A single low-energy, iron-poor supernova as the source of metals in the star SMSS J031300.36-670839.3.

The element abundance ratios of four low-mass stars with extremely low metallicities (abundances of elements heavier than helium) indicate that the gas out of which the stars formed was enriched in each case by at most a few--and potentially only one--low-energy supernova. Such supernovae yield large quantities of light elements such as carbon but very little iron. The dominance of low-energy supernovae seems surprising, because it had been expected that the first stars were extremely massive, and that they disintegrated in pair-instability explosions that would rapidly enrich galaxies in iron. What has remained unclear is the yield of iron from the first supernovae, because hitherto no star has been unambiguously interpreted as encapsulating the yield of a single supernova. Here we report the optical spectrum of SMSS J031300.36-670839.3, which shows no evidence of iron (with an upper limit of 10(-7.1) times solar abundance). Based on a comparison of its abundance pattern with those of models, we conclude that the star was seeded with material from a single supernova with an original mass about 60 times that of the Sun (and that the supernova left behind a black hole). Taken together with the four previously mentioned low-metallicity stars, we conclude that low-energy supernovae were common in the early Universe, and that such supernovae yielded light-element enrichment with insignificant iron. Reduced stellar feedback both chemically and mechanically from low-energy supernovae would have enabled first-generation stars to form over an extended period. We speculate that such stars may perhaps have had an important role in the epoch of cosmic reionization and the chemical evolution of early galaxies.

Efficacy and safety of ultrasound-guided high intensity focused ultrasound ablation of symptomatic uterine fibroids in Black women: a preliminary study.

OBJECTIVE: To evaluate the therapeutic effect and safety of ultrasound-guided high-intensity focused ultrasound (USgHIFU) treatment on symptomatic uterine fibroids in Black women. DESIGN: A feasibility study. SETTING: Gynaecological department in a teaching hospital in South Africa. POPULATION: Premenopausal women with uterus fibroids. METHODS: Twenty-six patients with 53 fibroids who underwent USgHIFU treatment were enrolled. The USgHIFU treatment information was recorded, including treatment time, sonication time and total energy. Adverse events were also observed and recorded during and after treatment. MAIN OUTCOME MEASURES: Safety and efficacy of USgHIFU for the treatment of uterine fibroids in Black women. RESULTS: The median volume of fibroids was 52.7 (interquartile range, 18.6-177.4) cm3 . According to USgHIFU treatment plan, total energy of 298.6 ± 169.3 kJ (range, 76.0-889.2) within treatment time of 90.3 ± 43.3 minutes (range, 14.0-208.0), in which sonication time of 774.0 ± 432.9 seconds (range, 190.0-2224.0) was used to ablate fibroids. The average ablation rate was 80.6 ± 9.7% (range, 46.5-94.5%). During the procedure, 69.2% of the patients reported lower abdominal pain, 57.7% sciatic/buttock pain, 38.5% burning skin, and 34.6% transient leg pain. No severe complications were observed. CONCLUSIONS: USgHIFU is feasible and safe to use to treat symptomatic uterine fibroids in Black women. TWEETABLE ABSTRACT: Multiple uterine fibroids are more frequently detected in Black women. USgHIFU is feasible and safe for the treatment of uterine fibroids in Black women.

A multisite community-based health literacy intervention for Spanish speakers.

The National Action Plan to Improve Health Literacy emphasizes the importance of community-based opportunities for education, such as English as a second language (ESL) programs. It recommends collaborations among the adult literacy and ESL communities. However, limited attention has been given to researching the effectiveness of community-based interventions that combine ESL and health literacy. The purpose of this study was to explore the feasibility of using different community settings for improving health literacy among adult Spanish speakers through an English language program. The study used a pre-experimental, single arm pretest-posttest design, and implemented the Health Literacy and ESL Curriculum. A collaborative was established between the community and university researchers. Participants were recruited at three distinctive sites. Health literacy was assessed using the Spanish version of the Test of Functional Health Literacy in Adults (TOFHLA). Analysis included descriptive and paired-group t test. Forty-nine participants completed the intervention and post-tests (92% retention rate). Overall--all sites--posttest scores significantly improved for total TOFHLA, raw numeracy, and reading comprehension (p < 0.0001). Similarly, all three sites yielded significantly better mean differences for the total TOFHLA score while numeracy and reading comprehension significantly improved in some sites. Results suggest that community sites are viable venues for delivering health literacy/language instruction to Spanish speaking adults. The study also points to community engagement and ESL programs as two essential components of effective health literacy interventions among Spanish speakers.

Low central venous oxygen saturation in haemodynamically stabilized trauma patients is associated with poor outcome.

BACKGROUND: Central venous oxygen saturation (ScvO(2)) is suggested to reflect the adequacy of oxygen delivery, and the main objective of the present study was to determine whether ScvO(2) is associated with outcome in haemodynamically stabilized trauma patients. METHODS: Haemodynamically unstable trauma patients receiving a central venous line within 1 h of admission were eligible for inclusion in this prospective observational study. The mean arterial pressure (MAP), lactate and ScvO(2) were recorded at inclusion and every 6 h for 36 h or until lactate was <2.0 mmol/l and ScvO(2) was >75% in two consecutive measurements. Patients with a MAP of ≥70 mmHg were considered to be haemodynamically stabilized. The outcome measure was complications defined as infections, delta sequential organ failure assessment score of >0, and mortality. RESULTS: Fifty patients with a median new injury severity score of 27 (17-34) were analysed. Complications occurred in 33 patients. An association between ScvO(2) following resuscitation to MAP ≥70 mmHg and complications was detected with an odds ratio of 0.94 (95% confidence interval; 0.89-0.99). This association was also significant when adjusted for injury severity. The result implies that a low ScvO(2) value is associated with more complications. The optimal cut-off for ScvO(2) to discriminate between patients who did or did not develop complications was found to be 66.5% (56-86%). CONCLUSIONS: These data suggest that low ScvO(2) in haemodynamically stabilized patients is associated with a poor outcome and that ScvO(2) represents a potential endpoint of resuscitation in trauma patients.

Estrogen receptor in the rabbit corpus luteum.

The rabbit corpus luteum contains a cytoplasmic estrogen receptor substance in concentrations comparable to those found in the uterus. Receptor in corpora lutea of nonpregnant rabbits was highest at midluteal phase and decreased before physical regression of corpora was evident. These results support the view that the corpus luteum in this species is an estrogen-responsive tissue.

Medullary collecting duct acidification. Effects of potassium, HCO3 concentration, and pCO2.

The medullary collecting duct (MCD) from renal outer medulla possesses significant HCO3 absorptive capacity. In vitro microperfusion studies have shown that HCO3 absorption in this segment is carbonic anhydrase dependent, affected by peritubular and luminal chloride concentrations, is independent of the presence of Na or the presence of Na transport, and is stimulated by mineralocorticoid hormone. The present in vitro microperfusion studies defined regulatory influences on MCD acidification as assessed by acute changes in the extracellular K and HCO3 concentrations and pCO2. These studies showed that acute changes in the peritubular K concentration to either 0 mM K or 50 mM K have no significant effect on HCO3 absorption in MCD. Intracellular voltage recordings showed that elevation of peritubular K concentration from 5 to 50 mM produced only a 2.8 mV depolarization of the basolateral cell membrane of MCD cells. In addition, acute reduction of peritubular K from 5 to 0 mM had no significant effect on intracellular voltage. Studies that were designed to assess the effects of HCO3 concentration and pCO2 on acidification showed that acute reduction of peritubular HCO3 concentration from 25 to 5 mM (pH change from 7.4 to 6.8) increased lumen-positive voltage from 30.2 +/- 3.8 to 40.0 +/- 4.4 mV, and simultaneously increased net HCO3 absorption from 15.6 +/- 1.9 to 22.9 +/- 2.9 pmol X mm-1 X min-1. Elevation of peritubular HCO3 concentration from 25 to 50 mM (pH change from 7.4 to 7.8) significantly decreased lumen-positive voltage from 33.8 +/- 2.4 to 26.7 +/- 1.5 mV and simultaneously decreased net HCO3 absorption from 17.9 +/- 1.2 to 12.8 +/- 1.3 pmol X mm-1 X min-1. In addition, acute reduction of peritubular pCO2 from 40 to less than 14 mmHg (final pH 7.8-7.9) significantly decreased lumen-positive voltage from 31 +/- 4.4 to 15.7 +/- 1.0 mV. Coincidentally, HCO3 absorption decreased significantly from 11.0 +/- 3.7 to 5.3 +/- 0.7 pmol X mm-1 X min-1. We conclude that: alteration of peritubular K concentration from 0 to 50 mM in vitro does not affect HCO3 absorption in the MCD, and that this lack of effect appears to be related to a low basolateral cell membrane K conductance; net HCO3 absorption and the associated lumen-positive voltage can be modulated by in vitro changes in peritubular HCO3 and pCO2 (or pH); and the MCD perfused in vitro appears to be a good model for studying the mechanisms and regulation of distal nephron acidification.

Characteristics of volume reabsorption in rabbit superficial and juxtamedullary proximal convoluted tubules.

Segments of superficial and juxtamedullary proximal convoluted tubules of the rabbit were perfused in vitro to examine the mechanisms responsible for net volume reabsorption. The very early postglomerular segments were not studied. Fluid reabsorptive rates and transepithelial potential differences were compared under various conditions: (a) with perfusate that simulated glomerular filtrate; (b) with perfusate that lacked glucose, amino acids, and acetate and that had HCO(3) and Cl concentrations of 5 and 140 mM, respectively; (c) with perfusate that lacked glucose, amino acids, and acetate but with 20 meq of NaHCO(3) replaced with 20 meq of Na cyclamate; (d) with the same perfusate as in b but in the presence of ouabain in the bath; (e) with ultrafiltrate of rabbit serum titrated with HCl to final HCO(3) and Cl concentrations of 2 and 134 mM, respectively. Tubules were perfused with this titrated ultrafiltrate at 37 degrees C, 21 degrees C, and in the presence of 0.1 mM ouabain in the bath. Bath fluid in all experiments was regular rabbit serum. Under conditions a and b superficial proximal convoluted tubule (SFPCT) and juxtamedullary proximal convoluted tubule (JMPCT) behaved similarly with the exception that SFPCT exhibited a lumen-positive and JMPCT a lumen-negative electrical potential under condition b. However, under condition c SFPCT failed to exhibit net volume reabsorption, whereas reabsorption in JMPCT continued unchanged. Ouabain did not affect volume reabsorption in SFPCT under condition d, whereas neither ouabain nor hypothermia affected SFPCT under condition e. In contrast, ouabain and hypothermia totally inhibited volume reabsorption in JMPCT under conditions d and e. These studies document heterogeneous mechanisms responsible for volume reabsorption in the major portions of SFPCT and JMPCT with passive forces predominating in SFPCT and active forces in JMPCT.

Intrinsic differences in various segments of the proximal convoluted tubule.

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.

Regulation of rabbit medullary collecting duct cell pH by basolateral Na+/H+ and Cl-/base exchange.

The collecting duct of the inner stripe outer medulla (OMCDi) is a major site of distal nephron acidification. Using the pH sensitive fluorescent dye 2'-7'-bis(carboxyethyl)-5,6,-carboxyfluorescein (BCECF) and quantitative spectrofluorometry to measure intracellular pH in isolated perfused OMCDi, we have characterized basolateral transport processes responsible for regulation of intracellular pH. Experiments suggesting the existence of basolateral Cl-/base exchange were performed. In HCO3- containing buffers, bath Cl- replacement resulted in reversible alkalinization of the OMCDi from 7.22 +/- 0.05 to 7.57 +/- 0.12. Similarly 0.1 mM bath 4',4'-diisothiocyanostilbene-2,2'-disulphonic acid (DIDS) alkalinized the OMCDi from 7.14 +/- 0.09 to 7.34 +/- 0.09 and blocked further alkalinization by bath Cl- removal (delta = + 0.02 pH units). The concentration dependence kinetics of Cl-/base exchange revealed a K1/2 of 10 mM for external Cl- with a Vmax of 0.50 pH U/min. Experiments suggesting the existence of basolateral Na+/H+ exchange were also performed. Replacement of bath Na+ by tetramethylammonium resulted in reversible cell acidification (7.14 +/- 0.09 to 6.85 +/- 0.1). Tubules that were acidified by a brief exposure to NH4Cl displayed recovery of cell pH back to baseline at a rate that was highly dependent on bath Na+ concentration. Half maximal recovery rate was achieved at 7 mM bath Na+ and Vmax was 0.605 pH U/min. The Na+-dependent rate of cell pH recovery after acidification was blocked by 0.2 mM bath amiloride. These results suggest that intracellular pH in the OMCDi is regulated by parallel basolateral Na+/H+ exchange and Cl-/base exchange.

Epidermal growth factor inhibits the hydroosmotic effect of vasopressin in the isolated perfused rabbit cortical collecting tubule.

Epidermal growth factor (EGF) is a 53-amino acid polypeptide which is a potent mitogen for cultured cells. The kidney has recently been shown to be a major site of synthesis for the EGF precursor. EGF infusions in sheep result in a diuresis and natriuresis despite a fall in GFR, suggesting a direct tubular effect. Using in vitro microperfusion of rabbit cortical collecting tubules (CCTs) at 37 degrees C, we examined the effect of EGF on the transepithelial voltage (Vt) and arginine vasopressin (AVP)-stimulated hydraulic conductivity (Lp). Pretreatment with peritubular EGF at concentrations from 10(-8) to 10(-12) M resulted in a 50% inhibition of both AVP- and 8-chlorophenythio-cyclic AMP-stimulated peak Lp. This effect was reversed by the protein kinase C inhibitor, staurosporine, but unaffected by indomethacin. CCTs with an initially negative Vt, depolarized after exposure to bath EGF. 10(-8) M EGF applied from the lumen had no effect on either Lp or Vt. Specific binding of 20 nM 125I-EGF to microdissected CCTs was also demonstrated. These results suggest that EGF can modulate both salt and water transport in the CCT via a receptor linked to protein kinase C activation.

Prostaglandin E2 inhibits sodium transport in rabbit cortical collecting duct by increasing intracellular calcium.

The mechanism by which prostaglandin E2 (PGE2) inhibits sodium absorption (JNa) in the rabbit cortical collecting duct (CCD) was explored. PGE2 activates at least three signaling mechanisms in the CCD: (a) by itself PGE2 increases cAMP generation (b) PGE2 also inhibits vasopressin-stimulated cAMP accumulation, and (c) PGE2 raises intracellular calcium([Ca++]i). We tested the contribution of these signaling pathways to PGE2's effect on Na+ absorption, measuring 22Na flux (JNa) and [Ca++]i (using fura-2) in microperfused rabbit CCDs. In control studies PGE2 reduced JNa from 28.2 +/- 3.4 to 15.6 +/- 2.6 pmol.mm-1.min-1. Lowering bath calcium from 2.4 to 45 nM did not by itself alter JNa but in this setting PGE2 failed to inhibit JNa (28.6 +/- 5.4 to 38.5 +/- 4.0). In separate tubules, PGE2 raised [Ca++]i in a spike-like fashion followed by a sustained elevation. However, in 45 nM bath Ca++, PGE2 failed to produce a sustained [Ca++]i elevation. While pretreatment of CCDs with pertussis toxin blocked PGE2 inhibition of vasopressin-stimulated water permeability, it did not block the effect of PGE2 on JNa. To see if cAMP generation contributes to the effect of PGE2 on JNa, we tested the effect of exogenous cAMP, (8-chlorophenylthio(CPT)cAMP) on JNa. 0.1 mM 8-CPTcAMP reduced JNa from 35.75 +/- 2.3 to 21.6 +/- 2.2. However, the addition of PGE2 further blunted JNa to 15.9 +/- 1.3. In CCDs pretreated with indomethacin, 8-CPTcAMP did not significantly decrease JNa 33.6 +/- 2.8 vs. 28.4 +/- 2. However, superimposed PGE2 reduced JNa to 19.0 +/- 3.0. We conclude that PGE2 inhibits sodium transport predominantly by increasing intracellular calcium. This action is not mediated by a pertussis toxin-sensitive G protein. Finally, cAMP, through a cyclooxygenase-dependent mechanism, also inhibits CCD JNa and may contribute to the effects of PGE2 on JNa in the rabbit CCD.

Angiotensin II directly stimulates sodium transport in rabbit proximal convoluted tubules.

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.

Interactions of lysyl-bradykinin and antidiuretic hormone in the rabbit cortical collecting tubule.

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.

Phorbol myristate acetate, dioctanoylglycerol, and phosphatidic acid inhibit the hydroosmotic effect of vasopressin on rabbit cortical collecting tubule.

We explored the role for protein kinase C (PKC) in modulating vasopressin (AVP)-stimulated hydraulic conductivity (Lp) in rabbit cortical collecting tubule (CCT) perfused in vitro at 37 degrees C. In control studies, 10 microU/ml AVP increased Lp (mean +/- SE, X 10(-7) centimeters/atmosphere per second) from 4.4 +/- 0.9 to 166.0 +/- 10.4. Pretreatment with dioctanoylglycerol (DiC8) suppressed AVP stimulated peak Lp (peak Lp, 21.9 +/- 3.1). Pretreatment with 10(-9) and 10(-7) M 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (PMA) also blocked the increase in Lp in a dose-dependent fashion (peak Lp, 59.3 +/- 7.5 and 18.6 +/- 4.8, respectively). Inactive phorbol ester, 4 alpha-phorbol 12 beta,13 alpha-didecanoate (10(-7) M), had no effect. PMA also suppressed the increase in Lp induced by 10(-4) M 8-p-chlorophenylthio-cyclic AMP (CcAMP): peak Lp was 169.4 +/- 14.9 in control, 79.2 +/- 5.5 with 10(-9) M PMA, and 25.7 +/- 2.9 with 10(-7) M PMA. Furthermore, when 10(-7) M PMA was added to the bath 10 min after exposure to AVP, the Lp response to AVP was blocked. Peak Lp was 52.4 +/- 9.6 with PMA vs. 165.1 +/- 10.0 in control. Phosphatidic acid (PA), which is thought to stimulate phosphatidylinositol (PI) turnover, produced similar inhibitory effects on AVP as well as CcAMP-stimulated Lp: PA suppressed 10-microU/ml AVP-induced peak Lp from a control value of 159.6 +/- 7.9 to 88.9 +/- 15.8, and 10(-4) M CcAMP induced peak Lp from 169.4 +/- 14.9 to 95.5 +/- 7.7. We conclude that PMA, at concentrations known to specifically activate PKC, suppresses the hydroosmotic effect of AVP on CCT; This suppression is primarily a post-cAMP event; Inhibition of AVP-stimulated Lp by DiC8 and PA also suggests an inhibitory role for the PKC system; The ability of pre- and post-AVP administration of PMA to blunt the AVP response suggests that agents that act through modulation of PI turnover in CCT may regulate the hydroosmotic effect of AVP.

Regulation of net bicarbonate transport in rabbit cortical collecting tubule by peritubular pH, carbon dioxide tension, and bicarbonate concentration.

The effects of changes in peritubular pH, carbon dioxide tension (PCO2), and HCO3- concentration on net HCO3- transport was examined in in vitro perfused cortical collecting tubules (CCTs) from unpretreated New Zealand white rabbits. Lowering peritubular HCO3- concentration and pH by reciprocal replacement of HCO3- with Cl-, significantly stimulated net HCO3- absorption. Lowering peritubular HCO3- concentration and pH, by substitution of HCO3- with gluconate, while keeping Cl- concentration constant, also stimulated net HCO3- absorption. Raising peritubular HCO3- concentration and pH, by reciprocal replacement of Cl- with HCO3-, inhibited net HCO3- absorption (or stimulated net HCO3- secretion). When the tubule was cooled, raising peritubular HCO3- concentration had no effect on net HCO3- transport, suggesting these results are not due to the passive flux of HCO3- down its concentration gradient. The effect of changes in ambient PCO2 on net HCO3- transport were also studied. Increasing the ambient PCO2 from 40 mmHg to either 80 or 120 mmHg, allowing pH to fall, had no effect on net HCO3- transport. Similarly, lowering ambient PCO2 to 14 mmHg had no effect on net HCO3- transport. Simultaneously increasing peritubular HCO3- concentration and PCO2, without accompanying changes in peritubular pH, i.e., isohydric changes, stimulated net HCO3- secretion to the same degree as nonisohydric increases in peritubular HCO3- concentration. Likewise, isohydric lowering of peritubular HCO3- concentration and PCO2 stimulated net HCO3- absorption. We conclude that: acute changes in peritubular HCO3- concentration regulate acidification in the CCT and these effects are mediated by a transcellular process; acute changes in ambient PCO2 within the physiologic range have no effect on HCO3- transport in the in vitro perfused CCT; and acute in vitro regulation of CCT acidification is independent of peritubular pH.

Hormonal regulation of proton secretion in rabbit medullary collecting duct.

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.

Phorbol ester and A23187 have additive but mechanistically separate effects on vasopressin action in rabbit collecting tubule.

Activation of protein kinase C (PKC) and elevation of intracellular calcium ion concentration ([Ca++]i) result from phosphatidylinositol biphosphate (PIP2) breakdown. We previously demonstrated that PKC activation inhibits arginine vasopressin (AVP)-induced osmotic water flow in rabbit cortical collecting tubule (CCT) perfused in vitro at 37 degrees C. To estimate the potential significance of PIP2 turnover as a modulator of water transport in this nephron segment, we examined the effect of Ca on AVP action and explored the mechanisms of action of PKC and increased [Ca++]i. In rabbit CCTs perfused at 37 degrees C, pretreatment with bath A23187 (2 x 10(-8) M, 2 x 10(-6) M), a Ca ionophore, almost totally suppressed AVP (10 microU/ml)-induced peak hydraulic conductivity (Lp). The suppression by 2 x 10(-8) M A23187 was as potent as that by 2 x 10(-6) M A23187, and significant even when it was administered 10 min after AVP. When phorbol myristate acetate (PMA, 10(-9) M), a PKC activator, and A23187 (2 x 10(-8) M) were placed in the bath simultaneously, the combined suppressive effect on peak Lp was greater than that of either inhibitor alone. However, the mechanisms of inhibition by PMA and A23187 were different. While both 10(-7) and 10(-9) M PMA suppression are primarily post-cAMP, A23187 predominantly suppressed a pre-cAMP step: 10(-4) M chlorophenylthio-cAMP-induced peak Lp was not affected by 2 x 10(-8) M A23187, and only partially inhibited by 2 x 10(-6) M A23187. The PMA (10(-7) M) suppression of AVP-induced peak Lp was totally reversed by bath staurosporine (10(-7) M), a PKC inhibitor, but not attenuated by either bath indomethacin (5 x 10(-6) M) or low Ca (1-2 x 10(-6) M) bath medium. In contrast, the A23187 (2 x 10(-8) M) suppression of the peak Lp was not affected by staurosporine, but was significantly reversed by indomethacin or low Ca bath medium. We conclude: (a) Elevation of [Ca++]i, as well as activation of PKC, suppresses the hydroosmotic effect of AVP on CCT at 37 degrees C. (b) When stimulated simultaneously these two intracellular mediators are additive in their antagonism of AVP action. These results suggest that stimulated PIP2 breakdown may be an important modulator of water transport in CCT. (c) Different mechanisms underlie PKC and Ca-mediated suppression of the AVP-induced water transport. The inhibition of AVP action by increased [Ca++]i is primarily pre-cAMP, and involves a cyclooxygenase metabolite(s) of arachidonic acid, while the inhibition by PKC is post-cAMP, and independent of cyclooxygenase products of arachidonic acid.

Indomethacin secretion in the isolated perfused proximal straight rabbit tubule. Evidence for two parallel transport mechanisms.

We studied indomethacin as a probe of anion transport across the isolated perfused proximal straight tubule of the rabbit and discovered that a substantial component of transport may occur by anion exchange at the basolateral membrane. Various perturbations involving direct or indirect dissipation of the cellular sodium gradient (ouabain, sodium- or potassium-free solutions, cooling to 18 degrees C) resulted in only a 50% inhibition of indomethacin transport, which raised the question of a co-existent alternative pathway for secretion. Similarly, the anion exchange inhibitor, 4,4'-diisothiocyanostilbene (DIDS), diminished indomethacin secretion by only 50%. Cooling followed by DIDS or the reverse sequence resulted in additive inhibition such that the combination abolished active secretion of indomethacin. We conclude that active secretion of indomethacin by the proximal straight tubule appears to be in part sodium gradient dependent; the remainder may be driven by an anion exchanger on the basolateral membrane.

Mineralocorticoid modulation of rabbit medullary collecting duct acidification. A sodium-independent effect.

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.

Anion dependence of rabbit medullary collecting duct acidification.

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.