Torasemide significantly reduces thiazide-induced potassium and magnesium loss despite supra-additive natriuresis.

BACKGROUND: Resistance to high-dose loop diuretics can be overcome either by co-administration with thiazides or by treatment with medium-dose loop diuretics combined with thiazides. Combination therapy has been proven to be superior to high-dose loop diuretic monotherapy for cardiac and renal edema. However, such a strongly efficacious short-term regimen is often complicated by undesired effects, including circulatory collapse and electrolyte disturbances. The question of whether the loop diuretic/thiazide combinations are efficacious and safe when conventional doses are combined has not yet been answered. METHODS: The effects of hydrochlorothiazide (HCT) and torasemide (TO) given alone on the excretion of Na+, Cl-, K+, Mg2+, and Ca2+ were compared with the effects of combined administration of the diuretics in 12 healthy volunteers. RESULTS: The co-administration of HCT (25 mg) with TO (5 or 10 mg) strongly increased Na+ excretion. However, the combination significantly reduced K+ and Mg2+ excretion. The K+-sparing effect of the HCT/TO combination was shown to be due to a significant reduction in the HCT-induced increase in fractional K+ excretion by the loop diuretic. Total excretion of Ca2+ relative to Na+ excretion was less with the HCT/TO combination than with TO given alone. CONCLUSION: The enhancement of desired NaCl excretion by the HCT/TO combination with significant reduction of undesired loss of K+ and Mg2+ meets clinical requirements but has to be validated in long-term clinical trials.

Pharmacology of intracellular cytosine-arabinoside-5'-triphosphate in malignant cells of pediatric patients with initial or relapsed leukemia and in normal lymphocytes.

PURPOSE: The prodrug cytosinearabinoside (ara-C) is widely used in the treatment of acute leukemias. The active drug is the intracellular metabolite cytosine-arabinoside-5'-triphosphate (ara-CTP). The purpose of the present study was to investigate the relation between sensitivity and pharmacokinetic parameters Cmax, t1/2 and AUC of ara-CTP. The obtained results were compared to previous studies. EXPERIMENTAL DESIGN: Cmax, t1/2 and AUC of ara-CTP were assessed in leukemic cells of 17 pediatric patients with acute lymphoblastic leukemia (ALL) and in 6 lymphoblastic cell lines and compared with normal lymphocytes of 9 healthy donors by high pressure liquid chromatography (HPLC). The sensitivity of the cells against ara-C was determined by the MTT assay. RESULTS: The intracellular accumulation of ara-CTP was significantly lower in normal lymphocytes (Cmax 47.7-60.9 pmol/10(6) cells) compared to leukemic cell lines (Cmax 11-1128 pmol/10(6) cells) and leukemic cells of our patients (Cmax 85.9-631 pmol/10(6) cells). Similar results were found for the AUC. There was no significant difference between initial and relapsed leukemias in our small cohort. A correlation between sensitivity in terms of IC50 values and the intracellular ara-CTP accumulation was observed in cell lines, but not in leukemic cells and normal lymphocytes from healthy donors. CONCLUSIONS: Pharmacokinetic parameters varied tremendously in leukemic cells in contrast to normal lymphocytes without a difference in sensitivity. It is worthwhile to compare literature data to assess an optimal dosage of ara-C in pediatric patients.

Stereoselectivity of the interaction of muscarinic antagonists with their receptors.

The stereoselectivity of the interaction with muscarinic receptors of enantiomers of a series of chiral antagonists is receptor subtype dependent. There is no overall relationship between stereoselectivity and receptor affinity. Depending on the antagonist studied, receptor stereoselectivity may indeed reflect: (1) the weakening or loss of a single interaction involving one of the four groups bound to the asymmetric carbon; (2) steric hindrance preventing optimum interaction of the low affinity steroisomer with the receptor; and/or (3) the inversion of the relative positions of two moieties of the ligand with similar structural and electronic properties i.e. comparable affinities for the two corresponding subsites in the receptor.

Rate-limiting biotransformation of triamterene is mediated by CYP1A2.

OBJECTIVE: Triamterene (TA), a potassium-sparing diuretic, is extensively metabolized by hydroxylation in 4'-position and subsequent conjugation by cytosolic sulfotransferases. To identify the cytochrome P450 enzyme(s) catalyzing hydroxylation of triamterene (the rate-limiting step in the formation of the sulfate ester (STA)), in vitro incubation studies were performed with human liver microsomes. METHODS: Initial rates of TA hydroxylation (0 - 300 microM) were determined during a ten-minute-incubation period with liver microsomes of two donors. The role of individual CYP enzymes was determined by pre-incubation with selective inhibitors/alternative substrates. Vice versa, the effect of TA (0 - 500 microM) on 3-demethylation of caffeine (0 - 1,000 microM) was assessed. Metabolite concentrations were estimated by reversed-phase HPLC methods. RESULTS: TA Km values without inhibitors were 60 and 142 microM, Vmax was 177 and 220 pmol/min/mg protein, respectively. Mean inhibitor induced changes of 4'-hydroxy-TA formation were as follows: Furafylline 25 microM (CYP1A2), complete inhibition (-100%); omeprazole 250 microM (CYP1A2 inhibitor/CYP2C 19 substrate), -30%; coumarin 25 microM (CYP2A6), -11%; quinidine 25 microM (CYP2D6), -9%; ketoconazole 25 microM (CYP3A), -18%; and erythromycin 250 microM (CYP3A), -8%. In the reverse inhibition studies, TA competitively inhibited caffeine 3-demethylation with Ki values of 65 and 111 microM, respectively. CONCLUSION: 4'-hydroxylation of TA in humans appears to be mediated exclusively by CYP1A2. Inhibition or induction of CYP1A2 will change the time course of both TA and its active phase-II metabolite. The net pharmacodynamic effect of such changes is difficult to predict and needs to be evaluated in clinical studies.

Time course and extent of angiotensin II antagonism after irbesartan, losartan, and valsartan in humans assessed by angiotensin II dose response and radioligand receptor assay.

OBJECTIVES: To compare the angiotensin II antagonistic properties of the usual recommended oral starting doses of various angiotensin II receptor antagonists-150 mg irbesartan, 80 mg valsartan, and 50 mg losartan-in humans. SUBJECTS AND METHODS: Eighteen healthy men were enrolled in a double-blind, randomized crossover study. Angiotensin II dose-effect curves of diastolic blood pressure and radioreceptor assay were performed before and up to 47 hours after single and multiple doses of the antagonists. The rightward shift of the angiotensin II dose-effect curves (dose ratio-1) assessed the antagonistic effects in vivo. The degree of receptor occupancy in plasma was detected by a rat lung radioreceptor assay ex vivo in vitro. RESULTS: All of the drugs clearly showed antagonistic effects to angiotensin II in vivo (dose ratio-1) and in vitro (radioreceptor assay). Within the given doses the dose ratio-1 for irbesartan was greater than for valsartan and losartan after single and repetitive dosing, reaching statistical significance at various time points up to 36 hours versus valsartan and up to 47 hours versus losartan. The apparent half-lives of the decay of the effects were approximately 8 hours for valsartan and losartan, whereas 15 to 18 hours were obtained with irbesartan. These findings were supported by the radioreceptor assay data: the percentage of receptor occupancy for irbesartan was significantly greater than for valsartan and losartan up to 47 hours. CONCLUSION: Angiotensin II antagonistic effects of irbesartan, valsartan, and losartan were compared. Irbesartan showed the slowest decay and longest duration of its antagonistic effects. With the recommended initial doses used in this study, the following rank order of antagonistic intensity was obtained: irbesartan > valsartan > losartan. The findings of this study, specifically the longer-lasting effects of irbesartan, may have clinical implications.

Atenolol interaction with aspirin, allopurinol, and ampicillin.

Atenolol kinetics were investigated in six healthy subjects after 100 mg orally, as monotherapy a 6-day treatment began 48 hr later. After a therapy-free interval of 4 wk, the same subjects received the same dose of atenolol with 1 gm ampicillin, 500 mg aspirin, and with 300 mg allopurinol. Allopurinol and aspirin did not substantially alter the kinetics of atenolol. After a single oral dose of 100 mg atenolol combined with 1 gm ampicillin, the bioavailability of atenolol was reduced to 36 +/- 5% compared to 60 +/- 8% after monotherapy. During long-term treatment with atenolol and ampicillin the bioavailability of atenolol fell to 24% (P less than 0.01). Mean peak plasma levels were lowered from 511 +/- 59 ng/ml on monotherapy to 344 +/- 33 ng/ml after the combination with ampicillin. The area under the plasma level-time curve, mean steady-state concentration, and urinary recovery were reduced, also. Twelve hours after 100 mg atenolol and 1 gm ampicillin, exercise tachycardia was significantly higher than after atenolol alone. During the 4-wk treatment in six hypertensive patients blood pressure levels of those on atenolol alone were not different from those on the combination therapy with ampicillin.

Interaction of atenolol with furosemide and calcium and aluminum salts.

Six healthy subjects were treated with 100 mg atenolol. After a therapy-free interval of 4 wk the same subjects received the same dose of atenolol with furosemide, 40 mg, with calcium (as the lactate gluconate and carbonate), 500 mg, or with aluminum hydroxide, 5.6 gm. Atenolol alone and in combination was administered first as a single oral dose; a long-term 6-day treatment began 48 hr later. Addition of furosemide did not influence atenolol kinetics, but aluminum hydroxide led to an insignificant reduction (P greater than 0.05) of mean peak plasma levels of about 20% and of the area under the plasma concentration-time curve (AUC -infinity) from 5818 to 4353 ng ml-1 hr (P greater than 0.05). Calcium altered atenolol kinetics distinctly mean peak plasma levels of atenolol fell 51% (P less than 0.001), AUC0-infinity fell from 5818 to 3935 ng ml-1 hr (P less than 0.01) and elimination half-life (t1/2) increased to a mean of 11.0 hr (compared to 6.2 hr with atenolol alone). The prolongation of t1/2 induced by calcium coadministration led to atenolol cumulation during the long-term dosage. Twelve hours after atenolol, 100 mg, and calcium, 500 mg, exercise tachycardia was lower than with atenolol alone (P less than 0.001). During the 4-wk treatment in six hypertensive patients, blood pressure values of those on atenolol alone were not different from those on the combination therapies (P greater than 0.05).

Digoxin concentrations in serum and cantharides blister fluid: correlations with cardiac response.

The relationship between the pharmacokinetics and dynamics of digoxin was investigated using a skin blistering technique that allows experimental access to tissue fluid concentrations. Eight healthy volunteers received digoxin, 1.0 mg, and placebo intravenously according to a double-blind crossover design. Drug concentrations were determined during a 72-hour period in serum, urine, and cantharides blister fluid (CBF). Digoxin levels in the hypothetic peripheral compartments were calculated from serum concentrations. Digoxin effects (total electromechanical systole [QS2c], left ventricular ejection time [LVETc], preejection period [PEPc], QTc time, heart rate, and T wave amplitude) were measured simultaneously. Peak levels in the shallow and deep compartments occurred at 12 1/2 to 20 minutes and 3 hours and the maximum concentration in CBF (2.75 +/- 0.48 ng/ml) occurred at 1 hour. Digoxin effects on QS2c, PEPc, and the ratio PEP/LVET were not related to serum concentrations but were closely related to CBF concentrations (r = 0.90). CBF concentrations were then within the range of serum digoxin concentrations usually associated with the treatment of heart failure. Thus, CBF allows experimental access to active drug concentrations after a single intravenous dose.

Cardiovascular effects, pharmacokinetics, and converting enzyme inhibition of enalapril after morning versus evening administration.

The cardiovascular effects and pharmacokinetics of once-daily enalapril were studied after single-dose and subchronic treatment in eight patients with hypertension by use of ambulatory blood pressure monitoring. Enalapril, 10 mg, was given at either 7 AM or 7 PM in a randomized crossover design. In addition, inhibition of serum converting enzyme was studied. Subchronic treatment at 7 AM significantly reduced blood pressure during the day but was less effective at night. Subchronic dosing at 7 PM significantly further decreased nighttime blood pressure followed by a slow increase during the day, with no effect on elevated afternoon values. Peak concentrations of enalaprilat were found 3.5 hours (morning) and 5.6 hours (evening) after drug intake (p < 0.05), whereas peak effects occurred 7.4 hours (morning) and 12 hours (evening) after drug administration. In conclusion, 24-hour blood pressure profiles in patients with hypertension were significantly influenced by the time of enalapril dosing. Differences in effect profiles could not be attributed to similar changes in pharmacokinetics or to different time courses of angiotensin converting enzyme inhibition.

Xipamide disposition in liver cirrhosis.

The pharmacokinetics of the sulfonamide-type diuretic xipamide was studied in patients with liver cirrhosis and ascites and compared with healthy control subjects. After oral administration of 40 mg xipamide, the diuretic was rapidly distributed in the blood and the ascites. The ratio of the area under the concentration-time curve (AUC) of plasma and ascitic fluid was 7:2, as was the protein content in the respective compartments. The AUC in plasma of cirrhotic patients was significantly greater than in control subjects (p less than 0.001). The most striking finding was the increase of the amount (Ae) of parent drug and main metabolite excreted into the urine (p less than 0.001). The renal clearance of xipamide was only moderately reduced in patients with liver cirrhosis. Both AUC and Ae were positively correlated to the plasma concentration of direct bilirubin of the patients (p less than 0.05). We concluded that nonrenal drug clearance in patients with liver cirrhosis was reduced as a result of the blockade of hepatobiliary excretion during cholestatic conditions.

AF-DX 116, a cardioselective muscarinic antagonist in humans: pharmacodynamic and pharmacokinetic properties.

Effects of AF-DX 116, a cardioselective antagonist, on M cholinergic receptors (M-ChR) were studied in healthy volunteers. Occupancy of M-ChR subtypes by drug present in plasma samples (radioreceptor assay) was compared with these effects. After an intravenous dose of AF-DX 116 saturating greater than 90% of cardiac M2-ChR, an increase in heart rate by 25 beats/min was observed. This cardiac receptor occupancy and effect wore off with a parallel time course within 10 hours. No inhibition of salivary flow was observed, coinciding with a lack of M3-ChR blockade in the radioreceptor assay. Beta-adrenergic receptor blockade by propranolol did not affect either of the effects. No indication for active metabolites or stereoselective drug metabolism was found comparing HPLC and receptor assay for drug concentrations in plasma. We conclude that AF-DX 116 may be a useful drug for the treatment of bradycardia. Its lack of troublesome side effects is the result of its selectivity for cardiac M-ChR.

Zinc changes AMPA receptor properties: results of binding studies and patch clamp recordings.

The influence of zinc ions on alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors was investigated using binding studies with [3H]AMPA to rat cortical membranes and patch clamp recordings from cultured superior colliculus neurones. In Tris-HCl buffer, zinc (1-10 mM) significantly increased the specific binding of [3H]AMPA whereas this increase was negligible in the presence of CaCl2 (2.5 mM) and KSCN (100 mM). This effect was associated with a dramatic increase in Bmax but a decrease in both agonist and antagonist affinity. Association and dissociation experiments showed that equilibrium [3H]AMPA binding is reached with faster kinetics in the presence of zinc. At low concentrations (0.3 mM) zinc also concentration-dependently potentiated both peak and plateau components of whole cell current responses to AMPA (100 microM). This effect was accompanied by a reduction of the degree, and slowing of the rate, of AMPA receptor desensitisation. In contrast, higher concentrations of zinc (1-3.0 mM) inhibited AMPA responses to some degree, but slowed desensitisation further. This ability of zinc to change AMPA receptor properties may be relevant to neurotoxicity associated with AMPA receptor activation.

Different binding affinities of NMDA receptor channel blockers in various brain regions--indication of NMDA receptor heterogeneity.

The N-methyl-D-aspartate (NMDA) receptor-channel complex exists in multiple forms which probably have different physiological and pharmacological properties. To further evaluate this concept of different NMDA receptor subtypes, receptor binding and autoradiographic techniques were used to study the phencyclidine (PCP) binding site of the NMDA receptor ion-channel complex. [3H]MK-801 was employed to characterize binding properties of (+)-MK-801, (-)-MK-801, phencyclidine (PCP), (+/-)-ketamine, amantadine (1-amino-adamantane) and memantine (3,5-dimethyl-1-amino-adamantane) in different brain regions. Saturation experiments on homogenized membranes revealed the existence of single classes of binding sites in cortex and cerebellum but with significant different affinities between these regions (KD/Cortex = 4.59 nM, Bmax/Cortex = 0.836 pmol/mg protein; KD/Cereb. = 25.99 nM, Bmax/Cereb. = 0.573 pmol/mg protein) suggesting that the lower affinity in cerebellum indicates another population of NMDA receptor channels. In contrast, in striatum there was clear evidence for two binding sites (KD/high = 1.43 nM, Bmax/high = 0.272 pmol/mg protein; KD/low = 12.15 nM, Bmax/low = 1.76 pmol/mg protein). Displacement studies (autoradiography and binding) revealed a lower affinity for unlabeled (+)-MK-801 in striatum which was clearly not the case for memantine. In cerebellar membranes there was a significant decrease in the affinity for both MK-801 enantiomers and PCP but not for the 1-amino-adamantanes. In contrast, all compounds showed lowered affinity in the dentate gyrus. These findings support NMDA receptor heterogeneity which may be of particular relevance for the development of subtype-selective drugs.

Muscarinic ganglionic stimulants: conformationally restrained analogues related to [4-[[N-(3-chlorophenyl)carbamoyl]oxy]-2-butynyl]trimethylammonium chloride.

The synthesis of a series of tertiary and quaternary cyclic analogues (isoarecolinol, dihydroisoarecolinol, arecolinol, and 3-pyrroline-3-carbinol derivatives) of [4-[[N-(3-chlorophenyl)carbamoyl]oxy]-2-butynyl]trimethylammonium chloride (McN-A-343) (1), a selective stimulant of muscarinic receptors in sympathetic ganglia (so-called M1 receptors), is reported. The compounds 3-10 were tested for muscarinic ganglion-stimulating activity by recording blood pressure responses in pithed rats. All tertiary compounds tested had no ganglion-stimulating activity. Among the series of quaternary derivatives, only the isoarecolinol analogues 4a and 4b showed considerable ganglion-stimulating effects, whereas the dihydroisoarecolinol (8), the arecolinol (6a, 6b), and the 3-pyrroline-3-carbinol derivatives (10) were much less potent. Our experiments therefore demonstrate that in this series a quaternary nitrogen atom, unsaturation at C2 of the ammonium side chain, and a certain spatial arrangement of the ammonium and the phenylcarbamate groups are essential for potent M1-receptor stimulating activity.

The effect of cyclosporine on electrically paced isolated rat cardiomyocytes.

The acute cardiotoxicity of cyclosporine was investigated in isolated cardiomyocytes from adult rats. In a first study, myocytes were incubated with CsA ranging from 1 to 10 micrograms/ml and paced by electrical-field stimulation. After 30 min of stimulation the number of surviving rod-shaped myocytes was significantly reduced at 2.5 micrograms/ml (77.9%) and 5 micrograms/ml CsA (64.2%) as compared with the drug vehicle methanol (88.8%, P less than 0.05) with a further decrease at 10 micrograms/ml CsA (30.1% vs. 81.2%, P less than 0.005). In a second study, with the use of digital image processing of fura-2 fluorescence, the mean intracellular free calcium concentration, integrated over 1 sec, of single myocytes in the presence of 5 micrograms/ml CsA, the solvent methanol, or pure Krebs Ringer Hepes buffer was measured. Starting 2 Hz field stimulation increased the intracellular free calcium concentration from 100.1 to 177.9 nM in buffer and from 145.7 to 200.6 nM calcium with methanol. In contrast, there was a 3-fold increase of the intracellular free calcium concentration with 5 micrograms/ml CsA from 128.8 to 376.1 nM calcium. The intracellular free calcium during electrical stimulation was significantly higher with CsA than with the solvent (376.1 nM vs. 200.6 nM, P less than 0.001). In a further study, myocytes were incubated with calcium ranging from 0.5 to 8 mM calcium in the presence of 5 micrograms/ml CsA or the solvent methanol and electrically stimulated. Here, with increasing extracellular calcium the number of rod-shaped myocytes decreased significantly with CsA as compared with the solvent (P less than 0.02). The data suggest that CsA exerts a dose-dependent toxic effect on isolated rat cardiomyocytes that depends on the extracellular calcium concentration. There is direct evidence that CsA increases the intracellular free calcium concentration in rat cardiomyocytes.

Current status of sustained release formulations in the treatment of hypertension. An overview.

The principal advantages to be gained from controlling the variables of drug release in sustained release formulations are as follows: (i) a more uniform plasma drug profile with fewer occasions when super- or subtherapeutic concentrations of the drug, or its active metabolite(s), occur; and (ii) a smoother therapeutic response over the dosage interval (provided the time-course of drug effects reflects the plasma concentration-time profile). Clinically, this offers the potential to optimise drug therapy and decrease the occurrence of concentration-related adverse effects. In addition, sustained release formulations may increase the likelihood of patient acceptance of therapy, and a once-daily sustained release formulation of a shorter-acting drug that provides a 'residual' therapeutic response at the end of the dosage interval can provide additional 'cover' in comparison with a once-daily conventional (immediate release) formulation. In the treatment of hypertension, there are potential advantages to be gained from continuous 24-hour control of blood pressure (BP), particularly in view of epidemiological evidence linking the apparent underperformance of antihypertensive therapy in some major intervention trials in reducing the occurrence of coronary heart disease to predicted levels with a relative failure to control diurnal BP fluctuations. In this regard, the concept of the trough:peak ratio as a measure of antihypertensive efficacy has gained increasing acceptance during recent years. A sustained release antihypertensive formulation offering an improved plasma concentration-time profile and an adequately high trough:peak ratio may therefore provide more consistent 24-hour BP-lowering activity, with attenuation of early morning BP surges and maximal target organ protection. This, coupled with the fact that sustained release formulations can also provide economic advantages in cardiovascular therapeutics by lowering overall health expenditure (which more than offsets their usually higher acquisition costs in comparison with immediate release formulations), suggests that they may have an increasingly important role to play in the future.

Clinical pharmacokinetics and pharmacodynamics of torasemide.

The new loop diuretic torasemide belongs to the pyridine sulfonylurea class. It is well absorbed and yields a bioavailablity of about 80% in healthy individuals, even higher in patients with oedema. This is roughly double that of the 'classical' loop diuretic furosemide (frusemide) [26 to 65%]. Torasemide is highly bound to protein (99%) as is furosemide. The volume of distribution of torasemide was determined as 0.2 L/kg as compared with 0.11 to 0.18 L/kg for furosemide. Torasemide undergoes extensive hepatic metabolism; only 20% of the parent drug is recovered unchanged in the urine. For comparison only 10 to 20% of furosemide undergoes phase II metabolisation (to the glucuronide). In chronic renal failure the renal clearance of torasemide decreased in proportion to the decrease of the patients' glomerular filtration rate, whereas the total plasma clearance (3 times that of the renal clearance) appeared to be independent of renal function. As expected, the renal excretion of torasemide metabolites is significantly retarded in renal disease. The pharmacokinetics of torasemide are significantly influenced by liver disease. Total plasma clearance of torasemide was reduced to about half of that found in the control group, yielding an increase in elimination half-life. A greater than normal fraction of torasemide was recovered in the urine of patients with cirrhosis. In contrast, the kinetics of furosemide appeared to depend more on kidney function than on liver disease. The pharmacodynamics of torasemide are principally the same as those reported from conventional loop diuretics due to their interference with one binding site in the thick ascending limb of Henle's loop, the Na+:K+:2Cl- carrier. The maximum natriuretic effect of all loop diuretics amounts to about 3 mmol Na+/min. Members of this class differ, however, with respect to their intravenous potency or affinity for the receptor, respectively: bumetanide > piretanide > torasemide > furosemide. So far, the only loop diuretic which has been shown to effectively lower high blood pressure is torasemide. This effect occurs at the low dose of 2.5 mg/day.

Effects of a new pteridine derivative on urinary sodium, potassium and magnesium excretion in conscious saline-loaded rats.

1. Two recently synthesized pteridine derivatives (RPH 3036; RPH 3038) were tested in conscious saline-loaded rats and showed natriuretic and antimagnesiuretic properties but hardly reduced potassium excretion. 2. In the same model a dose-response curve was performed for RPH 3036. ED50 and Emax values were calculated for the natriuretic (ED50 = 13.4 mumol kg-1; Emax = 1.08 mmol kg-1) and antimagnesiuretic (ED50 = 11.3 mumol kg-1; Emax = -0.099 mmol kg-1) properties of RPH 3036. There were no significant changes of potassium and calcium excretion. 3. After a single dose of RPH 3036 (100 mumol kg-1) the time course of electrolyte excretion was analysed over 6 h. RPH 3036 did not show any significant effects on renal potassium and calcium excretion whereas a pronounced decrease (P less than 0.01) in renal magnesium excretion was evident during the 6 h. A moderate increase of sodium excretion was observed only after 3, 5 and 6 h. 4. A selective reduction of magnesium secretion in the late distal tubule and collecting duct was proposed as a possible mechanism of action of RPH 3036. This would explain the fast onset of action as well as the lack of antikaliuretic and anticalciuretic effects. The high selectivity of RPH 3036 makes it potentially valuable for the future investigation of renal magnesium transport.

Structure-activity relationships of new analogues of arecaidine propargyl ester at muscarinic M1 and M2 receptor subtypes.

1. The potency of arecaidine propargyl ester (APE) and of several analogues containing a modified ester side chain has been assessed at M1 and M2 muscarinic receptor subtypes. APE was shown to act as a potent agonist at ganglionic M1 receptors in the pithed rat, at M2 receptors in guinea-pig isolated atria (-log EC50 = 8.22) and ileum (-log EC50 = 7.77). 2. The arecaidine 2-butynyl and 2-pentynyl esters were approximately equipotent with APE at M1 and M2 receptors, whereas the 2-hexynyl derivative was found to be less potent than APE in atria (-log EC50 = 6.80) and ileum (-log EC50 = 6.70) by about one order of magnitude. The 2-heptynyl and 3-phenyl propargyl esters exhibited no agonist actions in atria and ileum. 3. Shifting the triple bond from the 2 to the 3 position and introducing a bulky group at position 1 of the ester side chain of APE and analogues resulted in competitive antagonists (pA2 ranging from 4.9 to 7.3). 4. APE and its 2-butynyl analogue showed some agonistic selectivity for cardiac M2 receptors (potency ratio, ileum/atria = 2.8 and 4.6 respectively). All antagonists in this series of compounds were not selective in terms of affinity since their pA2 values at cardiac and ileal M2 receptors were similar (potency ratios, ileum/atria = 0.4 to 1.2).

Selectivity profile of the novel muscarinic antagonist UH-AH 37 determined by the use of cloned receptors and isolated tissue preparations.

1. Functional in vitro experiments were carried out to determine the antimuscarinic potencies of the pirenzepine derivative UH-AH 37 (6-chloro-5,10-dihydro-5-[(1-methyl-4-piperidinyl)acetyl]-11H-dibenzo- [b,e] [1,4] diazepine-11-one hydrochloride) at M1 muscarinic receptors of rabbit vas deferens, M2 receptors of rat left atria and M3 receptors of rat ileum. Furthermore, N-[3H]-methylscopolamine competition binding experiments were performed to obtain its affinities for the five cloned human muscarinic receptors (m1-m5) stably expressed in CHO-K1 cells. Pirenzepine served as a reference drug throughout all experiments. 2. In all preparations used, UH-AH 37 interacted with muscarinic receptors in a fashion characteristic of a simple competitive antagonist. 3. In the functional studies, UH-AH 37, like pirenzepine, showed high affinity for M1 (pA2 8.49) and low affinity for M2 muscarinic receptors (pA2 6.63). In contrast to pirenzepine, UH-AH 37 also displayed high affinity for M3 receptors (pA2 8.04). 4. In agreement with its functional profile, UH-AH 37 bound with highest affinity to m1 (pKi 8.74) and with lowest affinity to m2 receptors (pKi 7.35). Moreover, it showed a 7 fold higher affinity for m3 (pKi 8.19) than for m2 receptors, whereas pirenzepine bound to both receptors with low affinities. 5. The binding affinity of UH-AH 37 for m4 and m5 receptors (pKi 8.32 for both receptors) was only ca. 2.5 fold lower than that for m1 receptors, while the corresponding affinity differences were 6 and 13 fold in case of pirenzepine. 6. In conclusion, the receptor selectivity profile of UH-AH 37 differs clearly from that of its parent compound, pirenzepine, in both functional and radioligand binding studies, the major characteristics being its pronounced M2 (m2)/M3 (m3) selectivity. UH-AH 37 thus represents a useful tool for the further pharmacological characterization of muscarinic receptor subtypes.