Low epinephrine levels and selective deficiency of ß2-adrenoceptor vasodilation at birth.

AIMS: Epinephrine is unique among biogenic catecholamines as a potent agonist of ß2-adrenoceptors. The ß2-adrenoceptor mediated effects during development might be linked to the increase of epinephrine synthesis. Our purpose was to characterize ß-adrenoceptor-mediated relaxation in the aorta of newborn and young rabbits (3 to 4months old), and to relate those responses with the epinephrine content of the adrenal gland. MAIN METHODS: The epinephrine levels and the tyrosine hydroxylase activity were determined in adrenal glands of newborn and young rabbits. Also, concentration-response curves to phenylephrine (selective α1-adrenoceptor agonist), dobutamine (selective ß1-adrenoceptor agonist), terbutaline (selective ß2-adrenoceptor agonist), and CL 316243 (selective ß3-adrenoceptor agonist) were determined in isolated aortic rings obtained from both groups. KEY FINDINGS: The adrenal gland content and the plasma concentrations of epinephrine were lower in newborn than in young rabbits. In contrast, the tyrosine hydroxylase activity was higher in newborn than in young rabbits. On the other hand, the maximal response to phenylephrine was lower in newborn than in young rabbits. Terbutaline at concentrations selective for ß2-adrenoceptors had no relaxing effects in neonates, in contrast to young rabbits. The potency and the maximal response of neither dobutamine nor CL 316243 were significantly different between the two groups. SIGNIFICANCE: In rabbits, as well as in humans, ß2-adrenoceptor-mediated responses and epinephrine synthesis are both immature at birth. On the other hand, the ß1 and ß3-adrenoceptor-mediated responses are fully developed. We conclude that epinephrine may influence the development of the ß2-adrenoceptor-mediated responses at birth and the rabbit is an excellent model to study these issues.

The role of adenylyl cyclase isoform 6 in ß-adrenoceptor signalling in murine airways.

BACKGROUND AND PURPOSE: Adenylyl cyclase (AC) is a key signalling enzyme for many GPCRs and catalyses the conversion of ATP to cAMP which, in turn, is a crucial determinant of many biological responses. ß-Adrenoceptor agonists are prescribed as bronchodilators for asthma and chronic obstructive pulmonary disease, and it is commonly assumed that they elicit their actions via AC-dependent production of cAMP. However, empirical evidence in support of this is lacking and the exact mechanism by which these drugs acts remains elusive. This is partly due to the existence of at least 10 different isoforms of AC and the absence of any truly selective pharmacological inhibitors. Here, we have used genetically modified mice and model systems to establish the role of AC isoforms in the airway responses to ß-adrenoceptor agonists. EXPERIMENTAL APPROACH: Receptors mediating responses to ß-adrenoceptor agonists in airway smooth muscle (ASM) and sensory nerve were identified in isolated tissue systems. Expression of mRNA for the AC isoforms in ASM and neurones was determined by qPCR. Functional responses were assessed in AC isoform KO mice and wild-type controls. KEY RESULTS: Airway and vagal tissue expressed mRNA for various isoforms of AC. AC6 was the most prominent isoform. Responses to ß-adrenoceptor agonists in tissues from AC6 KO mice were virtually abolished. CONCLUSIONS AND IMPLICATIONS: AC6 played a critical role in relaxation of ASM to ß1 -adrenoceptor agonists and in modulation of sensory nerves by ß1-3 -adrenoceptor agonists. These results further unravel the signalling pathway of this extensively prescribed class of medicine.

Profound and rapid reduction in body temperature induced by the melanocortin receptor agonists.

The melanocortin receptor 4 (MC4R) plays a major role in body weight regulation and its agonist MTII has been widely used to study the role of MC4Rs in energy expenditure promotion and feeding reduction. Unexpectedly, we observed that intraperitoneal (i.p.) administration of MTII induced a rapid reduction in both body temperature and energy expenditure, which was independent of its effect on feeding and followed by a prolonged increase in energy expenditure. The rapid reduction was at least partly mediated by brain neurons since intracerebroventricular (icv) administration of alpha melanocyte-stimulating hormone, an endogenous melanocortin receptor agonist, produced a similar response. In addition, the body temperature-lowering effect of MTII was independent of the presence of MC4Rs, but in a similar fashion to the previously shown effect on body temperature by 5'AMP. Moreover, ß-adrenergic receptors (ß-ARs) were required for the recovery from low body temperature induced by MTII and further pharmacological studies showed that the MTII's effect on body temperature may be partially mediated by the vasopressin V1a receptors. Collectively, our results reveal a previously unappreciated role for the melanocortin pathway in rapidly lowering body temperature.

ß2-Adrenergic agonists augment air pollution-induced IL-6 release and thrombosis.

Acute exposure to particulate matter (PM) air pollution causes thrombotic cardiovascular events, leading to increased mortality rates; however, the link between PM and cardiovascular dysfunction is not completely understood. We have previously shown that the release of IL-6 from alveolar macrophages is required for a prothrombotic state and acceleration of thrombosis following exposure to PM. Here, we determined that PM exposure results in the systemic release of catecholamines, which engage the ß2-adrenergic receptor (ß2AR) on murine alveolar macrophages and augment the release of IL-6. In mice, ß2AR signaling promoted the development of a prothrombotic state that was sufficient to accelerate arterial thrombosis. In primary human alveolar macrophages, administration of a ß2AR agonist augmented IL-6 release, while the addition of a beta blocker inhibited PM-induced IL-6 release. Genetic loss or pharmacologic inhibition of the ß2AR on murine alveolar macrophages attenuated PM-induced IL-6 release and prothrombotic state. Furthermore, exogenous ß2AR agonist therapy further augmented these responses in alveolar macrophages through generation of mitochondrial ROS and subsequent increase of adenylyl cyclase activity. Together, these results link the activation of the sympathetic nervous system by ß2AR signaling with metabolism, lung inflammation, and an enhanced susceptibility to thrombotic cardiovascular events.

Leucine deprivation stimulates fat loss via increasing CRH expression in the hypothalamus and activating the sympathetic nervous system.

We previously showed that leucine deprivation decreases abdominal fat mass largely by increasing energy expenditure, as demonstrated by increased lipolysis in white adipose tissue (WAT) and uncoupling protein 1 (UCP1) expression in brown adipose tissue (BAT). The goal of the present study was to investigate the possible involvement of central nervous system (CNS) in this regulation and elucidate underlying molecular mechanisms. For this purpose, levels of genes and proteins related to lipolysis in WAT and UCP1 expression in BAT were analyzed in wild-type mice after intracerebroventricular administration of leucine or corticotrophin-releasing hormone antibodies, or in mice deleted for three ß-adrenergic receptors, after being maintained on a leucine-deficient diet for 7 d. Here, we show that intracerebroventricular administration of leucine significantly attenuates abdominal fat loss and blocks activation of hormone sensitive lipase in WAT and induction of UCP1 in BAT in leucine-deprived mice. Furthermore, we provide evidence that leucine deprivation stimulates fat loss by increasing expression of corticotrophin-releasing hormone in the hypothalamus via activation of stimulatory G protein/cAMP/protein kinase A/cAMP response element-binding protein pathway. Finally, we show that the effect of leucine deprivation on fat loss is mediated by activation of the sympathetic nervous system. These results suggest that CNS plays an important role in regulating fat loss under leucine deprivation and thereby provide novel and important insights concerning the importance of CNS leucine in the regulation of energy homeostasis.

Direct control of peripheral lipid deposition by CNS GLP-1 receptor signaling is mediated by the sympathetic nervous system and blunted in diet-induced obesity.

We investigated a possible role of the central glucagon-like peptide (GLP-1) receptor system as an essential brain circuit regulating adiposity through effects on nutrient partitioning and lipid metabolism independent from feeding behavior. Both lean and diet-induced obesity mice were used for our experiments. GLP-1 (7-36) amide was infused in the brain for 2 or 7 d. The expression of key enzymes involved in lipid metabolism was measured by real-time PCR or Western blot. To test the hypothesis that the sympathetic nervous system may be responsible for informing adipocytes about changes in CNS GLP-1 tone, we have performed direct recording of sympathetic nerve activity combined with experiments in genetically manipulated mice lacking beta-adrenergic receptors. Intracerebroventricular infusion of GLP-1 in mice directly and potently decreases lipid storage in white adipose tissue. These effects are independent from nutrient intake. Such CNS control of adipocyte metabolism was found to depend partially on a functional sympathetic nervous system. Furthermore, the effects of CNS GLP-1 on adipocyte metabolism were blunted in diet-induced obese mice. The CNS GLP-1 system decreases fat storage via direct modulation of adipocyte metabolism. This CNS GLP-1 control of adipocyte lipid metabolism appears to be mediated at least in part by the sympathetic nervous system and is independent of parallel changes in food intake and body weight. Importantly, the CNS GLP-1 system loses the capacity to modulate adipocyte metabolism in obese states, suggesting an obesity-induced adipocyte resistance to CNS GLP-1.

Why we should put clothes on mice.

Mitochondrial uncoupling protein 1 (UCP1) is a key regulator of adaptive thermogenesis and energy expenditure. Mice lacking UCP1 are cold sensitive, but surprisingly not obese at room temperature. In this issue of Cell Metabolism, Feldmann et al. (2009) unmask an obesogenic phenotype by simply maintaining these mice at thermoneutrality.

Mice lacking beta-adrenergic receptors have increased bone mass but are not protected from deleterious skeletal effects of ovariectomy.

Activation of beta2-adrenergic receptors inhibits osteoblastic bone formation and enhances osteoclastic bone resorption. Whether beta-blockers inhibit ovariectomy-induced bone loss and decrease fracture risk remains controversial. To further explore the role of beta-adrenergic signaling in skeletal acquisition and response to estrogen deficiency, we evaluated mice lacking the three known beta-adrenergic receptors (beta-less). Body weight, percent fat, and bone mineral density were significantly higher in male beta-less than wild-type (WT) mice, more so with increasing age. Consistent with their greater fat mass, serum leptin was significantly higher in beta-less than WT mice. Mid-femoral cross-sectional area and cortical thickness were significantly higher in adult beta-less than WT mice, as were femoral biomechanical properties (+28 to +49%, P < 0.01). Young male beta-less had higher vertebral (1.3-fold) and distal femoral (3.5-fold) trabecular bone volume than WT (P < 0.001 for both) and lower osteoclast surface. With aging, these differences lessened, with histological evidence of increased osteoclast surface and decreased bone formation rate at the distal femur in beta-less vs. WT mice. Serum tartrate-resistance alkaline phosphatase-5B was elevated in beta-less compared with WT mice from 8-16 wk of age (P < 0.01). Ovariectomy inhibited bone mass gain and decreased trabecular bone volume/total volume similarly in beta-less and WT mice. Altogether, these data indicate that absence of beta-adrenergic signaling results in obesity and increased cortical bone mass in males but does not prevent deleterious effects of estrogen deficiency on trabecular bone microarchitecture. Our findings also suggest direct positive effects of weight and/or leptin on bone turnover and cortical bone structure, independent of adrenergic signaling.

Vagal tone dominates autonomic control of mouse heart rate at thermoneutrality.

It is generally accepted that cardiac sympathetic tone dominates the control of heart rate (HR) in mice. However, we have recently challenged this notion given that HR in the mouse is responsive to ambient temperature (T(a)) and that the housing T(a) is typically 21-23 degrees C, well below the thermoneutral zone ( approximately 30 degrees C) of this species. To specifically test the hypothesis that cardiac sympathetic tone is the primary mediator of HR control in the mouse, we first examined the metabolic and cardiovascular responses to rapid changes in T(a) to demonstrate the sensitivity of the mouse cardiovascular system to T(a). We then determined HR in 1) mice deficient in cardiac sympathetic tone ("beta-less" mice), 2) mice deficient in cardiac vagal tone [muscarinic M(2) receptor (M(2)R(-/-)) mice], and 3) littermate controls. At a T(a) of 30 degrees C, the HR of beta-less mice was identical to that of wild-type mice (351 +/- 11 and 363 +/- 10 beats/min, respectively). However, the HR of M(2)R(-/-) mice was significantly greater (416 +/- 7 beats/min), demonstrating that vagal tone predominates over HR control at this T(a). When these mice were calorically restricted to 70% of normal intake, HR fell equally in wild-type, beta-less, and M(2)R(-/-) mice (DeltaHR = 73 +/- 9, 76 +/- 3, and 73 +/- 7 beats/min, respectively), suggesting that the fall in intrinsic HR governs bradycardia of calorically restricted mice. Only when the T(a) was relatively cool, at 23 degrees C, did beta-less mice exhibit a HR (442 +/- 14 beats/min) that was different from that of littermate controls (604 +/- 10 beats/min) and M(2)R(-/-) mice (602 +/- 5 beats/min). These experiments conclusively demonstrate that in the absence of cold stress, regulation of vagal tone and modulation of intrinsic rate are important determinants of HR control in the mouse.

Effects of leptin on energy metabolism in beta-less mice.

OBJECTIVE: To investigate the impact of beta-adrenoceptor deficiency on the metabolic effects of leptin. MEASUREMENTS: Leptin was infused subcutaneously through an osmotic minipump in wild-type (WT) and beta(1)/beta(2)/beta(3)-adrenoceptor knockout (beta-less) mice and its effects on food intake, energy expenditure, carbohydrate and lipid utilization as well as on the levels of expression of the brown adipose tissue (BAT), thermogenic marker uncoupling protein-1 (UCP1) and type II deiodinase (D2) mRNAs were compared. RESULTS: Leptin treatment decreased food intake by 23% in both the WT and the beta-less mice. In pair-fed animals being used as controls, leptin treatment was found to increase energy expenditure in WT, but not in beta-less mice. No difference was observed in carbohydrate or fat utilization between leptin-treated WT and beta-less mice. Leptin increased UCP1 and D2 mRNA levels in WT mouse BAT 1.7- and 3-fold, respectively, but had no effect on the expression of these genes in beta-less mouse BAT. CONCLUSION: The stimulatory effects of leptin on oxygen consumption, BAT UCP1 and D2 expression require functional beta-adrenoceptors, but its inhibitory effect on food intake and its stimulatory effect on fat utilization is independent of the beta-adrenoceptor signalling.