Alpha 2-adrenoceptor control of ion and water transport in the newt renal distal tubule.

To study the nature of adrenergic stimulation of ions and water reabsorption in the newt renal distal tubule, stationary microperfusion of the nephron and electron probe analysis were used. After application of norepinephrine (NE 10(-6) M) to the tubule surface, the fractional reabsorption of fluid increased from 15.0 +/- 3.1 to 41.30 +/- 10.4% (n = 7, p < 0.01), of Na+ from 69.30 +/- 6.6 to 79.10 +/- 7.5% (p < 0.05), Cl- from 63.30 +/- 7.6 to 72.40 +/- 7.9% (p < 0.05). Instead of secretion (control), there was reabsorption of K+. Fractional reabsorption of Ca2+ decreased from 51.00 +/- 6.0 to 43.00 +/- 7.0% (p < 0.05). The nonspecific alpha-adrenergic antagonist dibenamine 10(-6) M completely inhibited the effect of NE while, under the action of propranolol (2 x 10(-6) M) NE increased ion and water reabsorption significantly. When applied alone, or with NE, the specific alpha 2-adrenoblocker idazoxan, 2 x 10(-6) M, did not interfere with reabsorption in the distal tubule. At the same time, under the action of alpha 1-adrenoblocker prazosin 2 x 10(-6) M NE, increased the fractional reabsorption of fluid from 24.10 +/- 3.4 to 44.40 +/- 4.0% (n = 6, p < 0.001). These results serve as evidence that there exist specific alpha 2-adrenoceptors in the newt distal tubule the stimulation of which increases membrane permeability of the distal tubule to water, Na+, K+, Cl-, but not to Ca2+.     

Effect of catecholamines on electrolyte and water transport in the nephron of lower vertebrates

Micropuncture technique and electron microprobe analysis have been used to investigate the role of noradrenalin in ion and water balance in the renal tubules of the lamprey Lampetra fluviatilis and newt Triturus vulgaris. Noradrenalin decreased Na, K, and Ca concentrations in the proximal lumen of the lamprey increasing the value of (TF/P)in from 1.1 +/- 0.1 to 1.3 +/- 0.1 (p less than 0.001). Regitin blocked these effects. Noradrenalin perfusion of the peritubular capillaries in newt kidney increased ion and water reabsorption in the proximal segment of the nephron and resulted in differential changes of ion transport in the distal tubule, increasing reabsorption of Na, Cl and K and inhibiting that of Ca and Mg. The rate of glomerular filtration in the nephron remained practically unaffected. The data obtained reveal direct effect of noradrenalin on the renal tubular function in lower vertebrates, this effect being realized presumably via alpha-adrenoreceptors.     

High affinity Ca2+ -Mg2+ ATPase in the distal tubule of the mouse kidney

The purpose of this study was to investigate whether Ca2+ -Mg2+ ATPase in the distal tubule (where calcium transport is active, against a gradient, and hormone dependent) presents some characteristics different from those observed in the proximal tubule, and whether these characteristics are likely to shed light on the respective roles of this enzyme at the two sites of the nephron. The Ca2+ - and Mg2+-dependent ATP hydrolysis was measured in microdissected segments of the distal nephron, the kinetic parameters were determined, and the influence of magnesium upon the sensitivity to calcium was examined. Results were compared with those obtained in the proximal tubule, and in purified membranes as reported by others. In the distal tubule, low concentrations of Mg2+ (less than 10(-7) M) did not influence ATP hydrolysis. At concentrations above 10(-7) M, Mg2+ increased ATP hydrolysis according to Michaelis kinetics (apparent Km = 11.3 +/- 2.4 microM, Vmax = 219 +/- 26 pmol.mm-1.20 min-1). The addition of 1 microM Ca2+ decreased the apparent Km for Mg2+ and the Vmax for Mg2+. Similar results were obtained in the proximal tubule. At low Mg2+ concentrations, Ca2+ also stimulated ATP hydrolysis according to Michaelis kinetics with an apparent Km value for Ca2+ of 0.18 +/- 0.06 and 0.10 +/- 0.03 microM Ca2+ (ns) and a Vmax of 101 +/- 12 and 89 +/- 9 pmol.mm-1.20 min-1 (ns) in the distal and proximal tubules, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Magnesium secretion by the distal segment of the Malpighian tubules of the black field cricket Teleogryllus oceanicus

The short distal segment of unstimulated Teleogryllus Malpighian tubules secreted hyperosmotic fluid containing primarily Mg (125mmoll(-1)), Cl (242mmoll(-1)) and Na (43mmoll(-1)). Remarkably, the volume secreted by the distal segment in unit time was independent of segment length, i.e. the volume was constant regardless of the length of the segment. Magnesium was secreted at a rate of 75.5pmolmin(-1)mm(-1); the highest rate recorded for any epithelium. Low concentrations of K (20mmoll(-1)) were present but almost no P or S. Ca (2.5mmoll(-1)) concentration was higher than in the main segment. The short distal segment secreted 100% of the Mg, 54% of the Cl and 23% of the Na secreted by the whole tubule. The main segment secreted fluid containing primarily K (199mmoll(-1)), Cl (149mmoll(-1)), Na (104mmoll(-1)) and P (48mmoll(-1)) with very low concentrations of Ca (1mmoll(-1)) and S. The main segment appeared to reabsorb a small fraction of the Mg secreted by the distal segment. The fluid secreted by the whole tubule was isosmotic and alkaline, approximately pH8.     

Effects of angiotensin on proximal tubular reabsorption

Effects of angiotensin II on rat, rabbit, and bovine proximal tubular reabsorption have been demonstrated with a variety of techniques, including in vivo microperfusion, free-flow micropuncture of surface and juxtamedullary nephrons, perfusion of isolated tubules in vitro, and cell culture. Blockade of endogenous angiotensin production in vivo with converting-enzyme inhibition, or of receptors with saralasin, consistently inhibits proximal reabsorption of fluid in both superficial and juxtamedullary proximal tubules. Angiotensin effects on the proximal tubule are not neurally mediated, for they persist in denervated kidneys and are seen in nerve-free isolated tubules. Physiological concentrations of angiotensin (10(-11)-10(-9) M) stimulate electroneutral sodium transport from the basolateral membrane, whereas pharmacological doses (10(-7) M and above) inhibit reabsorption. The stimulatory effects appear to be receptor mediated. In addition to these direct effects of angiotensin on the proximal tubule epithelium, endogenous angiotensin may also alter peritubular physical forces to further enhance proximal reabsorption. These effects of angiotensin may represent an important homeostatic mechanism during states of extracellular fluid volume depletion.     

Aminoglycosides inhibit hormone-stimulated Mg2+ uptake in mouse distal convoluted tubule cells

The clinical use of aminoglycosides often leads to renal magnesium wasting and hypomagnesemia. Of the nephron segments, both the thick ascending limb of Henle's loop and the distal tubule play significant roles in renal magnesium conservation but the distal convoluted tubule exerts the final control of urinary excretion. An immortalized mouse distal convoluted tubule (MDCT) cell line has been extensively used to study the cellular mechanisms of magnesium transport in this nephron segment. Peptide hormones, such as parathyroid hormone (PTH), glucagon, calcitonin, and arginine vasopressin (AVP) stimulate Mg2+ uptake in MDCT cells that is modulated by extracellular polyvalent cations, Ca2+ and Mg2+. The present studies determined the effect of aminoglycosides on parathyroid hormone (PTH)-mediated cAMP formation and Mg2+ uptake in MDCT cells. Gentamicin, a prototypic aminoglycoside, elicited transient increases in intracellular Ca2+ from basal levels of 102 +/- 13 nM to 713 +/- 125 nM, suggesting a receptor-mediated response. In order to determine Mg2+ transport, MDCT cells were Mg(2+)-depleted by culturing in Mg(2+)-free media for 16 h and Mg2+ uptake was measured by microfluorescence after placing the depleted cells in 1.0 mM MgCl2. The mean rate of Mg2+ uptake, d([Mg2+]i)/dt, was 138 +/- 24 nM/s in control MDCT cells. Gentamicin (50 microM) did not affect basal Mg2+ uptake (105 +/- 29 nM/s), but inhibited PTH stimulated Mg2+ entry, decreasing it from 257 +/- 36 nM/s to 108 +/- 42 nM/s. This was associated with diminished PTH-stimulated cAMP formation, from 80 +/- 2.5 to 23 +/- 1 pmol/mg protein x 5 min. Other aminoglycosides such as tobramycin, streptomycin, and neomycin also inhibited PTH-stimulated Mg2+ entry and cAMP formation. As these antibiotics are positively charged, the data suggest that aminoglycosides act through an extracellular polyvalent cation-sensing receptor present in distal convoluted tubule cells. We infer from these studies that aminoglycosides inhibit hormone-stimulated Mg2+ absorption in the distal convoluted tubule that may contribute to the renal magnesium wasting frequently observed with the clinical use of these antibiotics.     

Characterization of three types of calcium channel in the luminal membrane of the distal nephron

We previously reported a dual kinetics of Ca2+ transport by the distal tubule luminal membrane of the kidney, suggesting the presence of several types of channels. To better characterize these channels, we examined the effects of specific inhibitors (i.e., diltiazem, an L-type channel; omega-conotoxin MVIIC, a P/Q-type channel; and mibefradil, a T-type channel antagonist) on 0.1 and 0.5 mM Ca2+ uptake by rabbit nephron luminal membranes. None of these inhibitors influenced Ca2+ uptake by the proximal tubule membranes. In contrast, in the absence of sodium (Na+), the three channel antagonists decreased Ca2+ transport by the distal membranes, and their action depended on the substrate concentrations: 50 microM diltiazem decreased 0.1 mM Ca2+ uptake from 0.65 +/- 0.07 to 0.48 +/- 0.06 pmol. microg-1.10 s-1 (P < 0.05) without influencing 0.5 mM Ca2+ transport, whereas 100 nM omega-conotoxin MVIIC decreased 0.5 mM Ca2+ uptake from 1.02 +/- 0.05 to 0.90 +/- 0.05 pmol. microg-1.10 s-1 (P < 0.02) and 1 microM mibefradil decreased it from 1.13 +/- 0.09 to 0.94 +/- 0.09 pmol. microg-1.10 s-1 (P < 0.05); the latter two inhibitors left 0.1 mM Ca2+ transport unchanged. Diltiazem decreased the Vmax of the high-affinity channels, whereas omega-conotoxin MVIIC and mibefradil influenced exclusively the Vmax of the low-affinity channels. These results not only confirm that the distal luminal membrane is the site of Ca2+ channels, but they suggest that these channels belong to the L, P/Q, and T types.     

Cell nuclear DNases: multiplicity and heterogeneity

In order to determine the ratio of activities of major endonucleases of rat liver chromatin, a stepwise fractionation of cell nuclear extracts by chromatography on phosphocellulose and gel filtration through Toyopearl HW60 was carried out. This procedure resulted in partially purified preparations of Ca2+,Mg2+-dependent endonuclease (55 +/- 10 kD), Ca2+,Mg2+-dependent endonuclease (30 +/- 10 kD), Mn2+-dependent endonuclease (30 +/- 5 kD) and acid cation-independent endonuclease. The Ca2+,Mg2+-dependent endonuclease with Mr of 55 +/- 10 kD made up to 57% of the nuclear extract activity in the presence of Ca2+ + Mg2+ and revealed a high calcium-magnesium synergism. Under the same experimental conditions, the 30 +/- 10 kD enzyme made up to 33% of the nuclear extract activity and revealed a low synergism. The activity of Mn2+-dependent endonuclease made up to 26% of the total nuclear extract activity in the presence of Mn2+, that of acid endonuclease--11% of the extract activity in 1 mM EDTA at pH 5.0. It was assumed that the low molecular weight Ca2+,Mg2+-dependent endonuclease represents a product of limited proteolysis of high molecular weight Ca2+,Mg2+-dependent endonuclease.     

Stabilizing influence of calcium and magnesium ions on the decameric structure of chiton hemocyanins

The stabilizing effects of Ca2+ and Mg2+ ions on the decameric structure of hemocyanins from two representative chitons, Stenoplax conspicua and Mopalia muscosa were investigated by light-scattering molecular weight measurements, ultracentrifugation, absorbance, and circular dichroism methods. The dissociation profiles at any given pH resulting from the decrease in divalent ion concentration, investigated at a fixed protein concentration of 0.1 g.liter-1, could be fitted by a decamer-to-dimer-to monomer scheme of subunit dissociation. The initial decline in the light-scattering molecular weight curves required one or two apparent binding sites per hemocyanin dimer formed as intermediate dissociation product, with apparent dissociation constants (kD,2) for Ca2+ ions of 0.7 to 7 X 10(-4) M, not very different from the value of 2.5 X 10(-4) M obtained by Makino by equilibrium dialysis for the hemocyanin of the opistobranch, Dolabella auricularia. The binding of Mg2+ ion to S. conspicua and M. muscosa hemocyanins appears to be both weaker than the binding of Ca2+ and more pH dependent, with kD,2 values ranging from the 3 X 10(-4) to 4 X 10(-2) M at pH 8.5 to 9.5. The dissociation the decameric hemocyanin species (sedimentation coefficient ca. 60 S) is also observed in the ultracentrifugation with the initial appearance of 18-20 S dimers, followed by a shift in equilibrium to monomeric species of lower sedimentation rates of 11-12 S as the divalent ion concentration is reduced below 1 X 10(-4) M Ca2+ and Mg2+. The dissociation of dimers to monomers in the second step of the reaction is characterized by one or two binding sites per subunit and a somewhat stronger affinity for divalent ions, indicated by apparent dissociation constants (kD,1) of 0.7 X 10(-4) to 3 X 10(-3) M. Circular dichroism and absorbance measurements at 222 and 346 nm suggest no significant changes in the conformation of the hemocyanin subunits produced by the different stages of subunit dissociation.     

Composition of luminal fluid secreted by the seminiferous tubules and after reabsorption by the extratesticular ducts of the Japanese quail, Coturnix coturnix japonica

The present report examines the composition of luminal fluid in the seminiferous tubule (STF), rete testis (RTF), and ductus epididymidis of the Japanese quail (Coturnix coturnix japonica). This subject is of particular interest, both because the reproductive ducts are intra-abdominal and because sperm production is more rapid in birds than in mammals. It was interpreted that micropuncture samples of STF contain varying amounts of contamination with intracellular solute, particularly K and protein. The concentration of solute in samples was correlated with packed cell volume (spermatocrit), and when the latter was used to assess estimates of solute concentration in STF, the magnitude of the estimates were much the same as determinations in RTF. Consequently, it is concluded that the fluid entering the rete testis of the quail is the primary secretion of the seminiferous tubules. The composition of RTF in the quail was determined to be 148 mM Na, 126 mM Cl, 9.8 mM K, 2.7 mM Mg, 1.4 mM Ca, 2.1 mM glutamate, 3.4 mM glutamine, 20.2 mM bicarbonate, 1.8 microg microl(-1) of protein, pH 7.34, and 310 mmol kg(-1), and it is significantly different from the composition of blood plasma. Estimates of solute output by the testis and reabsorption by the extratesticular ducts indicate, first, that most of the solutes secreted into the seminiferous tubules are subsequently reabsorbed from the extratesticular ducts and, second, that sufficient solute of testicular origin (except for protein) exists to account for the concentrations of solutes throughout the lumen of the duct system. Changes in the concentration of solute in the extratesticular ducts probably result from different reabsorption rates of solute and water. The composition of fluid from the distal end of the ductus epididymidis was 133 mM Na, 125 mM Cl, 25 mM K, 1.0 mM Mg, 0.3 mM Ca, 6.7 mM glutamate, 4.0 mM glutamine, 19.5 mM bicarbonate, 6.0 microg microl(-1) of protein, pH 7.33, and 335 mmol kg(-1), and it is significantly different from those of RTF and blood.     

Sex different cytochrome-c uptake in the proximal tubule of the rat kidney

The present study deals with the dose- and time-dependent uptake of cytochrome c (CYT c) in the proximal tubule of the rat kidney, and shows that there are segment and sex differences in the reabsorption of CYT c. Rats of both sexes were intravenously injected with different doses of CYT c (0.75-9.0 mg per 100 g body weight), and the kidneys were investigated by light and electron microscopy at different times (3 min, 10 min, and 2 h) after the injection. After 3 and 10 min, CYT c was demonstrated in apical vacuoles of different sizes and in some lysosomes of the S1 and S2 segments, whereas after 2 h, CYT c was found only in lysosomes of all three segments of the proximal tubule. At these times, the S1 segment contained more CYT c than the S2 and S3 segments. However, 2 h after the injection of 6 or 9 mg CYT c, the differences between the S1 and S2 segments disappeared almost completely, due to a strong lysosomal accumulation of CYT c in the S2 segment. At all studied times and CYT-c doses, the S3 segment contained less CYT c than the S1 and S2 segments. On the whole, different levels of CYT-c reabsorption were found in the different segments of the proximal tubule, which was saturable with increasing CYT-c doses, i.e. firstly in the proximal and then in the distal parts of the proximal tubule. Two hours after the injection of CYT c, a difference between males and females was observed, with the lysosomes of the S1 and S2 segments of females containing more CYT c than those of males. Thus, more CYT c was reabsorbed in the proximal tubule of females than in that of males.     

The actions of cortisol on fetal renal function

Renal function was studied in 6 fetal sheep, aged 126-135 days, before and after 3 injection of 15 mg of cortisol given at intervals of 12 h. Cortisol caused a significant rise in both renal blood flow (P less than 0.05) and glomerular filtration rate (P less than 0.005), and in urine flow rate (P less than 0.02) but it did not consistently cause a natriuresis. The urinary pH was unchanged following cortisol treatment, but bicarbonate excretion increased. Urinary phosphate excretion was increased (P less than 0.005) because of a rise in filtered phosphate and a fall in phosphate reabsorption. The titratable acid excretion increased (P less than 0.005) but urinary ammonium excretion did not. The total amount of sodium reabsorbed increased after cortisol but the amount of sodium reabsorbed in the proximal tubule did not increase, so fractional reabsorption in the proximal tubule decreased from 61.7 +/- 4.1% to 47.3 +/- 4.2% (P = 0.01). The total amount of sodium reabsorbed in the distal tubule increased and distal fractional reabsorption increased from 33.3 +/- 2.4% to 47.3 +/- 4.2% (P less than 0.01). Cortisol may increase the capacity of the immature kidney to play a role in fluid and electrolyte homeostasis by increasing glomerular filtration rate and delivering more sodium and water to the distal nephron where the reabsorption of sodium and water can be modified independently and in accordance with need.     

Mg2+ transport in the kidney.

Magnesium is abundant in biological systems and an important divalent cation in the human body. Mg2+ helps mediate cellular energy metabolism, ribosomal and membrane integrity. Additionally Mg2+ modulates the activity of several membrane transport and signal transduction systems. Despite its importance however, little is known about the molecular mechanisms of Mg2+ transport and homeostasis in mammals. In mammals the amount of Mg2+ absorption is about the same as the amount of Mg2+ excretion in urine. Additionally, when total Mg2+ intake is deficient, the kidney is capable of reabsorbing all filtered Mg2+. This balance between intake and excretion indicates that the kidney plays a principal role in maintenance of total body Mg2+ homeostasis. Within the kidney, Mg2+ filtered by the glomerulus is handled in different ways along the nephron. About 10-20% of Mg2+ is reabsorbed by the proximal tubule. the bulk of Mg2+ (about 50-70%) is reabsorbed by the cortical thick ascending limb of the loop of Henle. In this region, Mg2+ moves across the epithelium through the paracellular pathway, driven by the positive lumenal transepithelial voltage. A recently cloned human gene, paracellin-1 was shown to encode a protein localized to the tight junctions of the cortical thick ascending limb and is thought to mediate Mg2+ transport via the paracellular space of this epithelium. The distal convoluted tubule reabsorbs the remaining 5-10% of filtered Mg2+. This segment seems to play an important role in determining final urinary excretion, since there is no evidence for significant Mg2+ absorption beyond the distal tubule. Although many renal Mg2+ transport activities have been characterized, no Mg2+ transporter cDNAs have been cloned from mammalian tissues. Recent research has certainly expanded our knowledge of Mg2+ transport in kidney; but details of the transport processes and the mechanisms by which they control Mg2+ excretion must await cloning of renal Mg2+ transporters and/or channels. Such information would provide new concepts in our understanding of renal Mg2+ handling.     

Parathyroid hormone stimulates ATP-dependent calcium pump activity by a different mode in proximal and distal tubules of the rat.

A new technique was developed to isolate basolateral membrane vesicles individually from proximal and distal tubules of the rat cortex. This new technique enabled us to study differences in their kinetics and mechanisms of hormonal regulation of Ca pump between proximal and distal tubules. The Ca pump in distal tubule has very high affinity (42.6 nM Ca2+) and the one in proximal tubule has relatively low affinity (75.6 nM Ca2+). Parathyroidectomy (PTX) decreased the Vmax of Ca pump activity in proximal tubule (4.68 +/- 0.99 vs. 9.08 +/- 2.21 nmol 45Ca2+/min per mg protein BLMV, P less than 0.05), while it increased Km in distal tubule (93.1 +/- 11.0 vs. 35.1 +/- 16.1 nM Ca2+, P less than 0.05). Restoration of serum Ca2+ concentration by 1,25(OH)2D3 supplement could not reverse these changes by PTX in Ca pump activity in either the proximal or the distal tubule. In conclusion, this study strongly suggested that parathyroid hormone stimulated Ca pump activity by increasing the Vmax in proximal tubule and by increasing the affinity in distal tubule. 1,25(OH)2D3 does not have a direct effect on the basolateral membrane Ca pump activity.     

Immunosuppressants inhibit hormone-stimulated Mg2+ uptake in mouse distal convoluted tubule cells

Immunosuppressants such as cyclosporinA and FK506 (tacrolimus) are widely prescribed to treat numerous disorders and to treat organ transplant recipients. However, cyclosporine A and FK506 are both known to produce hypomagnesaemia. The mechanism of this effect is still unclear. The present study determined the effects of immunosuppressant treatment on the parathyroid hormone (PTH) mediated Mg(2+) uptake and the mitogen-activated protein kinase (MAPK) activation in mouse distal convoluted tubule (MDCT) cells. The intracellular Ca(2+) and Mg(2+) concentrations in a single MDCT cell were measured by using the fluorescentdye Fura-2 AM and Mag-fura-2 AM, respectively. Cyclosporine A and FK506 illicited a transient increase of intracellular Ca(2+) from a basal level of 99 +/- 16 nM to 685 +/- 105 and 608 +/- 96 nM, respectively. In order to determine the Mg(2+) transport, the MDCT cells were Mg(2+)-depleted by culturing them in Mg(2+)-free media for 16 h, and the Mg(2+) uptake was measured by microfluorescence after placing the depleted cells in 1.5mM MgCl(2). The mean rate of Mg(2+) uptake, d([Mg(2+)](i))/dt, was 140 +/- 16 nM/s in the control MDCT cells. PTH increased the Mg(2+) uptake more than 2 times in this cell. Cyclosporine A (10 microM) and FK506 (0.1 microM) did not affect the basal Mg(2+)uptake (140 +/- 16 and 142 +/- 14 nM/s, respectively), but they inhibited the PTH-stimulated Mg(2+) entry, decreasing it from 248+/-12 to 147 +/- 7 and 148 +/- 14 nM/s, respectively. These effects were inhibited by L685818, which is a potent competitive antagonist of FK506. PTH stimulated the extracellular signal-regulated kinase1/2 (ERK1/2) protein synthesis. This PTH-stimulated ERK1/2 activation was inhibited by cyclosporine A and FK506. In the present study, the role of ERK1/2 activation on the PTH-dependent magnesium uptake was examined in MDCT cells, and we showed that immunosuppressants inhibit the hormone-stimulated Mg(2+) uptake into the MDCT cells by inhibiting the MAPK pathway.     

A search for a defect of proximal transport in denervated kidneys of conscious dogs

The influence of renal nerves on proximal Na+ reabsorption was studied in clearance experiments with unilaterally renal-denervated conscious dogs prepared by surgical bladder division. Two types of experiments were made : A. maximal water diuresis, and B. Total blockade of distal NaCl reabsorption with ethacrynic acid and chlorothiazide. In maximal water diuresis CH2O + CNa was used as a measure of fluid delivery to the distal nephron. At similar GFR on both sides, the proximal reabsorption estimated as GFR--(CH2O + CNa) was 38.4 +/- 5.6 ml/min for the intact and 35.9 +/- 4.2 ml/min for the denervated kidney (n = 6, difference NS). After distal tubular blockade, proximal Na+ reabsorption calculated as filtered load minus urinary excretion was 3.84 +/- 0.43 mmol/min for the intact and 3.91 +/- 0.36 mmol/min for the denervated kidney (n = 6, difference NS). The fractional reabsorption of NA+ was 64.9 +/- 1.0% for the intact and 66.9 +/- 1.1% for the denervated kidney (difference NS). In contrast to data from renal denervation studies with anaesthetized animals, the present experiments did not show any difference in proximal reabsorption between the innervated- and denervated kidney. We conclude that in absence of anaesthesia renal efferent nerves have no major effect on NaCl transport in dog proximal tubule.     

Association of calcium and sodium handling in the rabbit nephron. A micropuncture study.

Two-phase recollection micropuncture experiments were performed on female New Zealand rabbits to investigate the effect of flow rate (volume-expansion) compared to reabsorptive rate (furosemide) on calcium and sodium handling along the nephron. Group 1 (n = 6) rabbits represented nonvolume-expanded animals. Each experiment was conducted with a control phase followed by a second phase of furosemide administration (1 mg/kg/min). Group 2 rabbits (n = 6) were initially volume-expanded to 3% body weight with modified Ringers. The fractional excretion of sodium and calcium in the control phase of group I and II was 3 +/- 1 and 22 +/- 6% and 4 +/- 1 and 26 +/- 2%, respectively. Fractional excretion of sodium, calcium and magnesium rose after furosemide administration. The effect of volume expansion on sodium, calcium and magnesium remaining in the proximal tubule was relatively modest and not affected by furosemide. Our distal micropuncture data reveal that volume expansion has a greater inhibitory effect on fluid reabsorption at a site beyond the proximal micropuncture site (group 1, 9 +/- 2%, group 2,22 +/- 2%). After furosemide infusion, the amount of electrolytes remaining rose similarly in both groups; however, additional sodium and calcium reabsorption did not occur in the volume-expanded group in the final segment of the nephron. These results indicate that calcium reabsorption by the cortical terminal segment of the rabbits is passive similar to that suggested by the in vitro perfused study since no additional calcium reabsorption is seen in the volume-expanded rabbit.     

Regulation of renal proximal and distal tubule transport: sodium, chloride and organic anions

Renal tubular transport and its regulation are reviewed for Na(+) (and Cl(-)), and for fluid and organic anions (including urate). Filtered Na(+) (and Cl(-)) is reabsorbed along the tubules but only in mammals and birds does most reabsorption occur in the proximal tubules. Reabsorption involves active transport of Na(+) and passive reabsorption of Cl(-). The active Na(+) step always involves Na-K-ATPase at the basolateral membrane, but the entry step at luminal membrane varies among tubule segments and among vertebrate classes (except for Na(+)-2Cl(-)-K(+) cotransporter in diluting segment). Regulation can involve intrinsic, neural and endocrine factors. Proximal tubule fluid reabsorption is dependent on Na(+) reabsorption in all vertebrates studied, except ophidian reptiles. Fluid secretion occurs in glomerular and aglomerular fishes, reptiles and even mammals, but its significance is not always clear. A non-specific transport system for net secretion of organic anions (OAs) exists in the proximal renal tubules of almost all vertebrates. Net transepithelial secretion involves: (1) transport into the cells at the basolateral side against an electrochemical gradient by a tertiary active transport process, in which the final step involves OA/alpha-ketoglutarate exchange and (2) movement out of the cells across the luminal membrane down an electrochemical gradient by unknown carrier-mediated process(es). Regulation may involve protein kinase C and mitogen-activated protein kinase. Urate is net secreted in the proximal tubules of birds and reptiles. This process is urate-specific in reptiles but in birds, it may involve both a urate-specific system and the general OA system.     

Characterization of the Ca2+- or Mg2+-ATPase of transverse tubule membranes isolated from rabbit skeletal muscle

Transverse tubule membranes isolated from rabbit skeletal muscle have high levels of a Ca2+- or Mg2+-ATPase with Km values for Ca-ATP or Mg-ATP in the 0.2 mM range, but do not display detectable levels of ATPase activity activated by micromolar [Ca2+]. The transverse tubule enzyme is less temperature or pH dependent than the Ca2+-ATPase of sarcoplasmic reticulum and hydrolyzes equally well ATP, ITP, UTP, CTP, and GTP. Of several ionic, non-ionic, and zwitterionic detergents tested, only lysolecithin solubilizes the transverse tubule membrane while preserving ATPase activity. After extraction of about 50% of the transverse tubule proteins by solubilization with lysolecithin most of the ATPase activity remains membrane bound, indicating that the Ca2+- or Mg2+-ATPase is an intrinsic membrane enzyme. A second extraction of the remaining transverse tubule proteins with lysolecithin results in solubilization and partial purification of the enzyme. Sedimentation of the Ca2+- or Mg2+-ATPase, partially purified by lysolecithin solubilization, through a continuous sucrose gradient devoid of detergent leads to additional purification, with an overall 3- to 5-fold purification factor. The purified enzyme preparation contains two main protein components of molecular weights 107,000 and 30,000. Cholesterol, which is highly enriched in the transverse tubule membrane, copurifies with the enzyme. Transverse tubule membrane vesicles also display ATP-dependent calcium transport which is not affected by phosphate or oxalate. The possibility that the Ca2+- or Mg2+-ATPase is the enzyme responsible for the Ca2+ transport displayed by isolated transverse tubules is discussed.     

The renal connecting tubule: Resolved and unresolved issues in Ca(2+) transport

The renal connecting tubule (CNT) localizes to the distal part of the nephron between the distal convoluted tubule and the collecting duct, and consists of two different cell types: segment-specific and intercalated cells. The former reabsorb water (H(2)O), sodium (Na(+)) and calcium (Ca(2+)) ions to the blood compartment, while secreting potassium ions (K(+)) into the pro-urine. The latter cells contribute to the renal control of the acid-base balance. Several factors and hormones tightly regulate these transport processes. Although the CNT reabsorbs only ～15% of filtered Ca(2+) load, this segment is finally decisive for the amount of Ca(2+) that appears in the urine. Impaired Ca(2+) transport across CNT can provoke severe urinary Ca(2+) excretion, called hypercalciuria. This review mainly focuses on the activity, abundance and expression of the epithelial Ca(2+) channel named Transient Receptor Potential Vanilloid 5 (TRPV5) that is the gatekeeper of active Ca(2+) reabsorption in the CNT.