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The heart is involved in the regulation of blood pressure and body fluid homeostasis. As a blood volume sensor and effector for the regulation of the volume and pressure homeostasis, the atria are the central regulator to secrete humoral messenger cardiac natriuretic hormones into the circulation. The primary action of the atria in response to the volume change in the body is to control the secretion of atrial natriuretic peptide (ANP), a member of the family of cardiac natriuretic hormones. Although all cardiac chambers are able to secrete ANP, the major source of the cardiac hormone is the atria until reactivation of the synthesis of the ventricles. In heart disease including hypertension and cardiac hypertrophy, ventricular ANP synthesis and plasma levels of ANP are increased. However, the roles of the atria for the ANP secretion are not well defined in hypertension or heart failure. Under the high concentration of plasma levels of ANP by compensatory and/or pathophysiological reactivation of the ventricular synthesis and release of ANP, with activation of the renin-angiotensin system and changes in the atrial distensibility, the roles of the atria should be reevaluated in the heart disease. The purpose of the present review is to address modulation of the atrial role in the regulation of ANP secretion and its significance in the pathological changes in hypertension and cardiac disease and to strengthen the importance of the role of the interstitial fluid dynamics of the atrial wall in the regulation of ANP secretion.

Background: Oryeongsan (ORS), a herbal medicine formula, has long been used for the treatment of impaired body water balance in Asian countries. Recently, it was shown that ORS modulates the renin-angiotensin system (RAS). Purpose of the present study was to determine characteristics of atrial ANP secretion and effects of ORS on the secretion in the atria from spontaneously hypertensive rats (SHR). Methods: WKY-V treated with vehicle, SHR groups treated with vehicle (SHR-V), ORS (SHR-ORS), and losartan as a positive control (SHR-LOS). Experiments were performed in perfused beating atria (1.3 Hz) allowing atrial distension, acetylcholine (ACh) stimulation, and serial collection of atrial perfusates. The secreted ANP concentration was measured using radioimmunoassay. Interstitial fluid (ISF) translocation was measured using [3H]inulin clearance. Results: Stepwise increase in atrial distension by 1.1, 2.0, and 2.7 cmH2O above basal distension further increased ANP secretion proportionally in the atria from WKY-V, but the response was significantly suppressed in the atria from SHR-V. Cardiomyocyte ANP release, the first step of atrial ANP secretion, was suppressed in the atria from SHR-V compared to those from WKY-V (-8.02 ± 2.86, -15.86 ± 2.27, and -20.09 ± 3.62%; n = 8, vs. 8.59 ± 2.81, 15.65 ± 7.14, and 38.12 ± 8.28%; n = 8; p < 0.001 for all stepwise distension, respectively). Subacute treatment with ORS reversed the suppressed ANP release in atria from SHR-ORS (6.76 ± 3.92, 9.12 ± 2.85, and 28.79 ± 1.79% for SHR-ORS; n = 5 vs. SHR-V; n = 8; p = 0.01, p < 0.001, p < 0.001, respectively). The effects of ORS were comparable to those of losartan. ISF translocation was not significantly different between groups except SHR-V. ACh-induced ANP secretion and cardiomyocyte ANP release were also suppressed in the atria from SHR-V and ORS reversed the suppression. These findings were accompanied with accentuation of the AT1 receptor expression and suppression of the AT2/Mas receptor, M2 mACh receptor and GIRK4 expression in the atria from SHR-V. Further, treatment with ORS or losartan reversed the expressions. Conclusions: These results show that ANP secretion is suppressed in the atria from SHR in association with accentuation of AT1 receptor and suppression of AT2/Mas receptor and KACh channel expression. ORS ameliorates impaired ANP secretion through improving cardiomyocyte ANP release with modulation of the cardiac RAS and muscarinic signaling. These findings provide experimental evidence which supports the role of ORS in the regulation of atrial ANP secretion in the atria from SHR.

To investigate whether the methanol extract of Berberis amurensis Rupr. (BAR) augments penile erection using in vitro and in vivo experiments. The ex vivo study used corpus cavernosum strips prepared from adult male New Zealand White rabbits. In in vivo studies for intracavernous pressure (ICP), blood pressure, mean arterial pressure (MAP), and increase of peak ICP were continuously monitored during electrical stimulation of Sprague-Dawley rats. Preconstricted with phenylephrine (PE) in isolated endotheliumintact rabbit corus cavernosum, BAR relaxed penile smooth muscle in a dose-dependent manner, which was inhibited by pretreatment with NG-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase inhibitor, and 1H-[1,2,4]-oxadiazole-[4,3-α]-quinoxalin-1-one, a soluble guanylyl cclase inhibitor. BAR significantly relaxed penile smooth muscles dose-dependently in ex vivo, and this was inhibited by pretreatment with L-NAME 1H-[1,2,4]-oxadiazole-[4,3-α]-quinoxalin-1-one. BAR-induced relaxation was significantly attenuated by pretreatment with tetraethylammonium (TEA, P

Background Homocysteine (Hcy) induced vascular endothelial dysfunction is known to be closely associated with oxidative stress and impaired NO system. 1,8-Dihydroxy-3-methoxy-6-methylanthracene-9,10-dione (physcion) has been known to has antioxidative and anti-inflammatory properties. Purpose The purpose of the present study was to define the protective effect of physcion on Hcy-induced endothelial dysfunction and its mechanisms involved. Study Design and Methods Hyperhomocysteinemia (HHcy) rat model was induced by feeding 3% methionine. A rat thoracic aortic ring model was used to investigate the effects of physcion on Hcy-induced impairment of endothelium-dependent relaxation. Two doses, low (L, 30 mg/kg/day) and high (H, 50 mg/kg/day) of physcion were used in the present study. To construct Hcy-injured human umbilical vein endothelial cells (HUVECs) model, the cells treated with 3 mM Hcy. The effects of physcion on Hcy-induced HUVECs cytotoxicity and apoptosis were studied using MTT and flow cytometry. Confocal analysis was used to determine the levels of intracellular Ca2+. The levels of protein expression of the apoptosis-related markers Bcl-2, Bax, caspase-9/3, and Akt and endothelial nitric oxide synthase (eNOS) were evaluated by western blot. Results In the HHcy rat model, plasma levels of Hcy and malondialdehyde (MDA) were elevated (20.45 ± 2.42 vs. 4.67 ± 1.94 μM, 9.42 ± 0.48 vs. 3.47 ± 0.59 nM, p Conclusion Physcion prevents Hcy-induced endothelial dysfunction by activating Ca2+- and Akt-eNOS-NO signaling pathways. This study provides the first evidence that physcion might be a candidate agent for the prevention of cardiovascular disease induced by Hcy.

In the present study, the change in secretion of atrial natriuretic peptide (ANP) from the atria was defined in hypertension accompanied by ventricular hypertrophy and increased synthesis of ANP. To identify the change of the secretion and mechanisms involved, experiments were performed in isolated perfused beating atria from sham-operated normotensive and renovascular hypertensive rats. Expression of ANP, natriuretic peptide receptor (NPR)-C, components of the renin-angiotensin system, and muscarinic signaling pathway was measured in cardiac tissues. Basal levels of ANP secretion and acetylcholine (ACh)- and stretch-induced activation of ANP secretion were suppressed in the atria from hypertensive compared with normotensive rats. ACh increased ANP secretion via M2 muscarinic ACh receptor-ACh-sensitive K+ channel signaling. In hypertensive rats, ANP concentration increased in the left ventricle but decreased in the right ventricle. The atrial concentration of ANP was not changed in hypertensive compared with normotensive rats. ANP mRNA expression was accentuated in the left ventricle but suppressed in the other cardiac chambers in the hearts of hypertensive rats. NPR-C expression was inversely related to ANP mRNA levels. Angiotensin II type 1 receptor (AT1R) expression was accentuated in the cardiac chambers from hypertensive rats compared with normotensive rats, whereas angiotensin II type 2 receptor, M2 muscarinic receptor, and Kir3.4 channels were suppressed. AT1R blockade with losartan reversed the change observed in hypertensive rats. The present findings indicate that renovascular hypertension shifts the major site of ANP secretion and synthesis from the atria to the left ventricle through modulation of the expression of ANP, NPR-C, AT1R, and the M2 muscarinic signaling pathway. NEW & NOTEWORTHY Renovascular hypertension suppresses the atrial secretion of ANP and shifts the major site of the regulation of ANP secretion and synthesis from atria to the hypertrophied left ventricle possibly via modulation of the expression of ANP, natriuretic peptide receptor-C, angiotensin II subtype 1 receptor, and M2 muscarinic signaling pathway.

Oleanolic acid is known to possess beneficial effects on the regulation of the cardiovascular homeostasis. However, the exact nature of the role of oleanolic acid on the regulation of body fluid balance and blood pressure homeostasis and its mechanisms involved are not well defined. Experiments were performed to identify the effects of oleanolic acid on the renin-angiotensin system and cardiac natriuretic hormone (ANP) system, and also renal function and blood pressure in normotensive and renovascular hypertensive rats. The change in the plasma levels of hormones and the expressions of renin, angiotensin II receptors, ANP, natriuretic peptide receptor-C, M2 muscarinic receptor and GIRK4 were determined in the kidney, heart and aorta. Oleanolic acid was administered orally for 1 or 3 weeks. Here, we found that oleanolic acid suppressed plasma levels of renin activity and aldosterone and intrarenal levels of renin and angiotensin II type 1 receptor expression and increased angiotensin II type 2 receptor in normotensive and hypertensive rats. Also, oleanolic acid increased plasma levels of ANP. Further, oleanolic acid suppressed angiotensin II type 1 receptor and natriuretic peptide receptor-C expression and increased angiotensin II type 2 receptor and ANP expression in the heart and aorta. Along with these changes, oleanolic acid accentuated urinary volume, electrolyte excretion and glomerular filtration rate in normotensive rats and suppressed arterial blood pressure in hypertensive rats. These findings suggest that beneficial effects of oleanolic acid on the cardiorenal system are closely associated with its roles on the renin-angiotensin system and cardiac natriuretic hormone system.

Ursolic acid has recently been reported to increase both atrial natriuretic peptide (ANP) secretion and mechanical dynamics in rabbit atria.The present study was designed to clarify the regulatory effects of ursolic acid on the β-adrenergic or muscarinic receptor-mediated changes in ANP secretory and contractile function allowing measurement of atrial dynamics such as pulse pressure, stroke volume, and cAMP efflux in isolated perfused beating rabbit atria.Pretreatment with ursolic acid significantly attenuated the isoproterenol (β-adrenergic agonist)-induced decrease in ANP secretion and increases in cAMP levels and atrial dynamics. Interestingly, ursolic acid concentration-dependently accentuated the acetylcholine-induced increase in ANP secretion and decrease in pulse pressure in the presence of isoproterenol (p0.001). These findings indicate that acetylcholine-induced increase in ANP secretion is potentiated by ursolic acid; furthermore, acetylcholine-induced decrease in atrial dynamics is also potentiated by ursolic acid, suggesting that ursolic acid regulates muscarinic receptor-mediated secretory and contractile responses in perfused beating rabbit atria.This implicates for the beneficial effects of ursolic acid in the regulation of cardiovascular and body fluid homeostasis.

Homocysteine (Hcy) is an independent risk factor in the development of cardiovascular diseases (CVD). Hyperhomocysteinemia (HHcy), induces the injury of vascular endothelial cells via oxidative stress. Oxymatrine (OMT), one of the main components of Sophora flavescens, has displayed anti-inflammatory, anti-oxidant and anti-apoptotic activity. However, the effect of OMT on the Hcy-induced endothelial injury is not clearly defined yet. The aim of this study was to determine the protective effect of OMT on the Hcy-induced endothelial injury and its mechanisms involved. Human umbilical vein endothelial cells (HUVECs) were cultured in vitro. Methyl thiazolyl tetrazolium assay (MTT), fluorescence staining, flow cytometry and western blotting were used in this study. OMT prevented the Hcy-induced toxicity and apoptosis in HUVECs. Moreover, OMT suppressed Hcy-induced increases in reactive oxygen species, lactate dehydrogenase, malondialdehyde levels and increased superoxide dismutase levels. OMT reversed the Hcy-induced decrease in the protein expression of nuclear factor erythroid-2-related factor 2 (Nrf2). In addition, OMT reversed the Hcy-induced apoptosis related biochemical changes such as decreased mitochondrial membrane potential and Bcl-2/Bax protein ratio, and increased protein expression of caspase-9 and caspase-3. Furthermore, OMT elevated the phosphorylation levels of Akt and eNOS, and the formation of nitric oxide (NO) in injured cells. These results suggest that OMT prevents Hcy-induced endothelial injury by regulating mitochondrial-dependent apoptosis and Akt-eNOS-NO signaling pathways concomitantly with accentuation of Nrf2 expression.

Exogenous acetylcholine (ACh) is known to stimulate atrial natriuretic peptide (ANP) secretion concomitantly with a decrease in atrial pulse pressure. However, the role of intrinsic ACh in the regulation of ANP secretion remains unknown. Recently, it was shown that nonneuronal and neuronal ACh is present in the cardiac atria. From this finding we hypothesize that endogenously released ACh is involved in the regulation of ANP secretion in an autocrine or paracrine manner in the atria. Experiments were performed in isolated beating rat atria. ANP was measured using radioimmunoassay. To increase the availability of the ACh in the extracellular space of the atrium, its degradation was inhibited with an inhibitor of acetylcholinesterase. Acetylcholinesterase inhibition with physostigmine increased ANP secretion concomitantly with a decrease in atrial dynamics in a concentration-dependent manner. Inhibitors of M2 muscarinic ACh receptor (mAChR), methoctramine, and ACh-activated K+ (KACh+) channels, tertiapin-Q, abolished the physostigmine-induced changes. The effects were not observed in the atria from rats treated with pertussis toxin. Furthermore, the physostigmine-induced effects were attenuated by an inhibitor of high-affinity choline transporter, hemicholinium-3, which is a rate-limiting step of ACh synthesis. Inhibitors of the mAChR signaling pathway and ACh synthesis also attenuated the basal levels of ANP secretion and accentuated atrial dynamics. These findings suggest that endogenously released ACh tonically stimulates ANP secretion from atrial cardiomyocytes via activation of M2 mAChR-Gi/o-KACh+ channel signaling. It is also suggested that the ACh-ANP signaling is implicated in cardiac physiology and pathophysiology.

Oleanolic acid is known to have beneficial effects on the regulation of cardiovascular homeostasis. The present study was designed to identify the effects of oleanolic acid on plasma levels and atrial synthesis and secretion of atrial natriuretic peptide (ANP). Experiments were performed in rats and isolated perfused beating rat atria. ANP was measured using a selective radioimmunoassay. ANP mRNA expression was measured using real-time quantitative polymerase chain reaction. Administration of oleanolic acid increased plasma ANP levels in a dose-related manner. Similarly, oleanolic acid increased atrial ANP content and ANP mRNA expression. To evaluate the effects of oleanolic acid on ANP secretion, atrial stretch and muscarinic acetylcholine receptor activation were applied to the atria from rats chronically treated with oleanolic acid. Baseline levels of ANP secretion were higher in the atria from rats treated with oleanolic acid compared to rats treated with vehicle. Furthermore, oleanolic acid treatment enhanced the stretch-induced increase in ANP secretion. Acetylcholine in the presence of isoproterenol increased ANP secretion. The acetylcholine-induced increase in ANP secretion was also enhanced in the atria from rats treated with oleanolic acid compared to atria from rats treated with vehicle. The present findings indicate that oleanolic acid increases plasma ANP levels via increased ANP synthesis and secretion in rats. It is proposed that an accentuation of the ANP system is involved in the beneficial effects of oleanolic acid on the regulation of cardiovascular homeostasis.

The endothelins (ET) peptide family consists of ET-1, ET-2, ET-3, and sarafotoxin (s6C, a snake venom) and their actions appears to be different among isoforms. The aim of this study was to compare the secretagogue effect of ET-1 on atrial natriuretic peptide (ANP) secretion with ET-3 and evaluate its physiological meaning. Isolated nonbeating atria from male Sprague-Dawley rats were used to evaluate stretch-activated ANP secretion in response to ET-1, ET-2, ET-3, and s6C. Changes in mean blood pressure (MAP) were measured during acute injection of ET-1 and ET-3 with and without natriuretic peptide receptor-A antagonist (A71915) in anesthetized rats. Changes in atrial volume induced by increased atrial pressure from o to 1, 2, 4, or 6cm H2O caused proportional increases in mechanically-stimulated extracellular fluid (ECF) translocation and stretch-activated ANP secretion. ET-1 (10nM) augmented basal and stretch-activated ANP secretion in terms of ECF translocation, which was blocked by the pretreatment with ETA receptor antagonist (BQ123, 1μM) but not by ETB receptor antagonist (BQ788, 1μM). ETA receptor antagonist itself suppressed stretch-activated ANP secretion. As compared to ET-1- induced ANP secretion (3.2-fold by 10nM), the secretagogue effects of ANP secretion by ET-2 was similar (2.8-fold by 10nM) and ET-3 and s6C were less potent (1.7-fold and 1.5-fold by 100nM, respectively). Acute injection of ET-1 or ET-3 increased mean blood pressure (MAP), which was augmented in the presence of natriuretic peptide receptor-A antagonist. Therefore, we suggest that the order of secretagogue effect of ET family on ANP secretion was ET-1≥ET-2>>ET-3>s6C and ET-1-induced ANP secretion negatively regulates the pressor effect of ET-1.

Ethnopharmacological relevance Radix Paeoniae Rubra (RPR) is an important traditional Chinese medicine (TCM) commonly used in clinic for a long history in China. RPR is the radix of either Paeonia lactiflora Pall. or Paeonia veitchii Lynch. RPR has a wide variety of pharmacological actions such as anti-thrombus, anti-coagulation, and anti-atherosclerotic properties, protecting heart and liver. However, the mechanisms involved are to be defined. Aim of the study The aim of the present study was to define the effect of Paeonia lactiflora Pall. extracts on vascular tension and responsible mechanisms in rat thoracic aortic rings. Materials and methods Ethanol extract of Paeonia lactiflora Pall. (EPL) was examined for their vascular relaxant effects in isolated phenylephrine-precontracted rat thoracic aorta. Results EPL induced relaxation of the phenylephrine-precontracted aortic rings in a concentration-dependent manner. Vascular relaxation induced by EPL was significantly inhibited by removal of the endothelium or pretreatment of the rings with N G -nitro- l -arginine methylester ( l -NAME) or 1 H -[1,2,4]-oxadiazolo-[4,3- α ]-quinoxalin-1-one (ODQ). Extracellular Ca 2+ depletion or diltiazem significantly attenuated EPL-induced vasorelaxation. Modulators of the store-operated Ca 2+ entry (SOCE), thapsigargin, 2-aminoethyl diphenylborinate and Gd 3+ , and an inhibitor of Akt, wortmannin, markedly attenuated the EPL-induced vasorelaxation. Further, the EPL-induced vasorelaxation was significantly attenuated by pretreatment with tetraethylammonium, a non-selective K Ca channels blocker, or glibenclamide, an ATP-sensitive K + channels inhibitor, respectively. Inhibition of cyclooxygenases with indomethacin, and adrenergic and muscarinic receptors blockade had no effects on the EPL-induced vasorelaxation. Conclusions The present study suggests that EPL relaxes vascular smooth muscle via endothelium-dependent and Akt- and SOCE-eNOS-cGMP-mediated pathways through activation of both K Ca and K ATP channels and inhibition of L-type Ca 2+ channels.

Introduction Nitric oxide (NO) has been shown to mediate electrical field stimulation (EFS)‐caused smooth muscle relaxation. It is known that the neural control of penile erection involves adrenergic, cholinergic, and non‐adrenergic‐non‐cholinergic (NANC) neuro‐effector systems; however, the effects of EFS on adrenergic and cholinergic nerves are not clear. Aims To elucidate EFS‐induced signal transductions involved in adrenergic, cholinergic, and NANC neuro‐effector systems by using an in vitro penile perfusion model. Methods EFS was performed on penile corpus cavernosum smooth muscle from male New Zealand White rabbits, which was pre‐contracted with L‐phenylephrine (10 µM). We investigated the penile tracing elicited by EFS on tissues pre‐incubated with guanethidine (Guan, 50 µM), tetrodotoxin (TTX, 10 µM), Nω nitro‐L‐arginine‐methyl ester (L‐NAME, 1 mM), atropine (50 µM), or eserine (10 µM). Main Outcome Measures The time‐to‐peak of each phase, the percentage of relaxation, and the area under the curve (AUC). Results We discovered an extraordinary phenomenon: three distinct phases elicited by EFS. Phase I was abolished by L‐NAME. Phase II was decreased by eserine and Guan, but increased by L‐NAME. Phase III was abolished by atropine, but enhanced by eserine and Guan. TTX diminished all three phases. The time to reach the top of phase I was delayed by TTX. The time to attain the peak of phase II was shortened by L‐NAME, but delayed by TTX and atropine. The time to reach the top of phase III was shortened by L‐NAME, eserine, and Guan. AUC was significantly decreased by L‐NAME and TTX. Conclusions EFS stimulated adrenergic, cholinergic, and NANC neuro‐effector systems simultaneously. Phase I was related to the NO pathway. Phase II was multiply affected by self‐recovery properties, and adrenergic and cholinergic nerves. Phase III was related to cholinergic nerves. The corporal tracing elicited by EFS was the balanced result of multiple factors. Zhao C, Cho KW, and Park JK. Three phases of corporal tracing elicited by electrical field stimulation on rabbit corpus cavernosum smooth muscle in penile perfusion model.

Natriuretic peptides help to maintain sodium and fluid volume homeostasis in a healthy cardio-renal environment. Since the identification of Dendroaspis natriuretic peptide (DNP) as a new member of the natriuretic peptide family, DNP has been considered as an important regulator of natriuresis and dieresis. The present study was undertaken to investigate the presence of immunoreactive Dendroaspis natriuretic peptide (DNP) and its specific receptor in rabbit. DNP was detected in heart, kidney, liver, brain, and plasma by radioimmunoassay (RIA). DNP contents of cardiac atrium and ventricle, renal cortex and medulla, liver, and brain were 1.42 ± 0.15, 1.0 6 ± 0.08, 2.55 ± 0.21, 1.81 ± 0.16, 1.36 ± 0.22, and 0.69 ± 0.15 pg/mg of wet weight, respectively. The concentration of DNP in plasma was 235.44 ± 15.44 pg/ml. By quantitative in vitro receptor autoradiography, specific ¹²⁵I-DNP binding sites were revealed in glomeruli, interlobular artery, acuate artery, vasa recta bundle, and inner medulla of the kidney with an apparent dissociation constant (K(d)) of 0.29 ± 0.05, 0.36 ± 0.03, 0.84 ± 0.19, 1.18 ± 0.23, and 10.91 ± 1.59 nM, respectively. Basal rate of 3', 5'-cyclic guanosine monophosphate (cGMP) production by particulate guanylyl cyclase (GC) activation of glomerular membranes was basally 13.40 ± 1.70 pmol/mg protein/min. DNP caused an increment of cGMP production in similar magnitude to that caused by ANP, BNP, and urodilatin, while the production of cGMP by CNP was significantly lower than that by DNP. Our results show that plasma levels of DNP were higher when compared to other tissues. DNP produces cGMP via the NPR-A receptor subtype in the kidney, similarly to ANP and BNP, suggesting that plasma DNP could have similar functions as ANP and BNP.

Aim of the study The aim of the present study was to define the effect of Xanthoceras sorbifolia extracts (XS) on vascular tension and responsible mechanisms in rat thoracic aortic rings. Materials and methods Ethanol extract of the leaves of XS (EXS) was examined for their vascular relaxant effects in isolated phenylephrine-precontracted rat thoracic aorta. Results EXS (0.1–100 μg/ml) induced relaxation of the phenylephrine-precontracted aortic rings in a concentration-dependent manner. Endothelium-denudation abolished EXS-induced vasorelaxation. Pretreatment of the endothelium-intact aortic rings with N G -nitro- l -arginine methylester ( l -NAME) and 1H-[1,2,4]-oxadiazolo-[4,3-α]-quinoxalin-1-one (ODQ) inhibited EXS-induced vasorelaxation. Inhibition of Ca 2+ entry via l -type Ca 2+ channels failed to block the EXS-induced vasorelaxation. Extracellular Ca 2+ depletion significantly attenuated EXS-induced vasorelaxation. Modulators of the store-operated Ca 2+ entry (SOCE), thapsigargin, 2-aminoethyl diphenylborinate (2-APB) and Gd 3+ , and an inhibitor of Akt, wortmannin, markedly attenuated the EXS-induced vasorelaxation. EXS increased cGMP levels of the aortic rings in a concentration-dependent manner and the effect was blocked by l -NAME, ODQ, thapsigargin, Gd 3+ , 2-APB, and wortmannin. Further, EXS-induced vasorelaxation was significantly attenuated by tetraethylammonium, a non-selective K ca channels blocker, but not by glibenclamide, an ATP-sensitive K + channels inhibitor. Inhibition of cyclooxygenase with indomethacin, and adrenergic and muscarinic receptors blockade had no effects on EXS-induced vasorelaxation. Conclusions The present study suggests that EXS relaxes vascular smooth muscle via endothelium-dependent NO-cGMP signaling through activation of the Akt- and SOCE-eNOS-sGC pathways, which may, at least in part, be related to the function of K + channels.

Aim of the study The aim of the present study was to define the effects of extracts of leaves of Zanthoxylum piperitum (ZP) on the vascular tension and its mechanisms responsible in rat thoracic aortic rings. Materials and methods Methanol extract of ZP and aqueous fraction of the methanol extract (AZP) were examined for their vascular relaxant effects in isolated phenylephrine-precontracted aortic rings. Results Methanol extract of ZP and aqueous fraction of the methanol extract (AZP) induced relaxation of the phenylephrine-precontracted aortic rings in a concentration-dependent manner. Endothelium-denudation abolished the AZP-induced vasorelaxation. Pretreatment of the endothelium-intact aortic rings with NG-nitro- l -arginine methylester (L-NAME) and 1H-[1,2,4]-oxadiazolo-[4,3-α]-quinoxalin-1-one (ODQ) inhibited the AZP-induced vasorelaxation. Inhibition of Ca2+ entry via L-type Ca2+ channels failed to block the AZP-induced vasorelaxation. Extracellular Ca2+ depletion slightly but significantly attenuated the AZP-induced vasorelaxation. Thapsigargin significantly attenuated the AZP-induced vasorelaxation. Further, Gd3+ and 2-aminoethyl diphenylborinate (2-APB), inhibitors of store-operated Ca2+ entry (SOCE), markedly attenuated the AZP-induced vasorelaxation. Also, wortmannin, an inhibitor of Akt, an upstream signaling molecule of eNOS, attenuated the AZP-induced vasorelaxation. AZP increased cGMP levels of the aortic rings in a concentration-dependent manner and the effect was blocked by L-NAME, ODQ, thapsigargin, Gd3+, 2-APB, and wortmannin. K+ channel inhibition with glibenclamide and tetraethylammonium, cyclooxygenase inhibition with indomethacin, and adrenergic and muscarinic receptors blockade had no effects on the AZP-induced vasorelaxation. Conclusion Taken together, the present study suggests that AZP relaxes vascular smooth muscle via endothelium-dependent activation of NO-cGMP signaling through the Akt- and SOCE-eNOS pathways.

Aim of study Although Zanthoxylum schinifolium has long been used in the traditional oriental medicine, cardiac effects have not well been documented. The aim of the present study was to investigate the effects of aqueous extract of leaves and stems from Zanthoxylum schinifolium (AZS) on inotropic effect and atrial natriuretic peptide (ANP) secretion. Materials and methods The AZS-induced changes in atrial dynamics, cAMP efflux and atrial ANP secretion were determined in isolated perfused beating rabbit atria. Results AZS increased atrial pulse pressure, stroke volume, and cAMP efflux concomitantly with inhibition of ANP secretion in a concentration-dependent manner. The AZS-induced increases in atrial dynamics and cAMP efflux, and decrease in ANP secretion were attenuated by pretreatment with propranolol and CGP 20712 but not ICI 118,551. Also, the AZS-induced changes in atrial dynamics and ANP secretion were attenuated by diltiazem and KT 5720. Diltiazem and KT 5720 had not significant effect on the AZS-induced increase in cAMP efflux. Conclusion These results suggest that AZS elicits a positive inotropic effect and decrease in ANP secretion via β1-adrenoceptor-cAMP-Ca2+ signaling in beating rabbit atria.

It has been shown that histamine inhibits atrial natriuretic peptide (ANP) release. Because cardiac mast cells are the principal source of histamine in the heart, we hypothesized that cardiac mast cells are involved in the regulation of atrial ANP release. To test the hypothesis, experiments were performed in perfused beating rabbit atria allowing atrial pacing and measurements of changes in atrial stroke volume, intraatrial pulse pressure and myocyte ANP release. Mast cell degranulation with Compound 48/80 decreased atrial myocyte ANP release, and the response was blocked by a selective histamine H 2 receptor blocker, cimetidine, indicating that histamine was responsible for the decrease in ANP release. Mast cell stabilization with cromolyn blocked the Compound 48/80-induced decrease in ANP release. These data suggest that mast cell-derived histamine is involved in the regulation of cardiac ANP release. Thus, the cardiac mast cell–cardiomyocyte communication via the histamine–ANP pathway may implicate in the cardiac disorder associated with mast cell degranulation such as in acute coronary syndrome or cardiac hypertrophy.

To assess the role of peptides of the angiotensin (ANG) on the regulation of clitoral cavernosum tone and changes in ANG binding affinity in the rabbit with diabetes mellitus.The isometric tension measurement and in vitro autoradiography were used in sham and diabetic clitoral cavernosum.In tension study, contractility in response to ANG I, ANG II, ANG III and ANG IV was enhanced in diabetic clitoral cavernosum strips (EC50 was 67.6 +/- 27.2, 4.3 +/- 0.4, 189.3 +/- 37.3, 443.2 +/- 0.4 nM for diabetic versus 155.2 +/- 76.1, 38.3 +/- 0.1, 528.0 +/- 75.2, 616.9 +/- 69.5 nM for sham, respectively). Contractile responses to ANG II was significantly inhibited by type 1 ANG II receptor (AT1) antagonist but not by type 2 ANG II receptor (AT2) antagonist in both groups. Percentages in contractions by ANG II (1 nM) in the presence of Dup 753 decreased significantly 36.2 +/- 4.6 to 6.3 +/- 2.4% in sham and 56.1 +/- 7.7 to 6.0 +/- 4.8% in diabetic group. The binding affinities were enhanced in diabetic clitoral cavernosum for ANG II (dissociation constant, 4.9 +/- 1.0 for sham versus 0.9 +/- 0.2 nM for diabetic) and for ANG I, ANG III, and ANG IV (inhibitory constant, 28.6 +/- 1.5, 398.7 +/- 157.2, and 3966.5 +/- 1524.1 nM for sham versus 20.6 +/- 5.7, 78.5 +/- 23.7, and 1098.7 +/- 195.5 nM, for diabetic, respectively, all p0.05). Sensitivities of AT1 and AT2 receptors to ANG II enhanced in diabetic than sham clitoral cavernosum tissue.This results suggest that the contractile responses to all four ANG peptides are enhanced in the diabetic clitoral cavernosum. Enhancement of contractility in diabetic clitoral cavernosum may be related to the increased affinity to ANG II receptors for ANG peptides.

Hyperosmolality has been known to increase ANP release. However, its physiological role in the regulation of atrial myocytic ANP release and the mechanism by which hyperosmolality increases ANP release are to be defined. The purpose of the present study was to define these questions. Experiments were performed in perfused beating rabbit atria. Hyperosmolality increased atrial ANP release, cAMP efflux, and atrial dynamics in a concentration-dependent manner. The osmolality threshold for the increase in ANP release was as low as 10 mosmol/kgH2O (∼3%) above the basal levels (1.55 ± 1.71, 17.19 ± 3.11, 23.15 ± 5.49, 54.04 ± 11.98, and 62.00 ± 13.48% for 10, 20, 30, 60, and 100 mM mannitol, respectively; all P < 0.01). Blockade of sarcolemmal L-type Ca2+channel activity, which increased ANP release, attenuated hyperosmolality-induced increases in ANP release (−13.58 ± 4.68% vs. 62.00 ± 13.48%, P < 0.001) and cAMP efflux but not atrial dynamics. Blockade of the Ca2+release from the sarcoplasmic reticulum, which increased ANP release, attenuated hyperosmolality-induced increases in ANP release (13.44 ± 7.47% vs. 62.00 ± 13.48%, P < 0.01) and dynamics but not cAMP efflux. Blockades of Na+-K+-2Cl−cotransporter, Na+/H+exchanger, and Na+/Ca2+exchanger had no effect on hyperosmolality-induced increase in ANP release. The present study suggests that hyperosmolality regulates atrial myocytic ANP release and that the mechanism by which hyperosmolality activates ANP release is closely related to the cross-talk between the sarcolemmal L-type Ca2+channel activity and sarcoplasmic reticulum Ca2+release, possibly inactivation of the L-type Ca2+channels.

We defined whether extracorporeal shock wave therapy (ESWT) to the kidney activates the nitric oxide (NO)-cyclic 3',5'-guanosine monophosphate (cGMP) pathway.A total of 90 male rabbits were randomly divided into group 1--pretreated with normal saline, group 2--pretreated intravenously (i.v.) with Nomega nitro-L-arginine-methyl ester (NAME) (100 mg/kg), group 3--pretreated i.v. with NAME and L-arginine (300 mg/kg) with ESWT to 1 kidney, group 4--pretreated IV with ODQ (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one) (20 microg/kg) and group 5--pretreated with normal saline with ESWT to the bladder. Plasma nitrite, NO metabolite and cGMP were analyzed in peripheral blood samples before, immediately after, and 30 and 60 minutes after ESWT.ESWT to the kidney but not to the bladder caused an increase before, immediately after, and 30 and 60 minutes after ESWT in plasma nitrite in group 1 (186.1 +/- 20.6, 217.5 +/- 21.6, 241.9 +/- 28.4 and 230.5 +/- 25.3 nM) and group 5 (149.0 +/- 14.7, 155.6 +/- 18.4, 131.8 +/- 13.6 and 140.0 +/- 15.7 nM), and in cGMP in group 1 (24.2 +/- 1.9, 33.8 +/- 3.2, 32.9 +/- 2.2 and 29.4 +/- 1.9 pmol/ml) and group 5 (25.5 +/- 2.1, 27.5 +/- 2.5, 28.7 +/- 3.1 and 25.5 +/- 2.6 pmol/ml, respectively). In group 2 NAME significantly inhibited the production of nitrite (113.4 +/- 18.6, 118.2 +/- 19.9, 114.6 +/- 18.3 and 112.5 +/- 17.6 nM) and cGMP (19.4 +/- 2.6, 20.6 +/- 2.8, 19.3 +/- 2.7 and 18.6 +/- 2.6 pmol, respectively). In group 3 inhibited nitrite and cGMP production caused by NAME was recovered with L-arginine. In group 4 ODQ significantly inhibited cGMP production.The results show that ESWT increases the level of NO and cGMP released by the kidney in an animal model.

Lysophosphatidylcholine (LPC) is an endogenous phospholipid released from the cell membrane during ischemia, and it has potent, local effects on cardiac tissues. LPC has been implicated in arrhythmogenesis during ischemia by increasing intracellular Ca 2+ . However, it is not known whether LPC influences atrial release of atrial natriuretic peptide (ANP). The aim of this study was to investigate the effect of LPC on ANP secretion from isolated, perfused, beating rat atria. LPC (10 and 30 μmol/L) caused decreases in ANP secretion in a dose-dependent manner, with slight increases in intra-atrial pressure and extracellular fluid (ECF) translocation. Therefore, the ANP secretion in terms of ECF translocation was markedly decreased by LPC. The order of the suppressive effect of ANP release was stearoyl-LPC>LPC>myristoyl-LPC=lauroyl-LPC. Staurosporine and wortmannin significantly attenuated suppression of the ANP release and an increase in intra-atrial pressure by LPC. High extracellular Mg 2+ also attenuated the LPC-induced suppression of ANP release. However, other protein kinase C inhibitors such as chelerythrine, GF 109203X, and tamoxifen citrate did not affect LPC-induced suppression of ANP release. In single atrial myocytes, LPC caused increases in intracellular Ca 2+ in a dose-dependent manner. The order of an increase in intracellular Ca 2+ by LPC was stearoyl-LPC>LPC>myristoyl-LPC=lauroyl-LPC. An increase in intracellular Ca 2+ by LPC was attenuated by staurosporine. These results suggest that LPC-induced suppression of ANP release through protein kinase C/Ca 2+ and phosphoinositol-3-kinase might in part play an important role in the development of hypertension.

We investigated the role of nitric oxide (NO)-guanosine 3′,5′-cyclic monophosphate (cGMP) signaling in the regulation of rabbit clitoral cavernosum (CC) tone. Tension measurements, reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting, and NADPH-diaphorase staining were performed in CC. In the precontracted CC strips with phenylephrine (10–5 M), acetylcholine (ACh) relaxed, dependent on dosage. Pretreatment with atropine, Nω nitro-l-arginine-methyl ester (NAME) or 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), guanylate cyclase inhibitor abolished the ACh-induced relaxations, but tetrodotoxin (TTX) did not. Sodium nitroprusside relaxed the strips in the presence of atropine and NAME, but not in the presence of ODQ. Electrical field stimulation (EFS) relaxed the strips dependent on stimulus strength. Pretreatment with TTX, NAME, or ODQ abolished the EFS-induced relaxation, but atropine did not. l-Arginine partially restored the inhibited response to ACh and EFS. The inducible NO synthase (iNOS) and neuronal NOS (nNOS) mRNAs and iNOS and endothelial NOS (eNOS) proteins were identified in the CC. NADPH-diaphorase staining revealed the positivity on the nerve trunks and fine nerve fibers in the CC. Finally, results demonstrate that the nNOS, ENOS, and the NO-cGMP signaling pathway are involved in the regulation of clitoral tumescence.

The role of extracorporeal shock wave lithotripsy (ESWL, Dornier Medizintechnik GmbH, Wessling Munchen, Germany) on plasma nitric oxide (NO) and cyclic nucleotides was investigated. It is known that ESWL increased plasma levels of vasoactive substances.A total of 28 patients undergoing ESWL for renal (14) and lower ureteral (14) stones participated in a prospective study. Peripheral blood samples were analyzed for plasma nitrite, NO metabolite, cyclic 3', 5'-guanosine monophosphate (cGMP), cyclic 3',5'-adenosine monophosphate and plasma renin activity before, immediately after, 30 and 60 minutes after ESWL. Urine samples were also analyzed for nitrite before, immediately after, 30 and 60 minutes after ESWL. Blood pressure and pulse rate were measured before and after ESWL.ESWL increased mean plasma nitrite levels. Before, immediately after, 30 and 60 minutes after ESWL mean plasma nitrate plus or minus standard error in patients with renal versus lower ureteral stones was 43.6 +/- 8.5, 52.8 +/- 7.4, 54.0 +/- 8.0 and 52.2 +/- 9.1 versus 27 +/- 5.8, 24.6 +/- 4.7, 23.6 +/- 4.8 and 28.0 +/- 7.1 nM., respectively. Before, immediately after, 30 and 60 minutes after ESWL mean plasma cGMP in patients with renal versus lower ureteral stones was 1.21 +/- 0.1, 1.64 +/- 0.16, 1.52 +/- 0.12 and 1.58 +/- 0.14 versus 1.45 +/- 0.18, 1.52 +/- 0.13, 1.52 +/- 0.15 and 1.51 +/- 0.11 fmol., respectively. Cyclic 3',5'-adenosine monophosphate was not increased by ESWL in patients with renal stones. Urine nitrite increased slightly but not significantly after ESWL. Plasma cyclic nucleotides and nitrite, and urine nitrite were not changed by ESWL in patients with lower ureteral stones. Plasma renin activity decreased significantly in patients with renal stones 60 minutes after ESWL. Blood pressure and pulse rates increased mildly without statistical significance after ESWL for renal stones.ESWL increases plasma nitrite and cGMP in patients with renal stones, suggesting that shock waves stimulate the NO-cGMP signaling pathway.

Abnormal intracellular Ca 2+ -handling has been described in various heart diseases associated with cardiac hypertrophy. The crucial role of Ca 2+ in the excitation–secretion coupling in atrial cardiomyocytes is not well established. To investigate modulation of atrial natriuretic peptide (ANP) secretion regulated by Ca 2+ in hypertrophied atria, responsiveness of stretch-induced ANP to Ca 2+ was studied using isolated perfused quiescent hypertrophied rat atria. Male Sprague–Dawley rats were given a single subcutaneous injection of 50 mg/kg monocrotaline (MCT) and were sacrificed at 5–6 weeks. In isolated perfused hypertrophied right atria from MCT rats, changes in atrial volume induced by increased atrial pressure caused proportional increases in mechanically stimulated extracellular fluid (ECF) translocation and stretch-induced ANP secretion. Stretch-induced ANP secretion was markedly increased by the depletion of extracellular Ca 2+ . However, an accentuation of stretch-induced ANP secretion by Ca 2+ depletion was markedly attenuated in hypertrophied right atria, as compared to control right atria. Therefore, stretch-induced ANP secretion in terms of ECF translocation by Ca 2+ depletion in hypertrophied atria was significantly lower than in control right atria. However, no significant differences were observed between nonhypertrophied and control left atria. Depletion of extracellular Ca 2+ caused a decrease in intracellular calcium in single beating atrial myocytes, which was significantly attenuated in hypertrophied atrial myocytes. The results suggest that attenuation of Ca 2+ -induced negative regulation of ANP secretion in hypertrophied atria may be due to the disturbance of intracellular Ca 2+ regulation.

Effect of Berberine on the Atrial Natriuretic Peptide Secretion in Cardiac Atria Hye Rim Yoo 1,2, Hye Yoom Kim 1,2, Oh Jeong Kwon 1,2, Kyung Woo Cho 2, Dae Gill Kang 1,2, and Ho Sub Lee 1,2 * 1College of Oriental Medicine and Professional Graduate School of Oriental Medicine, Wonkwang University, Iksan, Jeonbuk 570-749, Republic of Korea; 2Hanbang Body-fluid Research Center, Wonkwang University, Shinyong-dong, Iksan, Jeonbuk, 570-749, Republic of Korea The aim of the present study is to identify the effects of berberine on the secretion of atrial natriuretic peptide (ANP) from the atrium. Experiments were performed in perfused beating rat left atria paced electrically. Atrial dynamics, atrial pulse pressure, was measured with physiograph. ANP was measured using radioimmunoassay. As a result, berberine increased ANP secretion and atrial dynamics in a concentration- dependent manner. Atropine-non-selective muscarinic receptor antagonist, methoctramine-selective M2 muscarinic receptor antagonist, and hemicho...

Emodin, an active anthraquinone constituent isolated from the rhubarb, a traditional Chinese herbal medicine which is widely used in clinical treatment, has cardiovascular protective properties. However, it remains unclear whether the cardiovascular protective actions of emodin are related to an activation of cardiac natriuretic hormone secretion. The purpose of the present study was to explore the effect of emodin on the secretion of ANP, a member of the family of cardiac natriuretic hormones, and its mechanisms involved. Experiments were performed in isolated perfused beating rabbit atria allowing measurement of ANP secretion, atrial pulse pressure, and stroke volume. Emodin increased ANP secretion concomitantly with a decrease in atrial pulse pressure and stroke volume in a concentration-dependent manner. These effects were reversible. Inhibition of K(+) channels with tetraethylammonium and glibenclamide attenuated the emodin-induced changes in ANP secretion and atrial dynamics. Furthermore, the emodin-induced changes in ANP secretion and atrial dynamics were attenuated by inhibition of L-type Ca(2+) channels with nifedipine. Atropine, methoctramine, tertiapin-Q, and pertussis toxin had no significant effect on the emodin-induced changes in ANP secretion and mechanical dynamics. The present study demonstrates that emodin increases ANP secretion via inhibition of L-type Ca(2+) channels through an activation of K(+)ATP channel in isolated beating rabbit atria. The results also provide a rationale for the use of emodin in the treatment of impairment of the regulation of the cardiovascular homeostasis.

C-type natriuretic peptide (CNP) is mainly distributed in the brain and vascular endothelium and is considered to act as a local regulator in many tissues. The present study was aimed to determine the presence of CNP system and its biological function in rabbit colon. The serial dilution curves of tissue extracts were parallel to the standard curve of CNP-22. With gel permeation chromatography and reverse-phase HPLC, the major immunoreactive peak of CNP was observed at the same elution time corresponding to the synthetic CNP-53. The concentration of CNP in the mucosal layer of colon was 212.49 +/- 30.44 pg/g tissue wet weight (n = 7), which was significantly higher than that in the muscular layer. The presence of CNP mRNA was also detected by RT-PCR and Southern blot analysis. Production of cGMP by the activation of particulate guanylyl cyclase stimulated by BNP and CNP was higher in membranes obtained from the muscular layer than from mucosal layer. More cGMP was produced by CNP than by ANP. Both natriuretic peptide receptor-A and -B mRNAs were detected by RT-PCR and specific binding sites to 125I-[Tyr(0)]-CNP-22 were mainly localized to the muscular layer. Synthetic CNP inhibited basal tension, frequency and amplitude of basal motility of taenia coli of the right colon. This study showing the presence of CNP system and its biological function in colon suggests that endogenous CNP synthesized in the mucosal layer may have a paracrine function as a local regulator of colonic motility.

Purpose: Angiotensin (ANG) II has been known to be a potent modulator for the maintenance of smooth muscle tone of the penile cavernosum. However, its role in clitoral cavernosum is unknown. The clitoris is the homologue of the penis arising from the embryological genital tubercle. We investigated the presence of ANG II receptors, the function of ANG II, and its interaction with nitric oxide (NO) in rabbit clitoral cavernosum.Materials and Methods: The isometric tension was measured in the strips of clitoral cavernosum. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to evaluate expression of AT1a and AT1b ANG II receptor subtype mRNAs. In vitro autoradiography was used to localize ANG II receptors in the clitoral cavernosum.Results: The clitoral cavernosum was contracted dose-dependently by the addition of ANG II. Dup 753 (ANG II type 1 receptor antagonist) inhibited significantly ANG II induced contraction. PD 123,319 (ANG II type 2 receptor antagonist) did not affect the ANG II respon...

Natriuretic peptide (NP) receptors (NPRs) located at the endocardial endothelium are suggested to be involved in regulating myocardial contractility. However, the characteristics and modulation of NPRs in relation to cardiac failure are not well defined. This study examined the properties of NPRs in ventricular endocardium using quantitative receptor autoradiography, RT-PCR, Southern blot analysis, and activation of particulate guanylyl cyclase (GC) by NPs. In control rats, specific125I-labeled rat atrial NP (rANP)(1–28) binding sites were localized in right (RV) and left ventricular (LV) endocardium. Binding affinities of125I-rANP(1–28) were remarkably higher in RV than LV endocardium. Radioligand binding at these sites was mostly inhibited by des[Gln18,Ser19,Gly20,Leu21,Gly22]ANP(4–23), a specific NP clearance receptor ligand. mRNAs for all three recognized NPRs were detected in endocardial cells by RT-PCR and confirmed by Southern blot analysis. Production of cGMP by particulate GC in endocardial cell membranes was stimulated by NPs with a rank order of potency of C-type NP(1–22) >> brain NP (BNP)(1–26) > ANP(1–28). We also examined the modulation of these NPRs during cardiac hypertrophy induced by monocrotaline (MCT). In MCT-treated rats with pulmonary hypertension, specific125I-rANP(1–28) binding to hypertrophied RV endocardium almost disappeared and cGMP production by NPs was significantly decreased. In rats with pulmonary hypertension, plasma levels of ANP and BNP were increased by fivefold compared with controls. The results indicate that there is a differential distribution of NPRs in the cardiac chambers, with the most abundant binding sites in RV endocardium, that NPR-B is the predominant GC-coupled NPR in ventricular endocardium, and that endocardial NPRs are downregulated with ventricular hypertrophy. Downregulation of NPRs may be associated with an increment of endogenous NP production caused by mechanical overload in hypertrophied ventricle.

To investigate the presence of C-type natriuretic peptide 1-22 (CNP)-dependent guanylyl cyclase (GC)-coupled receptor and its biological function in the penile erectile smooth muscle.Experiments have been done in rabbit and rat to detect cyclic GMP (cGMP) generation by the activation of particulate GC by natriuretic peptides (NPs) in cavernosal membrane, to localize precise receptor using a quantitative in vitro autoradiography of the snap frozen sections, to define natriuretic peptide receptor (NPR) mRNA using a reverse transcriptase-polymerase chain reaction (RT-PCR) technique and to monitor changes of erectile smooth muscle tone by NPs in the penile tissue strips.Productions of cGMP by particulate GC in the corpus cavernosum membranes of rabbit and rat were stimulated by CNP, atrial natriuretic peptide 1-28 (ANP) and brain natriuretic peptide 1-26 (BNP) with a rank order of potency of CNPBNPANP. HS-142-1, a selective antagonist for the GC-coupled NPR, inhibited the CNP-stimulated cGMP production in corpus cavernosal membrane of rabbit and rat. Specific 125I-(Tyr[0])-CNP bindings were localized in the corpus cavernosal smooth muscle of rabbit with Kd of 19.92+/-3.38 nM. and Bmax of 734.64+/-139.63 amol./mm2. B-subtype of NPR mRNA was detected in the penile corpus cavernosum of rat using RT-PCR technique. CNP relaxed the smooth muscle contracted by Nomega-nitro-L-arginine methylester (L-NAME).These results suggest for the first time that CNP modulates the erectile smooth muscle tone of penis by predominant activation of B-subtype of NPR with augmentation of cGMP production via particulate GC.

The distribution of receptors for atrial natriuretic peptide (ANP) and angiotensin II (ANG II) in the kidney and adrenal gland of the freshwater turtle, Amyda japonica, was examined by quantitative in vitro autoradiography using 125 I-labeled rat (r)ANP 1–28 and 125 I-labeled human ANG II as labeled ligands. Receptor subtypes were characterized by competition with des[Gln 18 ,Ser 19 ,Gly 20 , Leu 21 , Gly 22 ] ANP (4–23) (C-ANP) as a selective ligand of the clearance receptors to specific 125 I-rANP (1–28) binding, and with DuP 753 and PD 123319 as nonpeptide antagonists for ANG II receptors type 1 (AT 1 ) and type 2 (AT 2 ) subtypes, respectively, to 125 I-ANG II binding. Specific 125 I-rANP (1–28) binding with a single binding site was found overlying glomeruli of the kidney and the outer zone of the adrenal gland with dissociation constants ( K d ) of 4.39 ± 0.33 and 6.07 ± 1.36 n M, respectively. C-ANP (10 μ M ) inhibited about 90% of the glomerular and adrenal 125 I-rANP (1–28) binding. Specific 125 I-ANG II binding was also localized in the glomeruli of the kidney and the outer zone of the adrenal gland with K d of 1.02 ± 0.22 and 0.37 ± 0.04 n M, respectively. DuP 753 (10 μ M ) potently inhibited about 80% of glomerular and 90% of adrenal 125 I-ANG II binding, whereas PD 123319 (10 μ M ) was very weak in competing for specific 125 I-ANG II binding in both tissues. Therefore, these results indicate that specific ANP and ANG II receptors with high affinities have an overlapping distribution in glomeruli of the kidney and the outer zone of the adrenal gland of the freshwater turtle. They also suggest that biological and clearance ANP receptor-like subtypes coexist in both tissues, and the predominant ANG II receptor subtype in these tissues is the AT 1 -like receptor. The overlapping distribution of specific receptors for both peptides in these tissues provides the basis for possible receptor interactions to exert a functional antagonism between ANP and ANG II in the freshwater turtle.

Purpose: The regulation of the corporal smooth muscle tone is important in the process of penile erection. Although specific angiotensin (ANG) II binding to and effects of ANG II on some reproductive structures have been studied, the presence of the renin-angiotensin system has not yet been defined in the corpus cavernosum. ANG II is formed from ANG I by angiotensin I-converting enzyme (ACE). ANG II and ANG I produce contractions in vascular smooth muscles. Two subtypes of ANG II receptors (AT1 and AT2) have been characterized. The purpose of the present experiments was to determine whether the renin-angiotensin system regulates rabbit corpus cavernosum smooth muscle tone.Materials and Methods: A strip of rabbit corpus cavernosum was mounted in an organ chamber to measure the isometric tension. The specific binding for125 I-ANG II was characterized by in vitro autoradiography.Results: ANG II and ANG I, precursor of ANG II, contracted corpus cavernosum smooth muscle dose-dependently, but the respon...

The atrial natriuretic peptide (ANP) gene is expressed in several extraatrial tissues in which ANP may be involved in the regulation of autocrine or paracrine functions. In the ovary, the synthesis of ANP, its binding sites, and a physiological role were found. The ANP system in the oviduct, however, is yet to be defined. The purpose of the present study was to determine whether the ANP system is present in the oviduct and then to define its function. The serial dilution curves of oviductal extracts in rat and rabbit were parallel to the standard curve of ANP. Molecular profiles using reverse phase HPLC indicated that the prohormone and processed circulating peptide were the main forms present. The immunoreactive ANP content of the oviduct was 27.07 +/- 4.41 pg/mg tissue wet wt (1.19 +/- 0.19 ng/oviduct; n = 10; at metestrus) in rats and 1.21 +/- 0.12 pg/mg tissue wet wt (0.15 +/- 0.01 ng/oviduct; n 12) in rabbits. In adult 4-day cycling rats, the immunoreactive ANP contents in oviducts had a cyclic change characterized by the lowest level at proestrus (14.59 +/- 3.24 pg/mg; n = 12). A distinct and strong ANP immunoreactivity was found in the mucosal layer of rat oviduct, and ANP messenger RNA was also detected in the oviduct by reverse transcriptase-PCR. Specific high affinity binding sites for iodinated rat ANP ([125I]rANP) were observed in the mucosal layer of the oviduct in rats and rabbits. Specific [125I]rANP bindings localized in the mucosal layer of rabbit oviduct showed an apparent dissociation constant (Kd) of 18.69 +/- 5.55 nM and a maximal binding capacity of 14.85 +/- 6.19 fmol/mm2. These specific [125I]rANP bindings were not reversed by des-[Gln18,Ser19,Gly20,Leu21,Gly22]ANP-(4- 23) as a selective ligand of clearance receptor. Synthetic ANP inhibited both the frequency and amplitude of basal motility of rabbit oviduct in a dose-dependent manner. These results suggest that the oviduct has its own ANP system, and the system is involved in the regulation of oviductal motility.

Lycopus lucidus Turcz has been widely used as a traditional Oriental medicine (TOM) in Korea and China and prescribed for the enhancement of heart function. However, the precise effects have yet to be defined. The purpose of the present study was, therefore, to address whether the ethanol extract of Lycopus lucidus Turcz (ELT) has a positive inotropic effect. ELT-induced changes in atrial mechanical dynamics (pulse pressure, dp/dt, and stroke volume), and cAMP efflux were measured in perfused beating rabbit atria. Three active components, rosmarinic acid, betulinic acid, and oleanolic acid were identified in ELT by high performance liquid chromatography analysis. ELT increased atrial dynamics in a concentration-dependent manner without changes in atrial cAMP levels and cAMP efflux. The ELT-induced positive inotropic effect was blocked by inhibition of the L-type Ca(2+) channels and sarcoplasmic reticulum (SR). Inhibitors of β-adrenoceptors had no effect on the ELT-induced positive inotropic effect. The results suggest that ELT exerts a positive inotropic effect via activation of Ca(2+) entry through L-type Ca(2+) channel and Ca(2+) release from the SR in beating rabbit atria.

Specific binding sites for atrial natriuretic peptide (ANP) were investigated by in vitro autoradiographic techniques in various tissues of the freshwater turtle, Amyda japonica. A high density of binding sites for 200 pM of 125I-labelled rANP(1-28) was located in the glomeruli of the kidney and the cortical portion of the adrenal gland. A moderate density of binding sites was seen in the arachnoid matter and choroid plexus of the third and lateral ventricles of the brain and the epididymis. A low density of binding sites was revealed in lamina propria of the mucosa of stomach and intestine, the seminiferous tubules of testes, and the epithelial layer of oviduct. In the presence of excess unlabelled rANP(1-28) (1 microM), binding to these structures were completely displaced. Therefore, specific ANP receptors exist in the kidney, adrenal gland, stomach, intestine, oviduct, epididymis, seminiferous tubules and brain. The ANP system may be involved in physiological regulatory function in the freshwater turtle, Amyda japonica.

OBJECTIVES The exact role of atrial natriutetic peptide (ANP) in the pathogenesis of edema in nephrotic syndrome (NS) has not been fully elucidated. We aimed to investigate the possible contribution of ANP to edema formation in NS. METHODS We subjected 18 nephrotic subjects and 20 healthy volunteers to supine bicycle exercise (SBE), a maneuver that seemed to increase venous return and to enhance the release of ANP. Plasma concentrations of immunoreactive-ANP were measured before and after SBE by radioimmunoassay. RESULTS There was a significant rise in the plasma concentration of ANP in the controls after SBE (from 31.1 +/- 6.16 to 42.0 +/- 6.01 pg/ml: p < 0.05). Meanwhile, there was no change in plasma concentration of ANP in the patients with NS (from 35.4 +/- 6.04 to 35.1 +/- 5.31 pg/ml). The change in plasma concentration of ANP in controls was significantly different from that in those with NS (p < 0.05). The mean baseline value of ANP in controls was the same as in NS. CONCLUSIONS These results show that SBE was a simple maneuver to stimulate the release of ANP in healthy controls. In contrast, it failed to stimulate the release of ANP in subjects with NS.

Ethnopharmacological relevance Euphorbia humifusa Willd. (EH) is an important traditional Chinese medicine that has commonly been used for treating bacillary dysentery and enteritis in many Asian countries for thousands of years. EH has a wide variety of pharmacological actions such as antioxidant, hypotensive, and hypolipidemic effects. However, the mechanisms involved are to be defined. Aim of the study The present study was performed to evaluate the cardiovascular effects of EH in rats. Materials and methods Methanol extract of EH (MEH) and ethylacetate fraction of the MEH (EEH) was examined for their vascular relaxant effects in phenylephrine-precontracted aortic rings. Effects of EEH on systolic blood pressure and heart rate were tested in Sprague–Dawley rats. Results MEH and EEH induced vasorelaxation in a concentration-dependent manner. Endothelium-denudation abolished the EEH-induced vasorelaxation. Pretreatment of the endothelium-intact aortic rings with N G -nitro- l -arginine methylester ( l -NAME) and 1 H -[1,2,4]-oxadiazolo-[4,3-α]-quinoxalin-1-one (ODQ) significantly inhibited the EEH-induced vasorelaxation. EEH increased cGMP levels of the aortic rings in a concentration-dependent manner and the effect was blocked by l -NAME or ODQ. Extracellular Ca 2+ depletion and treatments with thapsigargin, Gd 3+ , and 2-aminoethyl diphenylborinate significantly attenuated the EEH-induced vasorelaxation. Wortmannin markedly attenuated the EEH-induced vasorelaxation. In addition, tetraethylammonium, iberiotoxin, and charybdotoxin, but not apamin, attenuated the EEH-induced vasorelaxation. Glibenclamide, indomethacin, atropine, and propranolol had no effects on the EEH-induced vasorelaxation. Furthermore, EEH decreased systolic blood pressure and heart rate in a concentration-dependent manner in rats. Conclusions The present study demonstrates that EEH induces endothelium-dependent vasorelaxation via eNOS-NO-cGMP signaling through the modification of intracellular Ca 2+ , Ca 2+ entry, and large- and intermediate-conductance K Ca channel homeostasis. The data also suggest that the Akt-eNOS pathway is involved in the EEH-induced vasorelaxation. EEH induces hypotension and bradycardia in vivo .

Dendroaspis natriuretic peptide (DNP) is one of four members of the natriuretic peptide family sharing functional and structural properties. The purpose of the present study was to elucidate the physiological role of DNP on renal functions and its cellular mechanism in the rabbit kidney. DNP (5 μg/kg/min) infused intravenously increased urine volume and urinary excretion of electrolytes. These renal actions induced by DNP were more pronounced than those caused by atrial natriuretic peptide (ANP). We compared profiles of 125I-ANP and 125I-DNP by reverse-phase HPLC during incubation in rabbit plasma at 37 °C for 1, 2, and 4 h. While 125I-ANP was quickly degraded within 1 h, 125I-DNP was still stable in plasma for 4 h. DNP induced the greatest cyclic guanosine monophosphate (cGMP) production in the glomeruli in a dose-dependent manner, when compared to other renal structures including cortical tubules, outer medullary tubules, and inner medullary tubules. Affinity cross-linking analysis revealed NPR-A is selective receptor for DNP in glomeruli. Forskolin, a stimulator of adenylyl cyclase, significantly decreased cGMP production in the renal glomeruli but not in the renal medulla. In summary, DNP is a more effective activator of renal functions than ANP, possibly because of the degradation resistance of DNP against the endogenous peptidases in plasma or tissues. These findings suggest that DNP plays a pivotal role as a renal regulating peptide via specific natriuretic peptide receptors with a guanylyl cyclase domain.

Ethnopharmacological relevance Oryeongsan (ORS, Wulingsan), a formula composed of five herbal medicines, has long been used for the treatment of impairments of the regulation of body fluid homeostasis in China, Japan and Korea. Aim of the study The purpose of the present study was to test the effects of ORS on the renal function and the mechanisms involved in rats. Materials and methods Experiments were performed in rats caged individually. Renal function and plasma levels of renin activity and aldosterone concentration were measured. Results Treatment of ORS resulted in increases in urinary volume, excretion of Na+, K+, and Cl−, and glomerular filtration rate, and decreases in urinary osmolality and Na+ balance. Further, ORS decreased plasma renin activity and aldosterone concentration. An increase in urinay excretion of Na+ was a function of glomerular filtration rate, while the increase in the day-time period was related with the increase in the ratio of urinary Na+/K+. Conclusion Therefore, the present results suggest that ORS induces diuresis and natriuresis via inhibition of the renin–angiotensin–aldosterone system in rats.

Oryeong-san which was first recorded in Shanghanrun describing the treatments of acute febrile disease is one of the frequently used oriental medicines. Oryeong-san has been prescribed for the treatment of symptoms accompanied by edema. The purpose of this study was to examine the diuretic effects of Oryeong-san by different routes of administration. Oryeong-san (100 mg/kg body weight) was administrated by three different routes in Sprague-Dawley rats: intravenous infusion, intraperitoneal injection and oral intake. Oral intake of Oryeong-san significantly increased urinary volume and excretion of Na + , Cl - , and K + compared to vehicle-treated control group. The effects were concentration-dependent. Intravenously administrated Oryeong-san increased urinary volume and electrolyte excretion but without significance in hydrated (0.02 ml/min/rat for 90 min) anesthetized rats. Similarly, intraperitoneally injected Oryeong-san had no effects on water and urine electrolyte excretion compared with saline control group. These findings suggest that Oryeong-san has different effects on water and electrolyte balance by routes of administration.

1. 1. The presence and partial characterization of immunoreactive ANP (irANP) in eggs were investigated using high performance liquid chromatography (HPLC), immunohistochemistry and Northem-blot hybridization in mammalian and nonmammalian vertebrates. 2. 2. Serial dilution curves of egg extracts from frogs and freshwater teleostean fishes (silver crusian carp and loach) were parallel to the standard curve of atriopeptin III. Rat oocytes also contained irANP (25–30 Pg/egg). 3. 3. The HPLC profile of irANP showed two main peaks corresponding to low and high molecular weight of irANP. 4. 4. ANP mRNA was detected in the fish eggs. With immunohistochemical analysis of the rat ovarian follicles, this ANP-like material was localized mainly at the oocytes. 5. 5. We, therefore, suggest that the vertebrate oocytes synthesize ANP and that the presence of ANP in the oocytes may be linked with the cell differentiation.

Ursolic acid is reported to have beneficial effects on the regulation of cardiovascular homeostasis. However, the effects of ursolic acid on cardiac hormone secretion are yet to be defined. The present study was designed to test the effects of ursolic acid on the secretory and contractile functions of the atria. Experiments were conducted in isolated perfused beating rabbit atria. We measured the changes in atrial dynamics, pulse pressure, stroke volume, cAMP efflux, as well as the secretion of atrial natriuretic peptide (ANP). Ursolic acid increased ANP secretion and mechanical dynamics in a concentration-dependent manner. The inhibition of L-type Ca(2+) channels with nifedipine attenuated the ursolic acid-induced increase in ANP secretion but not mechanical dynamics. The inhibition of K(+)(ATP) channels with glibenclamide attenuated the ursolic acid-induced increase in ANP secretion-but not atrial dynamics-in a concentration-dependent manner. The selective Na(+)-K(+)-ATPase inhibitor ouabain blocked the ursolic acid-induced increase in atrial dynamics but not ANP secretion. These findings show that ursolic acid increases ANP secretion via its activation of K(+)(ATP) channels and subsequent inhibition of Ca(2+) entry through L-type Ca(2+) channels in rabbit atria. These data also suggest that ursolic acid increases atrial dynamics via its inhibition of Na(+)-K(+)-ATPase activity.

Aims Although a β-adrenoceptor (β-AR) blockade-induced increase in plasma atrial natriuretic peptide (ANP) levels is implicated in the therapeutic significance of β-AR antagonists, the role of β-AR in the regulation of ANP release is not clearly defined. The purpose of the present study was to define the role of β-AR subtypes and the mechanisms responsible for regulation of atrial ANP release. Main methods Experiments were performed in isolated perfused beating rabbit atria, including measurement of atrial contractile response, cAMP efflux, and atrial myocyte ANP release. Key findings β-AR activation with (–)-isoproterenol decreased ANP release concomitantly with increases in cAMP efflux concentration, atrial dynamics, stroke volume and pulse pressure in a concentration-dependent manner. The ANP response was inversely related to the change in cAMP efflux concentrations. The isoproterenol-induced decrease in ANP release was inhibited by β 1 -AR blockade with CGP 20712A but not by β 2 -AR blockade with ICI 118551. The isoproterenol-induced decrease in ANP release was attenuated by the L-type Ca 2+ channel antagonist nifedipine and the cAMP-dependent protein kinase inhibitor KT5720. Significance These findings suggest that β 1 -AR activation decreases ANP release via cAMP- and Ca 2+ -dependent mechanisms.

Atrial natriuretic peptide (ANP) has been reported to be locally synthesized in the ovary although its physiological roles are still unknown. To define the origin of ovarian ANP, we demonstrated the presence and release of immunoreactive (ir) ANP in pig granulosa cells and characterized its biochemical properties. Serial dilution curves made with the extracts of pig granulosa cells, their perfusates and follicular fluid were paralleled to the standard curve of ANP. The amount of irANP in the granulosa cell was 2 fg/cell. The total amount of irANP in granulosa cells significantly correlated with the levels of irANP in follicular fluid. Additionally, the total content of irANP in the follicle negatively correlated with the follicular size. On reverse phase HPLC, the major form of irANP in granulosa cells and follicular fluid was high molecular weight but that in perfusate was low molecular weight. In Northern blot analysis, ANP mRNA was detected in the pig granulosa cells. Immunohistochemistry showed ANP prohormone location in granulosa cells of rat ovary. These data strongly suggest that the granulosa cells synthesize and secrete ANP.

In order to investigate the regulatory mechanism for the atrial release of atrial natriuretic peptide (ANP), a perfused rabbit atrial model was devised. In the present experiments, the effect of a reduction in atrial distension on the immunoreactive ANP (irANP) secretion was investigated and compared in the perfused right and left atria of rats. Elevations in right and left atrial pressure resulted in proportional increases in the volume of atrial distension-reduction which was larger in the right than in the left atria. The basal rate of irANP secretion was higher in the right than in the left atria. Increases in the volume of atrial distension-reduction resulted in proportional increases in irANP secretion in both atria. Increment in irANP secretion in response to a reduction in atrial distension was significantly higher in the right than in the left atria. Higher rate of irANP secretion in response to unit volume change was observed in the right atria. Increases in the volume of atrial distension-reduction resulted in accentuated irANP responses in the right atrium. IrANP content was significantly higher in the right than in the left atria. The results suggest that the right atrium is a predominant site in ANP secretion in rats.