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Objectives: This report presents an overview of preclinical data available on ferumoxtran-10, an ultrasmall superparamagnetic iron oxide nanoparticular contrast agent proposed for lymph node magnetic resonance imaging., Materials and Methods: Pharmacokinetic, safety pharmacology, single- and repeat-dose toxicity, reproduction toxicity, and genotoxicity studies were performed with ferumoxtran-10 given intravenously (bolus injection) in mice, rats, rabbits, dogs, and monkeys., Results: Ferumoxtran-10 was taken up by macrophages, mostly in liver, spleen, and lymph nodes, within 24 hours after bolus injection and underwent progressive metabolism. Toxicity was observed only at very high exposure levels and mainly was linked to a massive iron load after repeated injections. Ferumoxtran-10 was not mutagenic but was teratogenic in rats and rabbits., Discussion: The preclinical pharmacokinetic and safety profile of ferumoxtran-10 appears to be satisfactory in view of its proposed use as a single-dose diagnostic agent in human for MR imaging of lymph nodes.

The ultrasmall superparamagnetic iron oxide colloid BMS 180549 can be found lymph nodes by either subcutaneous (SC) or intravenous (IV) injection. With an SC injection in the front extremities, the axillary and brachial nodes attain the highest accumulations of the agent. With an SC injection in the rear extremities, the popliteal, iliac, and axillary nodes attain highest accumulations of the agent. With IV injection of the agent, the iliac, mediastinal and mesenteric nodes attain highest accumulations of the agent. Though the spleen is not involved with the drainage of the interstitial space near the site of SC injections, the mobility of BMS 180549 from such injection sites increases splenic relaxation rates. Based on a knowledge of the lymphatic system, a route of administration of BMS 180549 can be chosen to maximize the delivery of the agent to specific lymph nodes.

The distal inner medullary collecting duct (IMCD) is critical in the urinary concentrating process, in part because it is the site of vasopressin (AVP)-regulated permeability to urea. The purpose of these experiments was to develop a cell culture model of the IMCD on permeable structure and to characterize the responsiveness to AVP. Rat IMCD cells were grown to confluence on collagen-coated Millipore filters glued onto plastic rings. To assess the time required to achieve confluence, the transepithelial resistance was measured periodically and was found to be stable after 2 weeks, at a maximal value of 595 +/- 22 omega cm2. In separate monolayers the effect of AVP on inulin and urea permeability was determined. While inulin permeability was unchanged after AVP, urea permeability increased from 6.0 +/- 0.4 to peak values of 16.0 +/- 3.8 (10 nM), 23.1 +/- 3.9 (1 microM) and 28.1 +/- 4.9 (10 microM) x 10(-6) cm s-1 (n = 24). In 10 other monolayers, after the addition of 1 mM 8-Br-cAMP, urea permeability increased from 5.1 +/- 0.3 to 8.1 +/- 1.6 x 10(-6) cm s-1 and, after 8-Br-cAMP + 3-isobutyl-1-methylxanthine, to 12.2 +/- 0.7 x 10(-6) cm s-1. We conclude that rat IMCD cells grown in culture exhibit the characteristics of a 'tight' epithelium. Inulin and urea permeability are not different in the absence of AVP, consistent with high resistance junctional complexes. Furthermore, IMCD cells retain the capacity for AVP-regulated urea permeability, a characteristic feature of this nephron segment in vivo.

An ultrasmall superparamagnetic iron oxide (USPIO) preparation was developed that is small enough to migrate across the capillary wall, a prerequisite in the design of targetable particulate pharmaceuticals. Seventy percent of particles were smaller than 10 nm; 26%, smaller than 5 nm. The blood half-life of USPIO in rats was 81 minutes, considerably longer than that of larger superparamagnetic iron oxide preparations such as AMI-25 (6 minutes). Electron microscopy demonstrated that USPIO particles transmigrate the capillary wall by means of vesicular transport and through interendothelial junctions. Twenty-four hours after intravenous administration, 3.6% of the injected dose per gram of tissue was found in lymph nodes, 2.9% per gram in bone marrow, 6.3% per gram in liver, and 7.1% per gram in spleen. The major potential applications for USPIO are as (a) an intravenous contrast agent for the lymphatic system, (b) a bone marrow contrast agent, (c) a long-half-life perfusion agent for brain and heart, and (d) the magnetic moiety in organ-targeted superparamagnetic contrast agents for magnetic resonance imaging.

The purpose of this study was to quantitate sodium balance during gestation in the rat (3 wk) to determine the time course of salt retention and to define the distribution of the retained salt. We have found that while sodium retention appears to begin during the 2nd wk, the bulk of retention occurs during the 3rd wk when the rat excretes only 67% of that ingested. Compared to that of nonpregnant, control rats, net retention for all of gestation was 9.8 meq. Despite a significantly higher sodium intake in pregnant rats, neither urinary nor fecal sodium excretion was different in pregnant and control rats. Fifty-nine percent of the retained sodium was found in the products of conception. Inulin space was increased by term from 19.7 to 25.4% of nonconceptus body weight. In pregnant rats a net retention of 6.3 meq of potassium was noted, 55% of which was found in the products of conception. We conclude that the rat retains sodium and potassium during pregnancy, predominantly in the third trimester. Over half of these salts are found within the conceptus products.

The effect of the absence of parathyroid hormone on nephron acidification was determined in rats after acute thyroparathyroidectomy (TPTX). Tubular fluid samples were obtained from the superficial late proximal tubule (LPT), the early distal tubule ( EDT ), and along the inner medullary collecting duct (IMCD), and the results were compared with those obtained from control rats. In the LPT after TPTX, pH was lower, 6.66 +/- 0.01 vs. 6.73 +/- 0.01, and ammonium and net acid delivery were increased significantly. In the EDT no differences in pH, bicarbonate, or net acid were found between groups, whereas ammonium and acid phosphate were significantly different. Along the IMCD in control rats, pH decreased from 6.58 to 5.21 and the addition of about 430 nmol/min of net acid was observed. After TPTX more net acid entered the duct and pH was lower, 5.66, but did not change; neither did the amount of bicarbonate, ammonium, acid phosphate, or net acid change significantly along the duct. Net acid excretion was not different, however, among groups. These results demonstrate that TPTX markedly affects nephron acidification, increasing net acid along the proximal tubule. In contrast to that in control rats, however, net acidification is completed prior to the IMCD. We conclude that the acute absence of parathyroid hormone may significantly affect local nephron acidification but does not alter acid excretion.

The effect of volume expansion on inner medullary collecting duct (IMCD) sodium transport remains controversial. Studies employing micropuncture of the IMCD base and tip were interpreted to demonstrate enhanced sodium and chloride reabsorption. Data obtained by microcatheterization evaluating only sodium transport revealed either no reabsorption or net addition. We have examined both sodium and chloride transport by microcatheterization. Volume expansion was comparable to the micropuncture studies: 0.9% saline equal to 10% body wt and then matched to urine flow. The fraction of filtered fluid, sodium, and chloride was analyzed as a function of IMCD length. In eight hydropenic rats 60% of the fluid, 71% of the sodium, and 48% of the chloride delivered to the IMCD was reabsorbed. In six volume-expanded rats no significant net reabsorption of fluid, sodium, or chloride was found. Accordingly, in contrast to the micropuncture results, we have demonstrated that net sodium chloride and fluid reabsorption are absent during volume expansion. We conclude that during volume expansion, fluid, sodium, and chloride excretion increase, in part, because of a reduction in net reabsorption along the IMCD. The degree of volume expansion does not account for the discrepancy between the two techniques.

These experiments were designed to compare the natriuretic ability of old(age 22-24 months) and young (4-6 months) rats after volume expansion. No difference in extracellular fluid volume was noted as estimated by inulin space; old 18.8 +/- 0.6% and young 18.2 +/- 0.7% of body weight. Standard clearance techniques were utilized in unanesthetized animals. The fraction of infused sodium excreted during and after expansion with isotonic saline equal to 7% BW was statistically lower in the old group 53 +/- 2 vs. 68 +/- 3% (p less than 0.01). Similar measurements were made during the infusion of whole blood equal to 2.3% BW. Again the old rats excreted a significantly lower fraction of the infused Na, 55 +/- 10 vs. young 112 +/- 12%. These differences do not appear to be explained by changes in glomerular filtration rate, blood pressure, hematocrit or serum protein concentration. We conclude that aged rats have an impaired ability to excrete sodium with volume expansion but the mechanism for this defect is yet to be determined.

Excess mineralocorticoid activity is thought to be responsible for the increased sodium reabsorption found after adrenal enucleation, but no known mineralocorticoid has been demonstrated in quantities sufficient to account for this antinatriuresis. 19-Hydroxydeoxycorticosterone (19-OH-DOC) has been synthesized by the incubated enucleate adrenal capsule and 19-nordeoxycorticosterone (19-nor-DOC), a possible metabolite, has been found in the urine of rats with regenerating adrenal glands. To evaluate the in vivo mineralocorticoid potency of these steroids, we studied glucocorticoid-replete adrenalectomized rats and measured the sodium and potassium excretion after administration of these steroids. Our results indicate that 19-nor-DOC has equipotent antinatriuretic activity compared to aldosterone but was less kaluretic. 19-OH-DOC had no significant antinatriuretic or kaluretic activity. We conclude that 19-nor-DOC is a potent mineralocorticoid and may be responsible for the enhanced sodium reabsorption found after adrenal enucleation.

Adrenal enucleation (removal of the adrenal gland, leaving the capsule intact) results in regeneration of the adrenal cortex. During the first 1-2 wk of adrenal regeneration, marked renal sodium avidity and positive sodium balance are noted. This renal sodium avidity appears mediated via adrenocorticotropin-stimulated secretion of a potent mineralocorticoid by the regenerating adrenal cortex. In this review, we have examined relationships between the histology and ultrastructure of the regenerating adrenal cortex, renal sodium handling, and adrenal steroid production at various times after the initiation of adrenal regeneration. Plasma levels of known mineralocorticoids are subnormal during the period of most intense sodium avidity, while urinary excretion of a potent mineralocorticoid, 19-nordeoxycorticosterone, has been found to be increased in rats with regenerating adrenals during this period of most intense sodium avidity. This hormone, however, is not elevated in rats with regenerating adrenals after resolution of the period of sodium avidity. In this article, we review the experimental evidence regarding the potency of this mineralocorticoid and its likely role in the sodium retention after adrenal enucleation.

Young (3 to 4 months) and old (21 to 22 months) rats were fed either a regular or high potassium (K) diet. After acute potassium chloride infusion, the fraction of infused K excreted (K efficiency) was similar in rats on a normal diet (57 +/- 3%, young, vs. 61 +/- 2%, old). With high K feeding there was a significant increase in the young, 69 +/- 4%, but not in the old rats, 62 +/- 2%. Na-K ATPase activity was markedly reduced in the renal medulla of old rats on a regular or high K diet. In addition, the response to acute K loading was compared in acutely nephrectomized rats. In the young rats on a regular diet plasma K increased from 3.72 +/- 0.09 to 5.28 +/- 0.16 mEq/liter while with K ingestion the increase was significantly less, 3.62 +/- 0.07 to 4.75 +/- 0.12 mEq/liter. In the old rats plasma K increased similarly on a regular or high K diet, 3.68 +/- 0.10 to 5.68 +/- 0.33 mEq/liter and 3.76 +/- 0.06 to 5.97 +/- 0.30 mEq/liter, respectively. Thus, old rats have impaired renal and extrarenal adaptation, but they have a normal response to an acute K challenge. A reduction in Na-K ATPase may account for the defect in renal adaptation in the aged rats.

The effect of acute respiratory acidosis (ARA) on inner medullary collecting duct (IMCD) acidification was studied and the results were compared with previously obtained data by our laboratory in rats with acute metabolic acidosis (AMA). We employed the microcatheterization technique to directly measure pH and PCO2 with glass-membrane electrodes, and fluid samples were obtained for measurement of bicarbonate, phosphate, and ammonium. Arterial pH was 7.18 +/- 0.01 and PCO2 was 88 +/- 2 mmHg. The IMCD data were analyzed as a function of IMCD length (approximately 6 mm). pH decreased from 5.78 +/- 0.07 to 5.27 +/- 0.03 and PCO2 increased from 55 +/- 4 to 75 +/- 2 mmHg between origin and tip. Bicarbonate delivery decreased from 154 +/- 34 to 25 +/- 3 nmol/min but no change was noted in acid phosphate, ammonium, or net acid addition along the IMCD. However, net acid excretion was not different from that found previously in AMA. We conclude that during ARA acidification is augmented prior to, but not along, the IMCD. In contrast, during AMA we previously found that IMCD plays a major regulatory role in urinary acidification, accounting for about 50% of the excreted hydrogen ion.

Aldosterone and dexamethasone stimulate sodium absorption in the rat colon in vivo. In vitro, increased amiloride inhibitable short-circuit current (ISC) has been demonstrated following aldosterone or dexamethasone treatment. Since ISC bears no relationship to sodium flux (JNa) in the untreated rat colon, we measured JNa in partially stripped voltage clamped segments of rat distal colon. Our results demonstrate directly that continuous infusion of aldosterone or dexamethasone for 4-7 days stimulated amiloride inhibitable JNa by stimulating JNaM -S. The amiloride inhibitable portion of JNaM -S was highly correlated with and approximately equal to the amiloride inhibitable ISC. Amiloride had no effect in controls. We conclude that JNaM -S in the rat distal colon is only sensitive to mucosal amiloride after treatment with aldosterone or dexamethasone. The amiloride sensitive ISC in these treated tissues was a good measure of the amiloride sensitive JNa. Small differences between aldosterone and dexamethasone treatment were noted in the effect on transepithelial resistance, potential difference, and the ISC after amiloride.

19-Nor-corticosteroids are potentially important mineralocorticoids and hypertensive agents. We tested the mineralocorticoid potency of 19-nor-progesterone (19-NOR-P) and 19-nor-corticosterone (19-NOR-B) compared with aldosterone using the toad bladder short-circuit current as a measure of sodium transport. 19-NOR-B (10(-7) M) increased sodium transport to a degree not different from that caused by aldosterone (10(-7) M). The onset of action and duration of activity also were not different from those of aldosterone. 19-NOR-P (10(-7) M), however, had no effect on sodium transport. We conclude that 19-NOR-B has significant mineralocorticoid activity, while under the conditions of these studies, 19-NOR-P exhibited no effect on sodium transport.

The purpose of this study was to define individual nephron function and salt and fluid reabsorption in the term pregnant rat utilizing standard micropuncture techniques. The studies were performed at the time of maximal sodium retention and when extracellular fluid volume was significantly increased. During micropuncture in 7 pregnant and 9 non-pregnant rats, we found no difference, respectively, in GFR 1.04 +/- 0.06 vs. 1.13 +/- 0.06 ml/min/kidney or nephron GFR 35 +/- 0.9 vs. 33 +/- 1 ml/min. Fractional and absolute Na reabsorption was also not significantly different in the proximal tubule or the loop of Henle. Along the distal tubule, fractional reabsorption was higher in pregnancy, 71 +/- 3 vs. 62 +/- 4 but not statistically different, 0.05 less than p less than 0.10. Beyond the late distal tubule absolute delivery and reabsorption were greater in the nonpregnant rats. Distal nephron potassium secretion was similar in both groups. In anesthetized rats during clearance studies but without surgery for micropuncture, GFR in the pregnant group was significantly higher than the nonpregnant rats, 1.65 +/- 0.06 vs. 1.28 +/- 1.10 ml/min/kidneys as was tubular reabsorption of sodium. We conclude that pregnant rats exhibit glomerular tubular balance and that potassium handling along the superficial nephron is unaffected by pregnancy. The sodium retention of pregnancy appears to occur because of the failure of inhibition of tubular sodium reabsorption, a response which is normally expected when extracellular volume is expanded.

Diuresis and natriuresis follow volume expansion by intravenous infusion of either blood or saline. However, the mechanisms underlying the renal responses seem different for the two types of expansion. It has been suggested that the response to saline expansion is due to changes in physical factors with alterations in the morphology of the lateral intercellular space (LIS). The present study examines the width of the LIS of the proximal tubule after a Ringer-Locke (1%, 2.3%, or 7% body weight) or whole blood (2.3% body weight) infusion and in nonexpanded control. After Ringer-Locke infusion, a significant widening of the LIS of the proximal tubule is seen at all levels of expansion. There is a linear relationship between the level of the Ringer-Locke infusion and the degree of expansion of the LIS. In contrast, after blood volume expansion, the width of the LIS is the same as nonexpanded controls. These studies are consistent with the view that the renal response to saline expansion is mediated at least in part by physical factors affecting the proximal tubule.

The renal concentrating ability of Fischer 344 rats was studied at 23 and 4 mo of age. Maximum urine concentration after 40 h of dehydration with or without vasopressin injection was significantly lower (P less than 0.01) in old (2,550 +/- 70 and 2,363 +/- 107 mosmol/kg H2O2, respectively) vs. young (3,242 +/- 50 and 3,162 +/- 50 mosmol/kg H2O, respectively) rats. Free water reabsorption (TcH2O/GFR) rose progressively as a function of osmolar clearance, and at similar values of distal solute delivery TcH2O was clearly reduced in the old group. Free water formation (CH2O/GFR) rose linearly as a function of urine flow and was not different between old and young rats. Glomerular filtration rate was also not different between age groups under the conditions studied. Nonurea (sodium + potassium + ammonium) x 2 and urea solute concentrations as well as total calculated osmolality in the cortex, outer medulla, or inner medulla were not different between age groups. Because the indices of ascending limb solute delivery and transport and the solute gradient for water reabsorption were similar, we conclude that the concentrating defect in aged rats is most likely secondary to a decrease in water permeability along the collecting duct.

We have measured calcium and magnesium transport along the inner medullary collecting duct (IMCD) in rats by the microcatheterization technique. The fraction of filtered divalent cation ((F/P)Ca,Mg/In) as a function of IMCD length was analyzed by linear regression. In eight control rats calcium reabsorption was demonstrated, inasmuch as there was a significant correlation and slope, P less than 0.001. (F/P)Ca/In decreased from 2.2 +/- 0.3% at the beginning of the IMCD to 0,84 +/- 0.1% in the urine. (F/P)Mg/In was not changed along the IMCD, After acute thyroparathyroidectomy (TPTX) (F/P)Ca/In increased to 4.6 +/- 0.4% at the beginning of the IMCD and 1.55 +/- 0.1% in the urine. The slope was not significantly different from the control group. (F/P)Mg/In was unchanged and was not different from the control group. Accordingly, net Ca reabsorption occurs along the IMCD and the fractional Ca reabsorption is not altered by TPTX. Net Mg transport along the IMCD was absent in both control and TPTX rats; The increased calciuresis after TPTX is not dependent on IMCD function nor is Mg excretion under the conditions studied.

Adrenal enucleation is followed by a period of increased sodium reabsorption thought to be due to excess mineralocorticoid activity. However, it has not been demonstrated that increased production of any known mineralocorticoid accounts for this antinatriuresis. Recently, 19-hydroxydeoxycorticosterone (19-OH-DOC) was found in incubates of regenerating adrenal capsules 3-4 days postenucleation and 19-nordeoxycorticosterone (19-nor-DOC) was identified in the urine of rats with regenerating adrenals. Because it was possible that these hormones might play a role in the sodium retention after adrenal enucleation, we compared the mineralocorticoid activity of these steroids to aldosterone using the toad bladder. Using short-circuit current as a measure of sodium transport, we found that 19-OH-DOC (10(-8) M) had no significant effect on sodium transport. However, 19-nor-DOC (10(-8) M) increased sodium transport to a degree not different from aldosterone (10(-8) M). Furthermore, the onset of action, duration of activity, and inhibition of effect of 19-nor-DOC by spironolactone were not different from that of aldosterone. We conclude that 19-nor-DOC exhibits a significant effect on sodium transport and thus has the potential to play a role in the sodium retention following adrenal enucleation. Under the conditions of these studies, 19-OH-DOC exhibited no effect on sodium transport.

After adrenal enucleation rats have an impaired ability to excrete a salt load. From micropuncture studies comparing data obtained from the late distal collection site and the urine, it has been suggested that this antinatriuretic effect occurs along the collecting duct. These studies are indirect, however, and cannot evaluate the contribution of deep nephrons. We have performed studies directly measuring inner medullary collecting duct (IMCD) function in saline-loaded rats 6 days after adrenal enucleation (AE). The fraction of filtered fluid, sodium, chloride, and potassium was analyzed as a function of IMCD length. In six AE rats 35% of the fluid, 35% of the sodium, and 31% of the chloride delivered to the IMCD was reabsorbed. In six saline-loaded control rats, however, no statistically significant net reabsorption of fluid sodium, or chloride was detected. Net potassium secretion along the IMCD was found in both AE and control rats. No difference between groups was noted, and net addition accounted for 17% of the potassium excreted. We conclude that after AE, the excretion of fluid, sodium, and chloride is impaired during saline expansion because of enhanced reabsorption along the IMCD. AE does not affect potassium handling along the IMCD or potassium excretion.

We believe that in pregnant rats, GFR is elevated at term, whereas renal plasma flow appears not to be increased. The mechanism for this increment is unclear, as is the specific time during gestation when renal hemodynamics increase. Rats increase their salt appetite and significantly positive sodium balance occurs, which is most pronounced during the last week of gestation. The retained sodium is distributed in part to the products of conception and in part to the mother, particularly in increasing her extracellular fluid volume. Although chronic volume expansion is present, absolute tubular sodium reabsorption remains elevated. The mechanism for this elevated tubular reabsorption appears no to be related to aldosterone because reduction or inhibition of this mineralocorticoid does not affect sodium balance significantly.

The inner medullary collecting duct (IMCD) of the rat is a major site of acidification. However, previous micropuncture studies have failed to demonstrate acidification along the terminal IMCD during chronic acid feeding. To more completely evaluate this question we used the microcatheterization method in rats fed ammonium chloride for 3-7 days. Arterial pH was 7.30 +/- 0.015, and PCO2 was set at 40 +/- 0.6 mmHg. The IMCD data were analyzed as a function of IMCD length between 40% and the tip. Equilibrium pH decreased from 6.21 +/- 0.11 to 5.47 +/- 0.03, whereas PCO2 was unchanged (28 +/- 1 mmHg between the deep samples and tip). Bicarbonate delivery decreased from 92 +/- 14 to 10 +/- 1 nmol/min, titratable acid increased from 462 +/- 33 to 762 +/- 40 nmol/min, and ammonium delivery increased from 2,235 +/- 121 to 3,528 +/- 140 nmol/min. Thus estimated net acid increased from 2,638 +/- 134 to 4,303 +/- 161 nmol/min. To determine whether increasing delivery of buffer to the IMCD would stimulate acid secretion in acute acidosis, rats were studied during the infusion of HCl and creatinine. Arterial pH was 7.18 +/- 0.02. IMCD acidification was not increased compared with our previously published studies during HCl infusion [Am. J. Physiol. 241 (Renal Fluid Electrolyte Physiol. 10): F669-F676, 1981]. We conclude that chronic ammonium chloride ingestion stimulates IMCD acidification and that this increase may be an intrinsic modification of the acidification mechanism of the IMCD.

Tubular handling of potassium was studied in the Charles River CD (cesarean derived) rat by clearance, micropuncture, and anatomic techniques. The following groups were evaluated: group I, hydropenia; group II, KCl-mannitol infusion; group III, 10% body wt saline loading; group IV, uninephrectomy, hydropenia; and group V, uninephrectomy, saline loading. Comparison of micropuncture samples from early and late distal collection sites (LDCS) and urine collections revealed no net K transport along the distal convoluted tubule in groups I and III-V but net addition of K in all groups beyond the LDCS. Absolute K secretion beyond the LDCS appeared to be flow dependent in groups I-III. The LDCS was noted by light and electron microscopy always to be lined with collecting tubule epithelium. We conclude that no net change in potassium transport occurs along the superficial distal convoluted tubule during hydropenia or saline loading in normal or uninephrectomized Charles River CD rats, but secretion is demonstrable during KCl infusion. Net addition of potassium beyond the LDCS was noted in all groups and this addition was enhanced by uninephrectomy.

The purpose of these experiments was to evaluate the effect of acute respiratory alkalosis (ARA) and chronic bicarbonate drinking (CBD) on inner medullary collecting duct (IMCD) acidification. Microcatheterization was used to measure pH and PCO2, and samples were simultaneously obtained for measurement of bicarbonate, titratable acid (TA), and ammonium. In ten ARA rats (arterial pH was 7.54 +/- 0.02; PCO2 was 20 +/- 1 mmHg), IMCD equilibrium pH was not different (deep pH was 5.65 +/- 0.06 and PCO2 was 20 +/- 1 mmHg; tip pH was 5.54 +/- 0.07 and PCO2 was 22 +/- 1 mmHg). Delivery of bicarbonate, TA, and ammonium also did not differ between collection sites. Thus net acidification along the IMCD was negligible. Nine rats drank NaHCO3 for 5-8 days (pH = 7.48 +/- 0.02) but did not receive NaHCO3 during the experiment so that arterial pH fell to 7.40 +/- 0.01. IMCD equilibrium pH was different at deep (pH was 5.68 +/- 0.06; PCO2 was 32 +/- 1 mmHg) and tip (pH was 5.57 +/- 0.04; PCO2 was 27 +/- 1 mmHg; P less than or equal to 0.05) collection sites. However, only minimal changes in the delivery of bicarbonate, TA, and ammonium were noted, and net acidification along the IMCD was negligible. In ten control rats, net acidification was 219 nmol/min between collection sites (P less than 0.001). We conclude that ARA and CBD abolish acidification along the IMCD. In addition, CBD produces an intrinsic modification along the IMCD, which suppresses acid secretion and persists after acute recovery from alkalemia.

Male rats were studied between 20 and 60 days of age. Standard renal clearance techniques were used. The efficiency of the diuretic response, the sodium response, and the potassium response were calculated as defined previously. Results indicate that both diuretic and sodium efficiency is lower between the ages of 20 and 30 days as compared to 40 and 60 days of age, whereas potassium efficiency shows no age dependence. In morphological studies, it has been observed that the proximal tubule cells of 22-day-old rats show changes in lateral intercellular spaces with comparable features to those of mature animals. The spaces widened during volume expansion. The change is quantitatively the same as that found in mature rats. Both blood pressure and hematocrit show continuous increase with age. The response of Ringer infusion in both immature and mature animals may be a mixed response involving changes in physical factors and renal responses to volume expansion per se.

We previously reported that acute respiratory acidosis (ARA) did not stimulate inner medullary collecting duct (IMCD) acidification. It was possible that the failure to find enhanced IMCD acidification was a function of insufficient buffer delivery. To answer this question we studied IMCD acidification in rats with ARA during the infusion of the buffer creatinine. We employed the microcatheterization technique to directly measure pH and PCO2 with glass membrane electrodes and also obtained fluid samples for the measurement of titratable acid and ammonium. Arterial pH was 7.19 +/- 0.01 and PCO2 was 93 +/- 2 mmHg. The IMCD data were analyzed as a function of IMCD length (approximately 6 mm). Equilibrium pH decreased from 5.99 +/- 0.05 to 5.58 +/- 0.02 and PCO2 increased from 71 +/- 11 to 132 +/- 6 mmHg between origin and tip. Bicarbonate delivery decreased from 111 +/- 14 to 38 +/- 2 nmol/min; titratable acid increased from 867 +/- 87 to 1,625 +/- 61 nmol/min, but ammonium delivery did not change along the duct. Thus, estimated net acid increased from 1,772 +/- 155 to 2,709 +/- 88 nmol/min. We conclude that during the presence of increased buffer delivery to the IMCD, rats with ARA markedly increased proton secretion along the terminal nephron.

To directly characterize acidification by the collecting duct, we developed pH and PCO2 microelectrodes suitable for microcatheterization of the inner medullary collecting duct (IMCD). In saline-infused control rats apparent in situ pH fell significantly along the IMCD, from 5.95 at 60% length to 5.49 at the papilla tip. Luminal PCO2 averaged 34 +/- 1 mmHg and PD averaged +3 mV. In rats acutely infused with 0.1 N HCl, apparent in situ pH also decreased significantly from 5.56 to 5.28, PD averaged +2 mV, and luminal PCO2 31 +/- 1 mmHg. The luminal PCO2 of HCl-infused rats was significantly less than controls and both levels were significantly below arterial PCO2. Corroborating the in situ pH profiles, equilibrium pH measured on collected IMCD samples also decreased significantly with percent length. In samples measured in situ and at equilibrium, a small but significant acid disequilibrium pH ws seen in both groups. We interpret these results to indicate that the IMCD actively participates in distal acidification. It is proposed that acidification by the IMCD is predominantly mediated by hydrogen ion secretion which simultaneously acidifies luminal fluid and generates a cellular sink for CO2, thereby inducing an acid disequilibrium pH by two mechanisms.

The purpose of this study was to test whether the absence or inhibition of aldosterone would alter the Na retention of pregnancy in the rat. We performed metabolic balance studies during the last 2 wk of the 3-wk rat pregnancy in the following four groups of saline-drinking rats: nonpregnant, sham adrenalectomized; pregnant, sham adrenalectomized; nonpregnant, adrenalectomized; and pregnant, adrenalectomized. Significant net Na retention was not found during the 2nd wk in any of the groups. During the final week, however, both pregnant groups retained between 8 and 11 meq Na more than their respective control groups. In addition, we measured plasma aldosterone concentration in these rats at term. The mineralocorticoid was not detectable in the nonpregnant adrenalectomized rats but was present in the pregnant adrenalectomized rats at a concentration of about 50% of that found in the pregnant sham-adrenalectomized rats. Serial measurements revealed that aldosterone was measurable by day 18 and nondetectable by 2 days postpartum in these pregnant adrenalectomized rats. Comparable balance studies were also performed in three groups of water-drinking rats: nonpregnant, pregnant, and pregnant rats that received large daily doses of spironolactone. During the 3rd wk both pregnant groups were in significant positive Na balance. Net Na retention for untreated pregnant rats was 8.4 meq and for the spironolactone treated, 6.9 meq, values that were not significantly different. We conclude that is is unlikely that aldosterone plays a critical role in the Na retention found during rat pregnancy.

The question of phosphate transport along the collecting duct remains controversial inasmuch as no data from direct in vivo evaluation of this nephron segment have been reported. We measured net phosphate transport along the inner medullary collecting duct (IMCD) using the collecting duct microcatheterization technique in five groups of rats. In control rats no net phosphate transport was found and 9.4% of the fraction of filtered phosphate (FFP) entered the IMCD and was excreted. After acute thyroparathyroidectomy (TPTX) there was a striking reduction in the FFP entering the IMCD, 1.8%, and significant reabsorption occurred, 0.5% being excreted. With acute TPTX and parathormone infusion, delivery increased to 33% without significant change along the IMCD. With acute TPTX and phosphate infusion, delivery was increased to control levels but no change was found in net phosphate transport. In rats studied 5-7 days after uninephrectomy alone, phosphate delivery was greater than in control, 25%, and no net phosphate transport was found. These studies demonstrate that phosphate absorption occurs along the IMCD in acutely TPTX rats when the delivery of phosphate to the IMCD is markedly reduced. The increase in phosphaturia which occurs after a reduction in renal mass cannot be accounted for by changes in net phosphate transport along the IMCD.

It is generally assumed that at least part of distal acidification occurs along the collecting duct. Complete and direct evaluation of acidification along this nephron segment is unavailable, however. The purpose of these experiments was to quantify the net acidification rate along the inner medullary collecting duct (IMCD) and to measure the effect of acute HCl acidosis. In 13 control rats (arterial pH, 7.39 +/- 0.01; PCO2, 39 +/- 1 mmHg) and 11 HCl-infused rats (arterial pH 7.18 +/- 0.01; PCO2, 40 +/- 1 mmHg) we obtained four to eight IMCD samples by a modified microcatheterization technique that also permitted measurement of in situ and in vitro pH. Tubular fluid pH decreased along the IMCD in both groups and was more acidic by 0.2-0.4 pH units in the acid-infused rats. Bicarbonate reabsorption was noted in both groups as delivery along the IMCD decreased from 205 +/- 127 to 26 +/- 6 nmol/min in control rats and from 219 +/- 118 to 17 +/- 4 nmol/min in the acidotic group. Ammonia delivery to and addition along the IMCD was significantly greater in the acidotic rats--from 193 +/- 59 to 462 +/- 53 nmol/min in control and from 887 +/- 126 to 1,396 +/- 90 nmol/min in acidotic rats. No significant change in total or titrated phosphate was seen. Net acid addition along the IMCD was over twice as great in acidotic rats, 450 vs. 970 nmol/min. Our results indicate that significant net acid addition occurs along the IMCD and that during acidosis this acidification rate increases.

Kidney slices of 7-17-week-old human fetuses were incubated for 90 min or less in Krebs-Ringer solution containing aminoisobutyric acid (AIB). Cell to medium ratios of AIB decrease with increasing fetal age. Uptake rate is slower than for mature human tissue. Both water content and extracellular space is high in embryonic tissue as compared to adult.

Potassium transport along the inner medullary collecting duct (IMCD) was evaluated by the microcatheterization technique in Charles River CD (cesarean derived) rats 7-9 days after sham operation (S) or uninephrectomy (UNPX). The fraction of filtered potassium (TF/P)K/In) as a function of IMCD length was analyzed by linear regression. In 13 S rats there was a significant correlation and slope (P less than 0.001) and (TF/P)K/In increased from 14% at the beginning of the IMCD to 25% in the urine. IMCD potassium secretion accounted for about half of the excreted potassium. In the UNPX rats a significant correlation and slope was also obtained (P less than 0.001); (TF/P)K/In at the beginning of IMCD was 24% and increased to 36% in the urine. No difference in slope was noted between the groups. There was a slightly greater absolute potassium secretion after UNPX (0.77 +/- 0.03 S vs. 0.93 +/- 0.04 mueq/min UNPX), but this did not account for most of the difference in potassium excretion noted. We conclude that net potassium secretion occurs along the IMCD in S and UNPX rats. After UNPX, there is significantly greater potassium delivery to the IMCD and a greater kaluresis. The increased kaluresis cannot be accounted for primarily by increased potassium secretion along the IMCD.

To better define the relationship between adrenal enucleation (AE), sodium retention and adrenal regeneration hypertension (ARH), we measured daily sodium balance in rats after bilateral adrenal enucleation with (AEN) and without uninephrectomy (AE), and before and after development of ARH, along with the appropriate control groups. We found that both AE and AEN groups retained sodium during 7 days after surgery; AE 1.16 +/- 0.24 and AEN 1.32 +/- 0.33 meq/100 g body wt. Sham-operated, uninephrectomized rats and AE rats treated with spironolactone were in zero sodium balance. In the AEN rats after the development of ARH (BP greater than 150 Torr), zero sodium balance was also noted. Only AEN rats developed hypertension while sham-operated, uninephrectomized rats and AE rats with both kidneys remained normotensive for 28 wk. In summary, we have demonstrated that for 1 wk after AE, rats are in positive sodium balance and this is reversed by spironolactone but not by uninephrectomy.

After adrenal enucleation, rats have an impaired ability to excrete a salt load because of enhanced collecting duct reabsorption. This antinatriuretic effect, thought to be secondary to a mineralocorticoid-like substance secreted by the enucleate gland, can be reversed by treatment with spironolactone or dexamethasone. To define the renal mechanisms involved in this drug-induced natriuresis we have utilized clearance and micropuncture techniques in enucleate saline-expanded rats that were treated with either spironolactone (S) or dexamethasone (D), or were untreated (U). Sodium excretion was clearly increased after S, 13.9, and D, 19.3 mueq/min vs. u, 5.9 mueq/min. The mechanisms of this natriuresis, however, were dissimilar. Spironolactone-treated rats were not different from untreated rats except with regard to function beyond the superficial late distal tubule, where U rats reabsorbed over 50% of the delivered sodium. In the S group 38% of the excreted sodium was added along this tubular locus, 5.2% of the filtered sodium reaching the late distal tubule and 7.3% appearing in the urine. These data demonstrate that the natriuresis after S is secondary to the net addition of sodium beyond the superficial late distal tubule. Spironolactone may work by inhibiting a mineralocorticoid-like product of the enucleate gland and, thereby, eliminate the sodium-retaining effect of this product. The natriuresis after D, however, can be explained solely on the basis of a markedly increased filtered load of sodium traversing the nephron.

During a bicarbonate diuresis, final urine Pco2 considerably exceeds systemic Pco2, an effect thought to reflect the postpapillary delayed dehydration of carbonic acid. To test this explanation, Pco2 tensions along the inner medullary collecting duct (IMCD) of bicarbonate-loaded rats were measured directly using Pco2 microelectrodes. With systemic Pco2 held at 40 mm Hg, IMCD Pco2 exceeded systemic Pco2 in every measurement by an average of 20 to 30 mm Hg. A significant increment in Pco2 was seen between 50% IMCD length and the papilla tip. During the infusion of carbonic anhydrase, IMCD Pco2 was reduced but not to systemic levels. Finding elevated Pco2 along the terminal IMCD deemphasizes the importance of postpapillary delayed dehydration and suggests the possibility that bicarbonaturia is associated with papillary accumulation of carbon dioxide.

After adrenal enucleation (AE) rats avidly retain sodium (early phase), but after 7-10 days they lose this sodium avidity (late phase). Although increased production of a mineralocorticoid, 19-nor-deoxycorticosterone (19-Nor-DOC), has been implicated, 19-Nor-DOC levels during the early and late phases of AE have not been systematically measured. Furthermore it is not known why 19-Nor-DOC production should increase during a time when production of 11 beta- and 18-hydroxylated corticosteroids are decreased in AE. The purpose of this study was to examine the 11 beta, 18-, and 19-hydroxylase pathways in the early and late phases of AE. The results demonstrate increased urinary 19-Nor-DOC and decreased 18-OH-DOC and corticosterone excretion in the early phase of AE at a time when adrenal mitochondrial 11 beta- and 18-hydroxylase activities were decreased but 19-hydroxylase activity was unchanged. During the late phase of AE, urinary 19-Nor-DOC had decreased and 18-OH-DOC and corticosterone had increased to levels indistinguishable from those in sham controls. This reduction in 19-Nor-DOC was associated with a decrease in 19-hydroxylase activity in AE. Since the 11 beta, 18-, and 19-hydroxylases have a common substrate (DOC), it is possible that differential flux of DOC through these pathways could account for the changes in steroid production in AE. These data suggest that the increased 19-Nor-DOC excretion in AE may be due to alterations in enzyme activity leading to a shunting of DOC into the 19-Nor-DOC pathway. In addition, the synchronicity of 19-Nor-DOC with sodium excretion suggests that it has an important role in the pathogenesis of the sodium retention in AE.

The zonula occludens (Z.O.) has been implicated as a major pathway for passive fluid movement from lateral intercellular spaces to lumen in response to an expansion of blood or fluid volume. The present study examines to Z.O. of adult rat nephrons in control and Ringer-Locke or while blood infused animals to determine whether varying the conditions of passive fluid movement would influence the structure of the Z.O. Rats were infused over 20 minutes with Ringer-Locke (7% body weight) or while blood (2.3% body weight). In the controls, 92%-94% of all Z.O. observed in proximal tubules and 100% of Z.O. from distal tubules and collecting ducts had fused outer leaflets. The approximate linear extent of the Z.O. was 140 A for proximal tubules, 2,500 A for distal tubules and 3,000 A for collecting ducts. There was no significant difference in any of these values following the infusion with either Ringer-Locke or whole blood. We conclude that expansion of blood or fluid volume causes no detectable alteration in the fine structure of Z.O. under the conditions used in the present study.