pH-Sensitive Gating Kinetics of the Maxi-K Channel in the Tonoplast of Chara australis

The most frequently observed K⁺ channel in the tonoplast of Characean giant internodal cells with a large conductance (ca. 170 pS; Lühring, 1986; Laver & Walker, 1987) behaves, although inwardly rectifying, like animal maxi-K channels. This channel is accessible for patch–clamp techniques by preparation of cytoplasmic droplets, where the tonoplast forms the membrane delineating the droplet. Lowering the pH of the bathing solution, that virtually mimicks the vacuolar environment, from an almost neutral level to values below pH 7, induced a significant but reversible decrease in channel activity, whereas channel conductance remained largely unaffected. Acidification (pH 5) on both sides of the membrane decreased open probability from a maximum of 80% to less than 20%. Decreasing pH at the cytosolic side inhibited channel activity cooperatively with a slope of 2.05 and a pK _a 6.56. In addition, low pH at the vacuolar face shifted the activating voltage into a positive direction by almost 100 mV. This is the first report about an effect of extraplasmatic pH on gating of a maxi-K channel. It is suggested that the Chara maxi-K channel possesses an S4-like voltage sensor and negatively charged residues in neighboring transmembrane domains whose S4-stabilizing function may be altered by protonation. It was previously shown that gating kinetics of this channel respond to cytosolic Ca²⁺ (Laver & Walker, 1991). With regard to natural conditions, pH effects are discussed as contributing mainly to channel regulation at the vacuolar membrane face, whereas at the cytosolic side Ca²⁺ affects the channel. An attempt was made to ascribe structural mechanisms to different states of a presumptive gating reaction scheme. Received: 8 May 1998/Revised: 18 September 1998     

Anion Channels in Chara corallina Tonoplast Membrane: Calcium Dependence and Rectification

Tonoplast K⁺ channels of Chara corallina are well characterized but only a few reports mention anion channels, which are likely to play an important role in the tonoplast action potential and osmoregulation of this plant. For experiments internodal cells were isolated. Cytoplasmic droplets were formed in an iso-osmotic bath solution according to a modified procedure. Ion channels with conductances of 48 pS and 170 pS were detected by the patch-clamp technique. In the absence of K⁺ in the bath solution the 170 pS channel was not observed at negative pipette potential values. When Cl⁻ on either the vacuolar side or the cytoplasmic side was partly replaced with F⁻, the reversal potential of the 48 pS channel shifted conform to the Cl⁻ equilibrium potential with similar behavior in droplet-attached and excised patch mode. These results showed that the 48 pS channel was a Cl⁻ channel. In droplet-attached mode the channel rectified outward current flow, and the slope conductance was smaller. When Chara droplets were formed in a bath solution containing low (10⁻⁸ m) Ca²⁺, then no Cl⁻ channels could be detected either in droplet-attached or in inside-out patch mode. Channel activity was restored if Ca²⁺ was applied to the cytoplasmic side of inside-out patches. Rectification properties in the inside-out patch configuration could be controlled by the holding pipette potential. Holding potential values negative or positive to the calculated reversal potential for Cl⁻ ions induced opposite rectification properties. Our results show Ca²⁺-activated Cl⁻ channels in the tonoplast of Chara with holding potential dependent rectification. Received: 30 March 1999/Revised: 10 August 1999     

Temperature Sensitivity of the Tonoplast Ca2+-Activated K+ Channel in Chara: The Influence of Reversing the Sign of Membrane Potential

A detailed temperature dependence study of a well-defined plant ion channel, the Ca²⁺-activated K⁺ channel of Chara corallina, was performed over the temperature range of their habitats, 5–36°C, at 1°C resolution. The temperature dependence of the channel unitary conductance at 50 mV shows discontinuities at 15 and 30°C. These temperatures limit the range within which ion diffusion is characterized by the lowest activation energy (E _a = 8.0 ± 1.6 kJ/mol) as compared to the regions below 15°C and above 30°C. Upon reversing membrane voltage polarity from 50 to −50 mV the pattern of temperature dependence switched from discontinuous to linear with E _a = 13.6 ± 0.5 kJ/mol. The temperature dependence of the effective number of open channels at 50 mV showed a decrease with increasing temperature, with a local minimum at 28°C. The mean open time exhibited a similar behavior. Changing the sign of membrane potential from 50 to −50 mV abolished the minima in both temperature dependencies. These data are discussed in the light of higher order phase transitions of the Characean membrane lipids and corresponding change in the lipid-protein interaction, and their modulation by transmembrane voltage. Received: 14 June 2000/Revised: 20 September 2000     

The Effect of Electrical Deformation Forces on the Electropermeabilization of Erythrocyte Membranes in Low- and High-Conductivity Media

Electrical breakdown of erythrocytes induces hemoglobin release which increases markedly with decreasing conductivity of the pulse medium. This effect presumably results from the transient, conductivity-dependent deformation forces (elongation or compression) on the cell caused by Maxwell stress. The deformation force is exerted on the plasma membrane of the cell, which can be viewed as a transient dipole induced by an applied DC electric field pulse. The induced dipole arises from the free charges that accumulate at the cell interfaces via the Maxwell-Wagner polarization mechanism. The polarization response of erythrocytes to a DC field pulse was estimated from the experimental data obtained by using two complementary frequency-domain techniques. The response is very rapid, due to the highly conductive cytosol. Measurements of the electrorotation and electrodeformation spectra over a wide conductivity range yielded the information and data required for the calculation of the deformation force as a function of frequency and external conductivity and for the calculation of the transient development of the deformation forces during the application of a DC-field pulse. These calculations showed that (i) electric force precedes and accompanies membrane charging (up to the breakdown voltage) and (ii) that under low-conductivity conditions, the electric stretching force contributes significantly to the enlargement of ``electroleaks' in the plasma membrane generated by electric breakdown. Received: 12 December 1997/Revised: 13 March 1998     

Membrane Potassium Channels and Human Bladder Tumor Cells. I. Electrical Properties

These experiments were conducted to determine the membrane K⁺ currents and channels in human urinary bladder (HTB-9) carcinoma cells in vitro. K⁺ currents and channel activity were assessed by the whole-cell voltage clamp and by either inside-out or outside-out patch clamp recordings. Cell depolarization resulted in activation of a Ca²⁺-dependent outward K⁺ current, 0.57 ± 0.13 nS/pF at −70 mV holding potential and 3.10 ± 0.15 nS/pF at 30 mV holding potential. Corresponding patch clamp measurements demonstrated a Ca²⁺-activated, voltage-dependent K⁺ channel (K_Ca) of 214 ± 3.0 pS. Scorpion venom peptides, charybdotoxin (ChTx) and iberiotoxin (IbTx), inhibited both the activated current and the K_Ca activity. In addition, on-cell patch recordings demonstrated an inwardly rectifying K⁺ channel, 21 ± 1 pS at positive transmembrane potential (V _m ) and 145 ± 13 pS at negative V _m . Glibenclamide (50 μm), Ba²⁺ (1 mm) and quinine (100 μm) each inhibited the corresponding nonactivated, basal whole-cell current. Moreover, glibenclamide inhibited K⁺ channels in inside/out patches in a dose-dependent manner, and the IC₅₀= 46 μm. The identity of this K⁺ channel with an ATP-sensitive K⁺ channel (K_ATP) was confirmed by its inhibition with ATP (2 mm) and by its activation with diazoxide (100 μm). We conclude that plasma membranes of HTB-9 cells contain the K_Ca and a lower conductance K⁺ channel with properties consistent with a sulfonylurea receptor-linked K_ATP. Received: 12 June 1997/Revised: 21 October 1997     

Ionic Permeability on Isolated Mouse Liver Nuclei: Influence of ATP and Ca2+

Patch-clamp experiments on isolated nuclei revealed the existence of ionic channels on the nuclear envelope, but their exact localization and function are still unknown. Recent studies have demonstrated that ATP and calcium ions play an important role in nucleocytoplasmic protein traffic. ATP is essential to allow big molecules in and out of the nucleus. However, a cytoplasmic rise of calcium ions above 300 nm decreases both ATP-dependent transport and passive diffusion through the nuclear envelope. The use of isolated nuclei placed in a saline solution provides the possibility for testing only the compounds added in the bath or in the recording pipette. In the present study, we show that ATP is responsible for an increase of nuclear ionic permeability on isolated nuclei. This result not only confirms data previously reported in in situ nuclei, but also suggests that ATP is directly involved in the modulation of passive ionic permeability. In these particular experimental conditions, calcium ions decrease the channel current starting from a concentration of 1 μm. The parallelism in the modulation action of ATP and Ca⁺⁺ between nuclear pores and ionic channels present on the nuclear envelope contributes to the support of the idea that an ionic pathway is associated with the pore complex. Received: 5 September 1996/Revised: 13 January 1997     

Kinetic Analysis of the Inhibitory Effect of Glibenclamide on KATP Channels of Mammalian Skeletal Muscle

We investigated the block of K_ATP channels by glibenclamide in inside-out membrane patches of rat flexor digitorum brevis muscle. (1) We found that glibenclamide inhibited K_ATP channels with an apparent K _i of 63 nm and a Hill coefficient of 0.85. The inhibition of K_ATP channels by glibenclamide was unaffected by internal Mg²⁺. (2) Glibenclamide altered all kinetic parameters measured; mean open time and burst length were reduced, whereas mean closed time was increased. (3) By making the assumption that binding of glibenclamide to the sulphonylurea receptor (SUR) leads to channel closure, we have used the relation between mean open time, glibenclamide concentration and K _D to estimate binding and unbinding rate constants. We found an apparent rate constant for glibenclamide binding of 9.9 × 10⁷ m ⁻¹ sec⁻¹ and an unbinding rate of 6.26 sec⁻¹. (4) Glibenclamide is a lipophilic molecule and is likely to act on sulfonylurea receptors from within the hydrophobic phase of the cell membrane. The glibenclamide concentration within this phase will be greater than that in the aqueous solution and we have taken this into account to estimate a true binding rate constant of 1.66 × 10⁶ m ⁻¹ sec⁻¹. Received: 7 July 1996/Revised: 4 October 1996     

Discontinuities in the Temperature Function of Transmembrane Water Transport in Chara: Relation to Ion Transport

The NMR (nuclear magnetic resonance) method of Conlon and Outhred (1972) was used to measure diffusional water permeability of the nodal cells of the green alga Chara gymnophylla. Two local minima at 15 and 30°C of diffusional water permeability (P _d ) were observed delimiting a region of low activation energy (E _a around 20 kJ/mol) indicative of an optimal temperature region for membrane transport processes. Above and below this region water transport was of a different type with high E _a (about 70 kJ/mol). The triphasic temperature dependence of the water transport suggested a channel-mediated transport at 15–30°C and lipid matrix-mediated transport beyond this region. The K⁺ channel inhibitor, tetraethylammonium as well as the Cl⁻ channel inhibitor, ethacrynic acid, diminished P _d in the intermediate temperature region by 54 and 40%, respectively. The sulfhydryl agent p-(chloromercuri-benzensulfonate) the water transport inhibitor in erythrocytes also known to affect K⁺ transport in Chara, only increased P _d below 15°C. In high external potassium (`K-state') water transport minima were pronounced. The role of K⁺ channels as sensors of the optimal temperature limits was further emphasized by showing a similar triphasic temperature dependence of the conductance of a single K⁺ channel also known to cotransport water, which originated from cytoplasmic droplets (putatively tonoplast) of C. gymnophylla. The minimum of K⁺ single channel conductance at around 15°C, unlike the one at 30°C, was sensitive to changes of growth temperature underlining membrane lipid involvement. The additional role of intracellular (membrane?) water in the generation of discontinuities in the above thermal functions was suggested by an Arrhenius plot of the cellular water relaxation rate which showed breaks at 13 and 29°C. Received: 12 August 1998/Revised: 13 November 1998     

Two Kinds of Transient Outward Currents, I A and I Adepol , in F76 and D1 Soma Membranes of the Subesophageal Ganglia of Helix aspersa

Transient outward currents were characterized with twin electrode voltage clamp techniques in isolated F76 and D1 neuronal membranes (soma only) of Helix aspersa subesophageal ganglia. In this study, in addition to the transient outward current (A-current, I _A ) described by Connor and Stevens (1971b), another fast outward current, referred to as I _Adepol here, is described for the first time. This is similar to the current component characterized in Aplysia (Furukawa, Kandel & Pfaffinger, 1992). The separation of these two current components was based on activation and steady-state inactivation curves, holding potentials and sensitivity to 4-aminopyridine (4-AP). In contrast to I _A , I _Adepol did not require hyperpolarizing conditioning pulses to remove inactivation; it was evoked from a holding potential of −40 mV, at which I _A is completely inactivated. I _Adepol shows noticeable activation at around −5 mV, whereas I _A activates at around −50 mV. The time courses of I _Adepol activation and inactivation were similar but slower than I _A . It was found that I _Adepol was more sensitive than I _A to 4-AP. 4-AP at a concentration of 1 mm blocked I _Adepol completely, whereas 5–6 mm 4-AP was needed to block I _A completely. This current is potentially very important because it may, like other A currents, regulate firing frequency but notably, it does not require a period of hyperpolarization to be active. Received: 12 May 2000/Revised: 12 October 2000     

The Actions of Calmodulin Antagonists W-7 and TFP and of Calcium on the Gating Kinetics of the Calcium-activated Large Conductance Potassium Channel of the Chara Protoplasmic Drop: A Substate-sensitive Analysis

The effects of the calmodulin antagonists W-7 and trifluoperazine have been measured on the Ca²⁺-activated potassium channel in the membrane surrounding protoplasmic drops expressed from internodal cells of charophyte plants. The large-conductance (170 pS), voltage- and Ca²⁺-dependent gating, and prominent conductance substrate of this channel shows a strong kinetic resemblance to those of the Maxi-K channel from animal cells. This is the first study of the action of calmodulin antagonists which measures their effects on the most populated substates as well as the closed and main open states of Maxi-K channels. The substate analysis provides new evidence for different modes of action of- and different bindings sites for these calmodulin antagonists. Neither antagonist produces the simple closure of the channel reported previously as its effect on the Maxi-K channel, though both do induce flicker-block, reducing the mean current to near zero at high concentrations following an inverted Michaelis-Menten curve. W-7 reduces residence time in the fully open state, thus raising, in the same proportions, the probabilities of finding the channel in the closed state or a pre-existing substate. Its binding to the channel is voltage- and calcium-dependent. In contrast, trifluoperazine reduces residence in the open state and promotes an apparently new substate which overlaps the closed state at −50 mV but is distinguishable from it at voltages more negative than −100 mV. This substate may represent times that trifluoperazine is bound to the channel. Both antagonists have effects clearly distinguishable from that of withdrawing calcium from the channel, which does not affect open state residence time but increases closed state residence time. Thus neither antagonist reverses the activating effect of Ca²⁻. This is good kinetic evidence against the view that the channel is activated by Ca²⁺-calmodulin and that the effect of a calmodulin antagonist is to reverse this process by making Ca²⁻-calmodulin less available. Received: 26 August 1996/Revised: 7 October 1996     

Role and Source of ATP for Activation of Nonselective Cation Channels by AlF Complex in Guinea Pig Chromaffin Cells

Intracellular dialysis with the solution containing the G protein activator, AlF complex, induced an inward nonselective cation current (I _NS) at −55 mV in chromaffin cells. Amplitudes of I _NS induced by dialysis with ATP-free AlF solutions progressively diminished as cells were pretreated with cyanide, a mitochondrial inhibitor. After a 10-min pretreatment, generation of I _NS by the AlF complex depended on exogenous ATP delivered from pipette solution. The relationship between amplitudes of I _NS and concentrations of MgATP was well expressed by a rectangular hyperbola with an EC₅₀ of 0.265 mm. This result suggests that the cyanide treatment almost depleted ATP near the plasma membrane. On the other hand, a similar cyanide treatment of adrenal medullary preparations did not induce a marked decrease in cellular ATP content. GTP, ITP, or UTP could not substitute for ATP in generation of I _NS by the AlF complex. Similarly, the substitution of ATP with non- or poorly hydrolyzable ATP analogues did not aid in generating I _NS. Bath application of the kinase inhibitor, H-7 (100 μm), suppressed AlF-induced I _NS in a manner depending on intracellular Mg²⁺. We conclude that ATP is a prerequisite for generation of I _NS as a phosphoryl donor and that mitochondria is the main source of ATP. Received: 17 April 1996/Revised: 26 July 1996     

Loss of the gene for the alpha subunit of ATP synthase (ATP5A1) from the W chromosome in the African grey parrot (Psittacus erithacus)

This study describes the results of an analysis using Southern blotting, the polymerase chain reaction, and sequencing which shows that the African grey parrot (Psittacus erithacus) lacks the W-chromosomal gene for the alpha subunit of mitochondrial ATP synthase (ATP5A1W). Additional evidence shows that in other psittacines a fragment of the ATP5A1W gene contains five times as many nonsynonymous nucleotide replacements as the homologous fragment of the Z gene. Therefore, whereas in these other psittacines the corresponding ATP5A1Z protein fragment is highly conserved and varies by only a few, moderately conservative amino acid substitutions, the homologous ATP5A1W fragments contain a considerable number of, sometimes highly nonconservative, amino acid replacements. In one of these species, the ringneck parakeet (Psittacula krameri), the ATP5A1W gene is present in an inactive form because of the presence of a nonsense codon. Other changes, possibly leading to an inactive ATP5A1W gene product, involve the substitution of arginine residues by cysteine in the ATP5A1W protein of the mitred conure (Aratinga mitrata) and the blue and gold macaw (Ara ararauna). The data suggest also that although the divergence of the psittacine ATP5A1W and ATP5A1Z genes preceeded the origin of the psittacidae, this divergence occurred independently of a similar process in the myna (Gracula religiosa), the outgroup used in this study. Received: 6 September 2000 / Accepted: 7 March 2001     

Cobra Venom Cytotoxin Free of Phospholipase A2 and Its Effect on Model Membranes and T Leukemia Cells

Membrane-active toxins from snake venom have been used previously to study protein-lipid interactions and to probe the physical and biochemical states of biomembranes. To extend these studies, we have isolated from Naja naja kaowthia (cobra) venom a cytotoxin free of detectable phospholipase A₂ (PLA₂). The amino acid composition, pI (10.2), and net charge of the cytotoxin compares well with membrane-active toxins isolated from venoms of other cobras. The cytotoxin, shown by a spin label method, associates with PLA₂ in buffers at pH values between 7.0 and 5.0, but not at pH 4.0. It is suggested that cytotoxin and PLA₂ (pI close to 4.8) associate electrostatically in the native venom. The effect of the cytotoxin on model phospholipid membranes was studied by EPR of spin probes in oriented lipid multilayers and ¹H-NMR of sonicated liposomes. The cytotoxin did not significantly affect the packing of lipids in pure phosphatidylcholine (PC) membranes and in PC membranes containing 10 mol% phosphatidic acid (PA) or cardiolipin (CL). However, the cytotoxin induced an increase in membrane permeability and formation of nonbilayer structures in PC membranes containing 40 mol% of PA or CL. The purified cytotoxin was cytocidal to Jurkat cells, but had little effect on normal human lymphocytes. However, both Jurkat cells and normal lymphocytes were killed equivalently when treated with 10⁻⁹ m PLA₂ and 10⁻⁵ m cytotoxin in combination. From its effect on model membranes and Jurkat cells, it is suggested that purified cytotoxin preferentially targets and disrupts membranes that are rich in acidic phospholipids on the extracellular side of the plasma membrane. Received: 20 March 1996/Revised: 25 September 1996     

Influence of Cys-130 S. aureus Alpha-toxin on Planar Lipid Bilayer and Erythrocyte Membranes

Replacement of an amino acid residue at position 130 -Gly by Cys- in the primary structure of Staphylococcus aureus alpha-toxin decreases the single-channel conductance induced by the toxin in planar lipid bilayers. Concomitantly, the pH value at which the channel becomes unable to discriminate between Cl⁻ and K⁺ ions is also decreased. By contrast, the pH dependence of the efficiency of the mutant toxin to form ion channels in lipid bilayers was unchanged (maximum efficiency at pH 5.5–6.0). The asymmetry and nonlinearity of the current-voltage characteristics of the channel were increased by the point mutation but the diameter of the water pore induced by the mutant toxin, evaluated in lipid bilayers and in erythrocyte membranes, was found to be indistinguishable from that formed by wild-type toxin and equal to 2.4–2.6 nm. Alterations at the ``trans mouth' were found to be responsible for all observed changes of the channel properties. This mouth is situated close to the surface of the second leaflet of a bilayer lipid membrane. The data obtained allows us to propose that the region around residue 130 in fact determines the main features of the ST-channel and takes part in the formation of the trans entrance of the channel. Received: 8 September 1995/Revised: 20 November 1996     

Interactions of Primary Amphipathic Vector Peptides with Membranes. Conformational Consequences and Influence on Cellular Localization

The conformations of two peptides produced by the combinations of a nuclear localization sequence and a sequence issued from the fusion protein gp41 of HIV 1 have been analyzed both in solution and in membranes or in membrane mimicking environments. Both are shown to be nonordered in water, α-helical when incorporated into SDS micelles where the helical domain concerns the hydrophobic part of the peptides. Interactions with lipids induce the formation of β-sheet and the lipid-peptide interactions are governed by the nature of the lipid polar headgroups. A monolayer study shows that replacement of the sequence separating the two sequences with an arginine favors the lipid-peptide interactions which may contribute to the understanding of the different, nuclear and membrane associated, cellular localizations of the peptides. Received: 10 October 1997/Revised: 15 January 1998     

Modeling the Current-Voltage Characteristics of Charophyte Membranes. II. The Effect of Salinity on Membranes of Lamprothamnium papulosum

Lamprothamnium is a salt-tolerant charophyte that inhabits a broad range of saline environments. The electrical characteristics of Lamprothamnium cell membranes were modeled in environments of different salinity: full seawater (SW), 0.5 SW, 0.4 SW, and 0.2 SW. The cells were voltage-clamped to obtain the I/V (current-voltage) and G/V (conductance-voltage) profiles of the cell membranes. Cells growing at the different salinities exhibited one of three types of I/V profiles (states): pump-, background- and K(+)-states. This study concentrates on the pump- and background-states. Curved (pump-dominated) I/V characteristics were found in cells with resting membrane PDs (potential differences) of -219 +/- 12 mV (in 0.2 SW: 6 cells, 16 profiles), -161 +/- 12 mV (in 0.4 SW: 6 cells, 7 profiles), -151 +/- 12 mV (in 0.5 SW: 6 cells, 12 profiles) and -137 +/- 12 mV (in full SW: 8 cells, 13 profiles). The linear I/V characteristics of the background-state were found in cells with resting PDs of -107 +/- 12 mV (in 0.4 SW: 7 cells, 12 profiles), -108 +/- 12 mV (in 0.5 SW: 7 cells, 10 profiles) and -104 +/- 12 mV (in full SW: 3 cells, 5 profiles). The resting conductance (G) of the cells progressively increased with salinity, from 0.5 S x m(-2) (in 0.2 SW) to 22.0 S x m(-2) (in full SW). The pump peak conductance only rose from 2 S x m(-2) (0.2 SW) to 5 S x m(-2) (full SW), accounting for the increasingly depolarized resting PD observed in cells in more saline media. Upon exposure to hypertonic medium, both the pump and an inward K+ rectifier were stimulated. The modeling of the I/V profiles identified the inward K+ rectifier as an early electrical response to hypertonic challenge.     

The Peperomia Mitochondrial coxI Group I Intron: Timing of Horizontal Transfer and Subsequent Evolution of the Intron

The Peperomia polybotrya coxI gene intron is the only currently reported group I intron in a vascular plant mitochondrial genome and it likely originated by horizontal transfer from a fungal donor. We provide a clearer picture of the horizontal transfer and a portrayal of the evolution of the group I intron since it was gained by the Peperomia mitochondrial genome. The intron was transferred recently in terms of plant evolution, being restricted to the single genus Peperomia among the order Piperales. Additional support is presented for the suggestion that a recombination/repair mechanism was used by the intron for integration into the Peperomia mitochondrial genome, as a perfect 1:1 correspondence exists between the intron's presence in a species and the presence of divergent nucleotide markers flanking the intron insertion site. Sequencing of coxI introns from additional Peperomia species revealed that several mutations have occurred in the intron since the horizontal transfer, but sequence alterations have not caused frameshifts or created stop codons in the intronic open reading frame. In addition, two coxI pseudogenes in Peperomia cubensis were discovered that lack a large region of coxI exon 2 and contain a truncated version of the group I intron that likely cannot be spliced out. Received: 29 May 1997 / Accepted: 1 November 1997     

Single-Channel Characterization of a Nonselective Cation Channel from Human Placental Microvillous Membranes. Large Conductance, Multiplicity of Conductance States, and Inhibition by Lanthanides

The rate-limiting step for the maternofetal exchange of low molecular-weight solutes in humans is constituted by transport across a single epithelial layer (syncytiotrophoblast) of the placenta. Other than the well-established presence of a large-conductance, multisubstate Cl⁻ channel, the ionic channels occurring in this syncytial tissue are, for the most part, unknown. We have found that fusion of apical plasma membrane-enriched vesicle fractions with planar lipid bilayers leads, mainly (96% of 353 reconstitutions), to the reconstitution of nonselective cation channels. Here we describe the properties of this novel placental conductance at the single-channel level. The channel has a large (>200 pS) and variable conductance, is cation selective (P _Cl /P _K ≅ 0.024), is reversibly inhibited (presumably blocked) by submillimolar La³⁺, has very unstable kinetics, and displays a large number (>10) of current sublevels with a ``promiscuous' connectivity pattern. The occurrence of both ``staircaselike' and ``all-or-nothing' transitions between the minimum and maximum current levels was intriguing, particularly considering the large number of conductance levels spanned at a time during the concerted current steps. Single-channel data simulated according to a multistate linear reaction scheme, with rate constants that can vary spontaneously in time, reproduce many aspects of the recorded subconductance behavior. The channel's sensitivity to lanthanides is reminiscent of stretch-sensitive channels which, in turn, suggests a physiological role for this ion channel as a mechanotransducer during syncytiotrophoblast-volume regulation. Received: 30 August 1999/Revised: 12 November 1999