Characterising epigenetic alterations following cocaine consumption.

Abstract

Complementary data from clinical and animal research have converged on the hypothesis that persistent use of psychostimulant drugs such as cocaine may not only involve pathological alterations in neural processes that subserve reward–related learning, but also complex interactions between genes and the environment through epigenetic modifications. Fosb and Crem (cAMP response element modulator) are among the central trans-factors suspected to mediate these long–term neurobiological changes due to their potential roles in drug reward. However, the critical question that concerns inheritance of epigenetic marks associated with parental cocaine experience in social settings, offspring vulnerability and modifying maternal–foetal environment by fostering, remains poorly understood. The present study therefore aimed to investigate possible associations between cocaine–induced behavioural changes in social contexts and DNA methylation patterns of inducible transcription factors in the prefrontal cortex (PFC) and hippocampus (HPC) of the exposed parent mice. We further examined whether the induced epigenetic changes were inheritable and then determined the impact of early postnatal (PN) fostering on associated neurobiological changes in the offspring of drug-exposed parents. In doing so, we were able to investigate the interaction between epigenetic and environmental factors in relation to drug consumption. Behavioural response of C57BL/6 mice to cocaine treatments were examined using conditioned place preference (CPP) and IntelliCage (IC) phenotyping techniques. In the CPP experiment, male mice received 6 cocaine injections (10mg/kg, i.p.) on alternate days followed by 6 days of extinction learning. A subthreshold dose of cocaine (5mg/kg, i.p) was later injected to reinstate CPP behaviour. In the IC, female mice were group–housed and initially had free access to drugs (300mg/L cocaine and 12% v/v ethanol in their drinking bottles) and water for 30 days to investigate consumption preference. Subsequently, withdrawal effect and alternate nose-poke learning tasks (ANT) were examined in the following 28 days with concurrent access to cocaine and water. In both experiments, locomotor activity and novelty exploration/recognition memory of mice were examined post cocaine treatments. DNA methylation status of Crem and Fosb gene promoters, within the HPC and PFC, were also assessed using quantitative real–time polymerase chain reaction. Thereafter, cocaine exposed or unexposed male and female mice were matched for mating to produce offspring with lineal phenotypes. At birth, some of the offspring were cross-fostered to further examine the impact of PN fostering on drug-induced epigenetic changes. Locomotion, memory competence and DNA methylation status were also evaluated in the offspring similar to the parent mice. In male mice, cocaine treatment resulted in significant changes in CPP during conditioning. After extinction learning, the subthreshold dose of cocaine did not reinstate the conditioned behaviour. The treatment increased locomotor activity in the open field but decreased novelty exploration in the object recognition task. These changes were characterized by significant hypomethylation in Crem and Fosb gene promoters only in the PFC. During the first 30 days of free access to cocaine, ethanol or water in the IC, the female mice spent significantly more time licking and consuming cocaine than ethanol, whereas consumption of either drugs was significantly less than water. Overall, the mice exhibited motivational deficits as manifested by their inability to learn the ANT. Our data also showed that prolonged access to cocaine in the IC decreased locomotor activity while recognition memory remained intact in the cocaine–experienced mice compared to their controls. These changes were accompanied by hypomethylation or hypermethylation in the promoters of Fosb and Crem genes in the PFC and HPC of the cocaine–experienced mice, respectively. In the offspring, memory performance and locomotor activity were not affected by parental cocaine exposure, except that recognition memory was impaired by early PN fostering in offspring lineally inclined to either paternal and/or combined parental cocaine experience. Crem was hypomethylated only in the PFC of offspring of cocaine–exposed parent mice, while fostering the offspring reversed the expression. Significant change in Fosb methylation was only observed in the HPC of fostered offspring. Together, these findings suggest differential responses of the substrate brain regions to the converging environmental stimuli and dynamic regulation of induced neurobiological changes via DNA methylation. Overall, the data also provide some evidence that cocaine–induced epigenetic marks can be inherited by the non-drug exposed offspring while early PN fostering may enhance molecular switching that may render the individual vulnerable to drug consumption. Key words: Cocaine; social environment; IntelliCage, conditioned place preference; Crem; Fosb; DNA methylation; hippocampus; prefrontal cortex; epigenetic inheritance; postnatal fostering.