Control of H(+)-HCO3- plasma membrane transporters by urea hyperosmolality in rat medullary thick ascending limb

1994 ◽  
Vol 266 (5) ◽  
pp. C1157-C1164 ◽  
Author(s):  
F. Leviel ◽  
M. Froissart ◽  
H. Soualmia ◽  
J. Poggioli ◽  
M. Paillard ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Calcium Concentration ◽  
Inhibitory Concentration ◽  
Initial Rate ◽  
Maximum Velocity ◽  
Membrane Transporters ◽  
Thick Ascending Limb ◽  
Michaelis Constant ◽  
Medullary Thick Ascending Limb ◽  
Experimental Protocols

Hyperosmolality inhibits bicarbonate absorption by the rat medullary thick ascending limb (MTAL) by unknown mechanisms. Intracellular pH (pHi) was monitored with use of 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein in rat MTAL tubule suspensions to specify the H(+)-HCO3- membrane transporters affected by hyperosmolality. Measurements were made after > or = 15-min incubation of the cells in media rendered hypertonic by urea to avoid any change in cell volume. Na(+)-H+ antiport activity, estimated from the Na(+)-induced initial rate of pHi recovery of Na(+)-depleted acidified cells in the presence of 0.1 mM furosemide to inhibit Na(+)-K(+)-2Cl- cotransport, was inhibited by 300 mM urea and 10(-8) M arginine vasopressin (AVP) in an additive manner. Na(+)-H+ antiport inhibition by urea hyperosmolality was maximal at 300 mM urea with a half-maximal inhibitory concentration of 75 mM and was due to a 28% decrease in maximum velocity (Vmax) with no effect on the Michaelis constant for sodium. Urea hyperosmolality (300 mM) did not affect steady-state intracellular calcium concentration ([Ca2+]i), assessed with use of fura 2 fluorescence, and still inhibited Na(+)-H+ antiport in MTAL cells loaded with 1,2-bis(2- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid to minimize any transient change in [Ca2+]i during the preincubation in urea medium. Furthermore, 300 mM urea did not stimulate basal or AVP-induced adenosine 3',5'-cyclic monophosphate (cAMP) accumulation. Plasma membrane H(+)-adenosinetriphosphatase (ATPase) activity and HCO3- transport, assessed by appropriate experimental protocols, were unaltered by 300 mM urea.(ABSTRACT TRUNCATED AT 250 WORDS)

Interactions of external and internal K+ with K(+)-HCO3- cotransporter of rat medullary thick ascending limb

1996 ◽  
Vol 271 (1) ◽  
pp. C218-C225 ◽  
Author(s):  
A. Blanchard ◽  
F. leviel ◽  
M. Bichara ◽  
R. A. Podevin ◽  
M. Paillard
Keyword(s):  
Maximum Velocity ◽  
Hill Coefficient ◽  
Kinetic Properties ◽  
Thick Ascending Limb ◽  
Medullary Thick Ascending Limb ◽  
Sigmoidal Curve ◽  
K Concentration ◽  
Physiological Variations ◽  
Allosteric Activator

We studied [K+]i and [K+]o, where subscripts i and o refer to intracellular and extracellular, respectively, concentration dependency of the kinetic properties of the electroneutral K(+)-HCO3-cotransport, using suspensions of rat medullary thick ascending limb (mTAL). With the use of nigericin and monensin, [K+]i was clamped at various values, while maintaining [Na+]i = [Na+]o = 37 mM, [HCO3-]i = [HCO3-]o = 23 mM, and pHi = pHo = 7.4. As indicated by 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein HCO3(-)-dependent rates of change in pHi, at constant [K+]i, increasing the magnitude of the outward K+ gradient by varying [K+]o saturated HCO3-efflux with a Michaelis-Menten curve (apparent Michaelis constant for [K+]o = 2 mM, Hill coefficient = 1). On the other hand, increasing [K+]i from 30 to 140 mM, while either [K+]o or the magnitude of the K+ concentration gradient was fixed, saturated HCO3- efflux with a sigmoidal curve and yielded a Hill coefficient of 3.4 and 50% of maximum velocity at 70 mM [K+]i. These results indicate that [K+]i, independent of its role as a transportable substrate for the cotransport with HCO3-, has a role as an allosteric activator of the K(+)-HCO3- cotransporter. Such an allosteric modulation may contribute to the maintenance of net HCO3- absorption despite large in vivo physiological variations of K+ concentration in the medullary interstitium.

An apical K+-dependent HCO3− transport pathway opposes transepithelial HCO3− absorption in rat medullary thick ascending limb

2004 ◽  
Vol 287 (1) ◽  
pp. F57-F63 ◽  
Author(s):  
Bruns A. Watts ◽  
David W. Good
Keyword(s):  
Initial Rate ◽  
Apical Membrane ◽  
Channel Blockers ◽  
Thick Ascending Limb ◽  
Acid Base ◽  
Transport Pathway ◽  
Transport Pathways ◽  
Medullary Thick Ascending Limb ◽  
And Function

Absorption of HCO3− in the medullary thick ascending limb (MTAL) is mediated by apical membrane Na+/H+ exchange. The identity and function of other apical acid-base transporters in this segment have not been defined. The present study was designed to examine apical membrane HCO3−/OH−/H+ transport pathways in the rat MTAL and to determine their role in transepithelial HCO3− absorption. MTALs were perfused in vitro in Na+- and Cl−-free solutions containing 25 mM HCO3−, 5% CO2. Lumen addition of either 120 mM Cl− or 50 mM Na+ (50 μM EIPA present) had no effect on intracellular pH (pHi). Lumen Cl− addition also had no effect on pHi in the presence of 145 mM Na+ or in the nominal absence of HCO3−/CO2. Thus there was no evidence for apical Cl−/HCO3− (OH−) exchange, Na+-dependent Cl−/HCO3− exchange, or Na+-HCO3− cotransport. In contrast, in tubules studied in Na+- and Cl−-free solutions containing 25 mM HCO3−, 5% CO2 and 120 mM K+, removal of luminal K+ induced a rapid and pronounced decrease in pHi (ΔpHi = 0.56 ± 0.06 pH U). pHi recovered following lumen K+ readdition. The initial rate of net base efflux induced by lumen K+ removal was decreased 85% at the same pHi in the nominal absence of HCO3−/CO2, indicating a dependence on HCO3−/CO2 and arguing against apical K+/H+ exchange. A combination of the apical K+ channel blockers quinidine (0.1 mM) and glybenclamide (0.25 mM) had no effect on the lumen K+-induced pHi changes, arguing against electrically coupled K+ and HCO3− conductances. The effect of lumen K+ on pHi was inhibited by 1 mM H2DIDS. In addition, lumen addition of DIDS increased transepithelial HCO3− absorption from 10.7 ± 0.7 to 14.9 ± 0.7 pmol·min−1·mm−1 ( P < 0.001) and increased pHi slightly in MTAL studied in physiological solutions (25 mM HCO3− and 4 mM K+). Lumen DIDS stimulated HCO3− absorption in the absence and presence of furosemide. These results are consistent with an apical membrane K+-dependent HCO3− transport pathway that mediates coupled transfer of K+ and HCO3− from cell to lumen in the MTAL. This mechanism, possibly an apical K+-HCO3− cotransporter, functions in parallel with apical Na+/H+ exchange and opposes transepithelial HCO3− absorption.

Alternatively spliced isoform of apical Na+-K+-Cl− cotransporter gene encodes a furosemide-sensitive Na+-Cl−cotransporter

2001 ◽  
Vol 280 (4) ◽  
pp. F574-F582 ◽  
Author(s):  
Consuelo Plata ◽  
Patricia Meade ◽  
Amy Hall ◽  
Rick C. Welch ◽  
Norma Vázquez ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Alternative Splicing ◽  
Immunohistochemical Analysis ◽  
Xenopus Laevis Oocytes ◽  
Thick Ascending Limb ◽  
Terminal Domain ◽  
Medullary Thick Ascending Limb ◽  
Slc12a1 Gene ◽  
Alternatively Spliced ◽  
Oocyte Cytoplasm

In the absence of vasopressin, medullary thick ascending limb cells express a K+-independent, furosemide-sensitive Na+-Cl− cotransporter that is inhibited by hypertonicity. The murine renal specific Na+-K+-2 Cl− cotransporter gene ( SLC12A1) gives rise to six alternatively spliced isoforms. Three feature a long COOH-terminal domain that encodes the butmetanide-sensitive Na+-K+-2 Cl−cotransporter (BSC1–9/NKCC2), and three with a short COOH-terminal domain, known as mBSC1-A4, B4, or F4 (19). Here we have determined the functional characteristics of mBSC1-A4, as expressed in Xenopus laevis oocytes. When incubated at normal oocyte osmolarity (∼200 mosmol/kgH2O), mBSC1–4-injected oocytes do not express significant Na+ uptake over H2O-injected controls, and immunohistochemical analysis shows that the majority of mBSC1–4 protein is in the oocyte cytoplasm and not at the plasma membrane. In contrast, when mBSC1–4 oocytes are exposed to hypotonicity (∼100 mosmol/kgH2O), a significant increase in Na+uptake but not in 86Rb+ uptake is observed. The increased Na+ uptake is Cl− dependent, furosemide sensitive, and cAMP sensitive but K+independent. Sodium uptake increases with decreasing osmolarity between 120 and 70 mosmol/kgH2O ( r = 0.95, P < 0.01). Immunohistochemical analysis shows that in hypotonic conditions mBSC1-A4 protein is expressed in the plasma membrane. These studies indicate that the mBSC1-A4 isoform of the SLC12A1 gene encodes a hypotonically activated, cAMP- and furosemide-sensitive Na+-Cl− cotransporter. Thus it is possible that alternative splicing of the BSC1 gene could provide the molecular mechanism enabling the Na+-Cl−-to-Na+-K+-2Cl−switching in thick ascending limb cells.

scholarly journals P2X receptors trigger intracellular alkalization in isolated perfused mouse medullary thick ascending limb

2014 ◽  
Vol 213 (1) ◽  
pp. 277-284 ◽  
Author(s):  
P. I. A. Bruijn ◽  
M. Bleich ◽  
H. A. Praetorius ◽  
J. Leipziger
Keyword(s):  
P2x Receptors ◽  
Thick Ascending Limb ◽  
Medullary Thick Ascending Limb

Calcium- and potassium-permeable plasma membrane transporters are activated by copper inArabidopsisroot tips: linking copper transport with cytosolic hydroxyl radical production

2012 ◽  
Vol 36 (4) ◽  
pp. 844-855 ◽  
Author(s):  
ANA RODRIGO-MORENO ◽  
NURIA ANDRÉS-COLÁS ◽  
CHARLOTTE POSCHENRIEDER ◽  
BENET GUNSÉ ◽  
LOLA PEÑARRUBIA ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Hydroxyl Radical ◽  
Copper Transport ◽  
Membrane Transporters ◽  
Radical Production

scholarly journals High sodium intake increases HCO3− absorption in medullary thick ascending limb through adaptations in basolateral and apical Na+/H+ exchangers

2011 ◽  
Vol 301 (2) ◽  
pp. F334-F343 ◽  
Author(s):  
David W. Good ◽  
Thampi George ◽  
Bruns A. Watts
Keyword(s):  
Absorptive Capacity ◽  
Sodium Intake ◽  
Thick Ascending Limb ◽  
Acid Base Balance ◽  
High Sodium ◽  
High Sodium Intake ◽  
Medullary Thick Ascending Limb ◽  
Nhe1 Activity ◽  
Exchange Activity

A high sodium intake increases the capacity of the medullary thick ascending limb (MTAL) to absorb HCO3−. Here, we examined the role of the apical NHE3 and basolateral NHE1 Na+/H+ exchangers in this adaptation. MTALs from rats drinking H2O or 0.28 M NaCl for 5–7 days were perfused in vitro. High sodium intake increased HCO3− absorption rate by 60%. The increased HCO3− absorptive capacity was mediated by an increase in apical NHE3 activity. Inhibiting basolateral NHE1 with bath amiloride eliminated 60% of the adaptive increase in HCO3− absorption. Thus the majority of the increase in NHE3 activity was dependent on NHE1. A high sodium intake increased basolateral Na+/H+ exchange activity by 89% in association with an increase in NHE1 expression. High sodium intake increased apical Na+/H+ exchange activity by 30% under conditions in which basolateral Na+/H+ exchange was inhibited but did not change NHE3 abundance. These results suggest that high sodium intake increases HCO3− absorptive capacity in the MTAL through 1) an adaptive increase in basolateral NHE1 activity that results secondarily in an increase in apical NHE3 activity; and 2) an adaptive increase in NHE3 activity, independent of NHE1 activity. These studies support a role for NHE1 in the long-term regulation of renal tubule function and suggest that the regulatory interaction whereby NHE1 enhances the activity of NHE3 in the MTAL plays a role in the chronic regulation of HCO3− absorption. The adaptive increases in Na+/H+ exchange activity and HCO3− absorption in the MTAL may play a role in enabling the kidneys to regulate acid-base balance during changes in sodium and volume balance.

Identification of ferrichrome- and ferrioxamine B-mediated iron uptake by Aspergillus fumigatus

2016 ◽  
Vol 473 (9) ◽  
pp. 1203-1213 ◽  
Author(s):  
Yong-Sung Park ◽  
Ju-Yeon Kim ◽  
Cheol-Won Yun
Keyword(s):  
Gene Expression ◽  
Plasma Membrane ◽  
Aspergillus Fumigatus ◽  
Virulence Factors ◽  
Membrane Transporters ◽  
Yeast Cells ◽  
Double Deletion ◽  
Ferrioxamine B ◽  
Uptake Activity ◽  
Opportunistic Fungal Pathogen

Aspergillus fumigatus is an opportunistic fungal pathogen for immunocompromised patients, and genes involved in siderophore metabolism have been identified as virulence factors. Recently, we identified the membrane transporters sit1 and sit2, which are putative virulence factors of A. fumigatus; sit1 and sit2 are homologous to yeast Sit1, and sit1 and sit2 gene expression was up-regulated after iron depletion. When expressed heterologously in Saccharomyces cerevisiae, sit1 and sit2 were localized to the plasma membrane; sit1 efficiently complemented ferrichrome (FC) and ferrioxamine B (FOB) uptake in yeast cells, whereas sit2 complemented only FC uptake. Deletion of sit1 resulted in a decrease in FOB and FC uptake, and deletion of sit2 resulted in a decrease in FC uptake in A. fumigatus. It is of interest that a sit1 and sit2 double-deletion mutant resulted in a synergistic decrease in FC uptake activity. Both sit1 and sit2 were localized to the plasma membrane in A. fumigatus. The expression levels of the sit1 and sit2 genes were dependent on hapX under low-but not high-iron conditions. Furthermore, mirB, and sidA gene expression was up-regulated and sreA expression down-regulated when sit1 and sit2 were deleted. Although sit1 and sit2 failed to affect mouse survival rate, these genes affected conidial killing activity. Taken together, our results suggest that sit1 and sit2 are siderophore transporters and putative virulence factors localized to the plasma membrane.

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