Jay J. Kuo

Inhibition of NO Synthesis Enhances Chronic Cardiovascular and Renal Actions of Leptin

Acute studies suggest that leptin has pressor and depressor actions, including stimulation of sympathetic activity as well as increased release of NO from the vascular endothelium. The goal of this study was to examine the role of NO in modulating the chronic blood pressure, heart rate, and renal responses to hyperleptinemia, comparable to that found in obesity-induced hypertension. Male Sprague-Dawley rats were implanted with arterial and venous catheters, and mean arterial pressure and heart rate were monitored continuously 24 h/d. After a 4-day control period, the rats were infused with isotonic saline vehicle (n=6) or NG-nitro-l-arginine methyl ester (L-NAME, 10 &mgr;g/kg per minute; n=9) to inhibit NO synthesis for 7 days. After 7 days of vehicle or L-NAME administration, leptin was infused intravenously for 7 days at a rate of 0.5 &mgr;g/kg per minute, followed by a leptin infusion at 1.0 &mgr;g/kg per minute for 7 days, along with vehicle or L-NAME. A 21-day infusion of L-NAME alone (n=6) served as a control for the L-NAME+leptin rats. Although the low dose of leptin alone did not significantly elevate arterial pressure, it raised the heart rate by 18±3 bpm. The higher leptin infusion rate raised arterial pressure from 96±3 to 104±3 mm Hg but did not increase the heart rate further. L-NAME+leptin increased arterial pressure by 40±6 mm Hg and heart rate by 79±19 bpm compared with pretreatment levels. In control L-NAME rats, mean arterial pressure increased by 31±4 mm Hg, whereas the heart rate was not altered significantly compared with pretreatment levels. Neither chronic leptin infusion alone nor L-NAME alone altered the glomerular filtration rate or renal plasma flow significantly, but L-NAME+leptin reduced glomerular filtration rate by 27±11% and renal plasma flow by 47±9%. These results indicate that impaired NO synthesis mildly enhances the chronic renal hemodynamic and hypertensive effects of leptin but markedly amplifies the tachycardia caused by hyperleptinemia.

Role of Hypothalamic Melanocortin 3/4-Receptors in Mediating Chronic Cardiovascular, Renal, and Metabolic Actions of Leptin

The present study examined whether blockade of melanocortin receptors subtypes 3 and 4 (MC3/4-R) inhibits chronic cardiovascular and dietary responses to leptin infusion. A cannula was placed in the lateral ventricle of male Sprague-Dawley rats for chronic intracerebroventricular (ICV) infusion via osmotic minipump, and arterial and venous catheters were implanted for measurement of mean arterial pressure (MAP) and heart rate (HR) 24 h/d and IV infusions. After a 5-day control period, rats received (1) 0.9% saline vehicle ICV for 12 days plus leptin (1 μg/kg per minute IV, n=5) during the final 7 days; (2) MC3/4-R antagonist SHU-9119 (1 nmol/h ICV) for 12 days plus leptin (1 μg/kg per minute IV, n=6) during the final 7 days; and (3) SHU-9119 (1 nmol/h ICV, n=8) for 12 days. Leptin infusion in vehicle-treated rats caused a small increase in MAP (5±1 mm Hg) despite reduced food intake (23±1 to 10±1 g/d) and decreased body weight (−6%±1%). SHU-9119 infusion completely prevented the cardiovascular and dietary actions of leptin, leading to increased food intake (23±1 to 49±4 g/d) and body weight (+30%±2%), markedly decreased HR (−77±9 bpm), and caused a decrease in MAP (−6±1 mm Hg). Similar results were observed when SHU-9119 was infused alone in vehicle-treated rats. Leptin decreased plasma insulin to 30% of control values, an effect that was also abolished by SHU-9119 treatment, which caused a 5-fold increase in plasma insulin concentration. Thus, MC3/4-R antagonism completely blocked the chronic cardiovascular, satiety, and metabolic effects of leptin, suggesting that the hypothalamic melanocortin system plays an important role in mediating these actions of leptin.

Hypothalamic Melanocortin Receptors and Chronic Regulation of Arterial Pressure and Renal Function

Abstract—This study examined control of cardiovascular and renal function during chronic melanocortin-3/4 receptor (MC3/4-R) activation or inhibition. Arterial and venous catheters were implanted in Sprague-Dawley rats for measurements of mean arterial pressure (MAP) and heart rate (HR) 24 h/d and for intravenous infusions, and the lateral ventricle was cannulated for chronic intracerebroventricular (ICV) infusions. In experiment 1, after a 5-day control period, rats were administered the MC3/4-R agonist MTII (n=7, 10 ng/h ICV) or 0.9% saline (n=6, ICV) for 14 days, followed by a 5-day recovery period. In experiment 2, after a 5-day control period, rats were administered the MC3/4-R antagonist SHU-9119 (n=7, 1 nmol/h ICV) or 0.9% saline vehicle (n=7, ICV), or pair-fed during SHU-9119 infusion (n=5, 1 nmol/h ICV) for 12 days, followed by a 5-day recovery period. MC4-R activation transiently decreased food intake from 23±1 to 10±2 g/d. Despite the hypophagia, MC3/4-R activation increased MAP by 7±1 mm Hg. MC3/4-R inhibition for 12 days increased food intake from 21±1 to 35±4 g/d, decreased HR by 53±11 bpm, and caused no change in MAP despite the marked weight gain. In rats that were pair-fed to prevent increased food intake, MC3/4-R inhibition further decreased HR (−87±9 bpm), whereas MAP was unchanged. Thus, chronic hypothalamic MC3/4-R activation raises arterial pressure despite decreased food intake, whereas MC3/4-R inhibition causes marked weight gain without raising arterial pressure. These observations are consistent with the hypothesis that an intact hypothalamic MC3/4-R may be necessary for excess weight gain to raise arterial pressure.

Role of Adrenergic Activity in Pressor Responses to Chronic Melanocortin Receptor Activation

Abstract—Acute studies have shown that MC3/4-R stimulation increases sympathetic activity, but the role of adrenergic activation in mediating the cardiovascular and renal responses to chronic melanocortin 3- and 4-receptor (MC3/4-R) activation is unknown. The present study tested whether chronic MC3/4-R activation raises blood pressure and whether these changes are attenuated by &agr;1+&bgr;-adrenergic blockade. Rats were instrumented with an intracerebroventricular (ICV) cannula and arterial and venous catheters for measurements of mean arterial pressure (MAP) and heart rate (HR) 24 hours per day, and intravenous infusions. After control measurements, rats were intravenously infused with either saline vehicle (n=7) or &;1+ &bgr;-adrenergic antagonists (n=6, terazosin+propranolol, 10 mg/kg per day each) for 21 days. Five days after starting the vehicle or adrenergic blockade, the MC3/4-R agonist, MTII (10 ng/h), was infused ICV for 11 days followed by a 5-day recovery period. Another group of rats was infused with the adrenergic antagonists for 21 days but received the saline vehicle ICV for 11 days (n=7). MC3/4-R activation decreased food intake from 21±1 to 8±2 g/d by day 3 of MC3/4-R activation, and increased MAP and HR by an average of 8±2 mm Hg and 9±5 bpm, respectively. Adrenergic blockade did not alter the MC3/4-R-mediated decrease in food intake but abolished the increases in MAP and HR (1±2 mm Hg and −12±5 bpm, respectively, compared with control). ICV vehicle infusion during adrenergic blockade did not alter food intake or MAP. Glomerular filtration rate was unchanged in both the vehicle-infused and adrenergic blocked rats during MC3/4-R activation. These results indicate that the chronic actions of MC3/4-R activation on MAP and HR are mediated by adrenergic activation.

Role of Endothelin-1 in Blood Pressure Regulation in a Rat Model of Visceral Obesity and Hypertension

Abstract—Endothelial dysfunction has been suggested to play an important role in the development of obesity-induced hypertension. Because endothelin release increases in response to endothelial damage, we examined whether endothelin-1 contributes to increased arterial pressure in a model of visceral obesity produced by feeding Sprague-Dawley rats a high-fat (HF) diet (40% fat w/w, n=6) for 12 months. Arterial and venous catheters were implanted for measurement of mean arterial pressure (MAP) and heart rate (HR) 24 hours per day and intravenous infusions. After a 5-day control period, rats were infused with the selective endothelin-1 type A receptor (ET-A) blocker ABT-627 (2.5 mg/kg per day, IV) for 9 days, followed by a recovery period. Rats fed a standard chow (normal fat, or NF, group: n=6) for 12 months were also infused with ET-A blocker and were used as controls. Compared with NF rats, HF rats had higher MAP (113±4 versus 98±2 mm Hg), increased visceral fat (18.7±2.0 versus 10.8±1.4 g), and 3.2-fold increase in plasma leptin despite similar total body weight gain. Long-term ET-A blockade markedly reduced MAP in HF (−14±3 mm Hg) and NF (−14±2 mm Hg), but it had no effect on HR, GFR, or PRA. These results indicate that a long-term HF diet may cause visceral obesity and increased MAP, even in the absence of major changes in total body weight. Endothelin-1 appears to play an important role in the maintenance of arterial pressure in rats fed HF and NF diets, but it does not appear to contribute to increased MAP in this model of diet-induced hypertension.

Does Obesity Induce Resistance to the Long-Term Cardiovascular and Metabolic Actions of Melanocortin 3/4 Receptor Activation?

Previous studies suggest that blockade of melanocortin 3 and 4 receptors (MC3/4-R) markedly attenuates the chronic hypertensive effects of leptin. Although obesity has been reported to be associated with leptin “resistance,” it is unclear whether obesity alters the cardiovascular and metabolic effects of chronic MC3/4-R activation. Therefore, we tested whether the cardiovascular and metabolic actions of MC3/4-R activation are attenuated in Sprague-Dawley rats fed a high-fat diet (HF, n=6) compared with rats fed a standard chow (NF, n=6) for 12 months. A 21G steel cannula was placed in the lateral ventricle for ICV infusion, and arterial and venous catheters were implanted for measurement of mean arterial pressure (MAP) 24 hours/day and IV infusions. After a 5-day control period, rats were infused with MC3/4-R agonist melanotan II (10 ng/h, ICV), for 10 days followed by a 5-day recovery period. HF rats were heavier (558±21 versus 485±13 g) with 140% more visceral fat than NF rats, hyperleptinemic (8.9±0.5 versus 2.7±0.5 ng/mL), and insulin resistant. HF rats also had higher MAP (109±3 versus 100±1 mm Hg). Chronic melanotan II infusion significantly increased MAP in HF and NF (7±2 and 6±1 mm Hg), decreased caloric intake (−32±2 and −25 ±2 kcal/day), and reduced insulin levels in both groups by ≈50%. Thus, the metabolic and cardiovascular actions of chronic MC3/4-R activation are preserved in diet-induced obesity, supporting a potential role for the hypothalamic melanocortin system in obesity hypertension.

Cardiovascular, Renal, and Metabolic Responses to Chronic Central Administration of Agouti-Related Peptide

Although excess hypothalamic agouti-related peptide (AGRP), an endogenous antagonist of the melanocortin 3/4 receptor, causes hyperphagia and obesity, its role in regulating cardiovascular function is unclear. This study examined control of mean arterial pressure (MAP), heart rate (HR), and metabolism during chronic central administration of AGRP in rats. A cannula was placed in the lateral ventricle for intracerebroventricular infusion, and arterial and venous catheters were implanted for monitoring MAP and HR 24 hours per day, as well as intravenous infusions. After a 5-day control period, rats received AGRP (n=6; 0.02 nmol per hour ICV) or artificial cerebrospinal fluid (aCSF; n=9; 0.02 nmol per hour ICV) for 12 days, followed by a 5-day recovery period. A third group was infused intracerebroventricularly with AGRP and pair-fed to match food intake of control rats (n=7). AGRP produced a peak decrease in MAP and HR of −7±2 mm Hg and −68±7 bpm, respectively, despite increased food intake (from 23±0.5 to 36±3 g per day) and weight gain (from 350±8 to 454±5 g). AGRP also increased glomerular filtration rate, plasma insulin, glucose, and leptin. AGRP infusion in pair-fed rats produced a peak decrease in HR of −70±8 bpm but did not alter MAP or other variables. The metabolic effects of AGRP may be secondary to hyperphagia because they were abolished in pair-fed rats. aCSF infusion did not change any of the variables studied. These results demonstrate that increased central nervous system AGRP levels produce chronic reductions in MAP and HR despite marked increases in food intake and weight gain that normally tend to raise arterial pressure.