scholarly journals Intracellular K+ activity and its relation to basolateral membrane ion transport in Necturus gallbladder epithelium.

1980 ◽  
Vol 76 (1) ◽  
pp. 33-52 ◽  
Author(s):  
L Reuss ◽  
S A Weinman ◽  
T P Grady
Keyword(s):  
Cell Membrane ◽  
Amphotericin B ◽  
Basolateral Membrane ◽  
Membrane Potentials ◽  
Equilibrium Potential ◽  
Gallbladder Epithelium ◽  
Bathing Medium ◽  
Necturus Gallbladder ◽  
Intracellular K Activity ◽  
K Activity

A study of the mechanisms of the effects of amphotericin B and ouabain on cell membrane and transepithelial potentials and intracellular K activity (alpha Ki) of Necturus gallbladder epithelium was undertaken with conventional and K-selective intracellular microelectrode techniques. Amphotericin B produced a mucosa-negative change of transepithelial potential (Vms) and depolarization of both apical and basolateral membranes. Rapid fall of alpha Ki was also observed, with the consequent reduction of the K equilibrium potential (EK) across both the apical and the basolateral membrane. It was also shown that, unless the mucosal bathing medium is rapidly exchanged, K accumulates in the unstirred fluid layers near the luminal membrane generating a paracellular K diffusion potential, which contributes to the Vms change. Exposure to ouabain resulted in a slow decrease of alpha Ki and slow depolarization of both cell membranes. Cell membrane potentials and alpha Ki could be partially restored by a brief (3-4 min) mucosal substitution of K for Na. Under all experimental conditions (control, amphotericin B, and ouabain), EK at the basolateral membrane was larger than the basolateral membrane equivalent emf (Eb). Therefore, the K chemical potential difference appears to account for Eb and the magnitude of the cell membrane potentials, without the need to postulate an electrogenic Na pump. Comparison of the rate of Na transport across the tissue with the electrodiffusional K flux across the basolateral membrane indicates that maintenance of a steady-state alpha Ki cannot be explained by a simple Na,K pump-K leak model. It is suggested that either a NaCl pump operates in parallel with the Na,K pump, or that a KCl downhill neutral extrusion mechanism exists in addition to the electrodiffusional K pathway.

Potassium distribution and membrane potential of sensory neurons in the leech nervous system

1984 ◽  
Vol 51 (4) ◽  
pp. 689-704 ◽  
Author(s):  
W. R. Schlue ◽  
J. W. Deitmer
Keyword(s):  
Nervous System ◽  
Membrane Potential ◽  
Sensory Neurons ◽  
Membrane Resistance ◽  
Equilibrium Potential ◽  
Bathing Medium ◽  
Membrane Hyperpolarization ◽  
K Concentration ◽  
Intracellular K Activity ◽  
K Activity

The intracellular K activity (aKi) and membrane potential of sensory neurons in the leech central nervous system were measured in normal and altered external K+ concentrations, [K+]o, using double-barreled, liquid ion-exchanger microelectrodes. In control experiments membrane potential measurements were made using potassium chloride-filled single-barreled microelectrodes. All values are means +/- SD. At the normal [K+]o (4 mM) the mean aKi of all cells tested was 72.6 +/- 10.6 mM (n = 40) and the average membrane potential was -47.3 +/- 5.2 mM (n = 40). When measured with single-barreled microelectrodes, the membrane potential averaged -45.3 +/- 2.9 mV (n = 12). Assuming an intracellular K+ activity coefficient of 0.75, the intracellular K+ concentration of sensory neurons would be 96.8 +/- 14.1 mM). With an extracellular K+ concentration of 5.8 mM in the intact ganglion compared to the K+ concentration of 4 mM in the bath, the K+ equilibrium potential was -71.5 mV. When the ganglion capsule was opened, the extracellular K+ concentrations in the ganglion were similar to that of the bathing medium and the calculated K+ equilibrium potential was -81 mV. The membrane of sensory neurons depolarized following the changes to elevated [K+]o (greater than or equal to 10-100 mM), whereas aKi changed only little or not at all. At very low [K+]o (0.2, 0 mM) aKi and membrane potential showed little short-term (less than 3 min) effect but began to change after longer exposure (greater than 3 min). Reduction of [K+]o from 4 to 0.2 mM (or 0 mM) produced first a slow, and then a more rapid decrease of aKi and membrane resistance, accompanied by a slow membrane hyperpolarization. Following readdition of normal [K+]o, the membrane first depolarized and then transiently hyperpolarized, eventually returning slowly to the normal membrane potential.(ABSTRACT TRUNCATED AT 400 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.

Measurements of the Intracellular Potassium Activity of Retzius Cells in the Leech Central Nervous System

1981 ◽  
Vol 91 (1) ◽  
pp. 87-101
Author(s):  
JOACHIM W. DEITMER ◽  
WOLF R. SCHLUE
Keyword(s):  
Cell Membrane ◽  
External Solution ◽  
Equilibrium Potential ◽  
The Mean ◽  
K Concentration ◽  
Retzius Cells ◽  
Intracellular K Activity ◽  
K Permeability ◽  
K Activity ◽  
External K

The intracellular K activity of leech Retzius cells was measured using double-barrelled, liquid ion exchanger, microelectrodes. At the normal external K+ concentration of 4 mm (equivalent to 3 mm-K activity, assuming an activity coefficient of 0.75) the mean K activity was 101.3 ± 7.6 mm (S.D., n = 14) in the cell bodies, and 4.35 ± 0.4 mV (n = 27) in the extracellular spaces surrounding them, indicating a K+ equilibrium potential of - 80 mV. The mean membrane potential was - 43.6 + 4.9 mV (n = 14). In a K-free external solution, or in the presence of 5 × 10−4m-ouabain, the intracellular K activity decreased by up to 14 mm min−1. This indicates an efflux of K+ ions across the cell membrane of approximately 2 × 10−10 mol cm−2s, and an apparent K+ permeability coefficient of 8 × 10−8 cms−1. The cell membrane depolarized upon removal of K+ and upon addition of ouabain, and transiently hyperpolarized beyond its initial level on return to the normal external K+ concentration. The recovery from this hyperpolarization paralleled the increase of the intracellular K activity following the re-addition of K+. Our results suggest that, despite the high K+ permeability of the Retzius cell membrane, the intracellular K activity is maintained at a high level by an electrogenic pump.

Interaction between carbachol and vasoactive intestinal peptide in cells of isolated colonic crypts

1992 ◽  
Vol 262 (5) ◽  
pp. G940-G944
Author(s):  
L. Greenwald ◽  
B. A. Biagi
Keyword(s):  
Vasoactive Intestinal Peptide ◽  
Chloride Transport ◽  
Basolateral Membrane ◽  
Distal Colon ◽  
Equilibrium Potential ◽  
Colonic Crypts ◽  
Intracellular K Activity ◽  
Crypt Cells ◽  
Basolateral Membrane Potential ◽  
K Activity

In a previous study [B. Biagi, Y.-Z. Wang, and H. J. Cooke, Am. J. Physiol. 258 (Gastrointest. Liver Physiol. 21): G223-G230, 1990], carbachol stimulated active chloride transport in rabbit distal colon, yet had no effect on the basolateral membrane potential (Vbl) of cells from isolated crypts from the same tissue. In the present study, crypt cells were first depolarized with vasoactive intestinal peptide (VIP; 1 x 10(-9) M) (control Vbl = -62 mV; VIP Vbl = -48 mV) and then exposed to carbachol in the presence of VIP. The VIP-induced depolarization of Vbl was completely reversed by carbachol (0.1 mM; repolarization to -65 mV). Similar repolarization was seen by applying carbachol to crypt cells depolarized by 10 mM aminophylline. Intracellular K+ activity (aiK), measured with K(+)-selective microelectrodes, was 64.3 mM (concn = 85 mM), yielding a K+ equilibrium potential (EK+) of -76 mV. Neither carbachol nor VIP application caused significant changes in aiK. These results demonstrate the presence of cholinergic receptors on colonic crypt cells. The magnitude of the carbachol effect on Vbl is greater when Vbl is depolarized relative to EK+. The results are consistent with the hypothesis that carbachol acts by increasing basolateral K+ conductance, driving the cell toward the EK+.

scholarly journals Electrophysiological effects of extracellular ATP on Necturus gallbladder epithelium.

1991 ◽  
Vol 97 (5) ◽  
pp. 949-971 ◽  
Author(s):  
C U Cotton ◽  
L Reuss
Keyword(s):  
Cell Membrane ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Active Agent ◽  
Membrane Conductance ◽  
Membrane Resistance ◽  
Initial Increase ◽  
Bathing Solution ◽  
Gallbladder Epithelium ◽  
Necturus Gallbladder

The effects of addition of ATP to the mucosal bathing solution on transepithelial, apical, and basolateral membrane voltages and resistances in Necturus gallbladder epithelium were determined. Mucosal ATP (100 microM) caused a rapid hyperpolarization of both apical (Vmc) and basolateral (Vcs) cell membrane voltages (delta Vm = 18 +/- 1 mV), a fall in transepithelial resistance (Rt) from 142 +/- 8 to 122 +/- 7 omega.cm2, and a decrease in fractional apical membrane resistance (fRa) from 0.93 +/- 0.02 to 0.83 +/- 0.03. The rapid initial hyperpolarization of Vmc and Vcs was followed by a slower depolarization of cell membrane voltages and a lumen-negative change in transepithelial voltage (Vms). This phase also included an additional decrease in fRa. Removal of the ATP caused a further depolarization of membrane voltages followed by a hyperpolarization and then a return to control values. fRa fell to a minimum after removal of ATP and then returned to control values as the cell membrane voltages repolarized. Similar responses could be elicited by ADP but not by adenosine. The results of two-point cable experiments revealed that ATP induced an initial increase in cell membrane conductance followed by a decrease. Transient elevations of mucosal solution [K+] induced a larger depolarization of Vmc and Vcs during exposure to ATP than under control conditions. Reduction of mucosal solution [Cl-] induced a slow hyperpolarization of Vmc and Vcs before exposure to ATP and a rapid depolarization during exposure to ATP. We conclude that ATP4- is the active agent and that it causes a concentration-dependent increase in apical and basolateral membrane K+ permeability. In addition, an apical membrane electrodiffusive Cl- permeability is activated by ATP4-.

Electrophysiological effects of mucosal Cl- removal in Necturus gallbladder epithelium

1989 ◽  
Vol 257 (3) ◽  
pp. C568-C578 ◽  
Author(s):  
J. S. Stoddard ◽  
L. Reuss
Keyword(s):  
Cell Membrane ◽  
Cell Membranes ◽  
Equilibrium Potential ◽  
Bathing Solution ◽  
Gallbladder Epithelium ◽  
Membrane Hyperpolarization ◽  
Necturus Gallbladder ◽  
Partial Pressure Of Co2 ◽  
K Concentration ◽  
Electrophysiological Effects

The factors responsible for the cell membrane hyperpolarization elicited in Necturus gallbladder epithelium on Cl- removal from the mucosal bathing solution were evaluated with conventional and ion-sensitive microelectrode techniques. Cl- removal causes reversal of apical Cl- -HCO3- exchange, resulting in a fall in intracellular Cl- activity (aiCl) and an increase in intracellular pH (pHi). Concomitantly, the cell membranes hyperpolarize to values close to the K+ equilibrium potential (EK), aiNa falls, and aiK rises. The observed changes in membrane voltage are not attributable to a pHi-dependent increase in cell membrane K+ permeability (PK), because 1) the cell membrane resistances increased and 2) elevating solution partial pressure of CO2 (PCO2) to counterbalance the cellular alkalinization on mucosal Cl- removal caused a further hyperpolarization of the cell membranes to values greater than EK. This additional hyperpolarization was related to the activity of the Na+ pump, inasmuch as it was accompanied by an increase in aiNa and was ouabain sensitive. These results are consistent with, but do not prove, pump electrogenicity. During the period of Cl- removal from the mucosal bathing solution, the cell membrane depolarization caused by raising serosal K+ concentration was increased, whereas the depolarization caused by lowering serosal Cl- concentration was decreased, compared with substitutions under control conditions. These results indicate that mucosal Cl- removal causes a decrease in basolateral PCl, which we speculate could be due to a decrease in cell volume. We conclude that the hyperpolarization of the cell membranes on mucosal Cl- removal is primarily due to the combined effects of the fall in basolateral PCl and the increase in basolateral ECl.

Artifactual expression of maxi-K+ channels in basolateral membrane of gallbladder epithelial cells

1993 ◽  
Vol 264 (5) ◽  
pp. C1128-C1136 ◽  
Author(s):  
J. Copello ◽  
F. Wehner ◽  
L. Reuss
Keyword(s):  
Epithelial Cells ◽  
Cell Membrane ◽  
Patch Clamp ◽  
Background Noise ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Gallbladder Epithelium ◽  
K Channels ◽  
Necturus Gallbladder ◽  
Apical Side

To patch clamp the basolateral cell membrane, sheets of Necturus gallbladder epithelium were stripped of the subepithelial tissue layers and affixed apical side down on cover slips coated with Cell-Tak [F. Wehner, L. Garretson, K. Dawson, Y. Segal, and L. Reuss. Am. J. Physiol. 258 (Cell Physiol. 27): C1159-C1164, 1990]. In 90% of the patches we observed K+ channels identical to the maxi-K+ channels previously demonstrated in the apical membrane (Y. Segal and L. Reuss. J. Gen. Physiol. 95: 791-818, 1990). To ascertain whether these channels were present in the native tissue, we carried out intracellular-microelectrode studies. We tested for activation of basolateral membrane K+ conductance by depolarization or by elevation of intracellular Ca2+ and for tetraethylammonium sensitivity of the basolateral membrane voltage and fractional resistance. The results were negative, indicating that maxi-K+ channels are not expressed in the basolateral membrane of the "intact" epithelium. Using the same intracellular-microelectrode protocol on the apical membrane, we demonstrated the presence of an apical K+ conductance attributable to maxi-K+ channels. Additional experiments revealed a Ba(2+)-sensitive basolateral K+ conductance in the native epithelium. We conclude that in the stripped preparation there is artifactual expression of maxi-K+ channels. In addition, the native basolateral membrane K+ channels either are not expressed in this preparation or have a low conductance and cannot be discerned from the background noise.

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.

Association between HCO3(-) absorption and K+ uptake by Amphiuma jejunum: relations among HCO3(-) absorption, luminal K+, and intracellular K+ activity

1984 ◽  
Vol 246 (6) ◽  
pp. G732-G744
Author(s):  
M. A. Imon ◽  
J. F. White
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Uptake Mechanism ◽  
Bathing Medium ◽  
The Media ◽  
Absorptive Flux ◽  
Free Media ◽  
Intracellular K Activity ◽  
K Activity

Titration techniques and K+- sensitive microelectrodes have been used to investigate the relations among HCO3(-) absorption, luminal K+, and intracellular K+ activity in in vitro Amphiuma jejunum. The HCO3(-) absorptive flux (JHCO3(-] measured by pH-stat under short circuit was reduced by removal of K+ from the medium but not by replacement of Na+ with choline. JHCO3(-) exhibited a seasonal variation when K+ was absent from the media and was increased to a maximum when K+ equaled 5 mM. Addition of K+ to a K+-free luminal medium stimulated JHCO3(-) much more than addition to the serosal medium. Acetazolamide (10(-4) M) blocked K+-stimulated HCO3(-) absorption while benzolamide reduced the short-circuit current associated with HCO3(-) absorption much more rapidly when added to the mucosal bathing medium. Intracellular K+ activity (aik) and mucosal membrane potential (psi m) of jejunal villus cells were measured with double-barreled microelectrodes. When bathed bilaterally with HCO3(-)-containing media, K+ was actively accumulated for many hours (aik = 58.5 mM) but in the presence of ouabain fell to equilibrium (16 mM) after 2 h. In contrast, when HCO3(-) absorption was induced by removal of serosal HCO3(-), aik was elevated to 83.6 mM and, after 4-h exposure to ouabain cell K+, remained far above electrochemical equilibrium at 33 mM. Tissues bathed in Na+-free (Tris) media containing ouabain retained cell K+ after 4 h at even higher levels (46 mM). Cell K+ activity was reduced by removal of K+ from either the mucosal or serosal medium. Acetazolamide reduced aik over 2 h in Na+-free media from 66 to 42 mM. The decline in aik was associated with a concomitant decline in the HCO3(-) absorptive current. It is concluded that K+ is actively accumulated across both luminal and serosal membranes of the jejunal absorptive cell and that the luminal uptake mechanism is linked to HCO3(-) absorption or an equivalent process.

Microelectrode measurements of K+ and pH in rabbit gastric glands: effect of histamine

1984 ◽  
Vol 246 (4) ◽  
pp. G433-G444
Author(s):  
K. Kafoglis ◽  
S. J. Hersey ◽  
J. F. White
Keyword(s):  
Membrane Potential ◽  
Ion Exchange ◽  
Basolateral Membrane ◽  
Free Medium ◽  
Na Transport ◽  
K Uptake ◽  
Gastric Glands ◽  
Microelectrode Measurements ◽  
Intracellular K Activity ◽  
K Activity

Conventional and liquid ion-exchange microelectrodes sensitive to K+ or pH were used to examine the response of isolated rabbit gastric glands to histamine. The epithelial cells were impaled across the basolateral membrane. The membrane potential averaged -6.1 +/- 0.6 mV and was unchanged after replacement of medium K+, Cl-, or Na+. The intracellular K+ activity (alpha iK) averaged 41.3 +/- 3.0 mM, indicating K+ accumulation by a factor of 6.8. Active accumulation of K+ was eliminated by ouabain. In contrast, histamine increased K+ activity to 55.3 +/- 3.9 mM. This stimulation was blocked by ouabain. In glands bathed in a Na+-free medium containing ouabain, addition of histamine elevated alpha iK from 12.5 +/- 0.7 to 17.1 +/- 1.1 mM. Isobutylmethylxanthine (10(-4) M) also elevated alpha iK. When impaled with pH-sensitive microelectrodes, glands exposed to histamine exhibited regions of acidity as low as pH 3. Acidification was also produced by histamine after medium Na+ had been replaced with choline. Picoprazole (H 149/94) blocked the effects of histamine on alpha iK and gland pH. The results are consistent with the view that histamine-induced acid secretion by gastric glands is associated with K+ uptake by a mechanism that is independent of Na+ transport but is inhibited by intracellular Na+. This is most likely the H+-K+-ATPase on the secretory surface of the gland cells. Evidence that some tissue K+ is bound or compartmentalized is also discussed.

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