Claude Amiel

Stimulation by glucagon and PTH of Ca and Mg reabsorption in the superficial distal tubule of the rat kidney

The effects of glucagon and PTH on electrolyte reabsorption in the distal tubule were investigated in rats deprived of vasopressin, calcitonin, PTH, and glucagon. Micropunctures of distal tubule, at a late and an early site of a same nephron, have been performed in 23 rats, nine control, seven infused with glucagon (5 ng·min−1·100 g−1 b.w.) and seven with PTH (5 mU·min−1·100 g−1 b.w.). The Ca and Mg reabsorptive capacity of the distal segment was increased by glucagon and by PTH. Moreover, fractional Na and Cl reabsorption was significantly higher than in control during PTH administration. A K secretion appeared during the administration of both hormones. No phosphate net transport was observed in any group. Finally, the data presented here, together with those previously reported, indicate that the increase of Ca and Mg renal reabsorption observed with glucagon and PTH results from an effect located in both Henles loop, where the bulk of Ca and Mg is reabsorbed, and the distal tubule.

Ethacrynic acid facilitates gentamicin entry into endolymph of the rat.

Influence of ethacrynic acid (EA) upon gentamicin kinetics in perilymph and endolymph was studied in rats that were given a constant-infusion of gentamicin (150 micrograms/min) and EA (140 micrograms/min). Inner ear fluids and plasma were sampled up to 5 h. The purity of the endolymph was ensured by measurement of sodium and potassium concentrations. Gentamicin assay was done with a modified radioimmunoassay. Results show that EA facilitates the entry of gentamicin into endolymph, while it does not affect the kinetics of the drug in perilymph. Although the mechanism of this facilitation remains unclear, this result may account for the ototoxic potentiation reported between EA and aminoglycoside antibiotics.

Benzyl alcohol increases membrane fluidity and modulates cyclic AMP synthesis in intact renal epithelial cells

To evaluate a possible modulation by membrane fluidity of hormonal, cAMP-mediated effects on renal epithelial cells, we studied the effect of the neutral local anesthetic, benzyl alcohol, on membrane fluidity and on basal and stimulated intracellular cAMP content in intact MDCK cells. Benzyl alcohol induced a dose-dependent decrease of lipid order which was measured by steady-state fluorescence anisotropy using trimethylammonium-diphenylhexatriene and propionyl-diphenylhexatriene as fluorescent probes. Benzyl alcohol induced a 2-fold increase in basal cAMP content, likely as a consequence of increased prostaglandin synthesis since this effect was abolished by indomethacin. The effect of benzyl alcohol on stimulated cAMP synthesis depended on the nature of the ligand: 10 mM benzyl alcohol increased significantly the stimulatory effect of prostaglandin E2, glucagon and forskolin but not of vasopressin. At higher concentrations (40 mM), benzyl alcohol did not affect significantly the glucagon-stimulated cAMP content, while it inhibited significantly the prostaglandin E2-, forskolin- and vasopressin-stimulated cAMP synthesis. The 40 mM benzyl alcohol-induced inhibition was reversed by 1 mM Mn2+, which is known to block the inhibitory GTP-binding protein Ni. These results suggest that: (i) the various components of the adenylate cyclase-cAMP system and their coupling are affected differently by changes in membrane fluidity, which might reflect differences in their lipid environment, (ii) changes in membrane fluidity can modulate responses of renal tubular cells to hormones, and thus tubular functions.

Increase in membrane fluidity modulates sodium-coupled uptakes and cyclic AMP synthesis by renal proximal tubular cells in primary culture

In order to evaluate the influence of membrane fluidization on three apical transport systems and on a basolateral enzyme, and to analyse the mechanisms involved, we studied, in cultured rabbit proximal tubular cells, the effect of increasing concentrations of the local anesthetic drug benzyl alcohol on Na(+)-dependent uptakes of phosphate (Pi), methyl alpha-D-glucopyranoside (MGP), and L-alanine, as well as on basal and stimulated cyclic AMP content. At 10 mM, benzyl alcohol increased the Vmax of Pi uptake by 31%, decreased that of MGP uptake by 24%, and did not affect alanine uptake. Km values were not affected. Benzyl alcohol, up to 40 mM, increased in a concentration-dependent manner basal, PTH-stimulated, and cholera toxin-stimulated, but not forskolin-stimulated cyclic AMP accumulation. In the presence of 40 mM benzyl alcohol, the magnitude of PTH-induced inhibition of Pi uptake was enhanced from 11% to 24%. It is concluded that: (i) fluidization of apical membranes affected differently Na+/Pi, Na+/MGP, and Na+/alanine cotransports, reflecting differences in the lipidic environments of these transport system; (ii) fluidization of basolateral membranes enhanced PTH-stimulated cyclic AMP generation through improved coupling between the receptor-GS complex and the catalytic subunit of adenylate cyclase; (iii) these variations may result in physiological and pathophysiological modulation of the renal handling of solutes and of the phosphaturic effect of PTH.

Effect of Locally Applied Drugs on the Endolymphatic Sac Potential

In Ménières disease, an inner ear disorder related to an endolymphatic hydrops, an alteration of the functioning of the endolymphatic sac has been proposed. The endolymphatic sac is assumed to be involved in the secretion/resorption of endolymph. The epithelial transport systems have been indirectly studied by the recording of the endolymphatic sac transepithelial potential (ESP) in control conditions and after the local injection of drugs such as diuretics that have been proposed in the treatment of Ménières disease. The ESP was recorded, in vivo, in guinea pigs up to 150 minutes after the perisaccular injection of 5 μL of a 150 mmol/L (mM) NaCl solution containing various drugs known to inhibit ionic transport systems. The initial ESP was +8.4 ± 0.3 mV (mean ± SEM, n = 78). The basolateral injection of 5 μL of 150 mM NaCl induced an ESP decrease of 64% ± 6.0% (n = 12), 5 minutes after the end of the injection. Then ESP increased, returning to its initial value at 60 minutes and surpassing it at 120 minutes. Diuretics such as acetazolamide (10−3 mol/L [M]), an inhibitor of carbonic anhydrase, and amiloride (10−4 M), an inhibitor of Na channel or Na/H exchanger, decreased the ESP recovery. At variance, bumetanide (10−6 M, 10−4 M), the Na‐K‐Cl cotransport inhibitor, and chlorothiazide (10−4 M), a Na‐Cl cotransporter inhibitor, failed to alter the ESP as compared with the control group. Ouabain (10−3 M), the Na+,K+‐adenosine triphosphatase (ATPase) inhibitor, prevented the ESP recovery otherwise observed 60 minutes after the NaCl injection. Bafilomycin A1, the inhibitor of the vacuolartype H+‐ATPase, prevented the recovery of the ESP with a log‐dose/effect (10−5 M, 10−6 M, 10−8 M). Disulfonic acid stilbene (DIDS) (10−4 M), an inhibitor of transporters involving HCO3−, also prevented the ESP recovery. These results suggest that the genesis of the ESP was highly dependent on acid‐base transport systems including carbonic anhydrase, a vacuolar‐type H+‐ATPase, and an anionic transport system blocked by DIDS. Further studies are needed to confirm the alteration of the acid‐base balance in this epithelium and its possible involvement in the pathogenesis of Ménières disease.

Calcium and the Inner Ear Fluids

Total calcium (Ca) concentration in inner ear fluids was determined by fluorimetry, emission and absorption spectrophotometry, and electron probe analysis. The ionized Ca was measured with selective microelectrodes. In perilymph, the total Ca concentration (1.2 mM) was similar to the ultrafiltrable Ca concentration in plasma. The fraction of ionized Ca was 80%. In endolymph, a total Ca concentration of 0.5 mM contrasted with a reported ionized Ca concentration of 0.02 mM, which suggests, as a working hypothesis, that most of the Ca could exist as bicarbonate and/or phosphate undissociated salts. The decrease in the endocochlear potential induced an increase of the ionized fraction of the Ca. The electrochemical potential of Ca across the perilymph-endolymph barrier implies an active entry of Ca into the endolymph.

Antidiuretic hormone stimulation of adenylate cyclase in semicircular canal epithelium

SummaryBasal adenosine 3′,5′-cyclic monophosphate (cAMP) content and the modulation of its production were studied in the frogs semicircular canal epithelium. This epithelium secretes endolymph, a K+-rich, positively polarized fluid. The basal cAMP content measured by microradioimmunoassay was 244 ± 14.2 fmol/structure per 5 min (n = 30). This content was increased about 8 times by 10−5M forskolin. Vasotocin, the frog antidiuretic hormone, increased the cAMP production by factors of 1.3 and 3.3 at concentrations of 10−8M and 10−7 M, respectively. This stimulatory effect of vasotocin was blunted by the addition of α2-adrenergic agonists, such as 10−8M-10−5M norepinephrine, in the presence of 10−5M propranolol, or 10−5M clonidine. Prostaglandin E2 at a concentration of 10−8M, which did not affect the cAMP production, did not modify the response to vasotocin. Glucagon (10−6M), calcitonin (10−6M), and parathyroid hormone (10 units/ml) did not affect the cAMP content. Prostaglandin E2 (10−7M) and the β-adrenergic agonist isoproterenol (10−6M) stimulated the cAMP production by a factor of 1.6. These results indicate that the frog semicircular canal is a target of both vasotocin and catecholamines and that catecholamines through α2-receptors modulate vasotocin-induced cAMP generation. Further, this interaction might be of physiological relevance in the modulation of ion transport in this structure.

Tolerance of once‐daily dosing of netilmicin and teicoplanin, alone or in combination, in healthy volunteers

The purposes of this study were to test the pharmacokinetics and renal and otologic tolerances of a once‐daily regimen of netilmicin and teicoplanin administered intramuscularly, alone or in combination (4.5 and 6 mg/kg, respectively), for 7 days in 30 healthy male volunteers. Teicoplanin induced only a mild increase in enzymuria. Nephrotoxicity was moderate and reversible with netilmicin; there was increased enzymuria and alteration in diluting ability, without significant changes in urinary β2‐microglobulin levels, concentrating ability, and glomerular filtration rate. Ototoxicity was not detected in any of the subjects. Our results suggest that (1) teicoplanin and netilmicin given once daily induced only slight, reversible tubular damage, without any sign of ototoxicity; (2) their combination was not more toxic; and (3) clinical studies can be envisaged to evaluate the efficacy and tolerance of once‐daily regimens in long‐term treatment.

Effect of glucagon on magnesium renal reabsorption in the rat

The recent localization, in the rat, of a glucagon-sensitive adenylate cyclase in these segments where the bulk of calcium and magnesium is reabsorbed suggests an effect of this hormone on calcium and magnesium tubular transport. Renal tubular handling of calcium and magnesium as well as of sodium and phosphate was therefore studied by clearance methods in anesthetized rats, either intact or thyroparathyroidectomized (TPTX), infused with glucagon at a rate of 25 ng·min−1/100 g bw just after a priming dose of 2.5 μg. The hormone administration resulted in a significant decrease of absolute and fractional magnesium excretion (from 16.3±0.7% to 9.7±1.7% for intact rats and from 20.9±1.8% to 6.9±1.0% for TPTX rats), associated with the well-known increase in sodium and phosphate fractional excretion. Moreover, a small and transient decrease of calcium fractional excretion was observed concomitantly with a decrease of plasma calcium concentration. The significant increase in magnesium absolute reabsorption, observed whatever the filtered load and independently of PTH and calcitonin, may be an evidence for a direct tubular effect of glucagon.

Normal renal function 8 to 13 years after Cyclosporin A therapy in 285 diabetic patients

Cyclosporin A (CyA) may induce acute nephrotoxicity. The question has been raised of the possible long‐term unfavorable course of CyA‐induced lesions. Advantage was taken of a large cohort of diabetic patients treated for several months using moderate CyA dosage to evaluate the long‐term evolution of renal function in such patients.