Renal bicarbonate reabsorption in the rat. IV. Bicarbonate transport mechanisms in the early and late distal tubule.

Bicarbonate transport was studied in vivo by separate microperfusion experiments of early and late distal tubules. Total CO2 was measured by microcalorimetry and fluid absorption by 3H-inulin. Significant bicarbonate absorption was observed in all experimental conditions. Bicarbonate transport was load-dependent upon increasing the luminal bicarbonate concentration from 15 to 50 mM in both early and late distal tubule segments and remained constant at higher concentrations at a maximum rate of 100-110 pmol/min per mm. At low lumen bicarbonate concentrations (15 mM), higher rates of bicarbonate absorption were observed in early (32.9 +/- 4.57 pmol/min per mm) as compared to late distal tubules (10.7 +/- 3.1 pmol/min per mm). Amiloride and ethyl-isopropylamiloride both inhibited early but not late distal tubule bicarbonate absorption whereas acetazolamide blocked bicarbonate transport in both tubule segments. Fluid absorption was significantly reduced in both tubule segments by amiloride but only in early distal tubules by ethyl-isopropylamiloride. Substitution of lumen chloride by gluconate increased bicarbonate absorption in late but not in early distal tubules. Bafilomycin A1, an inhibitor of H-ATPase, inhibited late and also early distal tubule bicarbonate absorption, the latter at higher concentration. After 8 d on a low K diet, bicarbonate absorption increased significantly in both early and late distal tubules. Schering compound 28080, a potent H-K ATPase inhibitor, completely blocked this increment of bicarbonate absorption in late but not in early distal tubule. The data suggest bicarbonate absorption via Na(+)-H+ exchange and H-ATPase in early, but only by amiloride-insensitive H+ secretion (H-ATPase) in late distal tubules. The study also provides evidence for activation of K(+)-H+ exchange in late distal tubules of K depleted rats. Indirect evidence implies a component of chloride-dependent bicarbonate secretion in late distal tubules and suggests that net bicarbonate transport at this site results from bidirectional bicarbonate movement.     

Angiotensin II stimulation of hydrogen ion secretion in the rat early proximal tubule. Modes of action, mechanism, and kinetics.

Physiologic concentrations of angiotensin II stimulate sodium transport by intestinal and renal early (S1) and late (S2) proximal tubule epithelial cells. We recently found that hydrogen ion secretion, which effects sodium bicarbonate absorption, was a transport function preferentially and potently increased by angiotensin II in S1 cells. S1 cells are normally responsible for half of the total renal hydrogen ion secretion. The mechanism by which angiotensin II regulates intestinal sodium transport is by potentiating sympathetic nerve activity and norepinephrine release. Direct control of hydrogen ion secretion by angiotensin II via receptors on epithelial cells has not been previously demonstrated. We now report that stimulation of in vivo hydrogen ion secretion in the rat early proximal tubule by angiotensin II was not mediated via change in nerve activity. Rather, enhanced hydrogen ion secretion by angiotensin II correlated with increased angiotensin II receptor density on epithelial cells in the early compared to late microdissected proximal tubule. Basolateral as well as luminal angiotensin II stimulated bicarbonate absorption. Angiotensin II reduced bicarbonate permeability and caused alteration in the apparent substrate affinity, but not maximal capacity, of the proximal hydrogen ion secretory system involving the Na+/H+ antiporter.     

Luminal secretion of ammonia in the mouse proximal tubule perfused in vitro.

A major portion of the total ammonia (tNH3 = NH3 + NH+4) produced by the isolated perfused mouse proximal tubule is secreted into the luminal fluid. To assess the role of Na+-H+ exchange in net tNH3 secretion, rates of net tNH3 secretion and tNH3 production were measured in proximal tubule segments perfused with control pH 7.4 Krebs-Ringer bicarbonate (KRB) buffer or with modified KRB buffers containing 10 mM sodium and 0.1 mM amiloride. Net tNH3 secretion was inhibited by 90% in proximal tubule segments perfused with the pH 7.4 modified KRB buffer while tNH3 production remained unaffected. The inhibition of net tNH3 secretion by perfusion with the modified KRB buffer was only partially reversed by acidifying the modified KRB luminal perfusate from 7.4 to as low as 6.2. These data indicate that the Na+-H+ exchanger facilitates a major portion of net tNH3 secretion by the proximal tubule and that luminal acidification may play only a partial role in the mechanism by which the Na+-H+ exchanger mediates net tNH3 secretion.     

Apical Na+/H+ antiporter and glycolysis-dependent H+-ATPase regulate intracellular pH in the rabbit S3 proximal tubule.

The apical transport processes responsible for proton secretion were studied in the isolated perfused rabbit S3 proximal tubule. Intracellular pH (pHi) was measured with the pH dye, 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein. Steady state pHi in S3 tubules in nominally HCO3(-)-free solutions was 7.08 +/- 0.03. Removal of Na+ (lumen) caused a decrease in pHi of 0.34 +/- 0.06 pH/min. The decrease in pHi was inhibited 62% by 1 mM amiloride (lumen) and was unaffected by 50 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (lumen) and Cl- removal (lumen, bath). After a brief exposure to 20 mM NH4Cl, pHi fell by approximately 0.7 and recovered at a rate of 0.89 +/- 0.15 pH/min in the nominal absence of Na+, HCO3-, organic anions, and SO4(2-) (lumen, bath). 1 mM N,N'-dicyclohexylcarbodiimide (lumen), 1 mM N-ethylmaleimide (lumen), 0.5 mM colchicine (bath), and 0.5 mM iodoacetic acid (lumen, bath) inhibited the Na+-independent pHi recovery rate by 73%, 55%, 77%, and 86%, respectively, whereas 1 mM KCN (lumen, bath) did not inhibit pHi recovery. Reduction of intracellular, but not extracellular chloride, also decreased the Na+-independent pHi recovery rate. In conclusion, the S3 proximal tubule has an apical Na+/H+ antiporter with a Michaelis constant for Na+ of 29 mM and a maximum velocity of 0.47 pH/min. S3 tubules also possess a plasma membrane H+-ATPase that can regulate pHi, has a requirement for intracellular chloride, and utilizes ATP derived primarily from glycolysis.     

Expression of rat renal Na/H antiporter mRNA levels in response to respiratory and metabolic acidosis.

The mammalian proximal tubule is an important mediator of the renal adaptive response to systemic acidosis. In chronic metabolic and respiratory acidosis the bicarbonate reabsorptive (or proton secretory) capacity is increased. This increase is mediated, at least in part, by an increase in Vmax of the luminal Na/H antiporter. To determine whether this adaptation involves increased mRNA expression, Na/H antiporter mRNA levels were measured by Northern analysis in renal cortex of rats with metabolic (6 mmol/kg body wt NH4Cl for 2 or 5 d) and respiratory (10% CO2/air balanced for 2 or 5 d) acidosis and of normal, pair-fed rats. Na/H antiporter mRNA levels were unchanged after 2 d of both metabolic and respiratory acidosis. After 5 d, however, Na/H antiporter mRNA expression was increased 1.76 +/- 0.12-fold in response to metabolic acidosis (P less than 0.005, n = 8), but was not different from normal in response to respiratory acidosis: 1.1 +/- 0.2 (NS, n = 8). Thus, the renal adaptive response to metabolic acidosis involves increased cortical Na/H antiporter mRNA levels. In contrast, the enhanced proximal tubule Na/H antiporter activity and bicarbonate reabsorption in respiratory acidosis seem to involve mechanisms other than increased Na/H antiporter gene expression.     

Parallel adaptation of the rabbit renal cortical sodium/proton antiporter and sodium/bicarbonate cotransporter in metabolic acidosis and alkalosis.

Recent studies have shown that the bicarbonate reabsorptive capacity of the proximal tubule is increased in metabolic acidosis. For net bicarbonate reabsorption to be regulated, there may be changes in the rate of apical H+ secretion as well as in the basolateral base exit step. The present studies examined the rate of Na+/H+ exchange (acridine orange method) and Na+/HCO3 cotransport (22Na uptake) in apical and basolateral membranes prepared from the rabbit renal cortex by sucrose density gradient centrifugation. NH4Cl loading was used to produce acidosis (arterial pH, 7.27 +/- 0.03), and Cl-deficient diet with furosemide was used to produce alkalosis (arterial pH, 7.51 +/- 0.02). Maximal transport rate (Vmax) of Na+/H+ antiporter and Na+/HCO3 cotransporter were inversely related with plasma bicarbonate concentration from 6 to 39 mM. Furthermore, the maximal transport rates of both systems varied in parallel; when Vmax for the Na+/HCO3 cotransporter was plotted against Vmax for the Na+/H+ antiporter for each of the 24 groups of rabbits, the regression coefficient (r) was 0.648 (P less than 0.001). There was no effect of acidosis or alkalosis on affinity for Na+ of either transporter. We conclude that both apical and basolateral H+/HCO3 transporters adapt during acid-base disturbances, and that the maximal transport rates of both systems vary in parallel during such acid-base perturbations.     

Proton secretion by the sodium/hydrogen ion antiporter in the human neutrophil.

The reducing equivalents used by the human neutrophil respiratory burst oxidase are derived from NADPH generated by the hexose monophosphate shunt. The CO2 generated by the HMP shunt is spontaneously hydrated and the protons (H+) are secreted upon the dissociation of carbonic acid. The mechanism and significance of H+ secretion by the resting and stimulated neutrophil was investigated. A basal rate of H+ secretion by resting neutrophils observed in a choline buffer was augmented with the addition of sodium (Na+) (Km for Na+ was 3.22 +/- 0.32 mM). Amiloride, a Na+/H+ antiporter inhibitor, reduced H+ secretion in Na+-containing buffers with a Ki = 1.02 microM. This Na+/H+ exchange mechanism was also operative in cells stimulated with a variety of agonists, and an increased H+ flux, relative to resting cells, was observed at higher Na+ concentrations. Cytoplasts incorporating acridine orange were also used to assess Na+-H+ flux. Cytoplasts were used to avoid alteration of the fluorescent pH probe by HOCl formed in intact neutrophils. Alkalinization of the cytoplasm was dependent on extracellular Na+ in concentrations similar to that found to augment H+ secretion in intact cells. Also, amiloride competitively inhibited H+ secretion by the cytoplasts. Both superoxide (O2-) production and lysozyme release in cells stimulated with opsonized zymosan or concanavalin A was significantly inhibited in the absence of Na+, restored to normal with the addition of Na+ in low concentrations, and inhibited again in the presence of amiloride. A Na+/H+ antiporter similar to that found in other cell types is present in the human neutrophil and appears linked to activation of the respiratory burst and degranulation.     

Neonatal rabbit juxtamedullary proximal convoluted tubule acidification.

The present in vitro microperfusion study examined apical membrane Na+/H+ antiporter and basolateral membrane Na(HCO3)3 symporter activity in newborn and adult juxtamedullary proximal convoluted tubules. Proton fluxes were determined from the initial rate of change of intracellular pH after a change in the luminal or bathing solution, buffer capacity, and tubular volume of newborn and adult tubules. Intracellular pH (pHi) was measured fluorometrically using the pH-sensitive dye (2',7')-bis (carboxyethyl)-(5,6)-carboxyfluorescein (BCECF). Apical Na+/H+ antiporter proton flux, assayed by the effect of sodium removal (147----0 meq/liter) on pHi, was one-third the adult level for the first 2 wk and doubled in the 3rd wk of life. Adult levels were achieved by 6 wk of age. Na+/H+ antiporter activity was not detected on the basolateral membrane of 1-wk-old newborns, indicating that polarity of this transporter was already present. Basolateral membrane Na(HCO3)3 proton flux, assayed by the effect of a bath bicarbonate change (25----5 meq/liter) and by a bath sodium change (147----0 meq/liter) on pHi, was 50-60% of adult values in 1-wk-old newborns. Basolateral membrane Na(HCO3)3 proton flux assayed by a bath bicarbonate change (25----5 meq/liter) remained at 50-60% of adult values for the 1st mo of life and increased to adult levels by 6 wk of age. This transporter not only plays a role in net acidification, but is an important determinant of cell pH in newborn juxtamedullary proximal convoluted tubules.     

Novel amiloride-sensitive sodium-dependent proton secretion in the mouse proximal convoluted tubule

The proximal convoluted tubule (PCT) reabsorbs most of the filtered bicarbonate. Proton secretion is believed to be mediated predominantly by an apical membrane Na(+)/H(+) exchanger (NHE). Several NHE isoforms have been cloned, but only NHE3 and NHE2 are known to be present on the apical membrane of the PCT. Here we examined apical membrane PCT sodium-dependent proton secretion of wild-type (NHE3(+/+)/NHE2(+/+)), NHE3(-/-), NHE2(-/-), and double-knockout NHE3(-/-)/NHE2(-/-) mice to determine their relative contribution to luminal proton secretion. NHE2(-/-) and wild-type mice had comparable rates of sodium-dependent proton secretion. Sodium-dependent proton secretion in NHE3(-/-) mice was approximately 50% that of wild-type mice. The residual sodium-dependent proton secretion was inhibited by 100 microM 5-(N-ethyl-N-isopropyl) amiloride (EIPA). Luminal sodium-dependent proton secretion was the same in NHE3(-/-)/NHE2(-/-) as in NHE3(-/-) mice. These data point to a previously unrecognized Na(+)-dependent EIPA-sensitive proton secretory mechanism in the proximal tubule that may play an important role in acid-base homeostasis.     

Mechanisms of adaptation to chronic respiratory acidosis in the rabbit proximal tubule.

The hyperbicarbonatemia of chronic respiratory acidosis is maintained by enhanced bicarbonate reabsorption in the proximal tubule. To investigate the cellular mechanisms involved in this adaptation, cell and luminal pH were measured microfluorometrically using (2",7')-bis(carboxyethyl)-(5,6)-carboxyfluorescein in isolated, microperfused S2 proximal convoluted tubules from control and acidotic rabbits. Chronic respiratory acidosis was induced by exposure to 10% CO2 for 52-56 h. Tubules from acidotic rabbits had a significantly lower luminal pH after 1-mm perfused length (7.03 +/- 0.09 vs. 7.26 +/- 0.06 in controls, perfusion rate = 10 nl/min). Chronic respiratory acidosis increased the initial rate of cell acidification (dpHi/dt) in response to luminal sodium removal by 63% and in response to lowering luminal pH (7.4-6.8) by 69%. Chronic respiratory acidosis also increased dpHi/dt in response to peritubular sodium removal by 63% and in response to lowering peritubular pH by 73%. In conclusion, chronic respiratory acidosis induces a parallel increase in the rates of the luminal Na/H antiporter and the basolateral Na/(HCO3)3 cotransporter. Therefore, the enhanced proximal tubule reabsorption of bicarbonate in chronic respiratory acidosis may be, at least in part, mediated by a parallel adaptation of these transporters.     

Effects of extracellular fluid volume and plasma bicarbonate concentration on proximal acidification in the rat.

The effects of systemic bicarbonate concentration and extracellular fluid volume status on proximal tubular bicarbonate absorption, independent of changes in luminal composition and flow rate, were examined with in vivo luminal microperfusion of rat superficial proximal convoluted tubules. Net bicarbonate absorption and bicarbonate permeability were measured using microcalorimetry. From these data, net bicarbonate absorption was divided into two parallel components: proton secretion and passive bicarbonate diffusion. The rate of net bicarbonate absorption was similar in hydropenic and volume-expanded rats when tubules were perfused with 24 mM bicarbonate, but was inhibited in volume-expanded rats when tubules were perfused with 5 mM bicarbonate. Volume expansion caused a 50% increase in bicarbonate permeability, which totally accounted for the above inhibition. The rate of proton secretion was unaffected by volume expansion in both studies. The rate of net bicarbonate absorption was markedly inhibited in alkalotic expansion as compared with isohydric expansion. Bicarbonate permeabilities were not different in these two conditions, and the calculated rates of proton secretion were decreased by greater than 50% in alkalosis. Net bicarbonate absorption was stimulated in acidotic rats compared to hydropenic rats. This stimulation was attributable to a 25% increase in the rate of proton secretion. We conclude that (a) proton secretion is stimulated in acidosis, inhibited in alkalosis, and is not altered by volume status; (b) bicarbonate permeability is increased by volume expansion but is not altered by increases in plasma bicarbonate concentration; (c) when luminal bicarbonate concentrations are similar to those of plasma, net bicarbonate absorption is dominated by proton secretion and is thus sensitive to peritubular bicarbonate concentrations, and insensitive to extracellular fluid volume; (d) when luminal bicarbonate concentrations are low and proton secretion is slowed, bicarbonate permeability and thus extracellular fluid volume have a greater influence on net bicarbonate absorption.     

Contributions of cellular leak pathways to net NaHCO3 and NaCl absorption.

Proton and formic acid permeabilities were measured in the in vivo microperfused rat proximal convoluted tubule by examining the effect on intracellular pH when [H] and/or [formic acid] were rapidly changed in the luminal or peritubular fluids. Apical and basolateral membrane H permeabilities were 0.52 +/- 0.07 and 0.67 +/- 0.18 cm/s, respectively. Using these permeabilities we calculate that proton backleak from the luminal fluid to cell does not contribute significantly to net proton secretion in the early proximal tubule, but may contribute in the late proximal tubule. Apical and basolateral membrane formic acid permeabilities measured at extracellular pH 6.62 were 4.6 +/- 0.5 X 10(-2) and 6.8 +/- 1.5 X 10(-2) cm/s, respectively. Control studies demonstrated that the formic acid permeabilities were not underestimated by either the simultaneous movement of formate into the cell or the efflux of formic acid across the opposite membrane. The measured apical membrane formic acid permeability is too small to support all of transcellular NaCl absorption in the rat by a mechanism that involves Na/H-Cl/formate transporters operating in parallel with formic acid nonionic diffusion.     

Basolateral membrane Na+/H+ antiport, Na+/base cotransport, and Na+-independent Cl-/base exchange in the rabbit S3 proximal tubule.

The basolateral membrane Na+ and Cl(-)-dependent acid-base transport processes were studied in the isolated perfused rabbit S3 proximal straight tubule. Intracellular pH (pHi) was measured with 2'7'-biscarboxyethyl-5,6-carboxyfluorescein (BCECF) and a microfluorometer coupled to the tubule perfusion apparatus. Reduction of basolateral HCO3- from 25 to 5 mM caused pHi to decrease at a rate of 0.81 pH/min. Approximately 50% of this rate was Na+-dependent, 30% Cl(-)-dependent and 20% Na+ and Cl(-)-independent. Two basolateral Na+-dependent acid base transport pathways were detected: (a) an amiloride-sensitive Na+/H+ antiporter and (b) a stilbene-sensitive Na+/base cotransporter. No evidence was found for a Na+-dependent Cl-/base exchanger. The Cl(-)-dependent component of basolateral base efflux was mediated by a stilbene-sensitive Na+-independent Cl-/base exchange pathway. The results suggest that the acid base transport pathways of the basolateral membrane of the S3 proximal tubule differ from more proximal nephron segments.     

Chronic metabolic acidosis causes an adaptation in the apical membrane Na/H antiporter and basolateral membrane Na(HCO3)3 symporter in the rat proximal convoluted tubule.

The effect of chronic dietary acid on the apical membrane Na/H antiporter and basolateral membrane Na(HCO3)3 symporter was examined in the in vivo microperfused rat proximal tubule. Transporter activity was assayed with the epifluorescent measurement of cell pH using the intracellular, pH-sensitive fluorescent dye, (2'7')-bis(carboxyethyl)-(5,6)-carboxy-fluorescein (BCECF). BCECF was calibrated intracellularly, demonstrating similar pH-sensitivity of the dye in control and acidotic animals. In subsequent studies, lumen and peritubular capillaries were perfused to examine Na/H and Na(HCO3)3 transporter activity in the absence of contact with native fluid. The initial rate of change in cell pH (dpHi/dt) was 97, 50, and 44% faster in tubules from acidotic animals when peritubular [HCO3] was changed from 25 to 10 mM in the presence or absence of chloride, or peritubular [Na] was changed from 147 to 50 mM, respectively. dpHi/dt was 57% faster in tubules from acidotic animals when luminal [Na] was changed from 152 to 0 mM. Buffer capacities, measured using NH3/NH+4 addition, were similar in the two groups. The results demonstrate that chronic metabolic acidosis causes an adaptation in the intrinsic properties of both the apical membrane Na/H antiporter and basolateral membrane Na(HCO3)3 symporter.     

Distal tubule bicarbonate reabsorption in NH4Cl acidotic rats

NH4Cl acidosis--a common experimental model of hyperchloremic metabolic acidosis--elicits complex intrarenal responses whereby the fall in plasma bicarbonate concentration can be restored to normal after the initial acid load. Using the technique of in vivo micropuncture of surface distal tubules of the rat kidney, we attempted to further define controlling mechanisms underlying the enhanced bicarbonate reabsorption in this setting. Specifically, we wished to determine the dependence of distal tubule bicarbonate reabsorption (JtCO2) on sodium transport, water reabsorption, and carbonic anhydrase activity. Surface distal tubules of Sprague-Dawley rats made acidotic by ammonium chloride gavage (arterial blood pH: 7.15 +/- 0.01, [HCO3]: 14.8 +/- 0.5 mM) were perfused in vivo at 8 and 24 nL/min with 4 different isoosmotic, 25 mM bicarbonate solutions: Group 1 was perfused with 60 mM Na, Group 2 with 60 mM choline, Group 3 with 60 mM choline + 3 x 10(-4) M amiloride, and Group 4 with 60 mM Na + 10(-3) M acetazolamide. At 8 nL/min, significant bicarbonate reabsorption occurred with all perfusates. JtCO2 was 65 +/- 4, 59 +/- 5, 58 +/- 6, and 40 +/- 4 pmol.min-1.mm-1, in Groups 1, 2, 3, and 4, respectively. However, JtCO2 in Group 4 was significantly less than that in Groups 1 and 2 (p less than 0.01 and p less than 0.05, respectively). Amiloride added to the low sodium perfusate did not reduce bicarbonate reabsorption. We conclude that bicarbonate reabsorption in distal tubules of acidotic rats is acetazolamide-sensitive and is not significantly sustained by sodium or water movements.     

Induction of hypertrophy in cultured proximal tubule cells by extracellular NH4Cl.

Ammonia production increases in several models of renal hypertrophy in vivo. The present study was designed to determine whether ammonia can directly modulate the growth of renal cells in the absence of a change in extracellular acidity. In serum-free media NH4Cl (0-20 mM) caused JTC cells and a primary culture of rabbit proximal tubule cells to hypertrophy (increase in cell protein content) in a dose-dependent fashion without a change in DNA synthesis. Studies in JTC cells revealed that the cell protein content increased as a result of both an increase in protein synthesis and a decrease in protein degradation. Total cell RNA content and ribosome number increased after NH4Cl exposure and the cell content of the lysosomal enzymes cathepsin B and L decreased. Inhibition of the Na+/H+ antiporter with amiloride did not prevent the hypertrophic response induced by NH4Cl. The results indicate that ammonia is an important modulator of renal cell growth and that hypertrophy can occur in the absence of functioning Na+/H+ antiport activity.     

Synergistic alpha-1 and alpha-2 adrenergic stimulation of rat proximal nephron Na+/H+ exchange

Both alpha-1 and alpha-2 adrenoceptors have been localized to the renal cortex, with the majority of binding sites on the proximal tubule. Because the major regulator of Na+ uptake into the proximal tubule is the Na+/H+ exchanger, and because alpha-1 and alpha-2 adrenoceptors stimulate it in other tissues, we tested the hypothesis that both alpha adrenoceptor subtypes can increase Na+ uptake into the proximal nephron by stimulating the Na+/H+ antiporter. Enhancement of Na+ transport by agonists was studied in isolated rat proximal tubules by determining the uptake of 22Na that was suppressible by the Na+/H+ inhibitor, 5-(N-ethyl-N-isopropyl)amiloride (EIPA). The phorbol ester, phorbol-12-myristate-13-acetate, (0.1 microM), directly stimulated the antiporter through protein kinase C and increased EIPA-suppressible 22Na uptake 250% above control. The alpha-1 adrenoceptor agonists, cirazoline and phenylephrine, in addition to the mixed agonist, norepinephrine, maximally stimulated uptake by 226 to 232% at 1 microM concentrations. alpha-2 agonists produced a range of maximal stimulations at 1 microM from 65% with guanabenz to 251% with B-HT 933. Increases in 22Na uptake by agonists were inhibited by selective adrenergic antagonists and by EIPA. The drugs did not change the EIPA-resistant component of 22Na uptake. Inasmuch as the adrenoceptor subtypes likely stimulated Na+/H+ exchange by differing intracellular pathways impinging upon common transport steps, we examined whether simultaneous stimulation of both pathways was additive. Submaximal concentrations (5 nM each) of alpha-1 and alpha-2 adrenoceptor agonists in combination synergistically enhanced 22Na uptake to a level similar to 1 microM concentrations of adrenoceptor agonists alone or in combination.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of amiloride and 5-(N,N-hexamethylene)amiloride on dopaminergic and cholinergic neurotransmission

In order to establish the role of the Na+/H+ exchange transport on neurotransmission, we investigated the effects of amiloride and of 5-(N,N-hexamethylene)amiloride (HMA) on dopamine (DA) and acetylcholine (ACh) release and on receptor-mediated modulation of DA and ACh release. Superfused rabbit striatal slices prelabeled with [3H]DA and [14C]choline were stimulated electrically in the presence and absence of several concentrations of these agents. Amiloride (3-10 microM) and HMA (0.3-10 microM) reduced the basal efflux and the stimulation evoked overflow of total 3H and of [3H]-3,4-dihydroxyphenylacetic acid and inhibited monoamine oxidase activity. The inhibition of stimulation evoked overflow of total 3H was blocked by pretreatment with nomifensine but not by sulpiride. Amiloride had no effect on the basal efflux and the stimulation evoked overflow of ACh or it did modify apomorphine-induced inhibition of DA and ACh release. However, at 3 to 10 microM, HMA enhanced the basal efflux of 3H; this effect was not prevented either by uptake inhibition with nomifensine or by low extracellular calcium. These results suggest that amiloride-sensitive Na+ transport and the amiloride and HMA-sensitive Na+/H+ antiporter play no role on the secretion of DA and ACh, or on the mechanisms by which activation of pre- and postsynaptic DA receptors lead to inhibition of neurotransmitter release. Amiloride- and HMA-induced monoamine oxidase inhibition accounts for the effects of amiloride and HMA on DA efflux and overflow. The guanidine moiety present in the amiloride and HMA molecules is most likely responsible for these effects.     

Acid-base disturbance in adrenal and parathyroid diseases]

Aldosterone stimulates sodium absorption in the collecting tubule, which makes the potential more lumen-negative, and secondarily stimulates proton and potassium secretion. Potassium depletion stimulates ammonia synthesis, which increases distal buffer delivery, increases luminal pH in the collecting tubule, and secondarily stimulates proton secretion. This stimulation of the distal nephron acidification by aldosterone can result in metabolic alkalosis. Inversely, aldosterone deficiency can produce characteristic metabolic acidosis with hyperkalemia (type IV renal tubular acidosis). Acute administration of parathyroidhormone (PTH) decreases renal bicarbonate reabsorption in the proximal tubule. Since the effect of PTH is overridden by other factors, such as PTH-induced base release from bone, hyperparathyroidism does not uniformly cause metabolic acidosis.