Inhibitory Effect Of Furosemide Research Articles

Inhibitory Effect of Furosemide on Non-Selective Voltage-Independent Cation Channels in Human Erythrocytes

Background: Furosemide, a loop diuretic inhibiting the renal tubular Na⁺,K⁺,2Cl^- cotransporter, has been shown to decrease cytosolic Ca²⁺ concentration ([Ca²⁺]_i) in platelets and erythrocytes. [Ca²⁺]_i in erythrocytes is a function of Ca²⁺ permeable cation channels. Activation of those channels e.g. by energy depletion or oxidative stress leads to increase of [Ca²⁺]_i, which in turn triggers eryptosis, a suicidal erythrocyte death characterized by cell membrane scrambling. The present study was performed to explore whether furosemide influences the cation channels and thus influences eryptosis. Methods: Cation channel activity was determined by whole-cell patch clamp, [Ca²⁺]_i utilizing Fluo3 fluorescence and annexin V binding to estimate cell membrane scrambling with phosphatidylserine exposure. Results: A 45 min exposure to furosemide (10 and 100 µM) slightly, but significantly decreased cation channel activity and [Ca²⁺]_i in human erythrocytes drawn from healthy individuals. ATP-depletion (> 3 hours, +37°C, 6 mM ionosine and 6 mM iodoacetic acid) enhanced the non-selective cation channel activity, increased [Ca²⁺]_i and triggered cell membrane scrambling, effects significantly blunted by furosemide (10 – 100 µM). Oxidative stress by exposure to tert-butylhydroperoxide (0.1 –1 mM) similarly enhanced the non-selective cation channels activity, increased [Ca²⁺]_i and triggered cell membrane scrambling, effects again significantly blunted by furosemide (10 – 100 µM). Conclusions: The present study shows for the first time that the loop diuretic furosemide applied at micromolar concentrations (10 – 100 µM) inhibits non-selective cation channel activity in and eryptosis of human erythrocytes.

Inhibitory Effect of Furosemide on Carbonic Anhydrase

This study investigated the inhibitory effect of a high efficiency diuretic, furosemide, on carbonic anhydrase (CA). First, comparing the inhibitory effect of acetazolamide, a low efficiency diuretic, on CA, shows that furosemide or acetazolamide can quickly make CA inactive when its concentration is close to the enzyme concentration, different from the usual inhibitory kinetics in which the concentration of the inhibitor is far higher than the enzyme concentration. Secondly, the reaction of the enzyme indicates that the inhibitory effect of furosemide or acetazolamide on carbonic anhydrase is quickly reversible. Finally, the degree of the inhibitory effect of furosemide and of acetazolamide on CA are compared. The results show that furosemide inhibits CA less than acetazolamide.

Inhibitory effect of furosemide on activation of human peripheral blood polymorphonuclear leukocytes stimulated with n-formyl-methionyl-leucyl-phenylalanine

The clinical efficacy of inhalatory furosemide (Fu) has been extensively studied in bronchial asthma patients but there are only a few studies addressing its action on cells participating in the underlying inflammatory process. Therefore, we investigated the effect of Fu on human peripheral blood polymorphonuclear leukocytes (PMNL) at concentrations that can be achieved in the bronchial lining fluid by inhalation, i.e. 10 −5, 10 −4 and 10 −3 M. The influence of Fu on the following PMNL parameters was investigated: intracellular calcium changes ([Ca 2+] i) as a part of signal transduction and luminol dependent chemiluminescence (LCL) as an indirect measure of NADPH-oxidase activation upon n-formyl-methionyl-leucyl-phenylalanine (fMLP) stimulation; chemotaxis to fMLP, phagocytosis and intracellular killing of Staphylococcus aureus. Incubation with Fu resulted in a concentration dependent reduction of Ca 2+ influx and Fu (10 −3 M) decreased the main Ca 2+ parameters to one half of the control values and to the level obtained in calcium-free buffer. In contrast, Fu had no effect if preincubated with the cells and then removed by washing. The LCL signal was reduced by Fu (10 −3 M) from 2000±870 to 550±440 arbitrary units [aU] ( p<0.05). In contrast to the [Ca 2+] i measurements, a slightly diminished LCL was also observed following preincubation with Fu and washing. No effect of Fu was found on phagocytosis and intracellular killing of St. aureus. Fu diminished chemotaxis to fMLP but at 10 −3 M it also displayed weak chemoattractant properties. The differential action of Fu on human PMNL may add to the understanding of its topical and restricted efficacy in bronchial asthma.

Furosemide inhibits thromboxane A2-induced contraction in isolated human internal mammary artery and saphenous vein.

Evidence suggests that, in addition to its diuretic property, furosemide also may exert direct vascular effects. Because thromboxane A2 (TXA2) has a role in the control of vascular tone, we investigated the effect of furosemide on the contraction induced by U46619 (a stable TXA2 mimetic) on isolated human internal mammary artery (IMA) and saphenous vein (SV). Concentration-response curves to U46619 were performed in the absence (vehicle) or the presence of furosemide (0.1-1 mM) on rings of IMA and SV. In addition, the relaxant effect of furosemide (0.1 microM-1 mM) also was evaluated on U46619-precontracted IMA and SV. The participation of cyclooxygenase derivatives was studied by pretreatment with indomethacin. Furosemide (0.1-1.0 mM) caused parallel rightward shifts of U46619 concentration-response curves without affecting the maximal responses in both IMA and SV. Treatment with indomethacin (1 microM) modified neither the inhibitory effect of furosemide on U46619-induced contractions, nor the relaxant effect of furosemide on U46619-induced contractions, nor the relaxant effect of furosemide on U46619-precontracted IMA and SV. In conclusion, furosemide at high concentrations inhibited U46619-induced contraction in human isolated IMA and SV and relaxed U46619-precontracted IMA and SV by mechanisms independent of the release of relaxant prostaglandins. These results suggest that blockade of TXA2 receptors by furosemide may contribute to explaining the therapeutic effects of furosemide in the treatment of severe heart failure.

EFFECT OF ANTI-ASTHMATIC MEDICATIONS ON INTERLEUKIN 6 (IL-6) PRODUCTION FROM STIMULATED HUMAN PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCs). 43

Although the mechanism is uncertain, elevated levels of IL-6 in bronchoalveolar lavage fluids in asthmatics suggest a possible role in the pathogenesis of pulmonary inflammation. It has been shown that aerosolized furosemide (FUR) may be helpful in the treatment of asthma, and an inhibitory effect of FUR on cytokine production in vitro has recently been shown. A comparison of its effect with other anti-asthmatic medications appears warranted. PBMCs from 25 normal subjects were isolated by density gradient centrifugation and pretreated for 20 min with varying concentrations of FUR, nedocromil sodium (NS), sodium cromoglycate (SC), or hydrocortisone acetate (HC) before LPS was added (10μg/ml). The cells (1×106 cells/ml) were incubated at 37°C for 2 days and supernatants analyzed for IL-6 by ELISA (Genzyme). FUR inhibited IL-6 production at high concentrations(>2.5×10-3 M). At a concentration of 10-2 M, IL-6 production was reduced by 98% (p<0.001) compared to 93% for HC(p<0.001). There was no effect of NS nor SC. Figure

Renal adaptations to continuous administration of furosemide and bendroflumethiazide in rats.

During continuous treatment with diuretics, the kidney adapts to the initial Na loss by activating antinatriuretic mechanisms which serve to prevent further Na and volume losses. To study the renal sites of adaptations to constant diuretic treatment, bendroflumethiazide (4 mg daily), furosemide (8 mg daily) or vehicle (0.24 ml daily) was infused intraperitoneally to female Wistar rats by implanted osmotic minipumps. Half of the animals (groups vol.) were randomized to receive a balanced saline solution to drink in addition to water in order to replace Na, K and volume losses. On the 6th day of treatment, clearances of inulin, Na, and Li were determined during four consecutive 6 hr periods. Circadian changes in renal excretions occurred in all groups with highest excretions of Na, Li and water in the dark period (6 p.m. to 6 a.m.). Renal changes induced by continuous infusion of diuretics were most pronounced in the dark period and would probably not have been disclosed if the clearance experiments had been restricted to the daytime. The average 24-hour clearance for inulin (glomerular filtration rate) was not different among groups, except for a 20% decrease in the furosemide group. The 24-hour fractional Na excretion, being approximately 0.5% in the vehicle group, increased to approximately 0.8% in group (bendroflumethiazide+vol) and to approximately 2.8% in group (furosemide+vol) but was not different from the vehicle group in the diuretic groups without volume replacement.(ABSTRACT TRUNCATED AT 250 WORDS)

The inhibitory effect of furosemide on the contractile response of equine trachealis to cholinergic nerve stimulation

The effects of furosemide on the responses of equine trachealis muscle with and without epithelium to electrical field stimulation (EFS) and exogenous acetylcholine (ACh) were investigated in organ baths. Tissues were pretreated with guanethidine and the parameters used for EFS were those previously demonstrated to activate postganglionic cholinergic neurons. In tissues with intact epithelium, furosemide (100 μ m) shifted the frequency-response curve to the right. In the preparations without epithelium, furosemide did not affect the response to EFS. Neither in epithelium-on nor in epithelium-off tissues was the ACh dose-response curve affected by the administration of furosemide. We conclude that furosemide (100 μ m) decreases the equine tracheal smooth muscle responses to EFS through inhibition of cholinergic neurotransmission, and that this effect is dependent on epithelium.

Effect of furosemide on water and urea transport in cortical and inner medullary collecting duct

The present in vitro microperfusion study examined whether furosemide has an effect on hydraulic conductivity (Lp X 10(-6) cm/sec.atm) and 14C-urea permeability (Pu X 10(-5) cm/sec) in inner medullary collecting ducts (IMCD) and cortical collecting tubules (CCT) isolated from rat and rabbit kidneys. Furosemide added to the bath fluid decreased arginine-vasopressin (AVP)-stimulated Lp of rat IMCD in a dose-dependent manner, with the threshold effect at 10(-6) M. Furosemide (10(-4) M) reduced Lp from 20.5 +/- 2.3 to 12.1 +/- 1.2 (P less than 0.01) reversibility, but had no effect when added to the perfusate. In addition, furosemide reduced dibutyryl cyclic AMP-stimulated Lp from 20.3 +/- 1.1 to 11.2 +/- 1.6 (P less than 0.01). This effect of furosemide was also observed with indomethacin, a PGE2 synthesis inhibitor. The addition of indomethacin (10(-4) M) to AVP (50 microU/ml) increased Lp from 24.7 +/- 2.3 to 29.7 +/- 2.8 (P less than 0.001), which was reduced to 20.3 +/- 2.6 (P less than 0.001) when furosemide was added to indomethacin in the bath. The inhibitory effect of furosemide on AVP-stimulated Lp was also observed in rabbit IMCD (Lp decreased from 12.8 +/- 0.8 to 5.15 +/- 1.46, P less than 0.02), but it was not observed in the CCT isolated from rabbit kidneys (7.96 +/- 1.87 with AVP vs. 7.94 +/- 1.41 with AVP + furosemide). Furthermore, in rat IMCD the stimulatory effect of AVP on Pu from 7.7 +/- 0.4 to 26.8 +/- 1.3 was reduced by furosemide to 19.7 +/- 1.2 (P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Different effects of furosemide on alpha-adrenoceptors and on platelet aggregation in man.

The effect of a long-term administration of furosemide (2 x 30 mg/day for 3 weeks) on platelet alpha 2-adrenoceptor density and the fraction of high-affinity binding sites, as well as on platelet aggregation induced by adrenaline and ADP, was studied ex vivo in 8 normotensive volunteers. For comparison the in vitro effect of furosemide on platelet aggregation was also evaluated. Furosemide decreased alpha 2-adrenoceptor-density (P less than 0.01) and the fraction of high-affinity binding sites (P less than 0.05). Adrenaline-induced platelet aggregation was not altered ex vivo and in vitro. Furosemide inhibited ADP-induced platelet aggregation ex vivo (P less than 0.05) and in parallel in vitro (P less than 0.01) in a dose-dependent manner. The reduction of the density of alpha 2-adrenoceptors in the high affinity state may be of functional importance for the hemodynamic effects of furosemide. The inhibitory effect of furosemide on ADP-induced platelet aggregation ex vivo and in vitro, which is not related to the effects on adrenoceptors, seems to involve direct effects of furosemide on platelet function. It remains to be seen whether the latter effect is of clinical importance.

Potentiation and Its Mechanism of Cochlear Damage Resulting from Furosemide and Aminoglycoside Antibiotics

The severe ototoxic interaction of the combined administration of furosemide and aminoglycoside antibiotics (kanamycin, streptomycin and gentamicin) was studied in rabbits, and its mechanism clarified. Severe damage occurred not only in the inner ear but also in the kidney when both furosemide and aminoglycoside antibiotics were administered to rabbits. Kanamycin concentration after a single injection of kanamycin with furosemide was much higher not only in the perilymph but also in the cerebrospinal fluid and serum than that after a single injection of kanamycin alone. The ototoxic interaction following the combined use of furosemide and aminoglycoside antibiotics seems to be caused mainly by the inhibitory effect of furosemide on the excretion of aminoglycoside antibiotics from the kidneys.

Effects of Diuretics on Calcium Excretion and Ca2+-Activated ATPase in Rat Kidney

Effects of three diuretics on the urinary Ca2+ excretion and on the microsomal Ca2+-activated ATPase were examined in the rat kidney. Furosemide and bumetanide increased Na+, K+, and Ca2+ excretion in the rats. Acetazolamide increased Na+ and K+ excretion but not Ca2+. Urinary inorganic phosphate excretion was not affected during the administration of acetazolamide. It is suggested that acetazolamide inhibits Na+ transport without affecting Ca2+ reabsorption in the rat nephron. Microsomal ATPase of the rat kidney cortex was stimulated by Ca2+ or Mg2+ alone and an additive effect of the two cations was not observed. Microsomal ATPase activated by Ca2+ or Mg2+ was not inhibited by furosemide, bumetanide, and acetazolamide. These data suggest that the inhibitory effect of furosemide and bumetanide on the Ca2+ reabsorption is not related to the inhibition of Ca2+-activated ATPase.

The effects of transport inhibitors on sodium outflux and influx in red blood cells: evidence for exchange diffusion.

Active sodium transport (outflux or efflux) in red blood cells generally has been measured by assessing the amount of outflux inhibited by digitalis glycosides (outflux-fraction I). The presence of a ouabain-uninhibited sodium outflux (outflux-fraction II) attributable either to a second active transport mechanism or to exchange diffusion has been the subject of recent investigations. In the present study a variety of transport inhibitors, including ouabain, ethacrynic acid, furosemide, oligomycin, and amiloride, were studied for their effects on these components of sodium transport in RBC. In the presence of ouabain both ethacrynic acid and furosemide exerted similar effects on sodium outflux, inhibiting approximately 0.5 mmoles/L of cells per hr. This component of sodium outflux has been called outflux-fraction II. Ethacrynic acid showed no inhibitory potency when ouabain and furosemide were present, thereby suggesting that the same outflux component (fraction II) was affected by ethacrynic acid and by furosemide. In addition, furosemide reduced sodium influx to the same extent that it reduced sodium outflux. Outflux-fraction II, as defined by furosemide, did not contribute a net sodium outflux. These results of sodium outflux and influx experiments confirm the existence of a transport pathway which does not contribute to net flux and which fits the definition of exchange diffusion. The inhibitory effect of furosemide on outflux-fraction II remained despite the use of a sulfhydryl protective reagent, whereas the effect of ethacrynic acid was obliterated. No combination of inhibitors was found which affected the residual or uninhibited sodium outflux (0.4-0.5 mmoles/liter of cells per hr). Oligomycin possessed an inhibitory potency less than that of ouabain, and it exerted no effect on sodium outflux if it was superimposed upon ouabain inhibition. Amiloride proved to be a very weak inhibitor of sodium outflux in human erythrocytes.

Inhibitory effect of furosemide in vitro glucose metabolism of rat epididymal fat pad

Inhibitory effect of furosemide in vitro glucose metabolism of rat epididymal fat pad