Chloride movement across basolateral membrane of Necturus gallbladder epithelium

1984 ◽  
Vol 247 (5) ◽  
pp. C495-C500 ◽  
Author(s):  
R. S. Fisher
Keyword(s):  
Basolateral Membrane ◽  
Membrane Voltage ◽  
Gallbladder Epithelium ◽  
Necturus Gallbladder ◽  
Concentration Changes ◽  
The Difference ◽  
K Concentration ◽  
Low Ionic Strength ◽  
Basolateral Membrane Potential

The relative Cl- and K+ sensitivity of the basolateral membrane potential of the in vitro Necturus gallbladder epithelium was determined. Tissues were punctured with two conventional glass microelectrodes to simultaneously measure the intracellular voltage (Vcs) and the voltage across the subepithelial connective tissue (Vse). Increasing the serosal K+ concentration from 2.5 to 25 mM caused a rapid monotonic depolarization of Vcs without changes of Vse. Reduction of serosal Cl- concentration (98 to 8 mM) caused a transient change of Vse. Thus the difference between Vcs and Vse more accurately reflected the basolateral membrane voltage (Vc) after Cl- concentration changes. The changes of Vc were small and biphasic in response to the decrease of serosal Cl- concentration. Perfusion of a low-ionic-strength solution in the mucosal chamber decreased the current that normally passes through the epithelium. Consistent with the notion that the basolateral voltage changes are attenuated by parallel pathways, the K+-induced depolarization increased by 80% under these conditions. The changes of Vc in response to Cl- substitutions were not different from those of tissue bathed in control solution. Thus the basolateral membrane voltage is relatively insensitive to changes of serosal Cl- concentration. I conclude that Cl- movement across the basolateral membrane is not attributable to simple electrodiffusion, and Cl- exit from these cells at this membrane must be electroneutral.

scholarly journals Electrophysiological effects of basolateral [Na+] in Necturus gallbladder epithelium.

1992 ◽  
Vol 99 (2) ◽  
pp. 241-262 ◽  
Author(s):  
G A Altenberg ◽  
J S Stoddard ◽  
L Reuss
Keyword(s):  
Apical Membrane ◽  
Basolateral Membrane ◽  
Membrane Resistance ◽  
Membrane Voltage ◽  
Gallbladder Epithelium ◽  
K Channels ◽  
Necturus Gallbladder ◽  
Electrophysiological Effects ◽  
Paracellular Diffusion ◽  
Cl Conductance

In Necturus gallbladder epithelium, lowering serosal [Na+] ([Na+]s) reversibly hyperpolarized the basolateral cell membrane voltage (Vcs) and reduced the fractional resistance of the apical membrane (fRa). Previous results have suggested that there is no sizable basolateral Na+ conductance and that there are apical Ca(2+)-activated K+ channels. Here, we studied the mechanisms of the electrophysiological effects of lowering [Na+]s, in particular the possibility that an elevation in intracellular free [Ca2+] hyperpolarizes Vcs by increasing gK+. When [Na+]s was reduced from 100.5 to 10.5 mM (tetramethylammonium substitution), Vcs hyperpolarized from -68 +/- 2 to a peak value of -82 +/- 2 mV (P less than 0.001), and fRa decreased from 0.84 +/- 0.02 to 0.62 +/- 0.02 (P less than 0.001). Addition of 5 mM tetraethylammonium (TEA+) to the mucosal solution reduced both the hyperpolarization of Vcs and the change in fRa, whereas serosal addition of TEA+ had no effect. Ouabain (10(-4) M, serosal side) produced a small depolarization of Vcs and reduced the hyperpolarization upon lowering [Na+]s, without affecting the decrease in fRa. The effects of mucosal TEA+ and serosal ouabain were additive. Neither amiloride (10(-5) or 10(-3) M) nor tetrodotoxin (10(-6) M) had any effects on Vcs or fRa or on their responses to lowering [Na+]s, suggesting that basolateral Na+ channels do not contribute to the control membrane voltage or to the hyperpolarization upon lowering [Na+]s. The basolateral membrane depolarization upon elevating [K+]s was increased transiently during the hyperpolarization of Vcs upon lowering [Na+]s. Since cable analysis experiments show that basolateral membrane resistance increased, a decrease in basolateral Cl- conductance (gCl-) is the main cause of the increased K+ selectivity. Lowering [Na+]s increases intracellular free [Ca2+], which may be responsible for the increase in the apical membrane TEA(+)-sensitive gK+. We conclude that the decrease in fRa by lowering [Na+]s is mainly caused by an increase in intracellular free [Ca2+], which activates TEA(+)-sensitive maxi K+ channels at the apical membrane and decreases apical membrane resistance. The hyperpolarization of Vcs is due to increase in: (a) apical membrane gK+, (b) the contribution of the Na+ pump to Vcs, (c) basolateral membrane K+ selectivity (decreased gCl-), and (d) intraepithelial current flow brought about by a paracellular diffusion potential.

Electrophysiological characterization of upper portion of descending limb of long-looped nephron

1991 ◽  
Vol 260 (3) ◽  
pp. F311-F316 ◽  
Author(s):  
K. Yoshitomi ◽  
M. Imai
Keyword(s):  
Apical Membrane ◽  
Basolateral Membrane ◽  
High Water ◽  
Membrane Resistance ◽  
Membrane Voltage ◽  
Major Pathway ◽  
K Concentration ◽  
Electrophysiological Characterization

The upper portion of the descending limb of long-looped nephron (LDLu) of the hamster is characterized by high water and ion permeabilities. Although the paracellular route is considered to be the major pathway representing cation permselectivity of this segment, ion transport mechanisms through the transcellular pathway are unknown. To study this issue; we applied cable analysis and conventional microelectrode technique to the hamster LDLu perfused in vitro. The transmural voltage (VT) was not different from zero, and transmural resistance (RT) was very low, 18.3 +/- 2.0 omega.cm2 (n = 12). The basolateral membrane voltage was -80 +/- 2 mV (n = 55), and fractional apical membrane resistance was 0.92 +/- 0.23 (n = 5). Ouabain (0.1 mM) in the bath decreased basolateral membrane voltage (VB) by 23 +/- 3 mV (n = 6, P less than 0.001). Increase in K+ concentration in bath and in lumen from 5 to 50 mM decreased VB by 39 +/- 2 (n = 7, P less than 0.01) and apical membrane voltage (VA) by 10 +/- 1 mV (n = 7, P less than 0.001), respectively. Addition of 2 mM Ba2+ to bath and to lumen decreased VB by -47 +/- 2 (n = 11, P less than 0.001) and decreased VA by 8 +/- 1 mV, respectively. Reduction of HCO3- in bath from 25 to 2.5 mM decreased VB by 4 +/- 1 mV (n = 7, P less than 0.005). Reduction of bath Cl- did not cause any rapid deflection of VB. No appreciable Na+ conductance was detected in the apical membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanism of bicarbonate exit across basolateral membrane of rabbit proximal straight tubule

1987 ◽  
Vol 252 (1) ◽  
pp. F11-F18 ◽  
Author(s):  
S. Sasaki ◽  
T. Shiigai ◽  
N. Yoshiyama ◽  
J. Takeuchi
Keyword(s):  
Negative Charge ◽  
Driving Forces ◽  
Basolateral Membrane ◽  
Disulfonic Acid ◽  
Exit Mechanism ◽  
Total Replacement ◽  
Straight Tubules ◽  
Proximal Straight Tubule ◽  
Basolateral Membrane Potential

To clarify the mechanism(s) of HCO3- (or related base) transport across the basolateral membrane, rabbit proximal straight tubules were perfused in vitro, and intracellular pH (pHi) and Na+ activity (aiNa) were measured by double-barreled ion-selective microelectrodes. Lowering bath HCO3- from 25 to 5 mM at constant PCO2 depolarized basolateral membrane potential (Vbl), and reduced pHi. Most of these changes were inhibited by adding 1 mM 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS) to the bath. Total replacement of bath Na+ with choline also depolarized Vbl and reduced pHi, and these changes were also inhibited by SITS. Reduction in aiNa was observed when bath HCO3- was lowered. Taken together, these findings suggest that HCO3- exists the basolateral membrane with Na+ and negative charge. Calculation of the electrochemical driving forces suggests that the stoichiometry of HCO3-/Na+ must be larger than two for maintaining HCO3- efflux. Total replacement of bath Cl- with isethionate depolarized Vbl gradually and increased pHi slightly, implying the existence of a Cl(-)-related HCO3- exit mechanism. The rate of decrease in pHi induced by lowering bath HCO3- was slightly reduced (20%) by the absence of bath Cl-. Therefore, the importance of Cl(-)-related HCO3- transport is small relative to total basolateral HCO3- exit. Accordingly, these data suggest that most of HCO3- exits the basolateral membrane through the rheogenic Na+/HCO3- cotransport mechanism with a stoichiometry of HCO3-/Na+ of more than two.

Relation of membrane potential to basolateral TEA transport in isolated snake proximal renal tubules

1995 ◽  
Vol 268 (6) ◽  
pp. R1539-R1545 ◽  
Author(s):  
Y. K. Kim ◽  
W. H. Dantzler
Keyword(s):  
Steady State ◽  
Membrane Potential ◽  
Potent Inhibitor ◽  
Basolateral Membrane ◽  
Potential Difference ◽  
Renal Tubules ◽  
K Concentration ◽  
Electrochemical Equilibrium ◽  
Membrane Potential Difference ◽  
Basolateral Membrane Potential

We measured the effects of changes in bath K+ concentration ([K+]) on basolateral membrane potential difference (PD) and [3H]tetraethylammonium (TEA) transport in isolated snake (Thamnophis) proximal renal tubules (25 degrees C; pH 7.4). Increasing bath [K+] from 3 to 65 mM decreased PD from -60 mV (inside of cells negative) to -20 mV and 2-min uptake of [3H]TEA by approximately 25%, indicating that PD influences TEA entry into the cells. Uptake of [3H]TEA was inhibited similarly at both K+ concentrations by unlabeled TEA, indicating that uptake is carrier mediated. Kt (approximately 18 microM) for 2-min uptake of [3H]TEA in 3 mM K+ increased significantly in 65 mM K+, suggesting that the decrease in PD or the increase in [K+] alters the affinity of the transporter for TEA. The steady-state cell-to-bath ratio for [3H]TEA with 3 mM K+ (-60 mV PD) was approximately 16, significantly above the ratio of 10 predicted for passive distribution at electrochemical equilibrium. With 65 mM K+ (-20 mV PD) this ratio decreased to approximately 6, again significantly above the predicted ratio of 2. These data suggest that the PD can account for much, but not all, of the steady-state uptake of TEA. Efflux of [3H]TEA across the basolateral membrane was identical with either 3 or 65 mM K+ in the bath but was almost completely inhibited in either case by tetrapentylammonium, a potent inhibitor of TEA uptake. These data indicate that virtually all TEA transport across the basolateral membrane is carrier mediated and that transport out of the cells is unaffected by PD.

Intracellular potentials in rabbit proximal tubules perfused in vitro

1981 ◽  
Vol 240 (3) ◽  
pp. F200-F210 ◽  
Author(s):  
B. Biagi ◽  
T. Kubota ◽  
M. Sohtell ◽  
G. Giebisch
Keyword(s):  
Membrane Potential ◽  
Basolateral Membrane ◽  
Proximal Tubules ◽  
Rabbit Kidney ◽  
Electrical Measurements ◽  
Proximal Tubular ◽  
The Mean ◽  
Number Of Cells ◽  
Basolateral Membrane Potential

Conventional microelectrodes were used to measure the basolateral membrane potential (VBL) in isolated perfused superficial proximal convoluted (sPCT) and superficial proximal straight (sPST) tubules of the rabbit kidney. Stable recordings for periods up to 2 h can be obtained. The mean +/- SE (n = number of cells) values of VBL were sPCT = -51.0 +/- 1.63 (24) and sPST = -47.0 +/- 0.97 (94) mV. Inhibitors of active transport, ouabain (10(-5) M) and low bath potassium (0.1 mM), caused a significant depolarization of VBL in sPST. In contrast, short-duration bath cooling (10 degrees C) had no significant effect. Removal of luminal glucose caused a larger hyperpolarization in sPCT (-13.9 +/- 1.77 (9) mV) than in sPST (-3.8 +/- 1.02 (5) mV). Removal of luminal glucose and alanine resulted in an even larger hyperpolarization of VBL in sPCT (-19.0 +/- 0.44 (6) mV). Perfusion of the lumen with a solution resembling late proximal tubular fluid in sPST resulted in hyperpolarization of VBL (-4.3 +/- 0.85 (4) mV). Reducing bath pH to 6.7 depolarized VBL (39.9 +/- 1.77 (13) mV). This effect can be associated with a decrease in the relative potassium permeability of the basolateral membrane. These results demonstrate the feasibility of using intracellular electrical measurements to determine both luminal and basolateral membrane characteristics in isolated proximal tubular segments.

Direct measurement of basolateral membrane potentials from cells of the macula densa

1989 ◽  
Vol 257 (3) ◽  
pp. F463-F468 ◽  
Author(s):  
P. D. Bell ◽  
J. Y. Lapointe ◽  
J. Cardinal
Keyword(s):  
Membrane Potential ◽  
Direct Measurement ◽  
Basolateral Membrane ◽  
Membrane Potentials ◽  
Macula Densa ◽  
Rabbit Kidney ◽  
Fluid Composition ◽  
Nacl Solution ◽  
Basolateral Membrane Potential

At the present time, little is known concerning the electrophysiology of the cells of the macula densa and whether or not these cells are electrically responsive to alterations in luminal fluid composition. To investigate this issue, cortical thick ascending limbs (CTAL) containing macula densa and attached glomeruli were dissected from rabbit kidney and the CTAL perfused in vitro. Basolateral membrane potential (Vbl) was measured with microelectrodes in macula densa cells and, for comparison, in cells of the CTAL. Macula densa Vbl averaged -56.5 +/- 7.6 mV (n = 4) at a (n = 22) at 20 mM NaCl, -35.6 +/- 3.9 mV (n = 16) at 45 mM NaCl, and -25.5 +/- 2.6 mV (n = 32) at 150 mm NaCl. Thus macula densa Vbl depolarized markedly (31 mV) when luminal perfusate [NaCl] was increased from low to high values. In contrast, Vbl measured in CTAL cells averaged -62 +/- 6.1 mV (n = 6) in 45 mM NaCl and did not change significantly as perfusate NaCl was increased to 150 mM. In the presence of 150 mM NaCl, luminal application of furosemide (50 microM) produced a small (3.5 +/- 1.1 mV, n = 16) but statistically significant (P less than 0.02) hyperpolarization in macula densa cells, whereas CTAL cell Vbl hyperpolarized markedly (20 +/- 5.7 mV, n = 6) with addition of furosemide. Finally, neither macula densa cells nor the CTAL cells changed Vbl when 45 mM NaCl solution was made hypotonic by removing mannitol.(ABSTRACT TRUNCATED AT 250 WORDS)

Potassium-induced cell swelling in Necturus gallbladder epithelium

1988 ◽  
Vol 254 (5) ◽  
pp. C643-C650 ◽  
Author(s):  
C. W. Davis ◽  
A. L. Finn
Keyword(s):  
Cell Volume ◽  
Volume Regulation ◽  
Basolateral Membrane ◽  
Cell Volume Regulation ◽  
Cell Swelling ◽  
Gallbladder Epithelium ◽  
Bathing Medium ◽  
Necturus Gallbladder ◽  
Per Se ◽  
Cl Conductance

In Necturus gallbladder epithelium, elevation of mucosal K+ to 95 mM in the presence of 10 mM Na+ resulted in cell swelling at a rate of 3.2% original volume per minute, followed by volume-regulatory shrinking. When Na+ was completely removed from or when amiloride (10(-4) M) was added to the mucosal medium, K+-induced cell swelling was abolished. In the presence of 10 mM Na+, 1 mM Ba2+ abolished and substitution of mucosal Cl- by NO-3 had no effect on K+-induced swelling. Thus solute entry following elevation of mucosal K+ is effected by separate K+ and Cl- pathways. Furthermore, substitution of 95 mM K+ for Na+ in the mucosal bathing medium leads to the development of a Cl- conductance in the basolateral membrane as long as some Na+ remains in the medium. However, cell swelling induced by mucosal dilution does not lead to the appearance of a Cl- conductance. Thus the activation of this conductance requires both swelling and membrane depolarization. These results show that 1) high mucosal K+ leads to cell swelling due to the entry of Cl- along with K+ and the Cl- can enter across either membrane, 2) the Cl- pathways require the presence of mucosal Na+, and 3) cell volume regulation is activated by an increase in volume per se, i.e., a hyposmotic exposure is not required for volume regulation to occur.

pH sensitivity of the basolateral membrane of the rabbit proximal tubule

1986 ◽  
Vol 250 (2) ◽  
pp. F261-F266 ◽  
Author(s):  
B. A. Biagi ◽  
M. Sohtell
Keyword(s):  
Potassium Concentration ◽  
Ph Dependence ◽  
Basolateral Membrane ◽  
Rabbit Kidney ◽  
Bathing Solution ◽  
Solution Ph ◽  
Absolute Value ◽  
Ph Dependent ◽  
Basolateral Membrane Potential

Conventional microelectrodes were used to study the effects of bath pH and bicarbonate concentrations on the basolateral membrane potential (Vbl) of cells from the superficial proximal convoluted (PCT) and proximal straight (PST) tubules of the rabbit kidney perfused in vitro. Bathing solution pH was varied over the range of 5.9-7.4 using either control (22-25 mM) or low bicarbonate (5.0-6.6 mM) Ringer solutions and the appropriate CO2 tensions. The results show a strong pH dependence of the steady-state values of Vbl in both the convoluted and straight tubule segments. The pH-dependent depolarization was approximately 35 mV/pH unit change of the bathing solution in the acid direction and could be demonstrated in CO2-free HEPES-buffered solutions. A depolarizing response to increased bath potassium concentration (HK) was observed that was linearly related to the absolute value of the Vbl under control conditions. Under acidotic conditions, reduced HK depolarizations indicate that a decrease in the relative potassium permeability of the basolateral membrane is the principle mechanism underlying the effects of bath pH on Vbl.

scholarly journals Maxi K+ channels and their relationship to the apical membrane conductance in Necturus gallbladder epithelium.

1990 ◽  
Vol 95 (5) ◽  
pp. 791-818 ◽  
Author(s):  
Y Segal ◽  
L Reuss
Keyword(s):  
Patch Clamp ◽  
Apical Membrane ◽  
Membrane Conductance ◽  
Membrane Depolarization ◽  
Membrane Voltage ◽  
K Channel ◽  
Gallbladder Epithelium ◽  
Patch Clamp Technique ◽  
K Channels ◽  
Necturus Gallbladder

Using the patch-clamp technique, we have identified large-conductance (maxi) K+ channels in the apical membrane of Necturus gallbladder epithelium, and in dissociated gallbladder epithelial cells. These channels are more than tenfold selective for K+ over Na+, and exhibit unitary conductance of approximately 200 pS in symmetric 100 mM KCl. They are activated by elevation of internal Ca2+ levels and membrane depolarization. The properties of these channels could account for the previously observed voltage and Ca2+ sensitivities of the macroscopic apical membrane conductance (Ga). Ga was determined as a function of apical membrane voltage, using intracellular microelectrode techniques. Its value was 180 microS/cm2 at the control membrane voltage of -68 mV, and increased steeply with membrane depolarization, reaching 650 microS/cm2 at -25 mV. We have related maxi K+ channel properties and Ga quantitatively, relying on the premise that at any apical membrane voltage Ga comprises a leakage conductance and a conductance due to maxi K+ channels. Comparison between Ga and maxi K+ channels reveals that the latter are present at a surface density of 0.09/microns 2, are open approximately 15% of the time under control conditions, and account for 17% of control Ga. Depolarizing the apical membrane voltage leads to a steep increase in channel steady-state open probability. When correlated with patch-clamp studies examining the Ca2+ and voltage dependencies of single maxi K+ channels, results from intracellular microelectrode experiments indicate that maxi K+ channel activity in situ is higher than predicted from the measured apical membrane voltage and estimated bulk cytosolic Ca2+ activity. Mechanisms that could account for this finding are proposed.

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