Characterization of orexin input to dopamine neurons of the ventral tegmental area projecting to the medial prefrontal cortex and shell of nucleus accumbens

Orexin neurons are involved in homeostatic regulatory processes, including arousal and feeding, and provide a major input from the hypothalamus to the ventral tegmental area (VTA) of the midbrain. VTA neurons are a central hub processing reward and motivation and target the medial prefrontal cortex (mPFC) and the shell part of nucleus accumbens (NAcs). We investigated whether subpopulations of dopamine (DA) neurons in the VTA projecting either to the mPFC or the medial division of shell part of nucleus accumbens (mNAcs) receive differential input from orexin neurons and whether orexin exerts differential electrophysiological effects upon these cells. VTA neurons projecting to the mPFC or the mNAcs were traced retrogradely by Cav2-Cre virus and identified by expression of yellow fluorescent protein (YFP). Immunocytochemical analysis showed that a higher proportion of all orexin-innervated DA neurons projected to the mNAcs (34.5%) than to the mPFC (5.2%). Of all sampled VTA neurons projecting either to the mPFC or mNAcs, the dopaminergic (68.3 vs. 79.6%) and orexin-innervated DA neurons (68.9 vs. 64.4%) represented the major phenotype. Whole-cell current clamp recordings were obtained from fluorescently labeled neurons in slices during baseline periods and bath application of orexin A. Orexin similarly increased the firing rate of VTA dopamine neurons projecting to mNAcs (1.99 ± 0.61 Hz to 2.53 ± 0.72 Hz) and mPFC (0.40 ± 0.22 Hz to 1.45 ± 0.56 Hz). Thus, the hypothalamic orexin system targets mNAcs and to a lesser extent mPFC-projecting dopaminergic neurons of the VTA and exerts facilitatory effects on both clusters of dopamine neurons.

Tactile Stimulation in Adult Rats Modulates Dopaminergic Molecular Parameters in the Nucleus Accumbens Preventing Amphetamine Relapse.

Amphetamine (AMPH) is a psychostimulant drug frequently related to addiction, which is characterized by functional and molecular changes in the brain reward system, favoring relapse development and pharmacotherapies have shown low effectiveness. Considering the beneficial influences of tactile stimulation (TS) in different diseases that affect the central nervous system (CNS), here we evaluated if TS applied in adult rats could prevent or minimize the AMPH-relapse behavior also accessing molecular neuroadaptations in the Nucleus accumbens (NAc). Following AMPH conditioning in the conditioned place preference (CPP) paradigm, male rats were submitted to TS (15-min session, 3 times a day, for 8 days) during the drug abstinence period, which were re-exposed to the drug in the CPP paradigm for additional 3 days for relapse observation and molecular assessment. Our findings showed that besides AMPH relapse; TS prevented the dopamine transporter (DAT), dopamine 1 receptor (D1R), tyrosine hydroxylase (TH), mu opioid receptor (MOR) increase and AMPH-induced delta FosB (ΔFosB). Based on these outcomes, we propose TS as a useful tool to treat psychostimulant addiction, which subsequent to clinical studies; it could be included in detoxification programs together with pharmacotherapies and psychological treatments already conventionally established.

Enhanced food motivation in obese mice is controlled by D1R expressing spiny projection neurons in the nucleus accumbens.

Obesity is a chronic relapsing disorder that is caused by an excess of caloric intake relative to energy expenditure. In addition to homeostatic feeding mechanisms, there is growing recognition of the involvement of food reward and motivation in the development of obesity. However, it remains unclear how brain circuits that control food reward and motivation are altered in obese animals. Here, we tested the hypothesis that signaling through pro-motivational circuits in the core of the nucleus accumbens (NAc) is enhanced in the obese state, leading to invigoration of food seeking. Using a novel behavioral assay that quantifies physical work during food seeking, we confirmed that obese mice work harder than lean mice to obtain food, consistent with an increase in the relative reinforcing value of food in the obese state. To explain this behavioral finding, we recorded neural activity in the NAc core with both in vivo electrophysiology and cell-type specific calcium fiber photometry. Here we observed greater activation of D1-receptor expressing NAc spiny projection neurons (NAc D1SPNs) during food seeking in obese mice relative to lean mice. With ex vivo slice physiology we identified both pre- and post-synaptic mechanisms that contribute to this enhancement in NAc D1SPN activity in obese mice. Finally, blocking synaptic transmission from D1SPNs decreased physical work during food seeking and attenuated high-fat diet-induced weight gain. These experiments demonstrate that obesity is associated with a selective increase in the activity of D1SPNs during food seeking, which enhances the vigor of food seeking. This work also establishes the necessity of D1SPNs in the development of diet-induced obesity, identifying a novel potential therapeutic target.

Metabolic sensing in AgRP neurons integrates homeostatic state with dopamine signalling in the striatum

Agouti-related peptide (AgRP) neurons increase motivation for food, however whether metabolic sensing of homeostatic state in AgRP neurons potentiates motivation by interacting with dopamine reward systems is unexplored. As a model of impaired metabolic-sensing, we used the AgRP-specific deletion of carnitine acetyltransferase (Crat) in mice. We hypothesized that metabolic sensing in AgRP neurons is required to increase motivation for food reward by modulating accumbal or striatal dopamine release. Studies confirmed that Crat deletion in AgRP neurons (KO) impaired ex vivo glucose-sensing, as well as in vivo responses to peripheral glucose injection or repeated palatable food presentation and consumption. Impaired metabolic-sensing in AgPP neurons reduced acute dopamine release (seconds) to palatable food consumption and during operant responding, as assessed by GRAB-DA photometry in the nucleus accumbens, but not the dorsal striatum. Impaired metabolic-sensing in AgRP neurons suppressed radiolabelled 18F-fDOPA accumulation after ~30 minutes in the dorsal striatum but not the nucleus accumbens. Impaired metabolic sensing in AgRP neurons suppressed motivated operant responding for sucrose rewards during fasting. Thus, metabolic-sensing in AgRP neurons is required for the appropriate temporal integration and transmission of homeostatic hunger-sensing to dopamine signalling in the striatum.

RNA-sequencing of human post-mortem hypothalamus and nucleus accumbens identifies expression profiles associated with obesity.

Obesity, the accumulation of body fat to excess, may cause serious negative health effects, including increased risk of heart disease, type 2 diabetes, stroke and certain cancers. The biology of obesity is complex and not well understood, involving both environmental and genetic factors and affecting metabolic and endocrine mechanisms in tissues of the gut, adipose, and brain. Previous RNA sequencing studies have identified transcripts associated with obesity and body mass index in blood and fat, often using animal models, but RNA sequencing studies in human brain tissue related to obesity have not been previously undertaken. We conducted both large and small RNA sequencing of hypothalamus (207 samples) and nucleus accumbens (276 samples) from individuals defined as consistently obese (124 samples), consistently normal weight as controls (148 samples) or selected without respect to BMI and falling within neither case nor control definition (211 samples), based on longitudinal BMI measures. The samples were provided by three cohort studies with brain donation programs; the Framingham Heart Study (FHS), the Religious Orders Study (ROS) and the Rush Memory and Aging Project (MAP). For each brain region and large/small RNA sequencing set, differential expression of obesity, BMI, brain region and sex was performed. Analyses were done transcriptome-wide as well as with a priori defined sets of obesity or BMI-associated mRNAs and microRNAs (miRNAs). There are sixteen mRNAs and five microRNAs that are differentially expressed (adjusted p < 0.05) by obesity or BMI in these tissues, several of which were validated with qPCR data. The results include many that are BMI-associated, such as APOBR and CES1, as well as many associated with the immune system and some with addiction, such as the gene sets 'cytokine signaling in immune system' and 'opioid signaling'. In spite of the relatively large number of samples, our study was likely under-powered to detect other transcripts or miRNA with relevant but smaller effects.