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Purpose: We aimed to comprehensively evaluate the curative effect of torasemide, tolvaptan, furosemide and azosemide on patients with heart failure. Methods: Relevant studies were retrieved by searching the electronic databases until May 2018. Quality assessment and data extraction of selected studies were evaluated by two reviewers. Heterogeneity across studies was assessed utilizing the I2 statistic and Q-test, and appropriate effect model was selected to calculate the pooled effect size. Network meta-analysis was conducted and the convergence degree of model was evaluated. Results: A total of 12 studies were enrolled in this study. Significant heterogeneity was not identified across the studies. Significantly greater differences were found in left ventricular ejection fraction (LVEF) for furosemide VS. azosemide, in brain natriuretic peptide (BNP) for furosemide VS. azosemide and furosemide VS. torasemide, and in adverse effects for furosemide VS. torasemide through Meta-analysis of direct comparison. In addition, network meta-analysis results suggested there were no significant differences in adverse effects, mortality, BNP and LVEF among these groups. However, the relatively low mortality and small improvement of BNP and LVEF were found in HF patients treated with torasemide. Conclusion: Torasemide might be an optimal treatment for HF patients considering its comprehensive curative effect.

A long-acting loop diuretic, azosemide, has been shown to improve long-term prognosis in patients with heart failure compared with a short-acting loop diuretic, furosemide. However, the therapeutic advantages of azosemide over furosemide have not been clearly established. In this study, we retrospectively analyzed clinical outcomes and laboratory data in patients with congestive heart failure treated with furosemide or azosemide, and the efficacy of these agents was compared. First, we screened 1900 patients and selected 124 (furosemide group: n = 40; azosemide group: n = 84) as the total study population. From these patients, we next selected 72 patients for the propensity score-matched analysis (furosemide group: n = 36; azosemide group: n = 36). The incidence of all-cause death and rehospitalization due to worsening heart failure during 24 months of follow-up was similar between the furosemide and azosemide groups in both the total study population and the propensity score-matched population. However, in the propensity score-matched analysis, the estimated glomerular filtration rate time-dependently decreased during 36 months of follow-up in the furosemide group (56.5 ± 19.5-43.2 ± 16.3 mL/min/1.73 m), whereas it did not change in the azosemide group (58.6 ± 22.0-50.3 ± 17.8 mL/min/1.73 m) (P = 0.032). Azosemide might have some potential advantage for renal protection over furosemide in patients with congestive heart failure.

BackgroundLoop diuretics are commonly used in clinical practice to manage high fluid loads and to control fluid balance. In this paper, a novel quantitative analysis method for multiple components with a single marker (QAMS) was developed for the simultaneous determination of 5 diuretic drugs furosemide, torasemide, azosemide, etacrynic acid, and bumetanide, by HPLC. Qualitative analysis was performed using relative retention time and ultraviolet (UV) spectral similarity as the double indicator. The QAMS method was conducted with etacrynic acid as an internal reference substance. The quantities of the other four diuretics were calculated by using the relative correction factors for etacrynic acid. The quantities of the 5 diuretic drugs were also determined by the external standard method (ESM). Chromatographic separation was achieved on a Shimadzu HC-C18column (150 mm × 4.6 mm, 5 µm) using 50 mM potassium dihydrogen phosphate (pH adjusted to 4.0 with phosphoric acid) with acetonitrile (64:36, v/v) as the mobile phase at a flow rate of 1.0 mL/min and a column temperature of 30 ℃.ResultsUnder these conditions, the 5 diuretic drugs were well separated, showing linear relationships within certain ranges. The quantitative results showed that there was no significant difference between the QAMS and ESM methods.ConclusionsOverall, the HPLC-QAMS analytical scheme established in this study is a simple, efficient, economical, and accurate method for the quantitative evaluation of 5 diuretic drugs.

The sodium-potassium-chloride (Na-K-Cl) cotransporter NKCC1 is found in the plasma membrane of a wide variety of cell types, including neurons, glia and endothelial cells in the brain. Increased expression of neuronal NKCC1 has been implicated in several brain disorders, including neonatal seizures and epilepsy. The loop diuretic and NKCC inhibitor bumetanide has been evaluated as an antiseizure agent alone or together with approved antiseizure drugs such as phenobarbital (PB) in pre-clinical and clinical studies with varying results. The equivocal efficacy of bumetanide may be a result of its poor brain penetration. We recently reported that the loop diuretic azosemide is more potent to inhibit NKCC1 than bumetanide. In contrast to bumetanide, azosemide is not acidic, which should favor its brain penetration. Thus, azosemide may be a promising alternative to bumetanide for treatment of brain disorders such as epilepsy. In the present study, we determined the effect of azosemide and bumetanide on seizure threshold in adult epileptic mice. A structurally related non-acidic loop diuretic, torasemide, which also blocks NKCC1, was included in the experiments. The drug effects were assessed by determing the maximal electroshock seizure threshold (MEST) in epileptic vs. nonepileptic mice. Epilepsy was induced by pilocarpine, which was shown to produce long-lasting increases in NKCC1 in the hippocampus, whereas MEST did not alter NKCC1 mRNA in this region. None of the three loop diuretics increased MEST or the effect of PB on MEST in nonepileptic mice. In epileptic mice, all three diuretics significantly increased PB's seizure threshold increasing efficacy, but the effect was variable upon repeated MEST determinations and not correlated with the drugs' diuretic potency. These data may indicate that inhibition of NKCC1 by loop diuretics is not an effective means of increasing seizure threshold in adult epilepsy.

The majority of patients with chronic heart failure (HF) receive long-term treatment with loop diuretics. The comparative effectiveness of different loop diuretics is unknown. We searched PubMed, clinicaltrials.gov , the Cochrane Central Register of Controlled Trials and the European Union Clinical Trials Register for randomised clinical trials exploring the efficacy of the loop diuretics azosemide, bumetanide, furosemide or torasemide in patients with HF. Comparators included placebo, standard medical care or any other active treatment. The primary endpoint was all-cause mortality. Secondary endpoints included cardiovascular mortality, HF-related hospitalisation and any combined endpoint thereof. Hypokalaemia and acute renal failure were defined as additional safety endpoints. Evidence was synthesised using network meta-analysis (NMA). Thirty-four trials reporting on 2647 patients were included. The overall quality of evidence was rated as moderate. NMA demonstrated no significant differences between loop diuretics with respect to all-cause mortality, cardiovascular mortality or hypokalaemia. In contrast, torasemide ranked best in terms of HF hospitalisation, and there was a trend towards benefits with torasemide with regard to occurrence of acute renal failure. Sensitivity analyses excluding trials with a follow-up

Background: Clinical evidence of the effects of loop diuretics in patients with heart failure with preserved ejection fraction (HFpEF) is lacking. Thus, we compared the impact of azosemide and furosemide, long- and short-acting loop diuretics, in patients with HFpEF. Methods and Results: A prospective multicenter cohort study was conducted between July 2014 and July 2018. We enrolled 301 consecutive patients with HFpEF (median age, 84 years; IQR, 79-88 years; 54.8% female). Azosemide was used in 127 patients (azosemide group), and furosemide in 174 (furosemide group). We constructed Cox models for a composite of cardiac death, non-fatal myocardial infarction, non-fatal stroke, and HF hospitalization (primary endpoints). During a median follow-up of 317 days (IQR, 174-734 days), the primary endpoint occurred in 112 patients (37.2%). On multivariate inverse probability of treatment weighted (IPTW) Cox modeling, the azosemide group had a significantly lower incidence of adverse events than the furosemide group (hazard ratio [HR], 0.46; 95% confidence interval [CI]: 0.27-0.80; P=0.006). Furthermore, on multivariate IPTW Cox modeling for the secondary endpoints, cardiac death (HR, 0.38; 95% CI: 0.17-0.89; P=0.025) and unplanned hospitalization for decompensated HF (HR, 0.50; 95% CI: 0.28-0.89; P=0.018) were also reduced in the azosemide group. Conclusions: Azosemide significantly reduced the risk of adverse events compared with furosemide in HFpEF patients.

Patients with stage D heart failure (HF) frequently become dependent on high doses of diuretics and inotropic agents. Recently, a sodium-glucose cotransporter 2 inhibitor (SGLT2i), an oral antidiabetic agent, has been demonstrated to have favorable effects in preventing HF. However, it remains unknown whether SGLT2i is reliable for patients with decompensated HF. We experienced a case of a patient with stage D HF for whom attempting intravenous dobutamine withdrawal was difficult even after the administration of all conventional pharmacological treatment. Administration of canagliflozin produced an additive diuretic action and correction of volume overload in combination with azosemide and tolvaptan, and resulted in successful withdrawal of dobutamine. Thus, SGLT2i might be promising for the treatment of patients with congestive HF who are refractory to conventional diuretic treatment.

There have been no reports evaluating the impact of long-acting loop diuretics (LLD) on the outcome of heart failure (HF) and arrhythmia treatment in HF with reduced ejection fraction (HFrEF) patients implanted with a cardiac resynchronization therapy (CRT) device.This was a prospective, single-blind, randomized crossover study. We allocated 21 consecutive CRT implanted patients into 2 groups. The furosemide group received furosemide as a first treatment and azosemide as a second treatment. The azosemide group received this treatment in the reverse order. The first treatment was given to each group for 6 months and the second treatment continued for an additional 6 months. We combined the data of each medication regimen in each group and analyzed it at baseline, 6 months, and 1 year. The primary endpoints were the variation of fluid index and thoracic impedance measured by CRT at 6 months.The baseline characteristics were similar for both groups. The difference in the primary endpoints was not statistically significant between the 2 medication arms (fluid index: -29.6 ± 64.4 versus 16.2 ± 48.2; P = 0.22, thoracic impedance: -0.49 ± 17.8 versus 2.45 ± 12.5; P = 0.56). Likewise, the clinical outcome of HF and the CRT derived parameters in both arms were comparable.HFrEF patients taking LLD after CRT implantation might be comparable to those taking short-acting loop diuretics in the treatment of HF and HF-associated arrhythmias.

Because of its potent inhibitory effect on the Na+-K+-2Cl- symporter isotype 1 (NKCC1) in brain neurons, bumetanide has been tested with varying results for treatment of seizures that potentially evolve as a consequence of abnormal NKCC1 activity. However, because of its physicochemical properties, bumetanide only poorly penetrates into the brain. We previously demonstrated that NKCC1 can be also inhibited by azosemide and torasemide, which lack the carboxyl group of bumetanide and thus should be better brain-permeable. Here we studied the brain distribution kinetics of azosemide and torasemide in comparison with bumetanide in mice and used pharmacokinetic-pharmacodynamic modelling to determine whether the drugs reach NKCC1-inhibitory brain concentrations. All three drugs hardly distributed into the brain, which seemed to be the result of probenecid-sensitive efflux transport at the blood-brain barrier. When fractions unbound in plasma and brain were determined by equilibrium dialysis, only about 6-17% of the brain drug concentration were freely available. With the systemic doses (10 mg/kg i.v.) used, free brain concentrations of bumetanide and torasemide were in the NKCC1-inhibitory concentration range, while levels of azosemide were slightly below this range. However, all three drugs exhibited free plasma levels that would be sufficient to block NKCC1 at the apical membrane of brain capillary endothelial cells. These data suggest that azosemide and torasemide are interesting alternatives to bumetanide for treatment of seizures involving abnormal NKCC1 functionality, particularly because of their longer duration of action and their lower diuretic potency, which is an advantage in patients with seizures.

The cellular mechanisms of calcification in sea urchin larvae are still not well understood. Primary mesenchyme cells within the larval body cavity form a syncytium to secrete CaCO3 spicules from intracellular amorphous CaCO3 (ACC) stores. We studied the role of Na(+)K(+)2Cl(-) cotransporter (NKCC) in intracellular ACC accumulation and larval spicule formation of Strongylocentrotus droebachiensis. First, we incubated growing larvae with three different loop diuretics (azosemide, bumetanide, and furosemide) and established concentration-response curves. All loop diuretics were able to inhibit calcification already at concentrations that specifically inhibit NKCC. Calcification was most effectively inhibited by azosemide (IC50=6.5 μM), while larval mortality and swimming ability were not negatively impacted by the treatment. The inhibition by bumetanide (IC50=26.4 μM) and furosemide (IC50=315.4 μM) resembled the pharmacological fingerprint of the mammalian NKCC1 isoform. We further examined the effect of azosemide on the maintenance of cytoplasmic cords and on the occurrence of calcification vesicles using fluorescent dyes (calcein, FM1-43). Fifty micromolars of azosemide inhibited the maintenance of cytoplasmic cords and resulted in increased calcein fluorescence within calcification vesicles. The expression of NKCC in S. droebachiensis was verified by PCR and Western blot with a specific NKCC antibody. In summary, the pharmacological profile of loop diuretics and their specific effects on calcification in sea urchin larvae suggest that they act by inhibition of NKCC via repression of cytoplasmic cord formation and maintenance.

Chloride was recently recognized to play an important role in the pathophysiology of heart failure (HF). Chloride manipulation, including the use of acetazolamide, may be a requisite therapeutic target in HF treatment. An 87-year-old male patient with advanced HF and hypertrophic cardiomyopathy was admitted to the hospital due to hypochloremia (94 mEq/L) and hyponatremia (134 mEq/L) under diuretic treatment with azosemide, spironolactone, and tolvaptan. On admission, HF-related signs of overhydration were lacking, but B-type natriuretic peptide was moderately elevated. The etiology of the hypochloremia/natremia state was depletion of both electrolytes based on serum analysis and spot urinary concentrations. Immediately after admission, acetazolamide (500 mg/d) was prescribed to correct the hypochloremia in parallel with cessation of the preceding administration of azosemide and spironolactone, and tapering off of the tolvaptan over 7 days. Under treatment, both serum chloride and sodium concentrations recovered to normal (108 mEq/L and 148 mEq/L, respectively), and the serum potassium concentration decreased from 3.9 mEq/L to 2.4 mEq/L. Urinary concentrations of sodium and potassium increased from 18 mEq/L to 31 mEq/L and from 19 mEq/L to 51.5 mEq/L respectively, in concordance with the changes in serum concentrations, but the chloride concentration decreased from 18 mEq/L to 12 mEq/L, opposite the changes in the serum concentration.

Background We previously reported that the long-acting loop diuretic azosemide improves cardiac sympathetic nerve activity (CSNA) in patients with chronic heart failure (CHF), compared with short-acting furosemide. However, its influence on mortality has not been determined. Methods The present study was a further analysis of our previously published study in which azosemide showed improved 123 I-metaiodobenzylguanidine scintigraphic findings compared with furosemide in CHF patients with reduced left ventricular ejection fraction (LVEF). Patients with CHF were identified according to their histories of acute decompensated heart failure requiring hospitalization. A total of 108 patients were selected and propensity score matching was used to compare patients treated with azosemide ( n =54) or furosemide ( n =54). Results During the median follow-up period of 5.22years, 24 out of 108 patients experienced cardiac death events. In multivariate Cox regression analysis (adjusted for age, non-beta-blocker treatments, and impaired CSNA), furosemide treatment was an independent predictor of cardiac death events ( p =0.034, hazard ratio 2.624, 95% confidence interval 1.074 to 6.047). On Kaplan-Meier analysis, the cardiac death-free rate in the azosemide group was significantly higher than that in the furosemide group ( p Conclusions These findings indicate the superior effectiveness of azosemide in reducing mortality compared with furosemide.

We have previously reported that a long-acting loop diuretic, azosemide, reduces cardiovascular risks in patients with chronic heart failure (CHF) as compared with a short-acting one, furosemide, in Japanese Multicenter Evaluation of LOng- versus short-acting Diuretics In Congestive heart failure (J-MELODIC). However, the mechanisms of the difference have not been elucidated. This study aimed to examine whether there is a difference in the reduction in plasma brain natriuretic peptide (BNP) level and in left ventricular (LV) functional recovery between the CHF patients treated with the long-acting diuretic (the azosemide group) and the short-acting diuretic (the furosemide group). We reviewed changes in plasma BNP level and echo-assessed LV functional parameters from baseline to a year after the entry in 288 CHF patients with New York Heart Association class II or III symptoms that joined J-MELODIC. The decrease in plasma BNP levels was larger in the azosemide group than in the furosemide group (p

Summary Background Loop diuretics have two different classes with different duration of activity: short-acting such as furosemide (duration of activity, 6 h) and long-acting such as azosemide (duration of activity, 10–12 h). We conducted a multicenter, randomized, controlled trial in order to compare the therapeutic effects of azosemide, a long-acting loop diuretic, and furosemide, a short-acting one, on neurohumoral factors and cardiac function in outpatients with chronic heart failure (CHF). Methods We enrolled 98 patients with CHF who were receiving furosemide and an angiotensin-converting enzyme inhibitor, and they were randomly divided into furosemide ( n = 49) and azosemide ( n = 49) groups. The furosemide group continued furosemide at the same dosage, and the azosemide group switched from furosemide to azosemide. At baseline and after 3 months, we measured body weight, and levels of brain natriuretic peptide (BNP), atrial natriuretic peptide (ANP), norepinephrine, active renin, creatinine, blood urea nitrogen, sodium, potassium, and hematocrit. Chest X-ray and echocardiography were also performed. Results Body weight and plasma levels of BNP and ANP significantly decreased after 3 months in the azosemide group compared to the furosemide group. There were no significant differences in changes of levels of creatinine, blood urea nitrogen, sodium, potassium, hematocrit, norepinephrine, and active renin after 3 months between the furosemide and azosemide groups. Echocardiography and chest X-ray did not demonstrate significant differences between the two groups. Conclusions Long-acting azosemide is suggested to be useful for the improvement of neurohumoral factors compared with short-acting furosemide in patients with CHF.

Background: Diuretics are the most prescribed drug in heart failure (HF) patients. However, clinical evidence about their long-term effects is lacking. The purpose of this study was to compare the therapeutic effects of furosemide and azosemide, a short- and long-acting loop diuretic, respectively, in patients with chronic heart failure (CHF). Methods and Results: In this multicenter, prospective, randomized, open, blinded endpoint trial, we compared the effects of azosemide and furosemide in patients with CHF and New York Heart Association class II or III symptoms. 320 patients (160 patients in each group, mean age 71 years) were followed up for a minimum of 2 years. The primary endpoint was a composite of cardiovascular death or unplanned admission to hospital for congestive HF. During a median follow-up of 35.2 months, the primary endpoint occurred in 23 patients in the azosemide group and in 34 patients in the furosemide group (hazard ratio [HR], 0.55, 95% confidence interval [CI] 0.32-0.95: P=0.03). Among the secondary endpoints, unplanned admission to hospital for congestive HF or a need for modification of the treatment for HF were also reduced in the azosemide group compared with the furosemide group (HR, 0.60, 95%CI 0.36-0.99: P=0.048). Conclusions: Azosemide, compared with furosemide, reduced the risk of cardiovascular death or unplanned admission to hospital for congestive HF. (Circ J 2012; 76: 833-842)

Patients with chronic kidney disease （CKD） were treated with a long-acting loop di- di-（di-） diuretic, Azosemide and a short-acting loop diuretic, Furosemide in a crossover fashion. The two drugs were compared after crossover switch using a multicenter study design focusing on evaluation of diuretic effects and symptoms associated with quality of life. There was no significant difference between Azosemide and Furosemide treatment after crossover switch in alleviation of edema, blood pressure, and body weight, or change in daily urinary volume or sodium excretion. On the other hand, change in urinary protein excretion from start to end of treatment was less in Azosemide treatment （－0.239g/day） than in Furo-（－Furo-－Furo-） Furo- Furosemide treatment （0.353g/day） after crossover switch （p=0.0556）. In a questionnaire sur-（sur- sur-） sur-（sur-）. sur-. survey of symptoms after treatment, the percentage of patients complaining of “hand cramp” was higher after Azosemide than after Furosemide treatment; however this difference was not statistically significant （p=0.0935）. When asked “Which drug do you wish to contin-（contin-contin-）. contin-. continue?”, 30% selected Furosemide, 35% selected Azosemide and 35% gave a neutral answer. In conclusion, Azosemide was similar to Furosemide in efficacy and tolerability.

Objective—To determine the diuretic effects and changes in plasma aldosterone concentration (PAC) following oral administration of a single dose of furosemide or azosemide in healthy dogs. Animals—8 mixed-breed dogs. Procedures—A single dose of furosemide (2 mg/kg), azosemide (1, 5, or 10 mg/kg), or placebo (bifidobacterium [1 mg/kg]) was administered orally (in random order at 7-day intervals) to each dog (5 treatments/dog). Urine and blood samples were collected before (2 hours after evacuation of the urinary bladder; baseline) and at intervals for 24 hours after drug treatment to assess urine volume and plasma and urine biochemical variables. Results—Compared with baseline values, treatment with furosemide and azosemide (5 and 10 mg/kg) increased urine output for 1 to 2 hours and 2 to 4 hours, respectively. The 24-hour urine volume and urinary sodium excretion were significantly increased following furosemide and azosemide (5 and 10 mg/kg) treatments, compared with effects of pla-cebo; these increases were dose dependent for azosemide, and increases were similar for furosemide and the 5 mg/kg dose of azosemide. Compared with other treatments, 24-hour urinary potassium excretion was significantly increased with azosemide at 10 mg/kg. Azosemide (5 and 10 mg/kg) significantly increased plasma total protein concentration and decreased plasma potassium concentration, compared with baseline values. Compared with the effect of placebo, PAC was significantly increased by furosemide and the 10 mg/kg dose of azosemide. Conclusions and Clinical Relevance—In healthy dogs, a moderate dose of azosemide caused sufficient diuretic action and increased PAC to a lesser extent than furosemide.

We tested the hypothesis that furosemide accelerates the progression of left ventricular systolic dysfunction in a tachycardia-induced porcine model of heart failure.Furosemide activates the renin-angiotensin-aldosterone system in patients with congestive heart failure (CHF). Such activation may contribute to CHF progression, but prospective data are lacking.Thirty-two Yorkshire pigs were randomized to furosemide (1 mg/kg intramuscularly daily, mean 16.1 +/- 0.9 mg) or placebo. Thereafter, a pacing model of heart failure was utilized to produce systolic dysfunction in both sets of animals (fractional shortening0.16 by echocardiogram). The goal was to determine if furosemide would accelerate the progression of left ventricular dysfunction in the "treated" group. After sacrifice, sodium-calcium exchanger currents and their responsiveness to isoproterenol were measured during voltage clamp. All investigators were blinded to treatment assignment.Furosemide shortened the time to left ventricular dysfunction (35.1 +/- 5.1 days in placebo versus 21.4 +/- 3.2 days for furosemide animals; p = 0.038, log-rank test). By day 14, aldosterone levels were significantly higher in furosemide animals (43.0 +/- 11.8 ng/dl vs. 17.6 +/- 4.5 ng/dl; p0.05). Serum sodium was reduced (133.0 +/- 0.9 mmol/l furosemide vs. 135.7 +/- 0.8 mmol/l placebo; p0.05), but no difference in norepinephrine, potassium, magnesium, creatinine, or urea nitrogen was present. Basal sodium-calcium exchanger currents were significantly increased and isoproterenol responsiveness depressed by furosemide.Tachycardic pigs given furosemide had significant acceleration of both contractile and metabolic features of CHF, including left ventricular systolic dysfunction, elevated serum aldosterone levels, and altered calcium handling in a controlled experimental model of heart failure.

Objectives We sought to determine whether non–potassium-sparing diuretics (PSDs) in the absence of a PSD may result in progressive heart failure (HF). Background Angiotensin-converting enzyme (ACE) inhibitors incompletely suppress ACE activity in HF patients. Furthermore, non-PSDs are activators of aldosterone secretion. We reasoned that non-PSDs, in the absence of a PSD, might result in progressive HF. Methods In the 6,797 patients in the Studies Of Left Ventricular Dysfunction (SOLVD), we compared the risk of hospitalization for, or death from, HF between those taking a PSD and those who were not, adjusting for known covariates. Results The risk of hospitalization from worsening HF in those taking a PSD relative to those taking only a non-PSD was 0.74 (95% confidence interval [CI] 0.55 to 0.99; p = 0.047). The relative risk for cardiovascular death was 0.74 (95% CI 0.59 to 0.93; p = 0.011), for death from all causes 0.73 (95% CI 0.59 to 0.90; p = 0.004), and for hospitalization for, or death from, HF 0.75 (95% CI 0.58 to 0.97; p = 0.030). Compared with patients not taking any diuretic, the risk of hospitalization or death due to worsening HF in patients taking non-PSDs alone was significantly increased (risk ratio [RR] = 1.31, 95% CI 1.09 to 1.57; p = 0.0004); this was not observed in patients taking PSDs with or without a non-PSD (RR = 0.99, 95% CI 0.76 to 1.30; p = 0.95). Conclusions The use of PSDs in HF patients is associated with a reduced risk of death from, or hospitalization for, progressive HF or all-cause or cardiovascular death, compared with patients taking only a non-PSD.

This paper reports 1) the increase in expression of CYP1A2 in mutant Nagase analbuminemic rats (NARs), 2) the role of globulin binding of azosemide in circulating blood in its urinary excretion and hence its diuretic effects in NARs, and 3) the significantly faster renal (CL(R)) and nonrenal (CL(NR)) clearances of azosemide in NARs. Azosemide (mainly metabolized via CYP1A2 in rats), 10 mg/kg, was intravenously administered to control rats and NARs. Northern and Western blot analyses revealed that the expression of CYP1A2 increased approximately 3.5-fold in NARs as compared with control. The plasma protein binding of azosemide in control rats and NARs was 97.9 and 84.6%, respectively. In NARs, plasma protein binding (84.6%) was due to binding to alpha- (82.6%) and beta- (68.9%) globulins. In NARs, the amount of unchanged azosemide excreted in 8-h urine was significantly greater (37.7 versus 21.0% of intravenous dose) than that in control rats due to an increase in intrinsic renal active secretion of azosemide. Accordingly, the 8-h urine output was significantly greater in NARs. The area under the plasma concentration-time curve of azosemide was significantly smaller (505 versus 2790 microg. min/ml) in NARs because of markedly faster CL(R) (7.36 versus 0.772 ml/min/kg, secondary to a significant increase in urinary excretion of azosemide and intrinsic renal active secretion). Additionally, CL(NR) was significantly faster (12.4 versus 3.05 ml/min/kg, because of approximately 3.5 fold increase in CYP1A2) in NARs compared with control. Based on in vitro hepatic microsomal studies, the intrinsic M1 [a metabolite of azosemide; 5-(2-amino-4-chloro-5-sulfamoylphenyl)-tetrazole] formation clearance was significantly faster (67.0% increase) in NARs than that in control rats, and this supports significantly faster CL(NR) in NARs. Renal sensitivity to azosemide was significantly greater in NARs than in control rats with respect to 8-h urine output (385 versus 221 ml/kg) and 8-h urinary excretions of sodium, potassium, and chloride. This study supports that in NARs, binding of azosemide to alpha- and beta-globulins in circulating blood play an important role in its diuretic effects.

In the original publication of the article, the corresponding author’s name was incorrectly published. The corresponding author’s name should appear as Ashok Kumar.

Gender differences in pharmacokinetics and pharmacodynamics of azosemide were evaluated after intravenous, 10 mg kg(-1), and oral, 10 mg kg(-1), administration to male and female rats. After intravenous administration to male rats, the percentages of intravenous dose of azosemide recovered from entire gastrointestinal tract at 24 h (13.2 versus 3.93%) was significantly greater than those in female rats. In male rats, the nonrenal clearance of azosemide tended (p

The pharmacokinetic and pharmacodynamic differences of azosemide were investigated after intravenous (i.v.) and oral administration of azosemide, 10 mg kg-1, to the control and uranyl nitrate-induced acute renal failure (U-ARF) rats. After IV administration, the plasma concentrations of azosemide were significantly higher in the U-ARF rats and this resulted in a significant increase in AUC (2520 versus 3680 micrograms min mL-1) and significant decrease in Cl (3.96 versus 2.72 mL min-1 kg-1) of azosemide. The significant decrease in Cl in the U-ARF rats was due to the significant decrease in Clr of azosemide (1.55 versus 0.00913 mL min-1 kg-1) due to the decrease in kidney function in the U-ARF rats. After IV administration, the urine output (38.5 versus 8.45 mL 100 g-1 body weight) and urinary excretion of sodium (4.60 versus 0.420 mmol 100 g-1 body weight) decreased significantly in the U-ARF rats. After oral administration, the AUC0-8 h of azosemide decreased significantly (215 versus 135 micrograms min mL-1) in the U-ARF rats possibly due to the decreased GI absorption of azosemide. After oral administration, the 24-h urine output decreased considerably (16.1 versus 11.2 mL 100 g-1 body weight, p < 0.098) and the 24-h urinary excretion of sodium (1.74 versus 0.777 mmol 100 g-1 body weight) decreased significantly in the U-ARF rats. The i.v. and oral doses of azosemide needed to be modified in the acute renal failure patients if the present rat data could be extrapolated to humans.

The effects of differences in the rate and composition of intravenous fluid replacement for urine loss on the pharmacokinetics and pharmacodynamics of azosemide were evaluated using rabbit as the animal model. Each rabbit received a 4h constant intravenous infusion of 1 mg kg-1 azosemide with 0% replacement (treatment I, n = 4), 50% replacement (treatment II, n = 5), and 100% replacement (treatment III, n = 5) with lactated Ringer's solution, as well as with 100% replacement with 5% dextrose in water (D-5-W, treatment IV; n = 5). Renal clearance and urinary excretion rate of the drug in treatment III were considerably higher than those in treatments I, II, and IV. In spite of the similarities in kinetic properties, diuretic and/or natriuretic effects of azosemide were markedly different among the four treatments. For example, the mean 8 h urine output values were 98.2, 178, 733, and 237 mL for treatments I-IV, respectively, and the corresponding values for sodium excretion were 11.1, 19.4, 76.4, and 14.2 mmol, and for chloride 13.4, 23.8, 78.9, and 17.1 mmol. Except for treatment III, diuresis and/or natriuresis were found to be time dependent, generally decreasing with time until reaching a low plateau during the later hours of infusion. The present findings also show that (i) no fluid replacement and 100% replacement with D-5-W both produce the same degree (not significantly different) of severe acute tolerance in natriuresis, indicating the insignificance of water compensation in tolerance development; (ii) in treatment II, where neutral sodium balance was achieved, the development of acute tolerance in diuresis can mainly be attributed to negative water balance under this special condition; and (iii) at steady state the hourly diuresis and natriuresis can differ up to about 6.87- and 5.21-fold between treatments. Some implications for the bioequivalence evaluation of dosage forms of azosemide are discussed.

The effects of pretreatment with the enzyme inducers phenobarbital (PB) and 3-methylcholanthrene (3-MC) and the enzyme inhibitor chloramphenicol (CM) on the pharmacokinetic and pharmacodynamic parameters of azosemide were examined after intravenous (IV) administration of azosemide, 10 mg kg−1, to rats. The nonrenal clearance (1·63 versus 3·30 mL min−1 kg−1) of azosemide increased significantly in 3-MC pretreated rats. This suggested that the nonrenal metabolism of azosemide increased by pretreatment with 3-MC. This relationship was supported by the significant decrease in 24 h urinary excretion of unchanged azosemide in 3-MC pretreated rats (54·1 versus 41·1% of IV dose). This relationship was also supported at least in part by the results of a liver homogenate study; the amount of azosemide remaining per gram of liver decreased significantly (48·2 versus 43·0 μg) and the amount of M1 formed increased significantly (4.88 versus 6.66 μg when expressed in terms of azosemide) in 3-MC pretreated rats after 30 min incubation of 50 μg azosemide in 9000 g supernatant fractions of liver homogenates. The content of hepatic cytochrome P-450 (0·751 versus 1·57 nmol/mg protein) and the weight of liver (3.53 versus 4·20% of body weight) increased significantly in 3-MC pretreated rats, suggesting that the metabolizing enzyme(s) for azosemide seemed to be induced by pretreatment with 3-MC. The 8 h urine output (29·2 versus 18·1 mL) and 8 h urinary excretion of sodium (4·02 versus 2·39 mmol) and chloride (4·01 versus 2·73 mmol) per 100 g body weight decreased significantly in 3-MC pretreated rats. However, the diuretic, natriuretic, kaluretic, and chloruretic efficiencies were not significantly different between the control and 3-MC pretreated rats. The pharmacokinetic and pharmacodynamic parameters of azosemide were not significantly different between the control and PB pretreated rats, and similar results were also obtained from the control and CM pretreated rats. The above data indicate that the metabolizing enzyme(s) for azosemide seem(s) to be neither induced by PB pretreatment nor inhibited by CM pretreatment. However, the content of hepatic cytochrome P-450 and the weight of liver increased significantly in PB pretreated rats, while the values were not significantly different between the control and CM pretreated rats. © 1997 John Wiley & Sons, Ltd.

The pharmacokinetics and pharmacodynamics of torasemide were evaluated after intravenous administration of the same total dose of torasemide at a dose of 1mg/kg to rabbits with different infusion times, 1 min (treatment I), 30 min (treatment II) and 2 h (treatment III). The loss of water and electrolytes in urine induced by torasemide was immediately replaced with infusion of an equal volume of lactated Ringer's solution. All the pharmacokinetic parameters of torasemide, such as total area under the plasma concentration-time curve from time zero to time infinity (AUC), total body clearance (CL), apparent volume of distribution at steady state (Vss), terminal half-life and mean residence time (MRT), were independent of infusion times. However, the 8 h urine output (235, 534 and 808 ml) and 8 h urinary excretion of sodium (24.2, 80.1 and 89.2 mmol) and chloride (27.1, 89.2 and 94.0 mmol) were significantly greater in treatments II and III than those in treatment I, although the total 8 h urinary excretion of unchanged torasemide (1210, 1210 and 1310 microg) were not significantly different among the three treatments. This could be due to the higher diuretic efficiencies in treatments II and III.

ObjectivesTo report (1) the pharmacokinetics of drugs that are mainly metabolized via hepatic cytochrome P450s (CYPs) or mainly excreted via the urine and bile, (2) the mechanism for the urinary excretion of drugs (such as glomerular filtration or renal active secretion or re-absorption), and (3) the diuretic effect of some loop diuretics in mutant Nagase analbuminaemic rats (NARs), an animal model for human familial analbuminaemia based on the pharmacokinetics of drugs reported in the literatures.Key findingsIn NARs, the changes in the time-averaged non-renal clearances (CLNRs) of drugs that are mainly metabolized via CYPs were explained in terms of changes in the hepatic intrinsic clearance (mainly because of changes in CYPs), free (unbound) fractions of drugs in the plasma (fp) and hepatic blood-flow rate (QH) depending on the hepatic excretion ratios of drugs.SummaryThe CLNR changes of drugs mainly metabolized via hepatic CYPs can be sufficiently explained by the three earlier mentioned factors. The plasma albumin (furosemide) or globulin (azosemide, bumetanide and torasemide) binding affects their diuretic effects.

Because physiological changes occurring in diabetes mellitus patients could alter the pharmacokinetics and pharmacodynamics of the drugs used to treat the disease, the pharmacokinetics and pharmacodynamics of azosemide were investigated after intravenous and oral administration of the drug (10 mg kg−1) to control and alloxan-induced diabetes mellitus rats (AIDRs). After intravenous administration of azosemide to the AIDRs, the area under the plasma concentration-time curve (AUC) increased considerably (3120 compared with 2520 μg min mL−1; P < 0.135) and the total body clearance decreased considerably (3.20 compared with 3.96 mL min−1 kg−1; P < 0.0593). The considerable reduction in time-averaged total body clearance in the AIDRs was a result of the significant decrease in renal clearance (1.01 compared with 1.55 mL min−1 kg−1) in the AIDRs, the non-renal clearance being comparable between the two groups of rats. After intravenous administration, the 8-h urinary excretion of azosemide (29.5 compared with 40% of intravenous dose; P < 0.0883) and one of its metabolites, M1 (2.15 compared with 2.60% of intravenous dose, expressed in terms of azosemide; P < 0.05) decreased in the AIDRs because of the impaired kidney function. The diuretic, natriuretic, kaliuretic and chloruretic efficiencies increased significantly in the AIDRs. After oral administration of azosemide, AUC decreased significantly in the AIDRs (115 compared with 215 μg min mL−1) possibly because of the reduced gastrointestinal absorption of azosemide in the AIDRs. After oral administration of azosemide, the 8-h urine output decreased significantly in the AIDRs (9.32 compared with 16.1 mL per 100 g body weight) because of the significantly reduced 8-h urinary excretion of azosemide (3.00 compared with 9.14% of oral dose). After both intravenous and oral administration some pharmacokinetic and pharmacodynamic parameters of azosemide were significantly different in AIDRs.

We report a case of successful treatment with tolvaptan (15 mg/day) in a 73-year-old female patient with chronic kidney disease (CKD) stage 5 due to diabetic nephropathy and renal sclerosis for volume control and loop diuretic-induced hyponatremia. Her creatinine clearance has remained at 7–10 ml/min for the last 6 months. She was treated by dietary and drug therapy, namely, antihypertensives (nifedipine: 40 mg/day, olmesartan: 20 mg/day) and loop diuretics (azosemide: 40–120 mg/day), for CKD and concomitant diseases of hypertension and diabetic mellitus. She developed loop diuretic-induced hyponatremia (120 mmol/l) by increased sodium excretion, but the diuretic was required for the control of volume overload. Hence, azosemide was suspended and tolvaptan (15 mg/day) was administered. After tolvaptan treatment, the plasma sodium level gradually increased to a normal level (135–140 mmol/l) and volume overload was improved. Urine volume was maintained at about 1000 ml/day with low sodium excretion (

High-performance liquid chromatographic methods were developed for the determination of azosemide and its metabolite, M1, in human plasma and urine and rabbit blood and tissue homogenates. The methods involved deproteinization of the biological samples: 2.5 volumes of acetonitrile were used for the determination of azosemide and 1 volume of saturated Ba(OH)2 and ZnSO4 for that of M1. A 50-μl aliquot of the supernatant was injected onto a C18 reversed-phase column in each instance. The mobile phases employed were 0.03 M phosphoric acid—acetonitrile (50:40, v/v) for azosemide and 0.03 M phosphoric acid/0.2 M acetic acid—acetonitrile (83:17, v/v) for M1. The flow-rate was 1.5 ml/min in both instances. The column effluent was monitored by ultraviolet detection at 240 and 236 nm for azosemide and M1, respectively. The retention times for azosemide and M1 were 6.0 and 8.3 min, respectively. The detection limits for both azosemide and M1 in both human plasma and urine were 50 ng/ml. The coefficients of variation of the assay were generally low (below 11.0%) for plasma, urine, blood and tissue homogenates. No interferences from endogenous substances or other diuretics tested were observed.

The pharmacokinetics of azosemide were evaluated simultaneously using both arterial and venous plasma data in six rabbits after a rapid 5 s intravenous bolus dosing. Initial arterial to venous ratios at 5 s after injection were the highest with values of 81.1, 67.3, 58.7, 530, 2660, and 10.5 for rabbits 1-6, respectively. Both curves decayed, paralleling each other at the terminal phase, with the venous levels higher than the arterial levels by 15.3, 31.9, 34.1, 40.7, 30.5, and 16.5% for rabbits 1-6, respectively. An exponential term with a negative coefficient was used to account for the short and steep rising phase of venous plasma levels after injection. Detailed analysis showed significant differences in various pharmacokinetic parameters, such as initial volume of distribution, apparent volume of distribution at steady state, and mean residence time based on arterial or venous data. A plot of 1/Q (urine flow rate) versus 1/CLR (renal clearance) of azosemide yielded a straight line in six rabbits, indicating that the CLR of azosemide is urine flow dependent in rabbits.

Bumetanide, 2, 8, and 20 mg/kg, was administered both intravenously and orally to determine the pharmacokinetics and pharmacodynamics of bumetanide in rats (n=10–12). The absorption of bumetanide from various segments of GI tract and the reasons for the appearance of multiple peaks in plasma concentrations of bumetanide after oral administration were also investigated. After iv dose, the pharmacokinetic parameters of bumetanide, such ast1/2 (21.4, 53.8 vs. 127 min),CL (35.8, 19.1 vs. 13.4 ml/min per kg),CLNR (35.2, 17.8 vs. 12.6 ml/min per kg) andVSS (392, 250 vs. 274 ml/kg) were dose-dependent at the dose range studied. It may be due to the saturable metabolism of bumetanide in rats. After iv dose, 8-hr urine output per 100g body weight increased significantly with increasing doses and it could be due to significantly increased amounts of bumetanide exreted in 8-hr urine with increasing doses. The total amount of sodium and chloride exreted in 8-hr urine per 100g body weight also increased significantly after iv dose of 8 mg/kg, however, the corresponding values for potassium were dose-independent. After oral administration, the percentages of the dose excreted in 24-hr urine as unchanged bumetanide were dose-independent. Bumetanide was absorbed from all regions of GI tract studied and approximately 43.7, 50.0, and 38.4% of the orally administered dose were absorbed between 1 and 24 hr after oral doses of 2, 8, and 20 mg/kg, respectively. Therefore, the appearance of multiple peaks after oral administration could be mainly due to the gastric emptying patterns. The percentages of bumetanide absorbed from GI tract as unchanged bumetanide for up to 24 hr after oral doses of 2, 8, and 20 mg/kg (96.2, 95.4 vs. 98.2%) were not significantly different, suggesting that the problem of precipitation of bumetanide in acidic gastric juices or dissolution may not contribute significantly to the absorption of bumetanide after oral administration. Urine output per 100g body wt increased at oral doses of 8 and 20 mg/kg.

Loop diuretics could adversely influence prognosis due to activation of neurohumoral mechanism in the long term. Previous study showed torasemide, a loop diuretic with anti-aldosteronergic properties, was associated with lower mortality in patients with chronic heart failure (CHF). We evaluated the effects of torasemide, in comparison with azosemide, in patients with CHF. Patients received oral diuretic therapy with torasemide (8 mg/d, n = 15) or azosemide (60 mg/d, n = 15) for 3 months. Torasemide and azosemide were then switched, and the patients were treated for another 3 months. Torasemide treatment induced significant decreases in left ventricular (LV) systolic wall stress (from 259 +/- 95 to 232 +/- 80 kdyn/cm2) and the plasma level of aldosterone (from 133 +/- 61 to 95 +/- 50 pg/mL) and was not associated with a change in the plasma level of norepinephrine. In contrast, the plasma level of norepinephrine was significantly increased (from 370 +/- 170 to 481 +/- 247 pg/mL), whereas LV systolic wall stress was unchanged after azosemide treatment. This study indicates that torasemide treatment reduced LV systolic wall stress without activation of the sympathetic nervous system in patients with CHF. The anti-aldosteronergic properties of torasemide may contribute to its favorable effects.

The present study was undertaken to examine whether sulfonamide-derived diuretics affect [Arg 8 ]vasopressin (AVP)-stimulated or atrial natriuretic peptide (ANP)-stimulated cyclic nucleotide formation in cells cultured from rat or dog kidney. In rat renal cells, all four sulfonamide-derived diuretics examined significantly suppressed 10 −9 M AVP-stimulated cAMP formation at concentrations of 10 −4 and 10 −3 M, while basal cAMP formation was unchanged by the diuretics. When cells were stimulated with 10 −7 M AVP, low ceiling diuretics (indapamide and trichlormethiazide) did not suppress cAMP formation, while high ceiling diuretics (furosemide and azosemide) significantly suppressed cAMP formation at concentrations of 10 −4 and 10 −3 M. The suppressive effect of the diuretics on AVP-stimulated cAMP formation in vitro paralleled the reported diuretic potency of the agents in vivo. In dog renal cells, all four diuretics significantly suppressed 10 −9 M AVP-stimulated cAMP formation at concentrations from 10 −6 to 10 −5 M, while these diuretics did not change basal cAMP levels. High ceiling diuretics suppressed 10 −7 M AVP-stimulated cAMP formation, whereas low ceiling diuretics did not. The difference in effective doses between rats and dogs seems to be consistent with the species difference observed in vivo. None of the diuretics affected basal levels of intracellular cGMP or ANP-stimulated cGMP formation in cultured rat renal cells. In addition to the inhibition of the Na/K/Cl co-transporter, it is suggested that most sulfonamide-derived diuretics act, at least in part, by inhibiting the actions of AVP.

Objective Short-acting loop diuretics are known to enhance cardiac sympathetic nerve activity (CSNA) in patients with chronic heart failure (CHF). The effects of two loop diuretics—long-acting azosemide and short-acting furosemide—on CSNA were evaluated using 123I-metaiodobenzylguanidine (MIBG) scintigraphy in patients with CHF. Methods The present study was a subanalysis of our previously published study, which had reported that serial 123I-MIBG studies were the most useful prognostic indicator in patients with CHF. Patients with CHF (n=208, left ventricular ejection fraction

Human monocarboxylate transporter 6 (MCT6) has recently been isolated, and its tissue distribution has been established at the mRNA level, but its functional properties remain unknown. The aim of this study is to investigate the transport properties of MCT6. When expressed in Xenopus laevis oocytes, MCT6 transported [3H]bumetanide in a pH- and membrane potential-sensitive but not proton gradient-dependent manner, with the K(t) value of 84 microM. Furthermore, MCT6 transported various drugs such as probenecid and nateglinide. Neither [14C]L-lactic acid nor [3H]L-tryptophan, typical substrates of other MCT isoforms, was transported by MCT6. Four loop diuretics, i.e., furosemide, piretanide, azosemide, and torasemide, thiazides, probenecid, glibenclamide, and nateglinide inhibited the MCT6-mediated uptake of [3H]bumetanide. In contrast, short-chain carboxylic acids, such as L-lactic acid and succinic acid did not inhibit the MCT6-mediated uptake of bumetanide. These results suggest that the substrate specificity of MCT6 is distinct from those of other MCTs. Bumetanide would be a good tool for investigating the functional properties of MCT6. It is probable that MCT6 is involved in the disposition of various drugs, including bumetanide.

Objectives: We compared therapeutic effects of furosemide, a short-acting loop diuretic, and azosemide, a long-acting one, in hypertensive heart failure rats to test the hypothesis that long-acting diuretics are superior to short-acting types in heart failure. Methods: Dahl salt-sensitive rats fed an 8% NaCl diet from age 8 weeks were divided at age 21 weeks (compensated hypertrophic stage) into three groups: rats treated with furosemide (40 mg/kg/day), those treated with azosemide (80 mg/kg/day) and untreated rats. Rats fed a 0.3% NaCl diet served as controls. Results: Both medications prevented left ventricular systolic dysfunction and enlargement at age 31 weeks, and attenuated macrophage infiltration, reactive oxygen species generation, and gelatinolytic activity to the same degree. Azosemide suppressed left ventricular fibrosis to the control level, but furosemide did not. Azosemide ameliorated myocardial catecholamine depletion and improved survival rate. Furosemide increased plasma norepinephrine levels and did not exert such beneficial effects. Conclusions: Azosemide provided better prognosis in heart failure rats compared with furosemide, partly through attenuation of the reflex increase in cardiac sympathetic neuronal activity caused by the development of heart failure. The current findings suggest a need for clinical trials examining whether long- and short-acting diuretics provide a different prognosis in patients with heart failure.

We evaluated the comparative effects of furosemide, a short-acting loop diuretic, and azosemide, a long-acting loop diuretic, on neurohumoral factors and quality of life (QOL) in patients with congestive heart failure (CHF). Twenty-five stable patients with mild chronic CHF who had been administered furosemide (n = 14) or azosemide (n = 11) orally for more than 3 months were studied. We changed furosemide to azosemide or azosemide to furosemide and followed for 3 months. Echocardiography was performed, and we also measured neurohumoral factors and assessed QOL by questionnaire. Blood pressure, body weight, renal function and echocardiographic findings were the same during the furosemide and azosemide treatments. Plasma levels of atrial natriuretic peptide and brain natriuretic peptide were not different between the two treatments. However, plasma concentrations of active renin and norepinephrine were significantly higher with furosemide treatment than with azosemide treatment. QOL score was significantly lower with azosemide than with furosemide. These findings suggest that long-acting loop diuretics may have fewer adverse effects on the neuroendocrine system and QOL than short-acting loop diuretics in patients with mild CHF.

Nocturia is one of the major problems of elderly people. The possible causes of nocturia include irritation of lower urinary tract obstruction, disturbance of the fluid balance, and sleep disturbance. In a significant proportion of patients, the mechanism of nocturia is still unclear and a definitive method of treatment has yet to be determined. This study investigated how to treat nocturia.We analyzed 51 patients who visited Kobe City General Hospital between January 1998 and June 1999 with nocturia (three or more nocturnal voidings) and no daytime urological problems. Twenty-two of these patients were given daytime diuretic therapy (azosemide 60 mg), while 29 other patients received a minor tranquilizer. The method of treatment was selected randomly. Correlations between the plasma level of atrial natriuretic peptide (hANP) and the effect of treatment were analyzed.Daytime diuretics decreased the nocturnal frequency of voiding in 10 out of 22 patients. The plasma hANP level at the first visit was significantly higher in the patients with improvement (p = 0.0026). Furthermore, the plasma hANP level was significantly decreased after daytime diuretic therapy in patients who showed improvement of nocturia with diuretic therapy (p = 0.0180). Minor tranquilizer administration decreased nocturia in 22 out of 29 patients. The plasma hANP level at the first visit was significantly lower in the patients who improved (p = 0.0021).These findings suggest that patients with higher plasma hANP levels should be treated as having subclinical heart failure, while nocturia in patients with a normal plasma hANP level might be caused by sleep disturbance.

The effects of cysteine on the pharmacokinetics and pharmacodynamics of azosemide were investigated after intravenous (10 mg/kg) and oral (20 mg/kg) administration to male Sprague-Dawley rats fed on 23% protein diet (control rats), and 5% protein diet with (rats with PCMC) or without (rats with PCM) oral cysteine (250 mg/kg, twice daily for the fourth week) for 4 weeks. After intravenous administration to rats with PCMC, some pharmacokinetic parameters restored fully or more than the level of control rats; the time-averaged nonrenal clearance (2.70 versus 2.32 ml/min/kg) and apparent volume of distribution at steady state (160 versus 189 ml/kg) were comparable to those in control rats, however, the terminal half-life (34.7 versus 57.2 min) and mean residence time (73.3 versus 99.3 min) were significantly shorter, area under the plasma concentration-time curve from time zero to time infinity (AUC, 1930 versus 2680 μg min/ml) was significantly smaller, and time-averaged renal (2.24 versus 1.21 ml/min/kg) and total body (CL, 4.98 versus 3.65 ml/min/kg) clearances were significantly faster than those in control rats. This could be mainly due to significantly faster renal clearance and at least partly due to increased cytochrome P450 1A2 activity by cysteine supplementation. After intravenous administration to rats with PCMC, the total amount of 8-hr urinary excretion of unchanged azosemide was significantly greater (457 versus 305 μg/g body weight), however, the 8-hr urine output (15.3 versus 31.1 ml/g kidney) was not significantly different between control rats and rats with PCMC. This could be due to the fact that urine output seemed to reach an upper plateau from 10 mg/kg dose of azosemide in rats.

The Na+2Cl–K+ cotransporter accepts NH4 + at its K+-binding site. Therefore, the rate of cytosolic acidification after NH4 + addition to the bath (20 mmol/l) measured by BCECF fluorescence can be used to quantify the rate of this cotransporter. In isolated colon crypts of rat distal colon (RCC) addition of NH4 + led to an initial alkalinization, corresponding to NH3 uptake. This was followed by an acidification, corresponding to NH4 + uptake. The rate of this uptake was quantified by exponential curve fitting and is given in arbitrary units (Δ fluorescence ratio units/1000 s). In pilot experiments it was shown that the pH signal caused by the Na+2Cl–K+ cotransporter could be amplified if the experiments were carried out in the presence of bath Ba2+ to inhibit NH4 + uptake via K+ channels. Therefore all subsequent experiments were performed in the presence of 1 mmol/l Ba2+. In the absence of any secretagogue, preincubation of RCC in a low-Cl– solution (4 mmol/l) for 10 min enhanced the uptake rate significantly from 1.70±0.11 to 2.54±0.27 U/1000 s (n=20). The addition of 100 mmol/l mannitol (hypertonic solution) enhanced the rate significantly from 1.93±0.17 to 2.84±0.43 U/1000 s (n=5). Stimulation of NaCl secretion by a solution containing 100 µmol/l carbachol (CCH) led to a small but significant increase in NH4 + uptake rate from 2.06±0.34 to 2.40±0.30 U/1000 s (n=11). The increase in uptake rate observed with stimulation of the cAMP pathway by isobutylmethylxanthine (IBMX) and forskolin (100 µmol/l and 5 µmol/l, respectively) was from 2.39±0.24 to 3.06±0.36 U/1000 s (n=24). Whatever the mechanism used to increase the NH4 + uptake rate, azosemide (500 µmol/l) always reduced this rate to control values. Hence three manoeuvres enhanced loop-diuretic-inhibitable uptake rates of the Na+2Cl–K+ cotransporter: (1) lowering of cytosolic Cl– concentration; (2) cell shrinkage; (3) activation of NaCl secretion by carbachol and (4) activation of NaCl secretion by cAMP. The common denominator of all four activation pathways may be a transient fall in cell volume.