Search

Patients presenting with the combined finding of severe symptomatic hyponatremia and hypoxia have such high mortality rates that they should be admitted to an intensive care unit and intubated sooner rather than later. Without delay, these patients need rapid correction of their serum sodium by 8-10 mequiv. per liter and an increase in their partial pressure of arterial oxygen to values above 70 mm Hg.

Purpose: Diabetic ketoacidosis (DKA) and alcoholic ketoacidosis (AKA) are two medical emergencies characterized by elevated total ketone body concentration. We aimed to determine differences in pathogenesis of ketoacidosis and its metabolic consequences by comparing both at presentation and during treatment, the different metabolic products and hormones involved in the ketoacidotic state., Materials and Methods: We studied 12 patients with DKA and 8 patients with AKA. On admission and every 4 hours for 24 hours during treatment, samples were drawn for determination of serum ketone bodies, lactate and pyruvate, insulin, and counterregulatory hormones (glucagon, cortisol, growth hormone, and catecholamines)., Results: At presentation, with a similar beta-hydroxybutyrate concentration, patients with DKA had a higher plasma glucose (32 mmol/L vs. 6.6 mmol/L), lower beta-hydroxybutyrate/acetoacetate ratio (3:1 vs. 7:1), and a lower lactate/pyruvate ratio (11:1 vs. 19:1) than patients with AKA (all, P < .01). The mean time to resolve ketoacidosis in patients with AKA (6 +/- 1 hour) was significantly shorter than in patients with DKA (16 +/- 2 hours). At presentation, the mean insulin concentration in patients with DKA and AKA were similarly decreased (7.8 +/- 2 and 10.3 +/- 3 microU/mL, P = not significant [NS]). The mean glucagon level before therapy was 203 +/- 15 pg/mL and 188 +/- pg/mL for patients with DKA and AKA, respectively (P = NS). Levels of cortisol, growth hormone, and epinephrine at presentation and during the first 8 hours of treatment were higher in patients with DKA; however, the difference in these values did not reach statistical significance. During therapy, levels of counterregulatory hormones declined at similar rates and returned to normal values after resolution of ketoacidosis., Conclusions: Our results indicate that, in addition to a history of diabetes or alcoholism, patients with DKA and AKA differ in their metabolic parameters more than in their hormonal profile. The metabolic profile of DKA is characterized by a higher plasma glucose concentration, and lower beta-hydroxybutyrate to acetoacetate and lactate to pyruvate ratios compared with patients with AKA. The initial hormonal profile in both ketoacidotic states is characterized by similarly decreased insulin levels and elevated levels of counterregulatory hormones.

A 25-year-old man with acquired Bartter's syndrome, mitral valve prolapse, and supraventricular tachycardia secondary to a low atrial focus was diagnosed with asthma. The unique aspects of managing these coexisting diseases are evaluated. Calculation of free-water clearance in the diagnosis of Bartter's syndrome and the etiology and characteristics of the syndrome are discussed.

The production of a concentrated urine is achieved by countercurrent multiplication in the renal medulla. While the single effect in the outer medulla is known to be active NaCl reabsorption in the thick ascending limb, the single effect in the inner medulla is not definitively established. However, the passive model of Kokko and Rector [1] and Stephenson [2] remains the most widely accepted mechanism for the single effect in the inner medulla. Continued experimental studies of transport in perfused inner medullary nephron segments and mathematical simulations that incorporate these new experimental values and anatomic complexity will be needed to fully elucidate the process of urinary concentration. In addition, the availability of molecular reagents will permit investigation into the molecular mechanisms that regulate transport proteins which play crucial roles in the urinary concentrating mechanism.

Water homeostasis is regulated in large part by the proper operation of the urinary concentrating mechanism. In the renal inner medulla, urea recycling from the inner medullary collecting duct to the inner medullary interstitium is thought to be essential for the production of a concentrated urine; however, it has not been possible to test this hypothesis in humans. Recently, a unique combination of genetic abnormalities has been described: absence of Kidd blood group antigens and absence of carrier-mediated urea transport in erythrocytes. Because animal studies indicate a similarity between urea transport in red blood cells and the nephron, it was postulated that patients without the Kidd antigen might lack facilitated urea transport in their kidneys. Hence, their ability to concentrate urine maximally was measured. Current models of nephron function would predict that in the complete absence of urea transport, the maximal concentrating ability would be around 800 to 900 mosM/kg H2O. Two homozygous patients had a moderate decrease in maximal concentrating ability (UosM,max = 819 mosM/kg H2O); a heterozygote also had some limitation. These studies raise the possibility that the erythrocyte urea transporter and the kidney urea transporter are encoded by a single gene (detected by the mutational loss of the Kidd antigen) and that a lack of facilitated urea transport impairs urea recycling in the kidney and, hence, maximal urinary concentrating ability.

Urinary concentration is achieved by countercurrent multiplication in the inner medulla. The single effect in the outer medulla is active NaCl absorption from the thick ascending limb. While the single effect in the inner medulla is not definitively established, the majority of experimental data favors passive NaCl absorption from the thin ascending limb. Continued experimental studies in inner medullary nephron segments will be needed to elucidate fully the process of urinary concentration.

Recent studies have shown that in situ PCO2 in rat renal cortical structures far exceeds systemic arterial PCO2. These results were opposite to previous assumptions that renal proximal tubule fluid PCO2 approximated arterial PCO2. The present studies examined the species and organ specificity of the elevated PCO2 in 39 New Zealand White rabbits studied under normal acid-base conditions. In situ PCO2 was measured in renal cortex, superficial hepatic parenchyma, skeletal muscle, superficial cerebral cortex, and femoral nerve, artery, and vein. The results showed rabbit renal cortical PCO2 (57.2 +/- 1.2 mmHg) to be higher than both systemic arterial (39.1 +/- 2.0 mmHg) and venous PCO2 (45.4 +/- 2.1 mmHg). Similarly, liver PCO2 (64.1 +/- 3.5 mmHg) was found to be significantly higher than systemic arterial and venous PCO2 and also higher than portal and hepatic vein PCO2. Skeletal muscle, cerebral cortex, and femoral nerve PCO2 levels were usually greater than systemic arterial PCO2 but less than systemic venous PCO2. These observations show that in situ PCO2 is significantly elevated above afferent and efferent blood PCO2 in the kidney and liver but not in muscle or brain. A possible explanation for these findings in the former two organs may be high CO2 production and/or trapping of CO2 by their vascular systems.

Studies were undertaken to examine the mechanism whereby changes in intraluminal flow rates after reabsorption in the isolated perfused proximal tubule of the rabbit. All protocols employed the technique of in vitro perfusion of isolated segments of the proximal convoluted tubule. Stepwise elimination of d-glucose and l-alanine from an artifical perfusate stimulating ultrafiltrate decreased the unidirectional flux of sodium, transtubular potential difference, and net water absorption. Using isosmolal ultrafiltrate as the perfusate, net fluid reabsorption and the unidirectional lumen-to-bath flux of sodium and chloride decreased with a decrease in flow rate below 11 nl/min, but neither net fluid reabsorption nor the unidirectional fluxes of sodium and chloride increased further as the perfusion rate was increased above 11 nl/min. The unidirectional flux of 14C-urea was not affected by changes of perfusion rate from 1.6 to 44 nl/min. The dependence of net fluid reabsorption and unidirectional fluxes of sodium and chloride on flow rate per se, and not on intraluminal hydrostatic pressure or geometry, was established by demonstrating their decrease despite a rise in intraluminal pressure and inside diameter produced by counterpressure at the collecting end of the tubule, while flow was decreased. Ouabain decreased net fluid reabsorption to near zero at all flow rates, but ouabain had no effect on the flow-dependency of unidirectional sodium anf sodium was eliminated with a decrease in bicarbonate concentration and removal of d-glucose and l-alanine from the perfusate. Thus, the present studies demonstrate that net water and unidirectional sodium and chloride fluxes are flow-dependent. At flow rates somewhere below 11 nl/min, unidirectional fluxes decreased with decreasing perfusion rates; however, at perfusion rates greater than 11 nl/min, there was no further effect of perfusion rate on either net water absorption or the unidirectional fluxes of sodium or chloride. These effects may be partly mediated through the flow-dependent changes in the intraluminal concentration of bicarbonate, d-glucose, and 1-alanine.

With the exception of aldosterone, little is known about the hormonal regulation of distal nephron acidification. These experiments investigated the effects of prostaglandin E2, indomethacin, lysyl-bradykinin, 8-bromo-cyclic AMP, and forskolin on proton secretion in the major acidifying segment of the distal nephron, the medullary collecting duct from inner stripe of outer medulla. Using in vitro microperfusion and microcalorimetry, net bicarbonate reabsorption (proton secretion) was measured in rabbit medullary collecting ducts before, during, and after exposure to each test substance. PGE2 reduced proton secretion 12.2%, while the following substances stimulated proton secretion: indomethacin 14.2%; 8-bromo-cyclic AMP 34.5%; forskolin 39%. Lysyl-bradykinin was without effect. These studies demonstrate that distal nephron acidification, in addition to being stimulated by aldosterone, is significantly inhibited by the hormone PGE2. The stimulation of proton secretion by cAMP suggests that other hormones known to activate adenylate cyclase may also influence distal nephron acidification.

The possibility that mineralocorticoids have a direct influence on renal Na-K ATPase activity has been the focus of intense research effort and some controversy for a number of years. Early studies were hindered by an inability to differentiate between possible glucocorticoid vs. mineralocorticoid effects on this enzyme within the multitude of cells that comprise the heterogeneous mammalian nephron. This study attempts to circumvent this problem by monitoring Na-K ATPase activity in the rabbit renal cortical collecting tubule (CCT), a proposed target epithelium for mineralocorticoids. Using an ultramicro assay, Na-K ATPase activity was measured in CCT from normal, adrenalectomized (adx), and adx rabbits subjected to one of several corticosteroid treatment protocols. The results indicate that Na-K ATPase activity in the CCT decreased by 86% subsequent to adrenalectomy. Injection of physiological doses of aldosterone (10 micrograms/kg) but not dexamethasone (100 micrograms/kg) restored CCT Na-K ATPase activity in adx rabbits to normal levels within 3 h after injection. An insignificant rise in activity was observed 1.5h after aldosterone treatment. In addition, spirolactone SC 26304, a specific mineralocorticoid antagonist, blocked the action of aldosterone on Na-K ATPase.. Therefore an acute increase in Na-K ATPase activity participates in the action of aldosterone on Na transport in this segment. To differentiate between primary vs. secondary activation of this enzyme, adx animals were treated with amiloride before the injection of aldosterone with the intent of blocking luminal membrane Na entry into CCT. In these animals, pretreatment with amiloride blocked the increase in CCT Na-K ATPase act activity seen with aldosterone alone at 3 h. Thus the increase in activity with aldosterone appears to be a secondary adaptation that is dependent on an aldosterone-enhanced increase in the passive entry of Na across the luminal membrane. The subcellular mechanism by which Na modulates Na-K ATPase activity remains obscure.

The purpose of this study was to determine and compare pH, PCO2, and fractional bicarbonate delivery in both superficial and juxtamedullary nephrons by microelectrode techniques and microcalorimetry in the rat in vivo in order to define more clearly the role of deeper nephron segments in urinary acidification. Values for pH and total CO2 concentration ([tCO2]) at the bend of Henle's loop (LOH) (7.39 +/- 0.04 units and 20.5 +/- 1.5 mM) were significantly greater and the PCO2 was significantly less (36.6 +/- 1.5 mmHg) than values for these same parameters in the superficial late proximal tubule (LPT) (6.78 +/- 0.03 units, 8.1 +/- 1.2 mM, and 63.2 +/- 1.0 mmHg, P less than 0.001). The fraction of filtered bicarbonate delivered to the LPT and LOH did not differ, however (12.2 +/- 2.5 vs. 9.0 +/- 0.8%). The pH and PCO2 values in the late distal tubule (6.59 +/- 0.04 units and 64.0 +/- 1.3 mmHg) were significantly greater than at the base (6.24 +/- 0.07 units and 34.5 +/- 1.5 mmHg) and tip (6.12 +/- 0.03 units and 35.2 +/- 1.2 mmHg) of the papillary collecting duct. The [tCO2] in the LOH and an adjacent vasa recta was compared and did not differ significantly (20.5 +/- 1.5 vs. 21.2 +/- 1.3 mM, P greater than 0.05). In summary, we have demonstrated significant alkalinization of tubule fluid in the deep LOH as a result of water abstraction and CO2 diffusion from the nephron. Our results suggest that a spontaneous disequilibrium pH may not exist in the LOH. Furthermore, similar values for [tCO2] in vasa recta and the LOH suggest that passive HCO-3 reabsorption in the thin ascending limb of Henle would be unlikely and does not contribute to the "loop" component of bicarbonate reabsorption.

In a recent micropuncture study electrodes with relatively large tips (3 to 5 mu O.D.) and, hence, low tip resistances were used to measure the transepithelial potential difference (PD) across the proximal tubule of the rat kidney. The present study reexamines the PD of the distal tubule of the rat kidney using such electrodes. In contrast to previous studies where a negative PD has been uniformly found in the distal tubule, the transtubular PD was found to be positively oriented (+3.7 mv) when particular efforts were made to puncture the earliest accessible segments. In accord with previous observations, the PD of the late segment was consistently negative (mean, -19.6 mv). Morphologic examination of the epithelium at the site of puncture suggests that in the very early distal tubule where positive potentials are recorded, the epithelium is characteristic of the distal convoluted tubule. By contrast, in the latter part of the distal tubule, where negative potentials are recorded, the epithelium displays the morphologic characteristics of the cortical collecting duct. The results of these studies suggest that the net transport properties of the distal tubule, that is the region of the nephron beginning just beyond the macula densa and extending to the first junction with another renal tubule, are a composite of activities of at least two types of epithelium.

Studies utilizing in vitro microperfusion were designed to examine whether urea is actively or passively transported across superficial and juxtamedullary straight segments of rabbit proximal tubules. With perfusate and bath solutions containing 1 mM urea and electrolytes similar to normal plasma, the efflux (lumen-to-bath) isotopic permeability (X 10(-5) cm s-1) of superficial segments was 1.37 +/- 0.16 and of juxtamedullary segments was 2.14 +/- 0.20. In the same tubules, the influx (bath-to-lumen) isotopic permeability was 3.70 +/- 0.35 in superficial segments and 4.75 +/- 0.37 in juxtamedullary segments. Despite net water movement in the opposite direction (0.5 nl mm-1 min-1), the influx rate was significantly higher than the efflux rate of urea in both groups. With a low perfusion rate (2 nl/min) and equivalent specific activities of [14C]urea in bath and perfusate, the collected-to-perfused ratio of [14C]urea, corrected for volume marker change, was 1.07 +/- 0.01 in superficial and 1.09 +/- 0.01 in juxtamedullary nephrons, thus indicating net secretion in both segments. In separate studies urea influx was inhibited by hypothermia (decrease from 37 degrees to 28 degrees C), by phloretin (0.1 mM in bath), by cyanide (1 mM), but not by probenecid (0.2 mM). In each case the inhibition was highly significant and reversible. These data suggest that urea is actively secreted by the straight segments of both the superficial and juxtamedullary proximal tubules. These segments may, therefore, contribute significantly to the high urea concentration found at the bend of Henle's loop by micropuncture.

Luminal pH in the proximal convoluted tubule may alter phosphate reabsorption in a variety of acid-base disturbances and in normal conditions as luminal pH decreases along the length of the proximal tubule. These studies address the influence of luminal pH on phosphate reabsorption in isolated perfused proximal convoluted tubules from normal rabbits. Initial perfusates were either pH 6.2 or 7.4, and the bath pH was 7.4. All solutions contained 10 mM total phosphate. The first experiments used HCO3-/CO2-free solutions to examine the effect of luminal pH independent of changes in [HCO3-] or Pco2. Lumen-to-bath phosphate flux increased from 0.63 +/- 0.23 with alkaline perfusate to 2.04 +/- 0.35 pmol X mm-1 X min-1 with acid perfusate. In a separate group of tubules, acetazolamide had no qualitative effect on this result. With HCO3-/CO2-containing solutions, phosphate reabsorption increased from 4.53 +/- 1.46 with alkaline perfusate to 9.67 +/- 1.77 pmol X mm-1 X min-1 with acid perfusate. Thus, an acid luminal fluid can enhance proximal phosphate reabsorption independent of the presence or absence of HCO3-/CO2. To examine the specificity of this effect, the influence of luminal pH on another solute (glucose), reabsorbed via a Na+-coupled mechanism, was studied. Lumen-to-bath glucose flux increased in the same direction: from 52.78 +/- 4.91 with alkaline perfusate to 57.13 +/- 4.70 pmol X mm-1 X min-1 with acid perfusate. The mechanism of the influence of luminal pH on phosphate and glucose reabsorption is not explained but could be direct or indirect from changes in intracellular pH, Na+ activity, metabolism, or basolateral transport. Since an acid luminal pH is expected to inhibit or decrease Na+-H+ exchange at the luminal membrane, these results are also consistent with a competition for the available Na+ gradient between phosphate and glucose transport and the Na+-H+ exchanger in the proximal tubule.

The possibility that the rabbit collecting tubule is heterogeneous with respect to electrical and morphologic properties was examined in isolated segments. Collecting tubules from rabbits treated with desoxycorticosterone acetate were dissected from various locations within the cortex and outer medulla, and the transepithelial potential was measured. The cortical collecting tubule always displayed a negative potential. In contrast, collecting tubules dissected from the inner stripe of the outer medulla displayed a positive potential. That this change is a function of position within the kidney is supported by the observation that some tubules generated a negative potential at the superficial (cortical) end and a positive potential at the deep (medullary) end of the same tubule. Histologic evaluation of tubules after perfusion resulted in a striking correlation between the existence of a negative potential and the presence of dark cells. The results of this study provide evidence that the collecting tubule is heterogeneous with respect to active transport processes and suggest that these functional differences may correlate with morphologic differences between the cortical segment and the medullary segment of the collecting tubule.

Diuretics have a central role in the treatment of edema and hypertension. This function is primarily an induction of a net negative balance of solute and water. Reviewed herein are the transport properties of each nephron segment that governs salt and water reabsorption with specific reference to the mechanisms by which the various diuretic agents affect those transport processes. Under normal circumstances, the proximal tubule reabsorbs about 50 to 66 percent of the filtered fluid by both active and passive mechanisms. However, diuretics that inhibit proximal reabsorption are "weak" diuretics since distal compensatory mechanisms can overcome their effect. The thin descending limb of Henle is highly permeable to water and relatively impermeable to solutes. Thus, its main physiologic function is to allow osmotic water abstraction. Although diuretics have no direct epithelial effect on this segment, many of the diuretics decrease fluid reabsorption from it by abolishing the papillary osmotic gradient. The decreased water absorption from the descending limb of Henle has a major role in over-all increased diuresis since nephron segments distal to the descending limb are impermeable to water in the absence of vasopressin. The thin ascending limb of Henle is impermeable to water while being highly permeable to sodium and chloride. Diuretics have no direct effect on the thin ascending limb of Henle. The medullary and cortical segments of the thick ascending limb of Henle absorb sodium chloride by active mechanisms as a result of a secondary active chloride transport mechanism that depends on the presence of sodium (co-transport mechanism). This transport mechanism is located on the luminal membrane. Most of the "loop" diuretics effect this process from the luminal side by having a direct inhibitory effect on this co-transport process. The diuretics that have a primary effect on the medullary segment (furosemide, bumetanide, ethacrynic acid) inhibit the concentrating mechanisms, whereas the diuretics that are effective primarily in the cortical segment (thiazides plus the diuretics affecting the medullary segment) inhibit the urinary diluting mechanism. The loop diuretics are physiologically the most potent family of diuretics. The cortical collecting duct segment reabsorbs sodium by active mechanisms. These processes are stimulated by aldosterone. The diuretics that affect these processes are considered weak diuretics, but they do have the metabolic effect of potassium sparing.(ABSTRACT TRUNCATED AT 400 WORDS)

Our previous in vitro studies have disclosed that the thin ascending limb of Henle (tALH) possesses some unique membrane characteristics. In those studies we failed to demonstrated active transport of sodium chloride by the tALH, although it was shown that the isotopic permeability to sodium and chloride was unusually high. However, we did not examine the mechanisms by which the apparent high permeation of sodium chloride occurs. Thus the purpose of the present studies was to elucidate the mechanism of sodium chloride transport across the isolated tALH of the rabbit by conducting four different types of studies: (1) comparison of the observed chloride and sodium flux ratios to those predicted by Ussing's equation under imposed salt concentration gradients; (2) kinetic evaluation of chloride and sodium fluxes; (3) examination of the effect of bromide on the kinetics of chloride transport; and (4) experiments to test for the existence of exchange diffusion of chloride. In the first set of studies the predicted and the theoretical flux ratios of sodium were identical in those experiments in which sodium chloride was added either to the perfusate or to the bath. However, the observed chloride flux ratio, lumen-to-bath/bath-to-lumen, was significantly lower than that predicted from Ussing's equation when 100 mM sodium chloride was added to the bath. In the second set of experiments the apparent isotopic permeability for sodium and for chloride was measured under varying perfusate and bath NaCl concentrations. There was no statistical change in the apparent sodium permeability coefficient when the NaCl concentration was raised by varying increments from 85.5 to 309.5 mM. However, permeation of 36Cl decrease significantly with an increase in Cl from 73.6 to 598.6 mM. These events could be explained by a two component chloride transport process consisting of simple diffusion and a saturable facilitated diffusion process with a Vmax = 3.71 neq mm-1 min-1. In the third set of studies it was shown that bromide inhibits transport of chloride and that the magnitude of inhibition is dependent on chloride concentrations. The fourth set of studies ruled out the existence of exchange diffusion. In conclusion, these studies indicate that sodium transport across tALH is by simple passive diffusion, while chloride transport across tALH involves at least two mechanisms: (1) simple passive diffusion; and (2) a specific membrane interaction process (carrier-mediated) which is competitively inhibited by bromide.

Until recently it has not been possible to compare directly the function of superficial and juxtamedullary nephrons. The present studies, using in vitro microperfusion, were designed to examine whether functional differences exist between proximal convoluted tubule segments of superficial and juxtamedullary nephrons. Electrophysiological studies showed that major differences exist between the relative chloride and sodium permeabilities of these segments. In the 1st mm of the superficial proximal convoluted tubule, the permeability to sodium was greater than that to chloride, whereas in the 2nd mm of the superficial proximal convoluted tubule and all later segments, the permeability to chloride was greater than that to sodium. The juxtamedullary proximal convoluted tubule was found to differ from the superficial proximal convoluted tubule in two respects: first, the relative permeabilities to chloride and sodium did not differ in the various segments of the juxtamedullary proximal convoluted tubule; second, the permeability to sodium was greater than to chloride throughout. When perfused with a solution lacking glucose and amino acids, the superficial and juxtamedullary convolutions exhibited the same transepithelial potential change, a reversible decrease to less than -- 1 mV. It thus appears that in both convolutions there exists electrogenic sodium transport coupled to the transport of these organic solutes. This differs from pars recta of both of these nephrons, which have been shown to exhibit electrogenic sodium transport independent of organic solutes. However, when perfused with a solution lacking glucose and amino acids but also containing high chloride and low bicarbonate concentrations, the superficial convolution developed a significantly more positive potential than the juxtamedullary. This difference reflects greater relative chloride permeability in the superficial proximal convolution. These studies show that intrinsic functional differences exist between proximal convoluted tubules obtained from the superficial and juxtamedullary nephron populations.

Calcium transport was studied in medullary and cortical segments of the thick ascending limb of Henle perfused in vitro. 45Ca was added to the perfusate for measuring lumen-to-bath flux (JlbCa), to the bath for measuring bath-to-lumen flux (JblCa), or to both perfusate and bath for measuring net flux (JnetCa). In the medullary segment JlbCa exceeding JblCa and the efflux:influx coefficient ratio was not different from the value predicted from the observed potential difference (PD). In the cortical segments, however, efflux:influx coefficient ratio was greater than the value predicted from the PD, suggesting that calcium transport in this segment may be active, while it is passive in the medullary segment. Furosemide, which reversibly decreases PD in both cortical and medullary segments, inhibited JlbCa only in the medullary segment. Parathyroid hormone (PTH), on the other hand, had no effect on JnetCa in the medullary segment, but it significantly augmented JnetCa in the cortical segment. These results indicate that calcium transport in the thick ascending limb is heterogeneous. In the medullary segment it is passive, inhibited by furosemide and not influenced by PTH. In the cortical segment, however, calcium transport appears to be active, not inhibited by furosemide and stimulated by PTH.

Using in vitro microperfusion of rabbit nephron segments we measured the effects of osmotically induced water flow on net transport of HCO3 and Cl. Measurements were made in superficial and juxtamedullary proximal convolutions and in superficial pars recta. In addition, measurements were taken in the presence and absence (hypothermia) of active transport. Using osmotic gradients of 25 mM raffinose in superficial and 50 mM in juxtamedullary segments, we observed increases in water flow equal to or greater than the normal rates of volume reabsorption observed in these tubule segments. However, there were no significant changes in HCO3 and Cl flux. This lack of significant solvent drag was seen both when osmotic water flow was in the lumen-to-bath direction and when osmotic flow was in the bath-to-lumen direction. The results of these studies suggest that solvent drag does not contribute significantly to NaCl and NaHCO3 reabsorption in proximal tubules. The lack of significant solvent drag of these salts can be interpreted as indicating either that osmotically induced transepithelial water flow in proximal tubules almost exclusively traverses transcellular pathways or that proximal tubule tight junction reflection coefficients for these salts are close to unity.

Cortical and outer medullary collecting duct segments were dissected from human kidneys and perfused in vitro. The transepithelial potential difference was measured and found to be lumen positive +6.8 +/- 0.6 mV (n= 20). This lumen-positive potential difference was inhibited by ouabain and furosemide but not by acetazolamide. Replacement of chloride in bath and perfusion fluids caused a reversible decrease of the potential difference to near zero. We conclude from these studies: (a) the lumen-positive potential difference is dependent upon the presence of chloride ion suggesting the existence of an active electrogenic chloride reabsorptive process in the human collecting duct and (b) it is possible to examine human renal physiology directly using in vitro microperfusion of tubule segments.

Rabbit medullary collecting duct (MCD) acidification has been demonstrated to occur by means of a sodium-independent, aldosterone-stimulated mechanism. We have examined the anionic dependence of this process by means of the isolated perfused tubule technique. Total replacement of perfusate chloride with gluconate enhanced tubular bicarbonate reabsorption (JHCO3), from a basal rate of 10.7 +/- 1.0 pmol X mm-1 X min-1 to a rate of 15.01 +/- 1.0 pmol X mm-1 X min-1. Removal of bath chloride, with and without removal of perfusate chloride completely abolished acidification. Bath, but not luminal 4-acetamido-4' isothiocyano-2,2'-disulfonic stilbene provoked a marked decrease in JHCO3 from 10.1 +/- 1.2 pmol X mm-1 X min-1 to 2.3 +/- 0.3 pmol X mm-1 X min-1. Measurement of chloride reabsorptive rate (JCl) revealed colinearity between JHCO3 (9.18 +/- 0.9 pmol X mm-1 X min-1) and JCl (9.75 +/- 1.18 pmol X mm-1 X min-1). We propose a model of mammalian distal nephron acidification in which (a) cellular base exit is effected by means of a basolateral membrane Cl-base exchanger and (b) net electroneutrality of electrogenic proton secretion is maintained by the parallel movement of an anionic species, functionally chloride.

Addisonian patients can maintain potassium homeostasis despite the absence of mineralocorticoid. The present in vitro microperfusion studies examine what role the cortical collecting tubule might play in this process. All studies were performed on tubules harvested from adrenalectomized rabbits, which were maintained on 0.15 M NaCl drinking water and dexamethasone 50 mug/d. Perfusion and bath solutions were symmetrical Ringer's bicarbonate with [K] of 5 meq/liter. Initial studies on cortical collecting tubules from adrenalectomized animals ingesting a high potassium chow (9 meq K/kg body wt) demonstrated net potassium secretion against an electrochemical gradient (mean collected fluid [K] 16.5+/-2.6 meq/liter with an observed transepithelial voltage of -6.3+/-4.1 mV; predicted voltage for passive distribution of potassium being -28.2 mV). To examine whether this active potassium secretion could be modulated by dietary potassium, independent of mineralocorticoid, two diets identical in all respects except for potassium content were formulated. Potassium secretion was compared in cortical collecting tubules harvested from adrenalectomized animals on low (0.1 meq K) and high (10 meq K) potassium intake. Mean net potassium secretion by cortical collecting tubules was 2.02+/-0.54 peq mm(-1) min(-1) in the low potassium diet group and 5.34+/-.74 peq.mm(-1).min(-1) in the high potassium group. The mean transepithelial voltages of the collecting tubules did not differ between the two dietary groups. While net Na reabsorption was significantly greater in tubules from the high K group, this could not account for the differences in K secretion. These data demonstrate that: (a) the cortical collecting tubule can actively secrete potassium and that the magnitude of this potassium secretion correlates with potassium intake; (b) this active potassium secretory process in independent of mineralocorticoid. These findings support the hypothesis that the cortical collecting tubule may contribute to K homeostasis in Addison's disease.

The drainage and instillation of poorly absorbable corticosteroids has recently been suggested as an laternative to the present modes of therapy for uremic pericarditis. One patient who underwent such a therapeutic approach subsequently had a left internal mammary artery to right internal mammary vein arteriovenous fistula develop. To our knowledge, this is the first report of the development of arteriovenous fistula after either pericardiocentesis or intrapericardial instillation of steroids.

The effect of tubular obstruction on renal function has been understood poorly at the tubular level and from the clinical standpoint. In our review the evidence for a direct influence of hydrostatic pressure on tubular transport and glomerular filtration is examined. The data generated to date indicate a direct influence of hydrostatic pressure on tubular transport only at the level of the distal convoluted tubule and collecting duct. With respect to glomerular filtration increased tubular pressure reduces the net driving force for filtration and reduces glomerular filtration rate in the absence of a compensatory increase in glomerular hydrostatic pressure. We next review physiological data concerning the mechanism of post-obstructive diuresis. Available information suggests 4 factors that play a significant role in the clinical syndrome of post-obstructive diuresis: 1) medullo-papillary washout, 2) decreased fractional and absolute salt and water reabsorption in the collecting duct, presumably secondary to direct influence of hydrostatic pressure on transport mechanisms, 3) osmotic diuresis secondary to retention of urea and other osmotic solutes during the period of obstruction and 4) prior salt and water administration in the absence of excretion, resulting in extracellular fluid volume expansion.

Rabbit medullary collecting duct (MCD) from inner stripe of outer medulla has been identified as a major distal nephron acidification site. The isolated, perfused tubule technique was used to examine the roles of mineralocorticoid and glucocorticoid in regulation of MCD acidification. Surgical adrenalectomy reduced bicarbonate reabsorptive rate (JHCO3, pmol X mm-1 X min-1) from the normal of 9.79 +/- 1.21 to 0.67 +/- 1.1. Chronic administration of deoxycorticosterone acetate (DOCA) increased JHCO3 of MCD significantly to 18.02 +/- 1.62 whereas chronic dexamethasone administration did not affect JHCO3. The direct effects of aldosterone and dexamethasone upon MCD acidification were examined by perfusing tubules harvested from adrenalectomized rabbits in the presence of aldosterone or dexamethasone. Aldosterone, at 5 X 10(-8) M, increased JHCO3 significantly from 1.27 +/- 0.28 to 3.09 +/- 0.34. At 10(-6) M, aldosterone produced a greater increase in JHCO3 from 0.67 +/- 1.1 to 9.39 +/- 1.59. In vitro dexamethasone treatment had no effect on JHCO3. Studies examining the sodium dependence of aldosterone-stimulated acidification demonstrated that JHCO3 in tubules harvested from normal and deoxycorticosterone acetate-treated animals was unaffected by total replacement of sodium with tetramethylammonium. Likewise, luminal amiloride (5 X 10(-5) M) had no effect on JHCO3 in tubules harvested from adrenalectomized and normal animals. Moreover, the acute, in vitro stimulatory effect of aldosterone was seen to occur in the presence of luminal amiloride. These studies define a mammalian distal nephron segment that possesses major acidifying capacity, which is modulated by mineralocorticoid but independent of luminal sodium.

This study was designed to examine whether prostaglandin E2 can directly affect sodium transport across isolated perfused rabbit renal collecting tubules. Changes in transepithelial potential and isotopic sodium fluxes in response to peritubular prostaglandin E2 were measured. In addition, changes in transepithelial potential of the outer medullary collecting tubule in response to prostaglandin E2 were also measured. With few exceptions, all rabbits received 5 mg/day desoxycorticosterone acetate for 4-11 days before experimentation. The results of the experiments show that: (a) prostaglandin E2 inhibits the negative transepithelial potential in the cortical collecting tubule as well as the outer medullary collecting tubule; (b) prostaglandin E2 inhibits net sodium transport out of the lumen by inhibiting efflux while backflux is unaffected; (c) prostaglandin E2 produces this inhibition within 15 min, and the effects are dose dependent and reversible. These results suggest that prostaglandin E2 may modulate sodium transport in vivo and may contribute to the final regulation of sodium excretion.

Numerous previous studies have proposed a role for angiotensin II (AII) in the renal regulation of salt balance. At least one nephron site, the proximal convoluted segment, has been implicated in this role. We used in vitro microperfusion of rabbit proximal convoluted tubules to further examine this question. To insure use of appropriate in vivo concentrations as well as potency of the hormone in vitro, we measured plasma AII levels by radioimmunoassay in normal, sodium-depleted, and adrenalectomized rabbits, and measured AII activity by bioassay after incubation in various microperfusion baths. Plasma levels ranged from approximately 2 X 10(-11) to 5 X 10(-11) M. AII activity was stable in Ringer's solution plus albumin, but not in rabbit serum or Ringer's solution plus fetal calf serum. In Ringer's solution plus albumin, physiologic concentrations of AII stimulated volume reabsorption (Jv). 10(-11) M AII increased Jv by 16% (P less than 0.01). 10(-10) M AII produced a lesser increase, 7.5% (P less than 0.05). At a frequently studied, but probably pharmacologic dose, 10(-7) M AII inhibited Jv by 24% (P less than 0.001). AII at 10(-11) M did not stimulate Jv in the presence of 10(-7) M saralasin. Though previous studies have suggested agonistic effects of saralasin alone in epithelia, we found no significant effect of 10(-7) M saralasin on Jv. None of the AII doses measurably changed transepithelial voltage. We conclude that AII in physiologic doses directly stimulates Jv in proximal convoluted tubules and this effect is probably receptor mediated and, within the limits of detection, electroneutral.

It has been previously demonstrated that superficial (SF) versus juxtamedullary (JM) proximal convoluted tubules (PCT) of rabbit have different intrinsic transport characteristics: Na is less permeable than Cl in the latter portions of the SFPCT, while Cl is less permeable than Na throughout the JMPCT. These permeability differences have major influences on the mechanism of salt reabsorption across the proximal tubules. However, both populations of PCT have the same embryological origin. Studies therefore were designed to examine whether two distinct populations of PCT exist at birth or whether a second population of tubules develops with delivery as a result of some unidentified acute change in humoral factor affecting epithelial transport properties. Both morphological and electrophysiological studies were conducted on PCTs from rabbits within 36 h of birth. Both transmission and scanning electron microscopy studies clearly disclosed that SFPCT are less mature than the JMPCT. Also the SFPCT had a lower Na permeability than Cl (0.55 +/- 0.06) while the JMPCT had a higher Na permeability than Cl (1.37 +/- 0.11). Thus these studies demonstrate that intrinsic heterogeneity of PCT is present at birth. Since the SF Na to Cl permeability approximates that of free diffusion these studies suggest that epithelial discrimination in PCT is part of a maturation process.

Several hormones induce phosphatidylinositol turnover in cell membranes and thus activate protein kinase C. Activation of protein kinase C can, in turn, have effects on epithelial transport. These experiments were designed to investigate the effects of two activators of protein kinase C, phorbol 12-myristate,13-acetate (PMA) and L-alpha-1,2-dioctanoylglycerol (L-alpha-1,2-DOG), and two inactive analogues, 4 alpha-phorbol and 4-O-methyl phorbol 12-myristate,13-acetate, on sodium, potassium, chloride, and total CO2 transport in the rabbit cortical collecting tubule. Utilizing in vitro microperfusion techniques, we found that activation of protein kinase C with either PMA or L-alpha-1,2-DOG significantly inhibited net sodium absorption, net potassium secretion and transepithelial voltage in a dose-dependent manner. There was no effect on net chloride or total CO2 transport. In contrast, the inactive phorbol analogues did not alter either sodium or potassium transport. These studies demonstrate that in the rabbit cortical collecting tubule sodium and potassium transport can be inhibited by compounds known to activate proteins kinase C. Thus, hormones that induce phosphatidylinositol turnover in the rabbit cortical collecting tubule may lead to inhibition of sodium transport by activation of protein kinase C.

Although intrarenal infusions of kinins produce diuresis, it is not clear to what extent this response is due to hemodynamically mediated medullary washout and/or to direct epithelial effects of kinins. Recent evidence has shown that bradykinin binds to collecting tubules in vitro. We therefore examined the interactions of lysyl-bradykinin and antidiuretic hormone (ADH) with respect to hydraulic conductivity (Lp) in the rabbit cortical collecting tubule perfused in vitro. To ensure adequate substrate for prostaglandin synthesis, the bath contained 2.5 microM arachidonic acid. Arachidonic acid produced no change in base-line Lp and had no effect on the subsequent response to a supramaximal dose of ADH (100 microU/ml). Therefore, all subsequent experiments were done in the presence of arachidonic acid. Lysyl-bradykinin (10(-9)M) added to either the lumen or bath had no effect on base-line Lp. Collecting tubules which were exposed for 1 h to bath lysyl-bradykinin (10(-9)M) had a significantly diminished subsequent Lp in response to ADH (P less than 0.02). In tubules exposed to bath lysyl-bradykinin plus indomethacin (5 microM), the subsequent ADH response was normal. Lysyl-bradykinin (10(-9)M) added to the lumen had no effect on subsequent ADH response. We conclude that lysyl-bradykinin from the basolateral side inhibits the hydroosmotic response of the cortical collecting tubule to ADH, and that this inhibition is probably prostaglandin-mediated. Lysyl-bradykinin does not affect water flow from the luminal surface. These data indicate that the diuresis seen with kinin infusions may result, at least in part, from a direct epithelial effect. They also suggest a role of the renal kallikrein-kinin system in modulating water transport in vivo.

31P-nuclear magnetic resonance (NMR) studies were conducted on split epithelial sheets of frog skins to examine the effects of hypoxia and respiratory pH variations on various phosphate-containing intracellular substrates. Frog skins were split into epithelial sheets from which the supporting tissue was removed. Epithelial sheets in either phosphate-free Cl--Ringer or phosphate-free SO2-4-Ringer were bubbled at room temperature with 100% N2, 100% O2, 2% CO2-98% O2, 5% CO2-95% O2, and 15% CO2-85% O2. The results show that the intracellular pH (pHi) with Cl- -Ringer was 7.19 and with SO2-4-Ringer was 7.42 with extracellular pH of 7.52 when bubbled with 100% O2. These pHiS indicate that H+ concentration is at least an order of magnitude less than predicted from the previously measured Nernst potential. With exposure to increasing extracellular PCO2, there is a polynomial decrease in pHi. The pHi tended to be more alkaline in SO2-4 -Ringer, suggesting the presence of a HCO-3/Cl- exchange mechanism. The ATP concentration is critically and reversibly dependent on PO2. ADP concentrations were consistently low in well-oxygenated conditions. Variable but small quantities of phosphocreatine were detected. These studies demonstrate further the potential importance in utilizing NMR spectroscopy to examine coupling of biochemical substrates to epithelial transport processes.

Electrical and permeability features of the distal convoluted tubule (DCT) and the cortical collecting tubule (CCT) were examined using the technique in which isolated segments of rabbit tubules were perfused in vitro. When rabbits were given a regular diet and tubules were perfused and bathed in artificial solutions simulating plasma ultrafiltrate, the potential difference (PD) was +3.7 plus or minus 1.9 mV in the CCT and -40.4 plus or minus 2.8 mV in the DCT. When rabbits were given a low sodium, high potassium diet plus i.m. deoxycorticosterone acetate (DOCA) (1 mg/kg per day), the PD in both the CCT (-30.8 plus or minus 3.9 mV) and the DCT (-33.8 plus or minus 5.5 mV) was negative. The PD in the CCT was quantitatively similar to that of diet plus DOCA when animals were given DOCA alone. The PD in both segments was inhibited by ouabain (10-minus 5 M) in the bath or by amiloride (10-minus 5 M) in the perfusate. Addition of vasopressin (200 muU/ml) to the bath caused a gradual decline of PD to zero in the CCT but failed to produce a potential response in the DCT. Osmotic water permeability was essentially zero in both segments in the absence of vasopressin. After addition of the vasopressin to the bath, osmotic water permeability in the DCT remained zero but increased to 71.9 plus or minus 25.5 X 10-minus 7 cm/s per atm in the CCT. We conclude that both segments are similar in that each possesses an electrogenic transport process but that these segments differ in that: (a) the CCT requires either exogenous or endogenous mineralocorticoid to maintain a maximal negative PD, whereas the PD in the DCT appears to be independent of mineralocorticoid effect; and (b) the CCT responds to vasopressin with a marked rise in water permeability, whereas the DCT is impermeable to water before and after addition of vasopressin.

We have previously shown that the transtubular potential of the rabbit cortical collecting tubule varies in concert with changes in plasma mineralocorticoid levels, while the potential of the distal convoluted tubule is invariant with such changes. In the present studies we have examined the effects of in vitro addition of d-aldosterone to isolated tubules, as well as the effects of triamterene and spirolactone. d-Aldosterone (0.2 mum added to the perfusate or 1 muM added to the bathing medium) resulted in a marked stimulation of the transtubular potential difference (lumen-negative) after a short latent period. d-Aldosterone had no effect on the potential difference of distal convoluted tubules of intact or adrenalectomized rabbits. Both the magnitude of the response and the length of the latent period in the cortical collecting tubule after aldosterone were markedly temperature-dependent. Triamterene caused a gradual but reversible inhibition of the potential difference in the cortical collecting tubule but had no effect in the distal tubule. Spirolactone, when added before aldosterone, blocked the electrical response to the hormone in the cortical collecting tubule, and produced a gradual inhibition of the potential difference in mineralocorticoid-stimulated tubules. Spirolactone had no effect on the potential difference of the distal tubule. We conclude that (a) the influence of aldosterone on the potential across the distal nephron is restricted to the distal convoluted tubule, (b) the electrical response to aldosterone and the latent period are temperature-dependent, (c) the response to aldosterone is blocked by spirolactone, and (d) triamterene inhibits the potential difference only in the cortical collecting tubule.