP) and systems-based pharmacology (SBP) methods, which represent the platform for target-based and system-based drug discovery (TBD and SBD), respectively, were compared to identify useful therapeutics. TBD has been favored, yet SBD now offers the promise of enhancing productivity by yielding higher value therapeutics.;In Chapter II, a systems pharmacology approach was used to evaluate the possible use of the neuroactive steroid pregnanolone hemisuccinate (3alpha5betaHS) in the treatment of cocaine addiction. 3alpha5betaHS has shown neuroprotective, analgesic and sedative effects in animal models of stroke and pain. This neurosteroid modulates excitatory and inhibitory neurotransmitter receptors and is a potent negative modulator at the AMPA GluR1 and NMDA NR2A-containing receptors. Pretreatment of rats self-administering cocaine with 10 mg/kg 3alpha5betaHS did not affect the animal's motivation to obtain the drug under a progressive ratio schedule of reinforcement. By contrast, 10 mg/kg 3alpha5betaHS reduced drug-seeking behavior in animals undergoing cocaine priming-induced reinstatement. Because 3alpha5betaHS also negatively modulates NMDA NR2B-containing receptors, the effects of the NR2B-specific antagonist Ro 25-6981 (2.5 or 5 mg/kg) on cocaine self-administration and cocaine priming-induced reinstatement were investigated but no inhibition was observed. The neuroactive steroid 3alpha5betaHS modulates several classes of receptor but may be a novel non-addictive therapeutic for the reduction of cocaine craving.;In Chapter III, the regulation of the GABAB receptor subunit gene promoter activity was investigated. The subcellular localization and developmental regulation of this inhibitory G-protein-coupled receptor were examined in cultured hippocampal neurons. GABAB receptor subunits GABAB1 and GABAB2 have been detected in the nucleus of neurons from the visual cortex, basal ganglia and/or neostriatum. Here, a combination of gel-shift and DNA pull-down analyses demonstrated that full length GABAB1 and some of its C-terminal fragments are present in neuronal nuclear extracts and interact with a short fragment of the Gabbr1 promoter-A. An analysis of the Gabbr1 promoter-A using chromatin immunoprecipitation demonstrates that GABAB1 interacts with its own promoter in hippocampal neurons. Moreover, overexpression of GABAB1a decreases the activity of its promoter, suggesting a novel role for the G-protein-coupled receptor GABAB1 subunit as an autologous transcriptional regulator of this inhibitory neurotransmitter subunit. |
