Influence of bile acids on the (Na+−K+)-activated- and Mg2+-activated ATPase of rat colon

Summary The influence of various bile acids on the (Na⁺−K⁺)-ATPase and Mg²⁺-ATPase activity of rat colon is described. At a concentration of 0.6 mmol/l C and TC did not inhibit the (Na⁺−K⁺)-ATPase activity in contrast to GC. The taurine derivates TC, TCDC and TDC did not influence or even enhanced the (Na⁺−K⁺)-ATPase activity. All bile acids except C, TC and CDC depressed the Mg²⁺-ATPase activity. At higher concentrations only C and TC did not influence the (Na⁺−K⁺)-ATPase activity. C, GC and TC at 2.5 mmol/l decreased the (Na⁺−K⁺)-activated phosphatase with ATP as substrate. All other substrates tested did not influence the enzymic activity significantly. The results indicate that bile acids can inhibit the Na⁺-absorbing system in rat colon. Hence this inhibition can cause diarrhea.     

Occurrence and properties of a (Na+−K+)-activated ATPase in the mucosa of the rat intestine

Summary The existence of an ouabain-sensitive (Na⁺–K⁺)-activated ATPase system has been demonstrated in the total intestine of the rat. The (Na⁺–K⁺)-ATPase activity was about 10–15% of the total ATPase in 4 equal parts of the small intestine; in the colon about 35% of the total ATPase was (Na⁺–K⁺)-activated ATPase. The highest (Na⁺–K⁺)-ATPase activity has been observed in the first and second part of the small intestine, while in the colon the activity was 2–4 times higher than in the ileum.The (Na⁺–K⁺)-ATPase of rat colon required both Na⁺ (K _m=8.3 mmoles/l) and K⁺ (K _m=0.6 mmoles/l). Maximal activation of the (Na⁺–K⁺)-ATPase system required 2 mM Mg²⁺ at an ATP concentration of 2 mM. The pH optimum for (Na⁺–K⁺)-ATPase of rat colon was 7.5, while the Mg²⁺-activated ATPase activity had a pH optimum of 8.6. The (Na⁺–K⁺)-ATPase was inhibited by ouabain (pI ₅₀=3.6).The relation between the differences in (Na⁺–K⁺)-ATPase activity and Na⁺-absorption on different parts of the intestine is discussed.     

Activity of (Na+K+)-stimulated adenosintriphosphatase in the rat nephron

Summary In 17 male Wistar rats in antidiuresis 10 different nephron segments and arteries are identified with the aid of Lowry's technique, dissected and total-and (Mg⁺⁺)-adenosintriphosphatase (=ATPase) determined. (Na⁺K⁺)-activated ATPase in the distal tubule is four to five times (max. eight times) more active than in the proximal segment. This difference of activity may speak for a high pump mechanism mediated by the way of a (Na⁺K⁺)-activated enzyme system in the distal nephron and for a partially passive reabsorption of sodium from the proximal convolution.With the support of the Schweiz. Nationalfonds zur Förderung der wissenschaftlichen Forschung (Nr. 4256 and Nr. 4809.3)     

Effect of olsalazine and mesalazine on human ileal and colonic (Na+ + K+)-ATPase

Summary Olsalazine (azodisalicylate) and mesalazine (5-aminosalicylic acid) have recently been developed as new treatment modalities for inflammatory bowel disease to avoid sulfasalazine-related side effects. However, there are reports regarding new and hitherto unexpected side effects in some patients receiving olsalazine or mesalazine, such as watery diarrhea. Since sodium pump activities play an important role in the pathogenesis of water and electrolyte disturbances, we investigated the influence of olsalazine and mesalazine on human ileal and colonic (Na⁺ + K⁺)-ATPase and its specific [³H]-ouabain binding. We found a concentration-dependent inhibition of ileal and colonic (Na⁺ + K⁺)-ATPase by olsalazine with an IC₅₀ of 4.1 mM and 4.8 mM, respectively. Mesalazine inhibited this enzyme in the ileum with an IC₅₀ of 4.5 mM and in the sigmoid colon with an IC₅₀ 3.5 mM. In addition, [³H]-ouabain binding was inhibited by mesalazine with an IC₅₀ of 3.6 mM. The maximal inhibition, however, did not exceed 80% under any conditions (up to 10 mM drug concentration). Olsalazine and mesalazine induce inhibition of the ileal and colonic sodium pump activities that may (in addition to other possible mechanisms) mediate impaired water and electrolyte absorption. This is possibly of clinical relevance in patients with severely damaged mucosa. In patients with milder forms of mucosal inflammation, this inhibition most likely is of minor importance because of the great capacitiy of the (Na⁺ + K⁺)-ATPase and the incomplete inhibition leaving at least 20% of the enzyme activity intact.Abbreviations 5-ASA 5-aminosalicylic acid - EDTA ethylenediaminetetracetic acid - IBD inflammatory bowel disease     

Intracellular Na+ modulates large conductance Ca2+-activated K+ currents in human umbilical vein endothelial cells

We studied the effects of Na⁺ influx on large-conductance Ca²⁺-activated K⁺ (BK_Ca) channels in cultured human umbilical vein endothelial cells (HUVECs) by means of patch clamp and SBFI microfluorescence measurements. In current-clamped HUVECs, extracellular Na⁺ replacement by NMDG⁺ or mannitol hyperpolarized cells. In voltage-clamped HUVECs, changing membrane potential from 0 mV to negative potentials increased intracellular Na⁺ concentration ([Na⁺]_i) and vice versa. In addition, extracellular Na⁺ depletion decreased [Na⁺]_i. In voltage-clamped cells, BK_Ca currents were markedly increased by extracellular Na⁺ depletion. In inside-out patches, increasing [Na⁺]_i from 0 to 20 or 40 mM reduced single channel conductance but not open probability (NPo) of BK_Ca channels and decreasing intracellular K⁺ concentration ([K⁺]_i) gradually from 140 to 70 mM reduced both single channel conductance and NPo. Furthermore, increasing [Na⁺]_i gradually from 0 to 70 mM, by replacing K⁺, markedly reduced single channel conductance and NPo. The Na⁺–Ca²⁺ exchange blocker Ni²⁺ or KB-R7943 decreased [Na⁺]_i and increased BK_Ca currents simultaneously, and the Na⁺ ionophore monensin completely inhibited BK_Ca currents. BK_Ca currents were significantly augmented by increasing extracellular K⁺ concentration ([K⁺]_o) from 6 to 12 mM and significantly reduced by decreasing [K⁺]_o from 12 or 6 to 0 mM or applying the Na⁺–K⁺ pump inhibitor ouabain. These results suggest that intracellular Na⁺ inhibit single channel conductance of BK_Ca channels and that intracellular K⁺ increases single channel conductance and NPo. GH Liang and MY Kim contributed equally to this publication and therefore share the first authorship.     

Effect of high NaCl intake on Na+ and K+ transport in the rabbit distal convoluted tubule

Morphological studies have demonstrated that a chronic increase in distal Na⁺ delivery causes hypertrophy of the distal convoluted tubule (DCT). To examine whether high NaCl-intake also causes functional changes in the well defined DCT, we measured transmural voltage (V _T), lumen-to-bath Na⁺ flux (J _Na(LB)), and net K⁺ secretion (J _K(net)) in DCTs obtained from control rabbits and those on high NaCl-intake diets. The lumen negativeV _T was significantly greater in the high NaCl group than in the control group. The net K⁺ secretion (pmol mm^–1 min^–1) was greater in the high NaCl-intake group (54.1±13.0 vs 14.7±5.6). The K⁺ permeabïlities in both luminal and basolateral DCT membranes, as assessed by the K⁺-induced transepithelial voltage deflection inhibitable with Ba²⁺, were increased in the experimental group. The lumen-to-bath²²Na flux (pmol mm^–1 min^–1) was also greater in the experimental group (726±119 vs 396±65). TheV _T component inhibitable with amiloride was also elevated in the high NaCl-intake group. Furthermore, Na⁺–K⁺-ATPase activity of the DCT was higher in the experimental than in the control group. We conclude that high NaCl intake increases both Na⁺ reabsorption and K⁺ secretion by the DCT. This phenomenon is associated with an increased Na⁺–K⁺-ATPase activity along with increased Na⁺ and K⁺ permeabilities of the luminal membrane, and an increase in the K⁺ permeability of the basolateral membrane. Cellular mechanisms underlying these functional changes remain to be established.     

Copper toxicity in cultured human skeletal muscle cells: the involvement of Na+/K+-ATPase and the Na+/Ca2+-exchanger

Copper (Cu²⁺) intoxication has been shown to induce pathological changes in various tissues. The mechanism underlying Cu²⁺ toxicity is still unclear. It has been suggested that the Na⁺/K⁺-ATPase and/or a change of the membrane permeability may be involved. In this study we examined the effects of Cu²⁺ on the Na⁺ and Ca²⁺ homeostasis of cultured human skeletal muscle cells using the ion-selective fluorescent probes Na⁺-binding benzofuran isophtalate (SBFI) and Fura-2, respectively. In addition, we measured the effect of Cu²⁺ on the Na⁺/K⁺-ATPase activity. Cu²⁺ and ouabain increase the cytoplasmic free Na⁺ concentration ([Na⁺]_i). Subsequent addition of Cu²⁺ after ouabain does not affect the rate of [Na⁺]_i increase. Cu²⁺ inhibits the Na⁺/K⁺-ATPase activity with an IC₅₀ of 51 M. The cytoplasmic free Ca²⁺ concentration ([Ca²⁺]_i) remains unaffected for more than 10 min after the administration of Cu²⁺. Thereafter, [Ca²⁺]_i increases as a result of the Na⁺/Ca²⁺-exchanger operating in the reversed mode. The effects of Cu²⁺ on the Na⁺ homeostasis are reversed by the reducing and chelating agent dithiothreitol and the heavy metal chelator N,N,N,N,-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN). In conclusion, SBFI is a good tool to examine Na⁺ homeostasis in cultured human skeletal muscle cells. Under the experimental conditions used, Cu²⁺ does not modify the general membrane permeability, but inhibits the Na⁺/K⁺-pump leading to an increase of [Na⁺]_i. As a consequence the operation mode of the Na⁺/Ca²⁺-exchanger reverses and [Ca²⁺]_i rises.The authors thank staff and coworkers of the Department of Neurology of the University Hospital Nijmegen, Nijmegen for their kind cooperation in obtaining muscle biopsies. Mr. Arie Oosterhof is gratefully acknowledged for culturing of the human muscle cells. The Prinses Beatrix Fonds and the Dutch-Chinese scientific exchange program contributed financial support for this study.     

Human colonic (Na++K+)-ATPase and specific ouabain binding are not influenced by lipoxygenase products or superoxide radicals

Summary The effects of lipoxygenase products (5-, 12-, 15-HETE, LTB₄) and superoxide radicals on human colonic (Na⁺+K⁺)-ATPase and specific ouabain binding were measured. No significant inhibition in concentrations up to 3 × 10^–5 M was observed. The results are discussed with regard to a possible role of lipoxygenase products and radicals in the pathogenesis of water and electrolyte disturbances in various diarrheal states including inflammatory bowel disease.Abbrevations IBD Inflammatory bowel disease - HETE Hydroxytetraenoic acid - LTB₄ Leukotriene B₄ Supported by DFG (Er 65/4-4)     

Influence of Mitragyna Ciliata (MYTA) on the Microsomal Activity of ATPase Na+/K+ Dependent Extract on a Rabbit Heart

Mitragyna ciliata (MYTA) (Rubiaceae) inhibits plasmodia activity. MYTA induces a cardiotonicity of the digitalic type on rat''s isolated heart. In this work we studied the effect of MYTA on microsomal Na⁺/K⁺ dependant ATPase (Na⁺, K⁺ ATPase) extracted from the heart of a rabbit since digitalics inhibit Na⁺, K⁺ ATPase. Our results revealed that the Na⁺/K⁺ ATPase has an optimum pH of 7.4 and temperature of 37°C respectively. There is a linear relationship between the organic phosphate formed and the incubation time over 25 mins incubation period. The ATP hydrolysis rate in the presence of MYTA was 0.775 µM/min. LINEWEAVER and BURK plots showed that MYTA did not alter K_M (1.31 mM) but decreased V_MAX. This study shows that MYTA exerts a non-competitive inhibition on the microsomal Na⁺/K⁺ ATPase extracted from rabbit heart with a Ci₅₀ of 48 µg / ml. We conclude that the mechanism of action of MYTA is linked to the inhibition of the Na⁺/K⁺ ATPase like cardiotonics of the digitalic type.     

Age differences in hormonal regulation of Na+, K+-ATPase activity in the rat renal cortex

Laboratory of Physiological Genetics, Institute of Cytology and Genetics, Siberian Brach, Russian Academy of Sciences, Novosibirsk. (Presented by Academician of the Russian Academy of Medical SciencesV. P. Lozov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 114, No. 8, pp. 150–153, August, 1992.     

Effect of extracellular volume expansion on renal cortical and medullary Na+−K+-ATPase

Summary The role of renal Na⁺–K⁺-ATPase in the acute changes in sodium reabsorption caused by isotonic volume expansion was evaluatedin vivo andin vitro in the rat and the dog. Duringin vivo volume expansion with isotonic saline in the rat, renal medullary Na⁺–K⁺-ATPase specific activity increased, while the simultaneously determined cortical Na⁺–K⁺-ATPase specific activity and kinetics remained unchanged. Furthermore, experimentsin vitro failed to demonstrate a circulating inhibitor of renal Na⁺–K⁺-ATPase both in plasma dialysates from volume-expanded rats and in plasma dialysates concentrated 20-fold by ultrafiltration from volume-expanded dogs. These results suggest that the decreased proximal tubular reabsorption of sodium during volume expansion is not mediated by inhibition of renal cortical Na⁺–K⁺-ATPase. The acute increment in medullary Na⁺–K⁺-ATPase observed could represent an adaptive response to increased sodium reabsorption by the loops of Henle, and raises the possibility that this enzyme may participate in relatively rapid adjustments in the transport of sodium by the renal tubule.     

Electrogenic Na+/K+-transport in human endothelial cells

Na⁺/K⁺ pump currents were measured in endothelial cells from human umbilical cord vein using the whole-cell or nystatin-perforated-patch-clamp technique combined with intracellular calcium concentration ([Ca²⁺]_i) measurements with Fura-2/AM. Loading endothelial cells through the patch pipette with 40 mmol/l [Na⁺] did not induce significant changes of [Ca²⁺]_i. Superfusing the cells with K⁺-free solutions also did not significantly affect [Ca²⁺]_i. Reapplication of K⁺ after superfusion of the cells with K⁺-free solution induced an outward current at a holding potential of 0 mV. This current was nearly completely blocked by 100 mol/l dihydroouabain (DHO) and was therefore identified as a Na⁺/K⁺ pump current. During block and reactivation of the Na⁺/K⁺ pump no changes in [Ca²⁺]_i could be observed. Pump currents were blocked concentration dependently by DHO. The concentration for half-maximal inhibition was 21 mol/l. This value is larger than that reported for other tissues and the block was practically irreversible. Insulin (10–1000 U/l) did not affect the pump currents. An increase of the intracellular Na⁺ concentration ([Na⁺]_i) enhanced the amplitude of the pump current. Half-maximal activation of the pump current by [Na⁺]_i occurred at about 60 mmol/l. The concentration for half-maximal activation by extracellular K⁺ was 2.4±1.2 mmol/l, and 0.4±0.1 and 8.7±0.7 mmol/l for Tl⁺ and NH₄ ⁺ respectively. The voltage dependence of the DHO-sensitive current was obtained by applying linear voltage ramps. Its reversal potential was more negative than –150 mV. Pump currents measured with the conventional whole-cell technique were about four times smaller than pump currents recorded with the nystatin-perforated-patch method. If however 100 mol/l guanosine 5-O-(3-thiotriphosphate) (GTPS) were added to the pipette solution, the currents measured in the ruptured-whole-cell-mode were not significantly different from the currents measured with the perforated-patch technique. We suppose that the use of the perforated-patch technique prevents wash out of a guanine nucleotide-binding protein (G-protein)-connected intracellular regulator that is necessary for pump activation.     

Substructure of membrane-bound Na+−K+-ATPase protein

Purified membrane-bound Na⁺–K⁺-ATPase from rat kidney outer medulla was studied by freeze-fracturing, by freeze-etching and by negative staining. Freeze-fracturing of purified Na⁺–K⁺-ATPase membranes shows intramembraneous particles with a diameter of about 100 Å. The frequency of these intramembraneous particles — as estimated from the particle densities on the two fracture faces — lies between 4700 and 5600 particles per m². Applying rotary shadowing a four partite substructure could be detected in these intramembraneous particles observed on the fracture planes. The same four partite substructure was detected in particles observed on freeze-fractured and rotary shadowed intact baso-lateral plasma membranes of the thick ascending limb of Henle's loop. Particles could be also detected on both membrane surfaces of the purified Na⁺–K⁺-ATPase. These surface particles have about the same diameter and are present at about the same frequency as those observed within the freeze-fractured membranes. Negative staining of isolated Na⁺–K⁺-ATPase membranes showed particles on both membrane surfaces with a diameter between 30 and 50 Å, at a frequency of about 19,000 per m². On aspects of membrane edges we observed structures which suggest a transmembraneous connection of the negatively stained particles on both membrane surfaces.Our results suggest that the Na⁺–K⁺-ATPase protein is composed of four units and that each unit spans the cell membrane. The native enzyme structure of the Na⁺–K⁺-ATPase protein seems to be preserved during freeze-fracturing and freeze-etching. It is proposed that the four enzyme units of the Na⁺–K⁺-ATPase complex are dissociated during the negative staining procedure.Part of this work was presented at the Frühjahrstagung of the Deutsche Physiologische Gesellschaft [6]     

Evidence for coupled sodium/hydrogen exchange in the rat superficial proximal convoluted tubule

Recent in vitro studies from the rat and rabbit have suggested a tightly coupled sodium/hydrogen ion exchanger on the luminal membrane of proximal tubules. The steep sodium gradient from the lumen to cell supplies indirect energy for hydrogen ions to be pumped from the cell to the lumen. However, a proton translocating pump has been demonstrated in other epithelia, which is independent of sodium transport and directly driven by ATP. To examine the role that sodium might play in the process of acidification, rat proximal convoluted tubules and their surrounding peritubular capillaries were perfused in vivo with artificial ultrafiltrate-like perfusion solutions. Total CO₂ absorption was measured by microcalorimetry during alterations in sodium transport by replacement of the sodium with an impermeant cation, choline, or by inhibition of the (Na⁺+K⁺)-ATPase by removing potassium from both perfusion solutions. Under control conditions the absolute rate of total CO₂ absorption was 140 pmol/mm·min. In the choline substitution and potassium removal experiments, absolute total CO₂ absorption fell to 23 and 28 pmol/mm·min, respectively. The data suggest that: 1) in the rat superficial proximal convoluted tubule approximately 80% of the bicarbonate absorption is tightly coupled to sodium transport; 2) this process is driven indirectly by the (Na⁺+K⁺)-ATPase system; and 3) the residual 20% of acidification appears to be mediated by another mechanism or may be a consequence of technical liminations.     

Training in inhibitory avoidance causes a reduction of Na+,K+-ATPase activity in rat hippocampus

Compelling evidence has indicated the involvement of Na(+),K(+)-ATPase in the mechanisms of synaptic plasticity. In the present study, we investigated the effect of inhibitory avoidance training on Na(+),K(+)-ATPase activity, at different times after training, in the rat hippocampus. Male adult Wistar rats were trained in a step-down inhibitory avoidance task and compared to those submitted to isolated footshock (0.4 mA) or placed directly onto the platform. Na(+),K(+)-ATPase activity decreased, by 60%, in hippocampus of rats sacrificed immediately after the isolated footshock, as well as immediately (0 min) and 6 h after training; this effect was not present 24 h after training. We also verified that enzyme activity was not altered in rats killed after just being on the platform. These findings suggest that Na(+),K(+)-ATPase activity may be involved in the memory consolidation of step-down inhibitory avoidance in the hippocampus.     

Erythrocyte cations and Na+,K+-ATPase pump activity in athletes and sedentary subjects

Summary The chronic effect of training on intraerythrocyte cationic concentrations and on red cell Na⁺,K⁺-ATPase pump activity was studied by comparing well-trained athletes with sedentary subjects at rest. Also the acute effect of a 50-min cross-country run on these erythrocyte measurements was studied in the athletes. At rest the intraerythrocyte potassium concentration was increased (P<0.01) in the athletes compared to that of the control subjects. The intraerythrocyte concentrations of sodium and magnesium and red cell Na⁺, K⁺-ATPase pump activity were, however, similar in the trained and the untrained subjects.As compared with the resting condition, the intraerythrocyte potassium concentration was decreased (P<0.05) after exercise in the athletes, and this was accompanied by a minor increase in the intraerythrocyte sodium concentration. Red cell Na⁺,K⁺-ATPase pump activity was slightly increased (P<0.05) after exercise.     

Interspersed normoxia during live high, train low interventions reverses an early reduction in muscle Na+, K+ATPase activity in well-trained athletes

Hypoxia and exercise each modulate muscle Na⁺, K⁺ATPase activity. We investigated the effects on muscle Na⁺, K⁺ATPase activity of only 5 nights of live high, train low hypoxia (LHTL), 20 nights consecutive (LHTLc) versus intermittent LHTL (LHTLi), and acute sprint exercise. Thirty-three athletes were assigned to control (CON, n = 11), 20-nights LHTLc (n = 12) or 20-nights LHTLi (4 × 5-nights LHTL interspersed with 2-nights CON, n = 10) groups. LHTLc and LHTLi slept at a simulated altitude of 2,650 m (F_IO₂ 0.1627) and lived and trained by day under normoxic conditions; CON lived, trained, and slept in normoxia. A quadriceps muscle biopsy was taken at rest and immediately after standardised sprint exercise, before (Pre) and after 5-nights (d5) and 20-nights (Post) LHTL interventions and analysed for Na⁺, K⁺ATPase maximal activity (3-O-MFPase) and content ([³H]-ouabain binding). After only 5-nights LHTLc, muscle 3-O-MFPase activity declined by 2% (P < 0.05). In LHTLc, 3-O-MFPase activity remained below Pre after 20 nights. In contrast, in LHTLi, this small initial decrease was reversed after 20 nights, with restoration of 3-O-MFPase activity to Pre-intervention levels. Plasma [K⁺] was unaltered by any LHTL. After acute sprint exercise 3-O-MFPase activity was reduced (12.9 ± 4.0%, P < 0.05), but [³H]-ouabain binding was unchanged. In conclusion, maximal Na⁺, K⁺ATPase activity declined after only 5-nights LHTL, but the inclusion of additional interspersed normoxic nights reversed this effect, despite athletes receiving the same amount of hypoxic exposure. There were no effects of consecutive or intermittent nightly LHTL on the acute decrease in Na⁺, K⁺ATPase activity with sprint exercise effects or on plasma [K⁺] during exercise.     

Heterogenous Na+, K+-ATPase expression in the epithelia of rabbit gut-associated lymphoid tissues

Na⁺, K⁺-ATPase expression in the epithelia of rabbit gut-associated lymphoid tissue was measured using indirect immunofluorescence and confocal laser scanning microscopy. All four major sites of aggregated lymphoid tissue, i. e. Peyer's patch, sacculus rotundus, caecal patch and appendix, were studied. Na⁺, K⁺-ATPase expression was localized to the basolateral surface of cells of the follicle-associated epithelium (FAE) and adjacent villous or surface epithelia (non-FAE), where increased expression during enterocyte migration was evident. In the FAE, expression of Na⁺, K⁺-ATPase appeared to be lower in the specialized M cells than in enterocytic-type cells, although expression in both cell types was lower than in adjacent non-FAE. Quantification of immunofluorescent staining of Na⁺, K⁺-ATPase by confocal laser scanning imaging showed a reduction of expression in the FAE to approximately 20–60% relative to that in the adjacent non-FAE. These results are consistent with a primary role of the FAE in mucosal immunity with minimal involvement in active solute absorption.     

Ouabain-induced isoform-specific localization change of the Na+, K+-ATPase alpha subunit in the synaptic plasma membrane of rat brain

Na+, K+-ATPase is one of major membrane proteins that has two subunits, alpha and beta. The alpha subunit has the ATPase activity and the ouabain binding site. Among four isoforms of the alpha subunit, expression of alpha1, alpha2, and alpha3, but not alpha4, is observed in matured rat brain. Ouabain is one of cardiac glycosides, and endogenous ouabain-like compounds have been recognized as a new class of steroid hormone. The alpha subunit is considered as their endogenous receptor. Recent studies envisaged the importance of membrane microdomains (MDs) as signaling platforms, which are recovered as a detergent-resistant membrane microdomain fraction (DRM). Although this ATPase has been considered as a non-DRM protein, some amount of the alpha subunit was found to be a component of the DRM prepared from the synaptic plasma membrane fraction (SPM) of rat brain. Ouabain treatment increased the amount of alpha3 isoform, but not alpha1, in the DRM derived from synaptosome fraction and SPM. These results suggest that the localization of the alpha subunit of Na+, K+-ATPase is regulated with isoform-specific mechanisms and the physiological importance of DRM in the signal transduction of the endogenous ouabain-like steroid hormone in neurons.     

Tubular transport processes in proximal tubules of hypothyroid rats. Lack of relationship between thyroidal dependent rise of isotonic fluid reabsorption and Na+−K+-ATPase activity

Hypothyroid rats reconstituted with 10 g/kg b.w. per day of tri-iodothironine (T₃) for 4 days resulted in normal free T₃ and TSH levels. FT₃ levels were: 0.53±0.3 pg/ml in hypothyroid rats; 2.78±1.21 pg/ml in hormone reconstituted rats and 2.90±0.90 pg/ml in euthyroid rats. TSH levels were 3,508±513 g/ml in hypothyroid rats; 1,008±204 g/ml in reconstituted rats and 270±184 ng/ml in euthyroid rats.When hypothyroid rats were reconstituted with 50 g T₃/kg b.w. per day, TSH levels were nearly normal after 4 days (1,157±621 ng/ml). However FT₃ levels after 1–4 days were always higher than in euthyroid rats.Hypothyroid rats show a decrease in isotonic fluid reabsorption (J _v) in the proximal tubule (1.50±0.08 versus 4.96±0.23 10^–2 nl·mm^–1·s^–1 in euthyroid animals). 1 day after T₃ (10 g/kg b.w./day) injectionJ _v was increased significantly to 2.05±0.20 10^–2 nl·mm^–1·s^–1 and continued to increase during 4 days of T₃ reconstitution.When 50 g T₃/kg b.w./day was used,J _v increased to 2.75±0.07 after 1 day and to 3.10±0.42 10^–2 nl·mm^–1·s^–1 after 4 days.J _v was never reaching a value close to that of euthyroid rats because the tubular radius in hypothyroid rats (14.7±1.8 m) is less than that of euthyroid rats (19.2±0.5 m). The radius in hypothyroid rats treated with T₃ was unchanged over a 4 day course with either high or low doses of T₃.Na⁺–K⁺-ATPase activity was found to be 2.91±0.16 M P_i/h×mg protein in homogenates of kidney cortex from hypothyroid rats. Treatment of hypothyroid rats with 10 g or 50 g of T₃ resulted in an initial decrease in ATPase activity, followed by an increase to base level in hypothyroid rats with 10 g and a significantly higher level with 50 g. This decrease in ATPase activity was contrasted to the increase inJ _v.These data indicate that there is a dissociation between the effects of physiological doses of thyroid hormones on proximal tubular reabsorption and the effects of T₃ on Na⁺–K⁺-ATPase activity of kidney cortex. This leads to question the relationship between sodium transport and ATPase activity under physiological doses of thyroid hormones. An early effect of physiological doses of thyroid hormones on brush border Na⁺ permeability is suggested.