Amygdala-Hippocampal phospholipase D (PLD) signaling as novel mechanism of cocaine-environment maladaptive conditioned responses

Background: Drug-environment associative memory mechanisms and the resulting conditioned behaviors are key contributors in relapse to cocaine dependence. Recently, we reported rat amygdala phospholipase D as a key convergent downstream signaling partner in the expression of cocaine-conditioned behaviors mediated by glutamatergic and dopaminergic pathways. In the present study, 1 of the 2 known upstream serotonergic targets of phospholipase D, the serotonin (5-hydroxytryptamine) 2C receptor, was investigated for its role in recruiting phospholipase D signaling in cocaine-conditioned behaviors altered in the rat amygdala and dorsal hippocampus. Methods: Using Western-blot analysis, amygdala phospholipase D phosphorylation and total expression of phospholipase D/5-hydroxytryptamine 2C receptor were observed in early (Day-1) and late (Day-14) withdrawal (cocaine-free) states among male Sprague-Dawley rats subjected to 7-day cocaine-conditioned hyperactivity training. Functional studies were conducted using Chinese Hamster Ovary cells with stably transfected human unedited isoform of 5-hydroxytryptamine 2C receptor. Results: Phosphorylation of phospholipase D isoforms was altered in the Day-1 group of cocaine-conditioned animals, while increased amygdala and decreased dorsal hippocampus phospholipase D/5-hydroxytryptamine 2C receptor protein expression were observed in the Day-14 cocaine-conditioned rats. Functional cellular studies established that increased p phospholipase D is a mechanistic response to 5-HT2CR activation and provided the first evidence of a biased agonism by specific 5-hydroxytryptamine 2C receptor agonist, WAY163909 in phospholipase D phosphorylation 2, but not phospholipase D phosphorylation 1 activation. Conclusions: Phospholipase D signaling, activated by dopaminergic, glutamatergic, and serotonergic signaling, can be a common downstream element recruited in associative memory mechanisms altered by cocaine, where increased expression in amygdala and decreased expression in dorsal hippocampus may result in altered anxiety states and increased locomotor responses, respectively.