[show abstract][hide abstract] ABSTRACT: The association between dietary obesity and mesolimbic systems that regulate hedonic aspects of feeding is currently unresolved. In the present study, we examined differences in baseline and stimulated central dopamine levels in obesity-prone (OP) and obesity-resistant (OR) rats. OP rats were hyperphagic and showed a 20% weight gain over OR rats at wk 15 of age, when fed a standard chow diet. This phenotype was associated with a 50% reduction in basal extracellular dopamine, as measured by a microdialysis probe in the nucleus accumbens, a projection site of the mesolimbic dopamine system that has been implicated in food reward. Similar defects were also observed in younger animals (4 wk old). In electrophysiology studies, electrically evoked dopamine release in slice preparations was significantly attenuated in OP rats, not only in the nucleus accumbens but also in additional terminal sites of dopamine neurons such as the accumbens shell, dorsal striatum, and medial prefrontal cortex, suggesting that there may be a widespread dysfunction in mechanisms regulating dopamine release in this obesity model. Moreover, dopamine impairment in OP rats was apparent at birth and associated with changes in expression of several factors regulating dopamine synthesis and release: vesicular monoamine transporter-2, tyrosine hydroxylase, dopamine transporter, and dopamine receptor-2 short-form. Taken together, these results suggest that an attenuated central dopamine system would reduce the hedonic response associated with feeding and induce compensatory hyperphagia, leading to obesity.
The FASEB Journal 06/2008; 22(8):2740-6. · 5.70 Impact Factor
[show abstract][hide abstract] ABSTRACT: The ability to create primary cell cultures of dopamine neurons allows for the study of the presynaptic characteristics of dopamine neurons in isolation from systemic input from elsewhere in the brain. In our lab, we use these neurons to assess dopamine release kinetics using carbon fiber amperometry, as well as expression levels of dopamine related genes and proteins using quantitative PCR and immunocytochemistry. In this video, we show you how we generate these cultures from rodent neonates. The process involves several steps, including the plating of cortical glial astrocytes, the conditioning of neuronal cell culture media by the glial substrate, the dissection of the midbrain in neonates, the digestion, extraction and plating of dopamine neurons and the addition of neurotrophic factors to ensure cell survival. The applications suitable for such a preparation include electrophysiology, immunocytochemistry, quantitative PCR, video microscopy (i.e., of real-time vesicular fusion with the plasma membrane), cell viability assays and other toxicological screens.
[show abstract][hide abstract] ABSTRACT: The ability to measure extracellular basal levels of neurotransmitters in the brain of awake animals allows for the determination of effects of different systemic challenges (pharmacological or physiological) to the CNS. For example, one can directly measure how the animal's midbrain dopamine projections respond to dopamine-releasing drugs like d-amphetamine or natural stimuli like food. In this video, we show you how to implant guide cannulas targeting specific sites in the rat brain, how to insert and implant a microdialysis probe and how to use high performance liquid chromatography coupled with electrochemical detection (HPLC-EC) to measure extracellular levels of oxidizable neurotransmitters and metabolites. Local precise introduction of drugs through the microdialysis probe allows for refined work on site specificity in a compound s mechanism of action. This technique has excellent anatomical and chemical resolution but only modest time resolution as microdialysis samples are usually processed every 20-30 minutes to ensure detectable neurotransmitter levels. Complementary ex vivo tools (i.e., slice and cell culture electrophysiology) can assist with monitoring real-time neurotransmission.
[show abstract][hide abstract] ABSTRACT: Cannabinoids, the active components of marijuana, stimulate appetite, and cannabinoid receptor-1 (CB1-R) antagonists suppress appetite and promote weight loss. Little is known about how CB1-R antagonists affect the central neurocircuitry, specifically the melanocortin system that regulates energy balance.
Here, we show that peripherally administered CB1-R antagonist (AM251) or agonist equally suppressed or stimulated feeding respectively in A(y) , which lack a functional melanocortin system, and wildtype mice, demonstrating that cannabinoid effects on feeding do not require melanocortin circuitry. CB1-R antagonist or agonist administered into the ventral tegmental area (VTA) equally suppressed or stimulated feeding respectively, in both genotypes. In addition, peripheral and central cannabinoid administration similarly induced c-Fos activation in brain sites suggesting mediation via motivational dopaminergic circuitry. Amperometry-detected increases in evoked dopamine (DA) release by the CB1-R antagonist in nucleus accumbens slices indicates that AM251 modulates DA release from VTA terminals.
Our results demonstrate that the effects of cannabinoids on energy balance are independent of hypothalamic melanocortin circuitry and is primarily driven by the reward system.
PLoS ONE 02/2008; 3(5):e2202. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: The contribution of mesolimbic dopamine to the development of dietary obesity is not yet well understood. Because this system responds to feeding, dietary and/or genetic obesity could be linked to central dopamine deficiencies. In this study, we use inbred obesity-prone and obesity-resistant Sprague-Dawley adult female rats (Charles River Laboratories) to show that obesity-prone animals have lower basal extracellular dopamine levels in the nucleus accumbens shell in vivo (0.029±0.002 vs. 0.040±0.001 pg/sample, p<0.05). Using carbon fiber amperometry to monitor real-time-evoked dopamine release in all terminals of the mesolimbic system, we find 50% lower dopamine signal throughout in the obesity-prone rats. Leptin-deficient mice also have low dopamine signal in nucleus accumbens slices, while lean melanin concentrating hormone-deficient mice show the opposite. Because these effects are potentially linked only to body weight phenotype differences, we also studied inbred obesity-prone and obesity-resistant rat neonates using immunocytochemistry and quantitative PCR. As predicted from differences in stimulated dopamine release in the adults, the number of VMAT2-immunopositive sites (248±1 vs. 648±1; p<0.01) and VMAT2 and TH mRNA levels (0.62±0.03 vs. 1.62±0.15; p<0.01, 0.52±0.15 vs. 1.07±0.17; p<0.05) were 50% lower in dopamine neurons from obesity-prone pups. Since TH and VMAT2 essentially regulate dopamine quantal size, we conclude that mesolimbic TH levels and VMAT2 transporter site number may predict obesity predisposition in neonates. Furthermore, different forms of obesity are linked to low levels of mesolimbic dopamine, which the obese animal may attempt to overcome through excess feeding. Supported by DK065872.