Functional distinction between two transport mechanisms in rabbit gall‐bladder epithelium by use of ouabain, ethacrynic acid and metabolic inhibitors.

Frederiksen, O.

doi:10.1113/jphysiol.1978.sp012389

Cited by 12 publications

(5 citation statements)

References 40 publications

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“…Therefore, the transepithelial PD of the rabbit gallbladder does not seem to be determined by any shunting effect in the paracellular pathway . Rather, the results support the previous conclusion (8,19,24) that the transepithelial transport mechanism in this epithe- Because the tight junctions in low resistance epithelia seem to comprise the pathway through which most of the passive diffusion of many smaller solutes take place, it has been postulated that this pathway may also serve as the route for a significant part of net transepithelial water transport (3,14,34). Recently, it has been suggested that all transepithelial net water flow in the Necturus gallbladder bypasses the epithelial cells (20) .…”

Section: Discussionsupporting

confidence: 86%

“…Control values of net mucosa-to-serosa fluid transport rates (To,), transepithelial PD, and RT obtained ---60 min after preparation were : Tvo , = -0.5 ttl -min-' -mg dryweight-'; PD = 2.9 ± 0.2 mV (mucosal side positive); and RT = 44.5 ± 2.6 ohm-cm'. These values are in agreement with previous observations and have been demonstrated to be nearly constant during at least 4 h of incubation (8,9) .…”

Section: Functional Observationssupporting

confidence: 93%

“…These values are in agreement with previous observations and have been demonstrated to be nearly constant during at least 4 h of incubation (8,9) . Fig.…”

Section: Functional Observationssupporting

confidence: 82%

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Lack of correlation between transepithelial transport capacity and paracellular pathway ultrastructure in Alcian blue-treated rabbit gallbladders.

Frederiksen¹,

Møllgård²,

Rostgaard³

1979

The Journal of Cell Biology

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The effects of mucosal application of 1 mg% Alcian blue (a trivalent cationic phthalocyanine dye) on functional and ultrastructural parameters of the isolated rabbit gallbladder have been studied. Apart from minor changes in the shape of the group of central microvilli observed in thin-section electron microscopy and scanning electron microscopy, the major ultrastructural change induced by Alcian blue was an almost complete collapse of intercellular spaces in the region above the tight junctions up to the bases of the marginal microvilli as revealed by thinsection electron microscopy . Freeze-fracture electron microscopy demonstrated a complete disappearance of intramembrane particles of neighboring cell membranes corresponding to the region of interspace collapse. Transepithelial electrical resistance (RT) increased from 44 .5 to 58.7 ohm-cm' upon treatment with Alcian blue . This increase could be well accounted for by the observed structural changes in the paracellular pathway if this pathway determines the low resistance of the rabbit gallbladder epithelium . Despite the increase in RT, net mucosa-to-serosa fluid transport and the spontaneous mucosa-positive potential difference of 3 mV were unaltered by Alcian blue treatment, supporting the hypothesis that the transepithelial transport mechanism per se is electroneutral . A calculation of the maximal paracellular mucosa-to-serosa waterflow in response to a lateral intercellular space hypertonicity of 20 mosM demonstrates that in the Alcian bluetreated gallbladder the resulting figure is about three orders of magnitude too low to keep up with the unaltered spontaneous transepithelial net fluid transport. It is therefore concluded that the tight junction pathway in rabbit gallbladders does not serve as a route for net fluid transport.KEY WORDS rabbit gallbladder epitheliumof proximal tubule, gallbladder, and small intestransport pathways . fluid absorption .tine is characterized by a very low transepithelial ultrastructure -Alcian blue electrical resistance compared with the resistances of apical and basolateral cell membranes (2, 12, Isosmotically transporting epithelium such as that 13). The low transepithelial resistance seems to be J. CELL BIOLOGY

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Section: Discussionsupporting

confidence: 86%

Section: Functional Observationssupporting

confidence: 93%

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Lack of correlation between transepithelial transport capacity and paracellular pathway ultrastructure in Alcian blue-treated rabbit gallbladders.

Frederiksen¹,

Møllgård²,

Rostgaard³

1979

The Journal of Cell Biology

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“…Gargus and Slayman (13) have reported that LM(TK-) cells contain a diuretic-sensitive Na+-K+ cotransport system, similar to that described in a variety of mammalian (14, 15, 17-23, [25][26][27] and avian (16,28) (11,21,29,30). It is therefore reasonable to expect that, at reduced extracellular K+ concentrations, the cotransport system should mediate the net efflux of KV.…”

Section: Effect Of Diuretics On Cation Fluxes In Lm(tk-) Cellsmentioning

confidence: 73%

Reduction of K+ efflux in cultured mouse fibroblasts, by mutation or by diuretics, permits growth in K+-deficient medium.

Jayme¹,

Adelberg²,

Slayman³

1981

Proc. Natl. Acad. Sci. U.S.A.

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The mouse fibroblastic cell line LM(TK-) is unable to grow at external K+ concentrations below a threshold value of 0.4 mM. At subthreshold K+ concentrations, LM(TK-) cells rapidly lose intracellular K+ and eventually lyse. We have analyzed the pathwayprimarily responsible for K+ efflux under these experimental conditions and report its specific inhibition by two diuretics, furosemide and bumetanide. Bumetanide, an analog of furosemide, was a more potent inhibitor (by several orders of magnitude) than was furosemide itself. The effects of ouabain and bumetanide were additive, suggesting independence of diureticsensitive K+ efflux from Na+/K+ pump-mediated fluxes. Characterization of K+ efflux in LTK-5, a mutant derived from LM(TK-) and selected for its ability to grow at 0.2 mM K+, indicated that the mutant had lost the diuretic-sensitive K+ efflux pathway. Net Mammalian cells maintain a high steady-state K+ concentration and a low steady-state Na+ concentration relative to their surrounding physiological fluids. The transmembrane electrochemical gradients established by these two cations have been implicated in several physiological responses, including the regulation of cell volume (1, 2), nutrient cotransport (3-6), and membrane excitability (7-10). Because the steady-state K+ concentration reflects the sum of influx and efflux components, both active and passive, an alteration in the activity of one component must be accompanied by a compensating alteration in another if the cationic steady state is to be maintained (11). Should a particular flux become altered so that antagonistic fluxes are unable to compensate, the resulting change in intracellular cation concentration may be lethal to the cell. Such considerations were the basis for the selection of two K+ transport mutants that gained the ability to grow at a K+ concentration below that minimally required for parental cell growth (12). Both mutants proved capable of maintaining high intracellular K+ concentrations at reduced external K+ through changes in ouabain-insensitive K+ transport.One of the mutants, LTK-5, exhibited altered activity of a Na+-K+ cotransport system (12, 13) similar to that described in human (14, 15) and avian (16) erythrocytes and in Ehrlich cells (17-21) and murine fibroblasts (22,23). Specifically, in characterizing the mutant relative to its parent strain under conditions of K+ depletion and high external K+, Gargus and Slayman (13) noted that LTK-5 demonstrated enhanced furosemidesensitive, Na+-dependent K+ influx. For two reasons, however, it seemed unlikely that the increase in K+ influx was causally related to the ability of the mutant to survive selection. (i) When furosemide-sensitive K+ influx was measured as a function of the extracellular K+ concentration in both parent and mutant cells, it reached a half-maximal rate at 6 mM K+, considerably higher than the Km for the ouabain-sensitive Na+/K+ pump (1.3 mM). At 0.2 mM K+, the concentration used in the selection, furosemide-sensitive influx was barely detecta...

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“…The diuretic-sensitive cation cotransport systems that have been described in avian (McManus and Schmidt, 1978;Bakker-Grunwald, 1981) and mammalian (Chipperfield, 1980;Dunham et al, 1980;Tosteson, 1981) erythrocytes, Ehrlich ascites cells (Bakker-Grunwald et al, 1980, Geck et a]., 1980), and a variety of epithelial tissues (Frederiksen, 1978;Rindler et al, 1980) show a specific activation by C1-; and in some cases, a coupled flux of C1-has been demonstrated (Dunham et al, 1980;Geck et al, 1980;Haas et al, 1982). It was therefore of interest to examine Kf efflux in LM(TK-) cells under Cl--free conditions.…”

Section: Role Of Cl-mentioning

confidence: 99%

Furosemide‐sensitive potassium efflux in cultured mouse fibroblasts

Jayme

Slayman

Adelberg

1984

Journal Cellular Physiology

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Transfer of LM(TK-) cells from normal growth medium to medium lacking K+ leads to a rapid loss of intracellular K+, which is 50-70% inhibited by furosemide or bumetanide. The diuretic-sensitive component of K+ efflux requires both Na+ and Cl-, and is presumably mediated by a K+, Na+, Cl- cotransport system of the kind described in avian erythrocytes and Ehrlich ascites cells. It can be calculated that such a system should be near equilibrium under normal growth conditions but should mediate net efflux (as observed) when the driving force is altered by reducing extracellular K+. The diuretic-sensitive component of net K+ efflux is also sensitive to amiloride. This effect is probably indirect, however, with amiloride acting to block the Na+ influx that supplies Na+ to the cotransport system. At the low extracellular K+ concentrations employed in these studies, the diuretic-sensitive system is a physiologically important pathway of K+ loss. The rate of growth in low-K+ medium can be increased (or the rate of cell lysis decreased) by adding diuretic or by reducing external Na+ or Cl-.

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Functional distinction between two transport mechanisms in rabbit gall‐bladder epithelium by use of ouabain, ethacrynic acid and metabolic inhibitors.

Cited by 12 publications

References 40 publications

Lack of correlation between transepithelial transport capacity and paracellular pathway ultrastructure in Alcian blue-treated rabbit gallbladders.

Lack of correlation between transepithelial transport capacity and paracellular pathway ultrastructure in Alcian blue-treated rabbit gallbladders.

Reduction of K+ efflux in cultured mouse fibroblasts, by mutation or by diuretics, permits growth in K+-deficient medium.

Furosemide‐sensitive potassium efflux in cultured mouse fibroblasts

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