Burnett, John C.

Endothelin (ET) is a potent vasoconstrictor and sodium-regulating peptide whose tissue and plasma concentrations are increased in congestive heart failure (CHF). ET may mediate its vasoconstrictor and sodium-regulatory actions secondary to an increase in intracellular calcium. Calcium influx may augment ET synthesis. Although felodipine, a dihydropyridine calcium-channel antagonist, is effective in reducing vascular resistance in generalized vasoconstriction, its actions in CHF on circulating and local tissue ET remain undefined. The current studies were designed to determine the modulating actions of felodipine (oral, 40 mg/day for 7 days; n= 6) in an experimental canine model of CHF produced by chronic thoracic inferior vena caval constriction (TIVCC) compared with normal ( n = 7) and TIVCC-alone ( n = 7) dogs. We hypothesized that felodipine would decrease circulating and renal ET. Plasma ET was significantly increased in TIVCC compared with normal dogs (26 ± 0.5 vs. 12 ± 0.7 pg/ml, P < 0.05) and was markedly decreased by felodipine compared with TIVCC alone (14 ± 3 vs. 26 ± 0.5 pg/ml, P < 0.05). Renal ET immunohistochemical staining demonstrated the presence of ET in normal kidney, which was markedly increased in renal cortex and medulla in TIVCC dogs. Renal cortical and medullary ET staining densities were markedly decreased with felodipine compared with those with TIVCC alone. In the TIVCC + felodipine group, cardiovascular hemodynamics also was markedly improved compared with the TIVCC-alone group [systemic vascular resistance: 27 ± 2 vs. 44 ± 3 resistance units (RU), P < 0.05; pulmonary vascular resistance: 3.3 ± 0.1 vs. 5.7 ± 0.4 RU, P < 0.05; cardiac output: 2.9 ± 0.2 vs. 1.7 ± 0.1 l/min, P < 0.05]. This study demonstrates important modulating inhibitory actions of felodipine on renal and plasma ET in an experimental model of CHF.

Myocardial actions of the vasodilator peptide adrenomedullin (ADM) in the intact animal are unknown. Negative and positive inotropic actions have been reported in ex vivo experiments. Myocardial and load-altering actions of ADM in dogs before and after development of heart failure were studied. With controlled heart rate (atrial pacing) and after β-blockade, ADM was administered to five normal dogs in doses of 20 ng · kg−1 · min−1iv, 100 ng · kg−1 · min−1iv, and 200 ng · kg−1 · min−1into the left ventricle (LV). LV peak systolic pressure and end-systolic volume decreased with each dose of ADM. End-systolic pressure decreased with the two higher doses. At the highest dose, arterial elastance and the time constant of LV isovolumic relaxation (τ) decreased, and LV end-systolic elastance ( Ees) increased. LV end-diastolic pressure and volume were unchanged. In five additional normal dogs receiving only the highest dose of ADM (200 ng · kg−1 · min−1intra-LV), to control for increased heart rate and sympathetic activation observed with the cumulative infusion, ADM produced arterial vasodilation but no change in Eesor τ. In four dogs with pacing-induced heart failure, ADM (200 ng · kg−1 · min−1intra-LV) was without effect on τ, Ees, and systolic or diastolic pressure and volume. In vivo, ADM appears to be a selective arterial dilator without inotropic or lusitropic effects. The vasodilatory actions are attenuated in heart failure.

C-type natriuretic peptide (CNP) is an endothelium-derived peptide that shares structural homology with atrial natriuretic peptide (ANP). CNP causes greater endothelium-independent relaxations in veins compared with arteries. Relaxations to CNP in porcine coronary arteries are mediated by hyperpolarization of the smooth muscle membrane. Experiments were designed to investigate the mechanism(s) by which CNP causes relaxation in canine femoral veins. Rings of canine femoral veins without endothelium were suspended for measurement of isometric force in organ chambers. Concentration-response curves to CNP were obtained in veins contracted with either endothelin-1 (10(-8) M), KCl (40 mM), phenylephrine (10(-6) M) or prostaglandin F2 alpha (2 x 10(-6) M) in the absence and presence of BQ-123 (10(-6) M), NG-monomenthyl-L-arginine (L-NMMA; 10(-4) M), HS-142-1 (10(-5) M), methylene blue (10(-5) M), or potassium channel blockers, tetraethylammonium chloride (TEA; 10(-3) M), charybdotoxin (10(-7) M), glibenclamide (10(-7) M), or apamin (10(-7) M). Relaxations to CNP were significantly attenuated when the tissue was contracted with KCl and endothelin-1. During contraction to either phenylephrine or prostaglandin F2 alpha, relaxations to CNP were inhibited by HS-142-1, methylene blue, TEA, and charybdotoxin, but not by L-NMMA, glibenclamide, or apamin. In separate experiments, guanosine 3',5'-cyclic monophosphate increased twofold within 10-60 s after the addition of CNP (10(-8) M). These data suggest that CNP mediates relaxation of canine femoral veins through activation of large-conduction, calcium-activated potassium channels and activation of particulate and soluble guanylate cyclase.

Although brain natriuretic peptide (BNP) of myocardial origin is important in cardiovascular and renal function and as a marker of cardiac dysfunction, the expression of BNP in atrial and ventricular myocardium remains controversial both under normal conditions and in heart failure. We therefore determined left atrial and left ventricular (LV) gene expression and tissue concentration as well as circulating BNP during the evolution of rapid ventricular pacing-induced congestive heart failure (CHF) in the dog. Early LV dysfunction after 10 days of pacing was characterized by impaired LV function but maintained arterial pressure, and overt CHF after 38 days of pacing was characterized by further impaired LV function and decreased systemic arterial pressure. Under normal conditions, cardiac BNP mRNA and cardiac tissue BNP were of atrial origin. In early LV dysfunction, BNP mRNA and tissue BNP were markedly increased in the left atrium in association with an increase in circulating BNP but remained below or at the limit of detection in the LV. In overt CHF, BNP mRNA was further increased in the left atrium and first increased in the LV, together with an increase in LV tissue BNP and a further increase in circulating BNP. In the progression of CHF, early LV dysfunction is characterized by a selective increase in atrial BNP expression in association with increased circulating BNP. Overt CHF is characterized by an additional recruitment of ventricular BNP expression and a further increase in circulating BNP. These studies provide important new insight into the local and temporal regulation of cardiac BNP gene expression during the progression of heart failure and underscore the predominant endocrine role of atrial myocardium under normal conditions and in early LV dysfunction.

Ventricular concentrations of atrial, brain (BNP) and C-type natriuretic peptide are enhanced in congestive heart failure (CHF). Natriuretic peptide receptors are present on ventricular myocytes and stimulate guanosine 3′,5′-cyclic monophosphate (cGMP) production. cGMP has been demonstrated to affect myocyte function in vitro. Thus we hypothesized that the intracardiac natriuretic peptide system may modulate myocardial and coronary function in CHF. To test this hypothesis, the effects of an intracoronary infusion of the natriuretic peptide receptor antagonist HS-142–1 on ventricular and coronary function were examined in anesthetized dogs with chronic CHF. To determine whether receptor stimulation had contrasting effects to those of receptor blockade, intracoronary BNP was infused in anesthetized normal and CHF dogs. Low-dose HS-142–1 delayed and slowed left ventricular (LV) relaxation and decreased coronary blood flow without changes in LV pressures. Higher doses further impaired LV relaxation without further decreases in coronary blood flow. In normal and CHF dogs, exogenous BNP produced the opposite effect with a quicker onset and faster rate of LV relaxation without effects on LV pressures or coronary blood flow. The endogenous natriuretic peptide system has an autocrine-paracrine role to modulate LV and coronary vascular function in CHF.

The present study was undertaken to investigate the presence of C-type natriuretic peptide (CNP) immunoreactivity in cultured human vascular endothelial cells and in human plasma. CNP immunoreactivity was present in cultured human aortic endothelial cells by both immunohistochemical staining and by radioimmunoassay. With the utilization of gel permeation chromatography, this immunoreactivity proved to be consistent with the higher molecular weight CNP-53. CNP immunoreactivity was also present in human plasma (n = 22) at low picogram concentrations (6.5 +/- 0.2 pg/ml) by specific radioimmunoassay. This immunoreactivity was consistent with the lower molecular weight CNP-22 by gel permeation chromatography. These findings suggest that the vascular endothelium may be the site of CNP production. The isolation of different molecular forms of CNP in tissue and plasma may be consistent with a storage form of the peptide in endothelial cells CNP-53, while CNP-22 circulates in plasma. In summary, the present study is consistent with CNP being a peptide of endothelial cell origin.

C-type natriuretic peptide (CNP) is a hormone that shares structural homology to atrial natriuretic peptide (ANP); however, distinct receptors have been found for each in cultured aortic endothelial and smooth muscle cells. CNP in vivo reduces arterial pressure, atrial pressures, and cardiac output. These actions are consistent with a decrease in cardiac preload. Therefore, the current studies were designed to test the hypothesis that CNP is a vasodilator distinct from ANP. Rings of canine renal and saphenous arteries and renal, saphenous, and femoral veins with and without endothelium were suspended to measure isometric force in an organ chamber. CNP caused significant concentration-dependent relaxations in veins with and without endothelium contracted with phenylephrine (10(-6) log M). In marked contrast, ANP caused no significant relaxation in veins either with or without endothelium. In arterial rings, responses to the peptides were heterogeneous. ANP caused relaxation in renal but not saphenous arteries. CNP induced modest and comparable relaxation in rings of saphenous but not renal arteries with and without endothelium. These results demonstrate that: 1) CNP and not ANP is a relaxing factor of isolated peripheral canine veins, and 2) the responses of arteries to ANP and CNP are heterogeneous. These findings support a distinct biological role for CNP in the regulation of cardiovascular tone.

Congestive heart failure (CHF) is a pathophysiological condition associated with increased plasma levels of atrial natriuretic factor (ANF), a peptide hormone of cardiac origin that participates in the homeostatic control of intravascular volume and vascular tone. Atrial myocytes serve as the principal source of ANF under physiological conditions, although recent studies have demonstrated that ventricular myocardium may also synthesize ANF in models of CHF associated with ventricular hypertrophy. The current study was designed to investigate the roles of atrial and ventricular myocardium to synthesize, store, and release ANF during the evolution of tachycardia-induced CHF in the dog. The present study demonstrates a persistent elevation of plasma ANF during the evolution of CHF. In acute CHF (3 h), plasma ANF increased independent of cardiac ANF synthesis. In chronic CHF (15 and 30 days), plasma ANF is maintained by an increase in atrial synthesis and release of the peptide, without recruitment of ventricular ANF synthesis. The present study demonstrates that in acute CHF the increase in plasma ANF is regulated by release of stored peptide, and in chronic CHF the persistent elevation of plasma ANF is maintained by an increase in atrial synthesis of ANF.

C-type natriuretic peptide (CNP) is a newly described 22-amino acid peptide of endothelial cell origin, which has selective cardiovascular actions and is structurally related to atrial natriuretic peptide (ANP). Recent in vitro studies have demonstrated that an important regulatory pathway for the clearance of natriuretic peptides involves binding to a common clearance receptor [natriuretic peptide C receptor (NPR-C)]. Although CNP has also been identified as a ligand for NPR-C in binding assays, no studies have defined the in vivo interaction of CNP with NPR-C. CNP (10 ng.kg-1.min-1) followed by C-ANP-(4-23), a specific ligand for NPR-C blockade, was infused intravenously in two groups (both n = 7) of anesthetized dogs at two different doses (0.1 or 1.0 micrograms.kg-1.min-1) to permit calculation of total metabolic clearance rate (TMCR). C-ANP-(4-23) increased circulating CNP and reduced TMCR in both groups. Pulmonary metabolic clearance rate was negative at baseline, suggesting a net secretion of CNP across the lung, which was increased during CNP infusion and was abolished with NPR-C blockade. Renal and femoral metabolic clearance rates were positive at baseline and increased with CNP infusion. A decrease in cardiac output and cardiac filling pressures in response to CNP administration was potentiated by NPR-C blockade. We conclude that 1) circulating CNP achieved by CNP infusion is regulated by NPR-C in vivo, 2) the pulmonary circulation is a possible site of CNP secretion, 3) the renal and peripheral circulations are sites of CNP clearance, and 4) NPR-C blockade potentiates the selective cardiovascular actions of CNP.

The endothelium is the production site of several potent vasoactive substances that modulate vascular tone and growth. The present study was undertaken to investigate the presence and secretion of atrial natriuretic peptide (ANP) immunoreactivity from vascular endothelial cells. ANP immunoreactivity was present in cultured human aortic endothelial cells by both immunohistochemical staining and radioimmunoassay. ANP immunoreactivity was also detectable in culture medium from human aortic endothelial cells in low picogram concentrations. These findings suggest that vascular endothelium is a site of ANP production and secretion of ANP. There was a differential distribution of ANP and endothelin-1 (ET-1), with a higher ANP concentration in cell extracts and a higher ET-1 concentration in cell culture media. Although ANP has been conceived as a circulating endocrine hormone, these findings are consistent with ANP functioning also as an autocrine and paracrine modulator in the regulation of vascular tone and growth.