Rose, Timothy2022-04-132022-04-132022-01https://hdl.handle.net/11299/226947University of Minnesota Ph.D. dissertation. 2022. Major: Pharmacology. Advisor: Kevin Wickman. 1 computer file (PDF); 160 pages.Drugs of abuse share the ability to enhance DA levels within the mesocorticolimbic system. This increased DA neurotransmission triggers persistent adaptations throughout the brain that are believed to underlie the detrimental behaviors that define addiction. For example, chronic cocaine exposure causes a suppression of inhibitory G protein-dependent signaling mediated by the GABAB receptor (GABABR) and G protein-gated inwardly rectifying K+ (GIRK/Kir3) channel in pyramidal neurons of the prelimbic cortex (PL), a cell population important for executive function. As GIRK-dependent signaling is crucial for tempering excitatory input in neurons, the loss of this “inhibitory brake" may drive neuronal hyperexcitability and foster the development of addiction-related behavior. The goal of this thesis is to examine the contribution of GIRK channels in PL pyramidal neurons to behaviors that may be relevant to addiction, and to further understand the regulatory mechanisms that control inhibitory signaling mediated by GABABRs and GIRK channels.To test the prediction that a loss of GIRK channel activity in pyramidal neurons promotes neuronal hyperexcitability, we employed a viral genetic approach to selectively ablate a critical GIRK channel subunit (GIRK1) in PL pyramidal neurons. GIRK channel ablation blunted GABABR-GIRK currents in, and elevated the excitability of, PL pyramidal neurons – electrophysiological outcomes that closely resemble the effects of repeated cocaine exposure. To examine the behavioral consequences of elevated PL pyramidal neuron excitability, we used complementary viral approaches to model the impact of acute (chemogenetic) and persistent (GIRK channel ablation) excitation of PL pyramidal neurons on PL-dependent behaviors, including acute cocaine-induced locomotion and trace fear conditioning. We found that GIRK channel ablation enhanced the motor-stimulatory effect of cocaine, but did not impact baseline activity or trace fear learning. In contrast, selective chemogenetic excitation of PL pyramidal neurons increased baseline and cocaine-induced activity and disrupted trace fear learning. These effects were mirrored in male mice by selective excitation of PL pyramidal neurons projecting to the ventral tegmental area, a brain region important for reward behavior. Collectively, these data show that manipulations enhancing the excitability of PL pyramidal neurons, and specifically those projecting to the VTA, recapitulate behavioral hallmarks of repeated cocaine exposure in mice. Withdrawal from prolonged cocaine exposure has been correlated with negative affective behaviors, as well as formation of persistent drug-related memories that drive drug-seeking behavior. Therefore, we next modeled the impact of viral-mediated GIRK, or GABABR, ablation in PL pyramidal neurons on mood-related behaviors and cocaine conditioned place preference (CPP). While GIRK ablation did not impact anxiety- or depression-related behavior, the manipulation impaired the extinction of cocaine CPP in male mice. In contrast, GABABR ablation was without effect. Since an impairment in extinction may result in prolonged drug-seeking behavior, we next assessed whether strengthening GIRK channel activity could enhance the extinction of cocaine CPP. As predicted, overexpression of GIRK2 in PL pyramidal neurons facilitated extinction of cocaine CPP in male mice. Together, these findings highlight a unique, sex-specific role for GIRK channels in PL pyramidal neurons in tempering cocaine conditioned responding. Despite established links between GIRK channel plasticity and disease, the basic mechanisms that regulate GIRK-dependent signaling in PL pyramidal neurons are not fully understood. One important regulator of GIRK channel activity is the regulator of G protein signaling (RGS) protein, and specifically RGS6 and RGS7 (RGS6/7). RGS6/7 facilitate the termination of inhibitory G protein-dependent signaling, and are thus critical for maintaining the high temporal resolution of GABABR-GIRK signaling. While both RGS6/7 are expressed in the PFC, little is known about their functional roles in the PL. After establishing that RGS6/7 are coexpressed in most PL pyramidal neurons, we next examined their contribution to synaptically-evoked and baclofen-activated GABABR-GIRK currents using constitutive RGS6–/– and RGS7–/– mice. We found that RGS6/7 differentially regulate GIRK channel activity; RGS6 regulates the amplitude, while RGS7 regulates the kinetics and sensitivity, of GIRK-dependent signaling. These shed light on the functional compartmentalization mechanisms that are critical for ensuring high temporal resolution of neuronal inhibitory G protein-dependent signaling. Overall, the work in this thesis suggests that GIRK-dependent signaling in PL pyramidal neurons represents an “inhibitory brake” on cellular excitability that is critical for excitation/inhibition balance and optimal behavioral function. Although the weakening of this inhibition following repeated cocaine exposure may promote neuronal hyperexcitability and addiction-related behavior, therapeutic interventions that restore inhibitory tone may confer resilience to these effects.enAddictionCocaineGABAB receptorGIRK channelPlasticityRGS proteinThesis or Dissertation