The Role of Prelimbic Corticotrophin Releasing Factor Type 1 Receptor and L-Type Calcium Channels in Stress-induced Relapse

Abstract: Cocaine use disorder (CUD) is a growing public health concern, with over 2.2 million individuals in the US regularly using cocaine, and cocaine-related overdoses accounting for 1 in 5 drug-related deaths in 2017. Relapse rates for CUD are approximately 50% within the first 12 weeks of rehabilitative therapy, and stress is a trigger for drug craving during abstinence. There's a pressing need to understand the mechanisms underlying stress-induced relapse. Recent studies have pointed to corticotrophin releasing factor type 1 receptor (CRF1R) and the L-type calcium channel (LTCC), Cav1.2, as players in stress-induced relapse. A risk allele in the Cav1.2 gene, CACNA1C, has been associated with altered fear and reward processing within stress-sensitive brain regions such as the nucleus accumbens (NAc) and prefrontal cortex (PFC). Additionally, variants in the CRF1R gene have been linked to higher rates of mood disorders, suicidality, and addiction. Decades of preclinical research demonstrated that CRF1 antagonists produced anxiolytic and antidepressant like effects, and reduced stress-induced relapse. However, clinical trials with LTCC and CRF1 blockers have seen mixed results in its efficacy for addiction treatment, suggesting deeper understanding of these compounds in stress-induced relapse is needed. Most studies have focused on Cav1.2 and CRF1R actions in various regions of the mesolimbic circuits, but very few have focused on the prelimbic cortex pyramidal neurons that project to the nucleus accumbens core (PrL-NAcc), a circuit known to be involved in stress and cocaine cue reactivity. My proposal will investigate the role of Cav1.2 and CRF1R during abstinence in the PrL-NAcc and its role in stress-induced relapse. The major goals of this proposal are to determine the regulation of Cav1.2 and CRF1R in the PrL-NAcc circuit, as well as to determine the role these channels and receptors play in mediating stress-induced relapse at different times in abstinence. I will investigate this question through two aims: In Aim 1 I will use retrograde tracing to isolate PrL-NAcc neurons and perform immunofluorescence experiments to examine the expression pattern of Cav1.2 and CRF1R within the prelimbic to see if they are co-localized on PrL-NAcc neurons or if they are on distinct cell populations (Aim 1.1). I will also use flow cytometry and cell sorting to isolate the specific population of PrL-NAcc cells and perform immunoblotting to investigate the dynamic regulation of Cav1.2 and CRF1R during cocaine abstinence (Aim 1.2). In Aim 2 I will determine the functional role of Cav1.2 or CRF1R activity during abstinence. During early and extended abstinence, I will perform stress-induced relapse tests in cocaine-abstinent rats and infuse an LTCC blocker (Aim 2.1) or CRF1R antagonist (Aim 2.2) in the PrL and measure the effect on stress-induced relapse in cocaine-seeking behavior. The proposed research will advance our understanding of LTCC and CRF1R mechanisms that support stress-induced relapse and guide clinicians on the most effective therapy to use at various points in cocaine addiction.