scholarly journals Tetraethylammonium transport by OK cells.

1990 ◽  
Vol 1 (6) ◽  
pp. 902-909
Author(s):  
T D McKinney ◽  
M B Scheller ◽  
M Hosford ◽  
J A McAteer
Keyword(s):  
Organic Cation ◽  
Epithelial Transport ◽  
Proton Exchange ◽  
Proximal Tubules ◽  
Organic Cations ◽  
Kidney Cells ◽  
Dependent Manner ◽  
Transport Pathway ◽  
Opossum Kidney ◽  
Opossum Kidney Cells

Mechanisms exist in renal proximal tubules for the mediated transepithelial secretion or reabsorption of endogenous and exogenous organic cations. In the studies presented here, the uptake of the organic cation tetraethylammonium (TEA) into confluent monolayers of opossum kidney cells was evaluated to determine if these cells might serve as an in vitro model of this transport pathway. 3H-TEA entered opossum kidney cells in a time-dependent manner. Uptake at early time points was saturable with an apparent Km of 59.1 +/- 11.2 microM and a Vmax of 1,292 +/- 210 fmol/micrograms of DNA. TEA uptake was inhibited in a dose-dependent manner by several other organic cations including amiloride, cimetidine, verapamil, procainamide, quinidine and N1-methylnicotinamide. With 1 mM concentrations of these compounds, uptake was virtually eliminated. However, another organic cation, N'-methylnicotinamide caused only minimal inhibition. TEA uptake was significantly reduced by sodium azide, suggesting dependence on oxidative phosphorylation. An alkaline medium pH enhanced TEA uptake, but, at the same pH, uptake was similar in the presence or absence of bicarbonate. When cellular pH was altered by ammonium chloride addition or removal, TEA uptake was not affected. Thus, organic cation/proton exchange, as has been demonstrated previously in apical membrane vesicles prepared from proximal tubules, is evidently not responsible for TEA uptake. Similarly, uptake does not appear to result from organic cation/organic cation exchange. These results indicate that the plasma membrane of opossum kidney cells contains a transport system(s) for the mediated uptake of organic cations and that these cells may be a useful mode for further study of renal epithelial transport of these solutes.

scholarly journals Basolateral transport of tetraethylammonium by a clone of LLC-PK1 cells.

1992 ◽  
Vol 2 (10) ◽  
pp. 1507-1515
Author(s):  
T D McKinney ◽  
M B Scheller ◽  
M Hosford ◽  
M E Lesniak ◽  
T S Haseley
Keyword(s):  
Organic Cation ◽  
Basolateral Membrane ◽  
Transport Processes ◽  
Proton Exchange ◽  
Metabolic Inhibitors ◽  
Organic Cations ◽  
Dependent Manner ◽  
Electrogenic Transport ◽  
The Media ◽  
Uptake Medium

In these studies, a clone of cells derived from the porcine renal epithelial line LLC-PK1 grown on porous filters was used to evaluate basolateral uptake of the organic cation tetraethylammonium (TEA). (3H) TEA (1 microM) entered cells in a saturable and time-dependent manner achieving a steady-state value at 2 to 2.5 h. Uptake was reduced by hypothermia and the metabolic inhibitors sodium azide and iodoacetate. Several other organic cations in 1 mM concentrations inhibited the majority of TEA uptake. In lower concentrations, the inhibitory potency of these was: verapamil greater than cimetidine approximately amiloride approximately quinidine greater than procainamide approximately N1-methylnicotinamide. When sodium was replaced with potassium in the uptake medium, TEA uptake was also reduced consistent with electrogenic transport. However, uptake was reduced further by 1 mM cimetidine in the presence of both NaCl and KCl buffers. TEA uptake was not significantly different when the media pH was varied from 6.0 to 8.0. In addition, results of experiments in which intracellular pH was altered with NH4Cl were not consistent with the presence of organic cation/proton exchange. TEA/TEA exchange could not be demonstrated in experiments in which cells were preloaded with 1 mM nonradioactive TEA and uptake of (3H)TEA was measured or in which nonradioactive TEA in the external medium failed to enhance efflux from cells preloaded with (3H)TEA. These results indicate that the basolateral membrane of LLC-PKc10 cells has one or more transport processes for the mediated uptake of organic cations. However, the precise mechanism(s) involved in this transport remains to be elucidated.

scholarly journals Phosphorylation of rat kidney Na-K pump at Ser938 is required for rapid angiotensin II-dependent stimulation of activity and trafficking in proximal tubule cells

2016 ◽  
Vol 310 (3) ◽  
pp. C227-C232 ◽  
Author(s):  
Katherine J. Massey ◽  
Quanwen Li ◽  
Noreen F. Rossi ◽  
Susan M. Keezer ◽  
Raymond R. Mattingly ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Rat Kidney ◽  
Proximal Tubules ◽  
Kidney Cells ◽  
Ang Ii ◽  
Wild Type ◽  
Stimulate Activity ◽  
Opossum Kidney ◽  
Opossum Kidney Cells ◽  
Stimulation Of

How angiotensin (ANG) II acutely stimulates the Na-K pump in proximal tubules is only partially understood, limiting insight into how ANG II increases blood pressure. First, we tested whether ANG II increases the number of pumps in plasma membranes of native rat proximal tubules under conditions of rapid activation. We found that exposure to 100 pM ANG II for 2 min, which was previously shown to increase affinity of the Na-K pump for Na and stimulate activity threefold, increased the amount of the Na-K pump in plasma membranes of native tubules by 33%. Second, we tested whether previously observed increases in phosphorylation of the Na-K pump at Ser938 were part of the stimulatory mechanism. These experiments were carried out in opossum kidney cells, cultured proximal tubules stably coexpressing the ANG type 1 (AT1) receptor, and either wild-type or a S938A mutant of rat kidney Na-K pump under conditions found by others to stimulate activity. We found that 10 min of incubation in 10 pM ANG II stimulated activity of wild-type pumps from 2.3 to 3.5 nmol K·mg protein−1·min−1 and increased the amount of the pump in the plasma membrane by 80% but had no effect on cells expressing the S938A mutant. We conclude that acute stimulation of Na-K pump activity in native rat proximal tubules includes increased trafficking to the plasma membrane and that phosphorylation at Ser938 is part of the mechanism by which ANG II directly stimulates activity and trafficking of the rat kidney Na-K pump in opossum kidney cells.

Regulation of K+ Channels in Proximal Tubules: Studies in Opossum Kidney Cells

Nephrology ◽  
1991 ◽  
pp. 1677-1684 ◽  
Author(s):  
Takako Ohno-Shosaku ◽  
Takahiro Kubota ◽  
Yoshiaki Mori ◽  
Jun Yamaguchi ◽  
Mamoru Fujimoto
Keyword(s):  
Proximal Tubules ◽  
Kidney Cells ◽  
K Channels ◽  
Opossum Kidney ◽  
Opossum Kidney Cells

D2-like receptor-mediated inhibition of Na+-K+-ATPase activity is dependent on the opening of K+ channels

2002 ◽  
Vol 283 (1) ◽  
pp. F114-F123 ◽  
Author(s):  
Pedro Gomes ◽  
P. Soares-da-Silva
Keyword(s):  
Protein Kinase ◽  
Atpase Activity ◽  
Receptor Activation ◽  
Inhibitory Effect ◽  
Skf 38393 ◽  
Kidney Cells ◽  
Dependent Manner ◽  
Opossum Kidney ◽  
Opossum Kidney Cells ◽  
Stimulation Of

This study examined the effects of D2-like dopamine receptor activation on Na+-K+-ATPase activity while apical-to-basal, ouabain-sensitive, amphotericin B-induced increases in short-circuit current and basolateral K+ ( I K) currents in opossum kidney cells were measured. The inhibitory effect of dopamine on Na+-K+-ATPase activity was completely abolished by either D1- or D2-like receptor antagonists and mimicked by D1- and D2-like receptor agonists SKF-38393 and quinerolane, respectively. Blockade of basolateral K+ channels with BaCl2 (1 mM) or glibenclamide (10 μM), but not apamin (1 μM), totally prevented the inhibitory effects of quinerolane. The K+ channel opener pinacidil decreased Na+-K+-ATPase activity. The inhibitory effect of quinerolane on Na+-K+- ATPase activity was abolished by pretreatment of opossum kidney cells with pertussis toxin (PTX). Quinerolane increased I K across the basolateral membrane in a concentration-dependent manner; this effect was abolished by pretreatment with PTX, S-sulpiride, and glibenclamide. SKF-38393 did not change I K. Both H-89 (protein kinase A inhibitor) and chelerythrine (protein kinase C inhibitor) failed to prevent the stimulatory effect of quinerolane on I K. The stimulation of the D2-like receptor was associated with a rapid hyperpolarizing effect, whereas D1-like receptor activation was accompanied by increases in cell membrane potential. It is concluded that stimulation of D2-like receptors leads to inhibition of Na+-K+-ATPase activity and hyperpolarization; both effects are associated with the opening of K+channels.

Expression of the angiotensinogen gene is synergistically stimulated by 8-BrcAMP and Dex in opossum kidney cells

1995 ◽  
Vol 268 (1) ◽  
pp. R105-R111 ◽  
Author(s):  
M. Ming ◽  
T. T. Wang ◽  
S. Lachance ◽  
A. Delalandre ◽  
S. Carriere ◽  
...  
Keyword(s):  
Fusion Gene ◽  
Dependent Manner ◽  
Fusion Genes ◽  
Opossum Kidney ◽  
Ok Cells ◽  
Angiotensinogen Gene ◽  
Opossum Kidney Cells ◽  
Dependent Protein ◽  
Flanking Region ◽  
Responsive Element

We transiently transfected fusion genes with the 5'-flanking region of the angiotensinogen gene linked to a bacterial chloramphenicol acetyltransferase (CAT) coding sequence as a reporter into opossum kidney (OK) cells. The addition of 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP) (10(-3)-10(-7) M) or forskolin (10(-9)-10(-5) M) stimulated the expression of the plasmid pOCAT [angiotensinogen nucleotide (N) -1498/+18] fusion gene in OK cells in a dose-dependent manner. The addition of dexamethasone (Dex) (10(-6) M) further enhanced the stimulatory effect of 8-BrcAMP or forskolin, whereas the addition of (R)-p-adenosine 3',5'-cyclic monophosphorothioate [(Rp)-cAMP[S], an inhibitor of cAMP-dependent protein kinase A, I and II] blocked the stimulatory effect of 8-BrcAMP. Furthermore, the addition of 8-BrcAMP (10(-3) M) or Dex (10(-6) M) or a combination of both stimulated the expression of pOCAT (angiotensinogen N -1138/+18), pOCAT (angiotensinogen N -960/+18), pOCAT (angiotensinogen N -814/+18), and pOCAT (angiotensinogen N -688/+18), but had no effect on the expression of pOCAT (angiotensinogen N -280/+18), pOCAT (angiotensinogen N -198/+18), pOCAT (angiotensinogen N -110/+18), pOCAT (angiotensinogen N -53/+18), and pOCAT (angiotensinogen N -35/+18). To further localize the putative cAMP-responsive element (CRE) in the angiotensinogen gene, we constructed fusion genes by inserting the DNA fragments angiotensinogen N -814 to N -689, angiotensinogen N -814 to N -761, and angiotensinogen N -760 to N -689 of the 5'-flanking region of the angiotensinogen gene upstream of the thymidine kinase (TK) promoter fused to a CAT gene and introduced them into OK cells.(ABSTRACT TRUNCATED AT 250 WORDS)

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