C. Cifani

Ventral medial prefrontal cortex neuronal ensembles mediate context-induced relapse to heroin

In a rat model of context-induced relapse to heroin, we identified sparsely distributed ventral medial prefrontal cortex (mPFC) neurons that were activated by the heroin-associated context. Selective pharmacogenetic inactivation of these neurons inhibited context-induced drug relapse. A small subset of ventral mPFC neurons formed neuronal ensembles that encode the learned associations between heroin reward and heroin-associated contexts; re-activation of these neuronal ensembles by drug-associated contexts during abstinence provoked drug relapse.

Amylin receptor components and the leptin receptor are co‐expressed in single rat area postrema neurons

Amylin is a pancreatic β‐cell hormone that acts as a satiating signal to inhibit food intake by binding to amylin receptors (AMYs) and activating a specific neuronal population in the area postrema (AP). AMYs are heterodimers that include a calcitonin receptor (CTR) subunit [CTR isoform a or b (CTRa or CTRb)] and a member of the receptor activity‐modifying proteins (RAMPs). Here, we used single‐cell quantitative polymerase chain reaction to assess co‐expression of AMY subunits in AP neurons from rats that were injected with amylin or vehicle. Because amylin interacts synergistically with the adipokine leptin to reduce body weight, we also assessed the co‐expression of AMY and the leptin receptor isoform b (LepRb) in amylin‐activated AP neurons. Single cells were collected from Wistar rats and from transgenic Fos‐GFP rats that express green fluorescent protein (GFP) under the control of the Fos promoter. We found that the mRNAs of CTRa, RAMP1, RAMP2 and RAMP3 were all co‐expressed in single AP neurons. Moreover, most of the CTRa+ cells co‐expressed more than one of the RAMPs. Amylin down‐regulated RAMP1 and RAMP3 but not CTR mRNAs in AMY+ neurons, suggesting a possible negative feedback mechanism of amylin at its own primary receptors. Interestingly, amylin up‐regulated RAMP2 mRNA. We also found that a high percentage of single cells that co‐expressed all components of a functional AMY expressed LepRb mRNA. Thus, single AP cells expressed both AMY and LepRb, which formed a population of first‐order neurons that presumably can be directly activated by amylin and, at least in part, also by leptin.

Regulation of hypothalamic neuropeptides gene expression in diet induced obesity resistant rats: possible targets for obesity prediction?

Several factors play a role in obesity (i.e., behavior, environment, and genetics) and epigenetic regulation of gene expression has emerged as a potential contributor in the susceptibility and development of obesity. To investigate the individual sensitivity to weight gain/resistance, we here studied gene transcription regulation of several hypothalamic neuropeptides involved in the control of energy balance in rats developing obesity (diet-induced obesity, DIO) or not (diet resistant, DR), when fed with a high fat diet. Rats have been followed up to 21 weeks of high fat diet exposure. After 5 weeks high fat diet exposure, the obese phenotype was developed and we observed a selective down-regulation of the orexigenic neuropeptide Y (NPY) and peroxisome proliferator-activated receptor gamma (PPAR-γ) genes. No changes were observed in the expression of the agouti-related protein (AgRP), as well as for all the anorexigenic genes under study. After long-term high fat diet exposure (21 weeks), NPY and PPAR-γ, as well as most of the genes under study, resulted not be different between DIO and DR, whereas a lower expression of the anorexigenic pro-opio-melanocortin (POMC) gene was observed in DIO rats when compared to DR rats. Moreover we observed that changes in NPY and POMC mRNA were inversely correlated with gene promoters DNA methylation. Our findings suggest that selective alterations in hypothalamic peptide genes regulation could contribute to the development of overweight in rats and that environmental factor, as in this animal model, might be partially responsible of these changes via epigenetic mechanism.

Epigenetic regulation of nociceptin/orphanin FQ and corticotropin-releasing factor system genes in frustration stress-induced binge-like palatable food consumption

Evidence suggests that binge eating may be caused by a unique interaction between dieting and stress. We developed a binge‐eating model in which female rats with a history of intermittent food restriction show binge‐like palatable food consumption after a 15‐minute exposure to the sight of the palatable food (frustration stress). The aim of the present study was to investigate the regulation of the stress neurohormone corticotropin‐releasing factor (CRF) system and of the nociceptin/orphanin FQ (N/OFQ) system genes in selective rat brain regions, using our animal model. Food restriction by itself seems to be responsible in the hypothalamus for the downregulation on messenger RNA levels of CRF‐1 receptor, N/OFQ and its receptor (NOP). For the latter, this alteration might be due to selective histone modification changes. Instead, CRF gene appears to be upregulated in the hypothalamus as well as in the ventral tegmental area only when rats are food restricted and exposed to frustration stress, and, of relevance, these changes appear to be due to a reduction in DNA methylation at gene promoters. Moreover, also CRF‐1 receptor gene resulted to be differentially regulated in these two brain regions. Epigenetic changes may be viewed as adaptive mechanisms to environmental perturbations concurring to facilitate food consumption in adverse conditions, that is, in this study, under food restriction and stressful conditions. Our data on N/OFQ and CRF signaling provide insight on the use of this binge‐eating model for the study of epigenetic modifications in controlled genetic and environmental backgrounds.

P.1.024 Epigenetic regulation of A2A gene transcription in a model of binge eating in the amygdala complex of female rats

Conclusions: Our data suggest a possible role for histone H3 phosphoacetylation in LPS induced sickness behavior as well as in the regulation of transcriptional mechanisms involved in the immune response. In this way, histone modification may represent the first of a cascade of genetic events that determine the behavioral alterations and sensitivity to stress observed after activation of the immune system.