Michael A. Cowley

Serotonin reciprocally regulates melanocortin neurons to modulate food intake

The neural pathways through which central serotonergic systems regulate food intake and body weight remain to be fully elucidated. We report that serotonin, via action at serotonin1B receptors (5-HT1BRs), modulates the endogenous release of both agonists and antagonists of the melanocortin receptors, which are a core component of the central circuitry controlling body weight homeostasis. We also show that serotonin-induced hypophagia requires downstream activation of melanocortin 4, but not melanocortin 3, receptors. These results identify a primary mechanism underlying the serotonergic regulation of energy balance and provide an example of a centrally derived signal that reciprocally regulates melanocortin receptor agonists and antagonists in a similar manner to peripheral adiposity signals.

Synaptic input organization of the melanocortin system predicts diet-induced hypothalamic reactive gliosis and obesity.

The neuronal circuits involved in the regulation of feeding behavior and energy expenditure are soft-wired, reflecting the relative activity of the postsynaptic neuronal system, including the anorexigenic proopiomelanocortin (POMC)-expressing cells of the arcuate nucleus. We analyzed the synaptic input organization of the melanocortin system in lean rats that were vulnerable (DIO) or resistant (DR) to diet-induced obesity. We found a distinct difference in the quantitative and qualitative synaptology of POMC cells between DIO and DR animals, with a significantly greater number of inhibitory inputs in the POMC neurons in DIO rats compared with DR rats. When exposed to a high-fat diet (HFD), the POMC cells of DIO animals lost synapses, whereas those of DR rats recruited connections. In both DIO rats and mice, the HFD-triggered loss of synapses on POMC neurons was associated with increased glial ensheathment of the POMC perikarya. The altered synaptic organization of HFD-fed animals promoted increased POMC tone and a decrease in the stimulatory connections onto the neighboring neuropeptide Y (NPY) cells. Exposure to HFD was associated with reactive gliosis, and this affected the structure of the blood-brain barrier such that the POMC and NPY cell bodies and dendrites became less accessible to blood vessels. Taken together, these data suggest that consumption of an HFD has a major impact on the cytoarchitecture of the arcuate nucleus in vulnerable subjects, with changes that might be irreversible due to reactive gliosis.

The arcuate nucleus mediates GLP-1 receptor agonist liraglutide-dependent weight loss

Liraglutide is a glucagon-like peptide-1 (GLP-1) analog marketed for the treatment of type 2 diabetes. Besides lowering blood glucose, liraglutide also reduces body weight. It is not fully understood how liraglutide induces weight loss or to what degree liraglutide acts directly in the brain. Here, we determined that liraglutide does not activate GLP-1-producing neurons in the hindbrain, and liraglutide-dependent body weight reduction in rats was independent of GLP-1 receptors (GLP-1Rs) in the vagus nerve, area postrema, and paraventricular nucleus. Peripheral injection of fluorescently labeled liraglutide in mice revealed the presence of the drug in the circumventricular organs. Moreover, labeled liraglutide bound neurons within the arcuate nucleus (ARC) and other discrete sites in the hypothalamus. GLP-1R was necessary for liraglutide uptake in the brain, as liraglutide binding was not seen in Glp1r(-/-) mice. In the ARC, liraglutide was internalized in neurons expressing proopiomelanocortin (POMC) and cocaine- and amphetamine-regulated transcript (CART). Electrophysiological measurements of murine brain slices revealed that GLP-1 directly stimulates POMC/CART neurons and indirectly inhibits neurotransmission in neurons expressing neuropeptide Y (NPY) and agouti-related peptide (AgRP) via GABA-dependent signaling. Collectively, our findings indicate that the GLP-1R on POMC/CART-expressing ARC neurons likely mediates liraglutide-induced weight loss.

Rational design of a combination medication for the treatment of obesity.

Existing obesity therapies are limited by safety concerns and modest efficacy reflecting a weight loss plateau. Here, we explore combination therapy with bupropion (BUP), a putative stimulator of melanocortin pathways, and an opioid antagonist, naltrexone (NAL), to antagonize an inhibitory feedback loop that limits sustained weight reduction. In vitro electrophysiologic experiments were conducted to determine the extent to which BUP+NAL stimulated hypothalamic pro‐opiomelanocortin (POMC) neurons in mouse brain. A subsequent study further characterized the effect of combination BUP+NAL treatment on food intake in lean and obese mice. Finally, a randomized, blinded, placebo‐controlled trial in obese adult subjects was conducted. Randomization included: BUP (300 mg) + NAL (50 mg), BUP (300 mg) + placebo (P), NAL (50 mg) + P or P+P for up to 24 weeks. BUP+NAL stimulated murine POMC neurons in vitro and caused a greater reduction in acute food intake than either monotherapy, an effect consistent with synergism. Combined BUP+NAL provided sustained weight loss without evidence of an efficacy plateau through 24 weeks of treatment. BUP+NAL completers diverged from NAL+P (P < 0.01) and P+P (P < 0.001) at week 16 and from BUP+P by week 24 (P < 0.05). The combination was also well tolerated. Translational studies indicated that BUP+NAL therapy produced synergistic weight loss which exceeded either BUP or NAL alone. These results supported the hypothesis that NAL, through blockade of β‐endorphin mediated POMC autoinhibition, prevents the classic weight loss plateau observed with monotherapies such as BUP. This novel treatment approach (BUP+NAL) holds promise for the treatment of obesity.

Leptin Resistance and Obesity

The prevalence of obesity, and the human and economic costs of the disease, creates a need for better therapeutics and better understanding of the physiological processes that balance energy intake and energy expenditure. Leptin is the primary signal from energy stores and exerts negative feedback effects on energy intake. In common obesity, leptin loses the ability to inhibit energy intake and increase energy expenditure; this is termed leptin resistance. This review discusses the evidence in support of leptin resistance in mouse models and humans and the possible mechanisms of leptin resistance.

Diet-Induced Obesity Causes Ghrelin Resistance in Arcuate NPY/AgRP Neurons

Circulating ghrelin is decreased in obesity, and peripheral ghrelin does not induce food intake in obese mice. We investigated whether ghrelin resistance was a centrally mediated phenomenon involving dysregulated neuropeptide Y (NPY) and agouti-related peptide (AgRP) circuits. We show that diet-induced obesity (DIO) (12 wk) suppresses the neuroendocrine ghrelin system by decreasing acylated and total plasma ghrelin, decreasing ghrelin and Goat mRNA in the stomach, and decreasing expression of hypothalamic GHSR. Peripheral (ip) or central (intracerebroventricular) ghrelin injection was able to induce food intake and arcuate nucleus Fos immunoreactivity in chow-fed but not high-fat diet-fed mice. DIO decreased expression of Npy and Agrp mRNA, and central ghrelin was unable to promote expression of these genes. Ghrelin did not induce AgRP or NPY secretion in hypothalamic explants from DIO mice. Injection of NPY intracerebroventricularly increased food intake in both chow-fed and high-fat diet-fed mice, indicating that downstream NPY/AgRP neural targets are intact and that defective NPY/AgRP function is a primary cause of ghrelin resistance. Ghrelin resistance in DIO is not confined to the NPY/AgRP neurons, because ghrelin did not stimulate growth hormone secretion in DIO mice. Collectively, our data suggests that DIO causes ghrelin resistance by reducing NPY/AgRP responsiveness to plasma ghrelin and suppressing the neuroendocrine ghrelin axis to limit further food intake. Ghrelin has a number of functions in the brain aside from appetite control, including cognitive function, mood regulation, and protecting against neurodegenerative diseases. Thus, central ghrelin resistance may potentiate obesity-related cognitive decline, and restoring ghrelin sensitivity may provide therapeutic outcomes for maintaining healthy aging.

Leptin Action in the Dorsomedial Hypothalamus Increases Sympathetic Tone to Brown Adipose Tissue in Spite of Systemic Leptin Resistance

Leptin regulates body weight in mice by decreasing appetite and increasing sympathetic nerve activity (SNA), which increases energy expenditure in interscapular brown adipose tissue (iBAT). Diet-induced obese mice (DIO) are resistant to the anorectic actions of leptin. We evaluated whether leptin still stimulated sympathetic outflow in DIO mice. We measured iBAT temperature as a marker of SNA. We found that obese hyperleptinemic mice have higher iBAT temperature than mice on regular diet. Conversely, obese leptin-deficient ob/ob mice have lower iBAT temperature. Additionally, leptin increased SNA in obese (DIO and ob/ob) and control mice, despite DIO mice being resistant to anorectic action of leptin. We demonstrated that neurons in the dorsomedial hypothalamus (DMH) of DIO mice mediate the thermogenic responses to hyperleptinemia in obese mammals because blockade of leptin receptors in the DMH prevented the thermogenic effects of leptin. Peripheral Melotan II (MTII) injection increased iBAT temperature, but it was blunted by blockade of DMH melanocortin receptors (MC4Rs) by injecting agouti-related peptide (AgRP) directly into the DMH, suggesting a physiological role of the DMH on temperature regulation in animals with normal body weight. Nevertheless, obese mice without a functional melanocortin system (MC4R KO mice) have an increased sympathetic outflow to iBAT compared with their littermates, suggesting that higher leptin levels drive sympathoexcitation to iBAT by a melanocortin-independent pathway. Because the sympathetic nervous system contributes in regulating blood pressure, heart rate, and hepatic glucose production, selective leptin resistance may be a crucial mechanism linking adiposity and metabolic syndrome.

Hypothalamic POMC neurons promote cannabinoid-induced feeding

Hypothalamic pro-opiomelanocortin (POMC) neurons promote satiety. Cannabinoid receptor 1 (CB1R) is critical for the central regulation of food intake. Here we test whether CB1R-controlled feeding in sated mice is paralleled by decreased activity of POMC neurons. We show that chemical promotion of CB1R activity increases feeding, and notably, CB1R activation also promotes neuronal activity of POMC cells. This paradoxical increase in POMC activity was crucial for CB1R-induced feeding, because designer-receptors-exclusively-activated-by-designer-drugs (DREADD)-mediated inhibition of POMC neurons diminishes, whereas DREADD-mediated activation of POMC neurons enhances CB1R-driven feeding. The Pomc gene encodes both the anorexigenic peptide α-melanocyte-stimulating hormone, and the opioid peptide β-endorphin. CB1R activation selectively increases β-endorphin but not α-melanocyte-stimulating hormone release in the hypothalamus, and systemic or hypothalamic administration of the opioid receptor antagonist naloxone blocks acute CB1R-induced feeding. These processes involve mitochondrial adaptations that, when blocked, abolish CB1R-induced cellular responses and feeding. Together, these results uncover a previously unsuspected role of POMC neurons in the promotion of feeding by cannabinoids.

Leptin mediates the increase in blood pressure associated with obesity

Summary Obesity is associated with increased blood pressure (BP), which in turn increases the risk of cardiovascular diseases. We found that the increase in leptin levels seen in diet-induced obesity (DIO) drives an increase in BP in rodents, an effect that was not seen in animals deficient in leptin or leptin receptors (LepR). Furthermore, humans with loss-of-function mutations in leptin and the LepR have low BP despite severe obesity. Leptin’s effects on BP are mediated by neuronal circuits in the dorsomedial hypothalamus (DMH), as blocking leptin with a specific antibody, antagonist, or inhibition of the activity of LepR-expressing neurons in the DMH caused a rapid reduction of BP in DIO mice, independent of changes in weight. Re-expression of LepRs in the DMH of DIO LepR-deficient mice caused an increase in BP. These studies demonstrate that leptin couples changes in weight to changes in BP in mammalian species.

Comparison of Combined Bupropion and Naltrexone Therapy for Obesity with Monotherapy and Placebo

CONTEXT The efficacy of current centrally acting obesity pharmacotherapies is limited by compensatory mechanisms that mitigate weight loss. OBJECTIVE Our objective was to determine whether opioid receptor antagonism (naltrexone) plus pro-opiomelanocortin activation (bupropion) causes greater weight loss than placebo or monotherapy. DESIGN/SETTING A randomized, placebo- and monotherapy-controlled, double-blind, dose-finding trial was conducted from August 2005 to December 2006 in seven U.S. outpatient clinics. PARTICIPANTS A total of 419 patients with uncomplicated obesity participated. INTERVENTIONS Interventions included 24 wk of sustained-release bupropion (400 mg/d), immediate-release naltrexone (48 mg/d), placebo, and three combination therapy [naltrexone/bupropion (NB)] groups consisting of immediate-release naltrexone, 16, 32, or 48 mg/d, plus sustained-release bupropion (400 mg/d) with a 24-wk extension. A minimal diet and exercise component was also included. MAIN OUTCOME MEASURES Percent weight change from baseline at wk 24 in the intent-to-treat population for NB48 vs. placebo and monotherapy was assessed. Other measurements included body mass index, waist circumference, fasting lipids, glycemic variables, safety, and tolerability. RESULTS At wk 24, placebo-subtracted weight loss was -4.62% [95% confidence interval (CI) -6.24 to -2.99; P < 0.001] for NB16, -4.65% (95% CI -6.20 to -3.09; P < 0.001) for NB32, and -3.53% (95% CI -5.15 to -1.90; P < 0.001) for NB48. Weight loss was statistically significant vs. monotherapy for all three NB combinations with the exception of NB48 vs. bupropion. Weight loss with NB continued after wk 24. The most common treatment-emergent adverse event was mild transient nausea. CONCLUSIONS NB caused gradual sustained weight loss over 48 wk; NB32 and NB16 demonstrated greater weight loss in the intent-to-treat population due to lower attrition rates. Further study is needed to demonstrate long-term efficacy and safety of NB.