Demonstration of atrial natriuretic peptide/cardiodilatin (ANP/CDD)-immunoreactivity in the salt gland of the pekin duck

Summary A novel peptide hormone, atrial natriuretic factor/cardiodilatin (ANP/CDD), was recently isolated and characterized from mammalian heart. Its presence has been demonstrated in several organs that contribute to water and sodium homeostasis, such as salivary glands. This study demonstrates the presence of ANP/CDD immunoreactivity in the salt gland of Pekin ducks by high performance liquid chromatography, radioimmunoassay and immunocytochemistry, using a specific antibody against atriopeptide I. A small number of distinct, ovoid or cuboid shaped ANP/CDD-immunoreactive cells were localized in the connective tissue surrounding and separating the central secretory tubules, whereas no immunostaining was observed in the peripheral tubules. Salt glands of ducks that were adapted to salt water revealed a significant hypertrophy of their secretory lobules. However, no differences were found between the number or localization of immunoreactive cells in the salt gland of salt water-acclimatized ducks and non-stimulated glands of ducks that were housed with ad libitum access to fresh water. Our results indicate that ANP/CDD may play a role in the regulation of sodium secretion in the salt gland of aquatic birds.Supported by the Walter-Schulz-Stiftung and Deutsche Forschungsgemeinschaft DFG, We 608/8-2Dedicated to Prof. Dr. med. Dr. phil. h.c. D. Starck on the occasion of his 80th birthday     

Increase in atrial natriuretic peptide in response to physical exercise

Summary Circulating atrial natriuretic peptide (ANP) level was determined during physical exercise to investigate the correlation between changes in ANP level and heart rate increases.Six subjects exercised at a work level of 75% for 30 min, two also performed two successive exercises at 75% while two more exercised for longer at 55% · Plasma ANP levels and heart rate increased in all the exercising subjects. At the end of the exercise, the ANP level fell immediately, suggesting an immediate reduction in ANP secretion by the heart. Pre-exercise values were reached after 30 min. Successive exercises gave the same heart rate related ANP patterns without previous secretory episodes having any effect. These results lead to the conclusion that ANP intervenes in the cardiovascular adjustments to exercise.     

Development of immunoreactive atrial natriuretic peptide in fetal hearts of spontaneously hypertensive and Wistar-Kyoto rats

Summary The development of immunoreactive atrial natriuretic peptide (ANP) was studied in fetal hearts of spontaneously hypertensive (SHR) and compared to normotensive Wistar-Kyoto (WKY) rats. While SHR fetal hearts were noticeably less developed than those of WKY at 10 and 11 days gestation, both strains showed ANP immunoreactive cells in some but not all primitive heart tubes. At 12 days additional ANP immunoreactive cells appeared in formative trabeculae of the ventricle and atrium. ANP cells were also observed in the myogenic layer of the truncus and bulbus arteriosus and their derivatives from 11 through 16 days, but not at 18 days. In both strains, there were more ANP cells in the left ventricle than in right beginning at day 13. There were no obvious strain differences in the developmental pattern and timing of ANP producing cells. However, on the day of birth, staining was reduced in hearts from some WKY newborn pups compared with hearts from SHR newborns and ventricular staining was reduced in both strains when compared to fetal hearts. These observations indicate that ANP is one of the earliest peptide hormones produced and that the predisposition to genetic hypertension does not appear to influence the development of ANP.     

Changes of molecular forms of atrial natriuretic peptide after treatment for congestive heart failure

Summary In the present study, an attempt was made to clarify whether ANP molecular forms in the plasma of severe congestive heart failure patients differ from those in healthy persons and whether ANP molecular forms in the plasma of the patients were changed by successful treatment of cardiac disease.Twenty patients with congestive heart failure were treated at Kitasato University Hospital. They were classified as class III or IV by New York Heart Association criteria at the time of admission. Plasma ANP concentrations decreased after treatment from 356 ± 58.2 to 72.3 ± 14.8 pg/ml. The gel permeation chromatograms from the plasma of healthy persons showed low, or low and high molecular weight ANP peaks which correspond to the elution positions of authentic -ANP or ribonuclease A (mol.wt., 13.7 kdalton). In patients with severe congestive heart failure at a severe stage, middle molecular weight ANP consisted with the elution position of authentic-ANP was particularly noted in addition of low and high molecular weight ANP peaks. This middle molecular weight peak disappears in most of cases by successful treatment. Molecular forms in the plasma obtained from the coronary sinus and the inferior or superior vena cava were essentially the same.These results indicate that the middle molecular weight ANP supposed as-ANP may particularly be secreted in severe congestive heart failure patients.Abbreviations ANP atrial natriuretic peptide - NYHA criteria New York Heart Association criteria - SIADH Syndrome of inappropriate secretion of ADH     

Increased brain natriuretic peptide and atrial natriuretic peptide plasma concentrations in dialysis-dependent chronic renal failure and in patients with elevated left ventricular filling pressure

Brain natriuretic peptide (BNP) and atrial natriuretic peptide (ANP) plasma concentrations were measured in patients with dialysis-dependent chronic renal failure and in patients with coronary artery disease exhibiting normal or elevated left ventricular end-diastolic pressure (LVEDP) (n = 30 each). Blood samples were obtained from the arterial line of the arteriovenous shunt before, 2 h after the beginning of, and at the end of hemodialysis in patients with chronic renal failure. In patients with coronary artery disease arterial blood samples were collected during cardiac catheterization. BNP and ANP concentrations were determined by radioimmunoassay after Sep Pak C₁₈ extraction. BNP and ANP concentrations decreased significantly (P < 0.001) during hemodialysis (BNP: 192.1 ± 24.9, 178.6 ± 23.0, 167.2 ± 21.8 pg/ml; ANP: 240.2 ± 28.7, 166.7 ± 21.3, 133.0 ± 15.5 pg/ml). The decrease in BNP plasma concentrations, however, was less marked than that in ANP plasma levels (BNP 13.5 ± 1.8%, ANP 40.2 ± 3.5%; P < 0.001). Plasma BNP and ANP concentrations were 10.7 ± 1.0 and 60.3 ± 4. 0 pg/ml in patients with normal LVEDP and 31.7 ± 3.6 and 118.3 ± 9.4 pg/ml in patients with elevated LVEDP. These data demonstrate that BNP and ANP levels are strongly elevated in patients with dialysis-dependent chronic renal failure compared to patients with normal LVEDP (BNP 15.6-fold, ANP 2.2-fold, after hemodialysis; P < 0.001 or elevated LVEDP (BNP 6.1-fold, ANP 2.0-fold, before hemodialysis; P < 0.001), and that the elevation in BNP concentrations was more pronounced than that in ANP plasma concentrations. The present results provide support that other factors than volume overload, for example, decreased renal clearance, are also involved in the elevationin BNP and ANP plasma levels in chronic renal failure. The stronger elevation in BNP concentrations in patients with chronic renal failure and in patients with elevated LVEDP and the less pronounced decrease during hemodialysis suggest a different regulation of BNP and ANP plasma concentrations.[/ p]Abbreviations ANP atrial natriuretic peptide - BNP brain natriuretic peptide - LVEDP left ventricular end-diastolic pressure Correspondence to: C. Haug     

Modulation of rat olfactory bulb mitochondrial function by atrial natriuretic peptide

Summary Atrial Natriuretic Peptide (ANP) and receptors for ANP are widely distributed in many tissues and cell types in vertebrates. ANP has been shown to be internalized into the cytoplasm in several cell types and thus it raises the possibility that it may act on intracellular receptors. Displacement experiments of [¹²⁵I]-ANP binding to rat olfactory bulb mitochondrial fraction demonstrated the presence of high affinity (Kd<10^–9M) binding sites (Bmax, 112 fmol/mg protein) in this preparation. The addition of ANP (10^–8M) to this mitochondrial preparation resulted in a 25% increase in TPP⁺ accumulation, signifying a striking hyperpolarization of the mitochondrial membrane. In contrast ANP did not increase TPP⁺ uptake to liver mitochondrial preparations. This direct effect of ANP on Olfactory bulb mitochondrial membrane potential may underly the known effects of this hormone on steroidogenesis.     

Central venous pressure – a physiological stimulus for secretion of atrial natriuretic peptide in humans?

A sensitive radio-immunoassay (RIA) for the measurement of human α-atrial natriuretic peptide (ANP) in extracted plasma was developed and used in a study of the possible effect of posture on the concentration of ANP in plasma. The least detectable quantity was less than 2 pg per tube equivalent to 5 pg ml^-1 plasma. In the middle sensitivity range (approximately 50 pg per tube), the within-assay and between-assay coefficients of variation were 4.0 and 2.8%, respectively. The recovery of ANP added to plasma prior to extraction was 95–101%. High pressure liquid chromatography (HPLC) of plasma extracts revealed that endogeneous ANP was eluted in the same fractions as synthetic ANP. In order to investigate the effect of posture on the concentration of ANP in plasma six healthy volunteers were exposed to five positions in the following sequence: supine, standing, sitting, supine and 10° head-down tilt on a tilt-table. The concentration of ANP was lower in the standing and sitting position than in the supine and head-down tilted position. In another study six healthy volunteers were subjected to passive tilting on a tilt-table in order to evaluate the effect of tilting on blood pressure (BP), heart rate, central venous pressure (CVP) and the concentration of ANP in plasma. It was found that a fall in CVP was accompanied by a significant decrease in the concentration of ANP and that a rise in CVP was accompanied by a rapid increase in the concentration of ANP in plasma. The results are in agreement with the hypothesis that CVP is a physiological stimulus for the secretion of ANP.     

Origin and characterization of atrial natriuretic peptide in the rat parotid gland

Summary The heart atria represent the major site of synthesis for atrial natriuretic peptide (ANP) which exerts potent natriuretic, diuretic and vasoactive functions. Recently, ANP-immunoreactivity has been detected in extracardial organs involved in water and electrolyte homeostasis, such as the intestine and certain exocrine glands. The present study investigates ANP in the parotid gland. It was found by immunohistochemical techniques that the peptide is localized in ductal cells of the gland. An analysis of the immunoreactive material by high-pressure liquid chromatography and radioimmunoassay revealed the prohormone of ANP (ANP 1-126) and the biologically active fragment (ANP 99-126). Furthermore, Northern blot hybridization disclosed the presence of mRNA coding for ANP. It is suggested that ANP is synthesized and released from the parotid gland and functions in the control of saliva production.     

Atrial natriuretic peptide response to endurance physical training in the rat

Summary The effect of an endurance physical training programme on the plasma and atrial natriuretic peptides (ANP) and on renal glomerular ANP receptors was evaluated in male normotensive Wistar rats. Maximal O₂ uptake was significantly greater in the endurance trained (117.1 Ml O₂ · kg^–1 · min^–1, SEM 6.18 versus the control rats 84.2 ml O₂ · kg^–1 · min^–1, SEM 4.88, P<0.01. In addition, various muscle oxidative enzymes were also significantly higher in endurance trained animals. An increase in resting plasma [ANP] was observed after 11 weeks of physical training (40.02 pg · ml^–1, SEM 7.07 vs 22.8 pg.ml^–1, SEM 3.83, P<0.05). Glomerular ANP receptor density was lower in trained rats (272 fmol · mg^–1 protein, SEM 3.1 vs 380 fmol · mg^–1 protein, SEM 6.1, P < 0.05), whereas atrial tissue [ANP] was not significantly different between controls and trained animals. However, in trained rats, circulating [ANP] was closely correlated with left atrial [ANP] (r = –0.92, P<0.05). Resting systolic blood pressure had not changed at the end of this physical training programme. It is considered that under physiological conditions ANP may be involved in long-term extracellular fluid volume homeostasis through the regulation of renal glomerular ANP receptors, and that the left atrium might play a significant role in this long term fluid volume control.     

Cellular orientation of atrial natriuretic peptide in the human brain

Many peptide hormones and neurotransmitters have been detected in human neuronal tissue. The localisation of atrial natriuretic peptide (ANP) in the human brain was considered to be both interesting and relevant to the understanding of neurochemistry and brain water–electrolyte homeostasis. This vasoactive peptide hormone has been localised in rat and frog neuronal tissue. In the present study, we report the immunohistochemical localisation of ANP in autopsy samples of human brain tissue employing the avidin–biotin–peroxidase complex technique, using an antibody against a 28 amino acid fragment of human ANP. The most intense staining of immunoreactive ANP was detected in the neurones of preoptic, supraoptic and paraventricular nuclei of the hypothalamus, epithelial cells of the choroid plexus and ventricular ependymal lining cells. Immunoreactive neurones were also observed in the median eminence, lamina terminalis, infundibular and ventromedial nuclei of the hypothalamus, and in neurones of the brain stem, thalamic neurones and some neurones of the caudate nucleus. The network of ANP cells in numerous hypothalamic centres may regulate the salt and water balance in the body through a hypothalamic neuro-endocrine control system. ANP in the brain may also modulate cerebral fluid homeostasis by autocrine and paracrine mechanisms.     

Diurnal rhythms of plasma renin activity,atrial natriuretic peptide and arterial pressure during head-down bed rest in humans

The effects of prolonged head-down bed rest on the rhythms of several parameters (blood pressure, heart rate, haematocrit, plasma renin activity (PRA), atrial natriuretic peptide (ANP) were assessed in six healthy men, aged 33 (SEM 2) years, who were submitted to bed rest for 28 days (D1-28). Systolic and diastolic blood pressure (BP_s and BP_d) and heart rate were measured at 0700 and 1900 hours; circulating PRA and ANP were determined from blood samples drawn at 0800, 1000, 1200, 1500, 1800 and 2200 hours before bed rest (D – 5), D1, 2, 7, 20, 27 during bed rest and post bed rest (D + 2). The BP_s was the lowest at 0700 hours and increased at 1900 hours. There was a significant difference between values during all the measurements. The BP_d and heart rate were lower at 0700 hours before and after bed rest and no significant difference appeared between these two values during the bed rest. The PRA and ANP concentrations were more stable during bed rest, and had not returned to original rhythmicity 2 days after bed rest. The mean daily concentration of ANP decreased during bed rest. It would seem from this study that changes occur in those rhythms during bed rest.     

Cellular signaling by cyclosporine A in contractile cells: interactions with atrial natriuretic peptide

Summary Immunosuppressive therapy with cyclosporine A (CyA) may be associated with severe side effects such as nephrotoxicity and arterial hypertension. The partial reversibility of these effects suggests that they are at least in part functional. The present study examined the effects of CyA on cellular signaling in vascular smooth muscle cells and in glomerular mesangial cells and the interactions with the endogenous vasodilator atrial natriuretic peptide (ANP). Intracellular free calcium concentrations ([Ca²⁺]_i) were measured using Fura-2. ⁴⁵Ca²⁺ was used to measure Ca²⁺ efflux and cellular Ca²⁺ influx. In the presence of cyclosporine (10 g/ml), the Ca²⁺-mobilizing effects of angiotensin II (10^–8 M) in smooth muscle cells and of arginine vasopressin (AVP) in mesangial cells were significantly enhanced. CyA significantly stimulated cellular Ca²⁺ uptake in both cell types. ANP blocked the Ca²⁺ mobilization by angiotensin II and AVP and also completely inhibited the potentiating effect of CyA on angiotensin II- and AVP-induced Ca²⁺ mobilization. ANP also completely blocked the CyA-stimulated Ca²⁺ uptake. These findings suggest that CyA stimulates transmembrane Ca²⁺ influx, thereby increasing vasopressor-sensitive intracellular Ca²⁺ stores and augmenting vasopressor-induced Ca²⁺ mobilization. This cellular effect of CyA in vitro was markedly diminished by ANP. The effects of CyA on intracellular signaling may directly enhance the contractile response of smooth muscle and the glomerular mesangium to vasopressor stimuli and may also contribute to other disturbances of cell metabolism associated with CyA.Abbreviations ANP atrial natriuretic peptide - AVP arginine vasopressin - AII angiotensin II - CyA cyclosporine A - EGTA ethyleneglycoltetraacetatic acid - MEM minimal essential medium - PBS phosphate-buffered saline - PSS physiologic saline solution - SDS sodium dodecyl sulfate Dedicated to Prof. Dr. N. Zöllner on the occasion of his 70th birthday     

人类心房利钠肽基因在自发性高血压大鼠体内表达及其对血压的影响

目的研究人类心房利钠肽（hANP）基因在自发性高血压大鼠（SHRs）体内表达、持续时间和对血压的影响。方法 12只8周龄雄性SHRs,随机分为两组,于左腿股四头肌注射pcDNA3.1-hANP质粒的为实验组,在同一部位注射pcDNA3.1空质粒的为对照组,每周测量大鼠尾动脉收缩压,及利用放射免疫方法监测血中hANP水平。结果转基因后第1周起与对照组比较,实验组血压开始下降,两组动物一直相差（13±3.1）mmHg（P〈0.05）,其作用可持续10周;且放射免疫方法监测实验组血中hANP水平较高;RT-PCR和Western印迹杂交技术检测显示实验组hANP基因在肌肉组织中高效表达,对照组则未见表达。结论肌肉注射法将hANP基因导入SHRs,hANP基因可在肌肉组织中高效表达,且hANP可释放入血,降低SHRs的血压,显示了hANP基因对高血压患者治疗的可能性。     

Concomitant increase in plasma atrial natriuretic peptide and cyclic GMP during volume loading

Summary To investigate the effects of fluid expansion on endogenous atrial natriuretic peptide (ANP) and cyclic 3,5-guanosine monophosphate (cGMP), four male volunteers were studied before, during and after intravasal volume loading. Volume expansion was performed by intravenous infusion of 2,000 ml isotonic saline solution within 30 min. Mean plasma ANP levels increased 2.5-fold from 31.2 pg/ml to 81.7 pg/ml 40 min after the start of infusion. Plasma cGMP levels paralleled the rise in ANP, shwoing a mean cGMP increment from 2.7 pmol/ml to a maximum of 8.2 pmol/ml. Both ANP and cGMP levels were back to basal levels 120 min after termination of the infusion. Stimulation of endogenous ANP release by volume loading suggests that ANP is involved in the regulation of fluid homeostasis in man. The parallel rise in plasma cGMP levels supports the idea that cGMP is a mediator for the effects of ANP.Abbreviations ANP atrial natriuretic peptid - cGMP cyclic 3,5-guanosine monophosphate - PRA plasma renin activity     

Atrial natriuretic peptide in human urine

Summary A highly sensitive radioimmunoassay to measure atrial natriuretic peptide (ANP) concentration in urine has been established, and its clinical usefulness is presented. ANP in urine was stable at 4° C for several days and was easily measured by our radioimmunoassay. The average ANP excretion in 65 healthy persons was 25.0±1.4 ng/day (mean ± SEM) and the fractional excretion of ANP was 0.7±0.05%. In 14 patients with congestive heart failure, the average ANP excretion was 119.2±29.4 ng/day, which decreased to 53.3±11.0 after successful treatment.Abbreviations ANP atrial natriuretic peptide - hANP human atrial natriuretic peptide - RIA radioimmunoasay - NSB non specific bound - FE_ANP the fractional excretion of atrial natriuretic peptide - FE_Na the fractional excretion of sodium - SIADH the syngrome of inappropriate secretion of antidiuretic hormone     

Atrial natriuretic peptide levels and pulmonary artery pressure awake, at exercise and asleep in obstructive sleep apnoea syndrome

Elevated nocturnal plasma atrial natriuretic peptide (ANP) levels were found in patients with obstructive sleep apnoea (OSA). The purpose of our study was to examine the secretion of ANP during the night and to measure changes in oxygen saturation, pulmonary artery pressure and intrathoracic pressure swings in patients with OSA. Moreover, we analysed the secretion of ANP and the pulmonary artery pressure in different behavioural states, e.g. awake, at exercise and asleep. Consecutive apnoeas in non-rapid eye movement (NREM) sleep at the beginning, middle and end of the sleep study were analysed in six patients with obstructive sleep apnoea. In addition, we measured the plasma levels of ANP. The apnoea duration was significantly longer (P< 0.05) at the middle of the sleep study than at the beginning or end. Correspondingly, the end-apnoeic oxygen saturation and end-apnoeic oesophageal pressure were both significantly lower (P< 0.05) in the middle of the sleep study than at the beginning or end. No significant differences were found in the end-apnoeic systolic transmural pulmonary artery pressure (P(PATM)) and the levels of ANP. Evaluation of the ANP levels during different behavioural states revealed that the asleep levels were slightly, but not significantly, higher than the awake levels (0.235+/-0.088 vs. 0.207+/-0.057 nmol/L). However, the highest levels were found during exercise (0.334+/-0.170 nmol/L) with a significant difference compared with the awake and asleep levels. These data suggest that volume effects may be a potent factor in liberating ANP during exercise, but the role of OSA in ANP secretion when asleep is questionable.     

Influence of isoproterenol on plasma immunoreactive atrial natriuretic peptide and plasma vasopressin in the anesthetized rabbit

Changes in levels of plasma immunoreactive atrial natriuretic peptide (IR-ANP) were measured in response to administration of isoproterenol in the anesthetized, vagotomized rabbit. A dose-dependent increase in plasma IR-ANP was seen in response to 10 min isoproterenol infusions between 0.1 and 10.0 g/kg/min. The time course of these responses showed the maximum levels of IR-ANP to be attained 10 min after the cessation of infusion. In rabbits in which plasma vasopressin (AVP) levels were also measured, the maximum levels of AVP were attained during the infusion period. There was no correlation between levels of AVP and IR-ANP suggesting that AVP released into the plasma did not affect directly the release of IR-ANP. The changes in IR-ANP in response to isoproterenol were significantly reduced in rabbits which had been administered the beta-1-adrenoceptor blocking agent, atenolol. In six rabbits in which the vagi remained intact, the increases in IR-ANP were reduced and became significant only with 10 g/kg/min isoproterenol infusion. The results demonstrate that isoproterenol infusion increases the level of plasma IR-ANP in the anesthetized rabbit and suggest that this is through an effect on the heart rather than on peripheral vessels.     

Plasma atrial natriuretic peptide (ANP) concentration and fluid balance during rapid intravenous saline infusions in camels

Human muscle samples were obtained with the percutaneous biopsy technique. The samples were membrane-hyperpermeabilized (skinned) using a chemical or freeze-drying technique. Short single fibre segments were dissected from the sample, transferred to an experimental chamber, connected to a force transducer and manipulator, and exposed to temperature-controlled solutions. The force generating-capacity, the sensitivity of the contractile apparatus to calcium and the caffeine threshold for calcium release from the sarcoplasmic reticulum could be studied in the short muscle fibre segments obtained from man with the percutaneous muscle biopsy technique. The average length of the fibre segments between the connectors was 0.44±0.21 mm. Thus, detailed studies of the contractile machinery can be made on human skinned muscle fibres with only minimal discomfort to the patient or subject during biopsy, which should be useful in studies of neuromuscular disease, muscle plasticity or in applied physiology.