pH-dependent electrical properties and buffer permeability of the Necturus renal proximal tubule cell

Necturus kidneys were perfused with Tris-buffered solutions at three different pH values, i.e. 7.5, 6.0 and 9.0. A significant drop in fluid absorption occurred at pH 6.0, whereas pH 9.0 did not increase volume flow significantly. When acute unilateral, i.e. either in the lumen or the peritubular capillaries, and bilateral pH changes were elicited in both directions from 7.5 to 9.0 at a constant Tris-butyrate buffer concentration, both peritubular membrane potential difference V1 and transepithelial potential difference V3 hyperpolarized, independently of the side where the change in pH was brought about. Acid perfusions at pH 6.0 caused a similar response but of opposite sign. Analysis of the potential changes shows that pH influences not only the electromotive force and resistance of the homolateral membrane, but also the electrical properties of the paracellular path. Interference of pH with Na, Cl or K conductance was assessed. Any appreciable role for sodium or chloride was excluded, whereas the potassium transference number (tK) of the peritubular membrane increased 16% in alkaline pH. However, this increase accounts only for 19 to 36% of the observed hyperpolarization. Since changes in Tris-butyrate buffer concentration at constant pH do not affect V1 or V3 considerably, the hyperpolarization in pH 9 cannot be explained by an elevation in internal pH only, or by a Tris-H+ ion diffusion potential only. The role of the permeability of the buffers: bicarbonate, butyrate and phosphate, in determining electrical membrane parameters was evaluated. Transport numbers of the buffer anions ranked as follows: tHCO3 greater than tbutyrate greater than tphosphate. It is concluded that modulation of membrane potential by extracellular pH is mediated primarily by a change in peritubular cell membrane tK and additionally by membrane currents carried by buffer anions.     

Regulation of the basolateral potassium conductance of theNecturus proximal tubule

Summary Two methods, the measurement of the response of the basolateral membrane potential (V _bl) of proximal tubule cells ofNecturus to step changes in basolateral K⁺ concentration, and cellular cable analysis, were used to assess the changes in basolateral potassium conductance (G _K) caused by a variety of maneuvers. The effects of some of these maneuvers on intracellular K⁺ activity (a _K ⁱ ) were also evaluated using double-barreled ion-selective electrodes. Perfusion with 0mm K⁺ basolateral solution for 15 min followed by 45 min of 1mm K⁺ solution resulted in a fall in basolateral potassium (apparent) transference number (t _K),V _bl anda _K ⁱ . Results of cable analysis showed that total basolateral resistance,R _b , rose. The electrophysiological effects of additional manipulations, known to inhibit net sodium reabsorption across the proximal tubular epithelium ofNecturus, were also investigated. Ouabain caused a fall int _K accompanied by large decreases ina _K ⁱ andV _bl. Lowering luminal sodium caused a fall int _K and a small reduction inV _bl. Selective reduction of peritubular sodium, a maneuver that has been shown to block sodium transport from lumen to peritubular fluid, also resulted in a significant decrease int _K. These results suggest thatG _K varies directly with rate of transport of the sodium pump, irrespective of the mechanism of change in pump turnover.Part of this material has been presented at the 10th International Conference on Biological Membranes (Cohen & Giebisch, 1984).     

A kinetic analysis of the electrogenic pump ofChara corallina: II. Dependence of the pump activity on external pH

Summary The current-voltage curve of theChara membrane was obtained by applying a slow ramp de- and hyperpolarization by use of voltage clamp. By inhibiting the electrogenic pump with 50m DCCD (dicyclohexylcarbodiimide), theI–V curve approached a steady state within 100 min, which gave thei _d -V curve of the passive diffusion channel. Thei _p -V curve of the electrogenic pump channel was obtained by subtracting the latter from the former. With the increase of external pH, thei _d -V curve showed only a slight change, while thei _p -V curve of the pump channel showed almost a parallel shift, in the hyperpolarizing direction, along the voltage axis in the pH range between 6.5 and 7.5. The sigmoidali _p -V curve in this pH range could be simulated satisfactorily with the five-state model reported previously (U. Kishimoto, N. Kami-ike, Y. Takeuchi & T. Ohkawa,J. Membrane Biol. 80:175–183, 1984) as well as with a lumped two-state model presented in this report. The analysis based on these models suggests that the electrogenic pump of theChara membrane is mainly a 2H⁺/1ATP pump. The forward rate constant in the voltage-dependent step increased with the increase of external pH, while the backward one decreased. On the other hand, the forward rate constant in the voltage-independent step remained almost unchanged with the increase of external pH, while the backward one increased markedly. The pump conductance at the resting membrane potential showed either a slight increase or a decrease with the increase of external pH, depending on the sample. Nevertheless, the pump current showed generally a slight increase with the increase of external pH.     

Effect of entomocidal proteins from <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> on ion permeability of apical membranes of <Emphasis Type="Italic">Tenebrio molitor</Emphasis> larvae gut epithelium

Effects of entomocidal Cry-type proteins, δ-endotoxins Cry3A and Cry11A produced by Bacillus thuringiensis, on ion permeability of the apical membranes of intestinal epithelium from Tenebrio molitor larvae midgut were studied. Using potential-sensitive dyes safranine O and oxonol VI and δpH indicator acridine orange, it was shown that placing brush border membrane vesicles (BBMV) (loaded with Mg²⁺ during their preparation) into a salt-free buffer medium resulted in spontaneous generation of transmembrane electric potential on the vesicular membrane (negative inside the vesicles) accompanied by acidification of the aqueous phase inside the vesicles. The generation of transmembrane ion gradients on the vesicular membrane was a result of an electrogenic efflux of Mg²⁺ from the vesicles as shown by abolishing of the membrane potential by such agents as MgSO₄ or CaCl₂ in centimolar concentrations, a highly lipophilic cation tetraphenylphosphonium, and some blockers of cell membrane Ca²⁺-channels in submillimolar concentrations. A passive generation of membrane potential on the vesicular membrane (but positive inside the vesicles) was also observed upon addition of centimolar concentrations of K₂SO₄. Addition of δ-endotoxins Cry3A and Cry11A to the vesicle suspension in a salt-free buffer medium or in the same medium supplemented with centimolar concentrations of K₂SO₄ exerted a pronounced hyperpolarization of the vesicular membrane. This hyperpolarization was sensitive to the same agents, which abolished the membrane potential generation in the absence of δ-endotoxin. It is concluded that Cry proteins induced in BBMV from T. molitor opening pores or ion channels, which were considerably more permeable for alkaline- and alkaline-earth metal cations than for the accompanying anions.     

Effects of carbon dioxide on the spatially separate electrogenic ion pumps and the growth rate in the hypocotyl ofVigna sesquipedalis

Carbon dioxide, introduced into the gas phase of the experimental chamber, has distinct effects on two spatially separate membrane potentials and the rate of elongation growth in hypocotyl segments ofVigna sesquipedalis Wight. Both membrane potentials (V _ps andV _px=the electric potential difference between the parenchyma symplast and the surface of the hypocotyl, and that between the parenchyma symplast and the xylem, respectively) hyperpolarized rapidly but transiently at the introduction of CO₂. Prolonged exposure of the hypocotyl to high concentrations of CO₂ (above 10%) caused depolarization of membrane potentials above the level before CO₂ introduction. When CO₂ was replaced with air, the membrane potentials exhibited a distinct depolarization response of transient nature. The growth rate of the hypocotyl segments exhibited similar responses to CO₂ as did the membrane potentials (the increase and the decrease of the growth rate were corresponded to the hyperpolarization and the depolarization, respectively), but these responses always followed the changes of the membrane potentials. The CO₂-induced maximum hyperpolarization ofV _ps and the maximum increase of the growth rate were closely correlated. All these responses were strictly dependent on aerobic metabolism. These results indicate that CO₂ may regulate elongation growth in two ways: by affecting the activity of the electrogenic ion pump via intracellular acidification, and also by acting via apoplastic acidification as a wall-loosening acid.Symbols and abbreviations V _sx electric potential difference between the surface (S) and the xylem (X) of the hypocotyl - V _px electric potential difference between the inside of a parenchyma cell (P) andX - V _ps electric potential difference betweenP andS - V _ps (CO₂, max) the maximum value of CO₂-induced hyperpolarization ofV _ps - GR(CO₂, max) the maximum value of CO₂-induced increase of the growth rate - IAA indole-3-acetic acid     

Evaluation of ion gradient-dependent H+ transport systems in isolated enterocytes from the chick

Summary The experiments reported here evaluate the capability of isolated intestinal epithelial cells to accomplish net H⁺ transport in response to imposed ion gradients. In most cases, the membrane potential was kept constant by means of a K⁺ plus valinomycin voltage clamp in order to prevent electrical coupling of ion fluxes. Net H⁺ flux across the cellular membrane was examined at pH 6.0 (the physiological lumenal pH) and at pH 7.4 using methylamine distribution or recordings of changes in media pH. Results from both techniques suggest that the cells have an Na⁺/H⁺ exchange system in the plasma membrane that is capable of rapid and sustained changes in intracellular pH in response to an imposed Na⁺ gradient. The kinetics of the Na⁺/H⁺ exchange reaction at pH 6.0 [K _t for Na⁺=57mm,V _max=42 mmol H⁺/liter 3OMG (3-O-methylglucose) space/min] are dramatically different from those at pH 7.4 (K _t for Na⁺=15mm,V _max=1.7 mmol H⁺/liter 3OMG space/min). Experiments involving imposed K⁺ gradients suggest that these cells have negligible K⁺/H⁺ exchange capability. They exhibit limited but measurable H⁺ conductance. Anion exchange for base equivalents was not detected in experiments performed in media nominally free of bicarbonate.     

Electrical effects of potassium and bicarbonate on proximal tubule cells ofNecturus

Summary The effects of stepwise concentration changes of K⁺ and HCO ₃ ^– in the basolateral solution on the basolateral membrane potential (V _bl) of proximal tubule cells of the doubly-perfusedNecturus kidney were examined using conventional microelectrodes. Apparent transference numbers were calculated from changes inV _bl after alterations in external K⁺ concentration from 1.0 to 2.5mm (t _{K, 1.0–2.5}), 2.5 to 10, and in external HCO ₃ ^– concentration (at constant pH) from 5 to 10mm (t _{HCO3, 5–10}), 10 to 20, or 10 to 50.t _{K, 2.5–10} was 0.38±0.02 under control conditions but was sharply reduced to 0.08±0.03 (P>0.001) by 4mm Ba⁺⁺. This concentration of Ba⁺⁺ reducedV _bl by 9±1 mV (at 2.5 external K⁺). Perfusion with SITS (5×10^–4 m) for 1 hr hyperpolarizedV _bl by 10±3 mV and increasedt _{K, 2.5–10} significantly to 0.52±0.01 (P<0.001). Ba⁺⁺ application in the presence of SITS depolarizedV _bl by 22±3 mV. In control conditionst _{HCO3, 10–50} was 0.63±0.05 and was increased to 0.89±0.07 (P<0.01) by Ba⁺⁺ but was decreased to 0.14±0.02 (P<0.001) by SITS. In the absence of apical and basolateral chloride, the response ofV _bl to bicarbonate was diminished but still present (t _{HCO3, 10–20} was 0.35±0.03). Intracellular pH, measured with liquid ion-exchange microelectrodes, increased from 7.42±0.19 to 7.57±0.17 (P<0.02) when basolateral bicarbonate was increased from 10 to 20mm at constant pH. These data show that the effects of bicarbonate onV _bl are largely independent of effects on the K⁺ conductance and that there is a significant current-carrying bicarbonate pathway in the basolateral membrane. Hence, both K⁺ and HCO ₃ ^– gradients are important in the generation ofV _bl, and their relative effects vary reciprocally.     

Electrical properties of the plasma membrane of microplasmodia ofPhysarum polycephalum

Summary Microplasmodia ofPhysarum polycephalum have been investigated by conventional electrophysiological techniques. In standard medium (30mm K⁺, 4mm Ca⁺⁺, 3mm Mg⁺⁺, 18mm citrate buffer, pH 4.7, 22°C), the transmembrane potential differenceV _m is around –100 mV and the membrane resistance about 0.25 m².V _m is insensitive to light and changes of the Na⁺/K⁺ ratio in the medium. Without bivalent cations in the medium and/or in presence of metabolic inhibitors (CCCP, CN^–, N ₃ ^– ),V _m drops to about 0 mV. Under normal conditions,V _m is very sensitive to external pH (pH _o ), displaying an almost Nernstian slope at pH _o =3. However, when measured during metabolic inhibition,V _m shows no sensitivity to pH _o over the range 3 to 6, only rising (about 50 mV/pH) at pH _o =6. Addition of glucose or sucrose (but not mannitol or sorbitol) causes rapid depolarization, which partially recovers over the next few minutes. Half-maximal peak depolarization (25 mV with glucose) was achieved with 1mm of the sugar. Sugar-induced depolarization was insensitive to pH _o . The results are discussed on the basis of Class-I models of charge transport across biomembranes (Hansen, Gradmann, Sanders and Slayman, 1981,J. Membrane Biol. 63:165–190). Three transport systems are characterized: 1) An electrogenic H⁺ extrusion pump with a stoichiometry of 2 H⁺ per metabolic energy equivalent. The deprotonated form of the pump seems to be negatively charged. 2) In addition to the passive K⁺ pathways, there is a passive H⁺ transport system; here the protonated form seems to be positively charged. 3) A tentative H⁺-sugar cotransport system operates far from thermodynamic equilibrium, carrying negative charge in its deprotonated states.     

The function of tonoplast ATPase in intact vacuoles of red beet is governed by direct and indirect ion effects

The pH gradient and the electric potential across the tonoplast in mechanically isolated beetroot vacuoles has been studied by following the uptake of [¹⁴C]methylamine and [¹⁴C]triphenyl-methylphosphoniumchloride. In response to Mg-ATP, the vacuolar interior is acidified by 0.8 units. This strong acidification is accompanied by a slight hyperpolarization of the membrane potential, which is probably caused by a proton diffusion potential. In preparations where only a small acidification (0.4 units) occurred, the membrane potential was depolarized by the addition of Mg-ATP. Different monovalent cations and anions were tested concerning their effect on the pH gradient and ATPase activity in proton-conducting tonoplasts. Chloride stimulation and NO ₃ ^- inhibition were clearly present. The observed decline of the pH gradient upon the addition of Na⁺ salts is probably caused by an Na⁺/H⁺ antiport system.Abbreviations and symbol CCCP carbonylcyanide-m-chlorophenylhydrazone - Mes 2(N-morpholino)ethanesulfonic acid - TPMP⁺ triphenylmethylphosphoniumchloride - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol - membrane potential Dedicated to Professor A. Betz on the occasion of his 65th birthday     

Effects of bafilomycin A1 and amiloride on the apical potassium and proton gradients in Drosophila Malpighian tubules studied by X-ray microanalysis and microelectrode measurements

The intracellular distribution of potassium in Malpighian tubules from Drosophila larva was measured by electron probe X-ray microanalysis of freeze-dried cryosections. Application of amiloride alone to the haemolymph space had no effect on the intracellular potassium concentration in the region of intermediate cytoplasm (between the basal region of basal membrane infoldings and the apical brush border), whereas a potassium increase as well as a chloride increase was observed after simultaneous blocking of the potassium conductance of the basal membrane with barium. Injected bafilomycin and amiloride applied in the haemolymph caused an increase of the potassium content in the basal cytoplasm but not in the microvilli. In addition, the intracellular water portion was decreased by bafilomycin. pH measurements in isolated larval anterior tubules with proton-selective microelectrodes showed that bafilomycin added to the bathing solution caused a decrease in intracellular pH. Addition of amiloride had no significant effect on intracellular pH, but the pH of the luminal fluid was decreased within 1 min by 0.5 pH units. The amiloride-induced luminal pH decrease could be inhibited by the metabolic blocker KCN as well as by bafilomycin. Furthermore, removing potassium from the bathing saline caused a slow luminal acidification, which could be blocked by KCN. Our results support the hypothesis of a functionally coupled transport system in the apical membrane consisting of a bafilomycin-sensitive V-ATPase and a K⁺-dependent, amiloride-sensitive K⁺/H⁺ exchange system.Abbreviation C _a element concentration related to water - C _d element content related to dry weight - dw dry weight - DMSO dimethylsulphoxide - emf electromotive force - NBD-Cl 7-chloro-4-nitrobenz-2-oxa-1,3-diazole - NEM N-ethylmaleimide - NMDG⁺ N-methyl-d-glucamine - PD potential difference - pH_i intracellular pH value - pH_lu luminal pH value - pmf protonmotive force - SD standard deviation - SE standard error - STEM scanning transmission electron microscopy - V _a apical potential difference - V _b basal potential difference - V _t transepithelial potential difference     

Effect of changes in pH of the medium on electrical activity of the Ranvier node

The effect of changes in pH of the medium from 4 to 10 on the action potential and its first derivative was studied at the original resting potential and during hyperpolarization of the membrane in experiments on single nodes of Ranvier. Raising the pH of the medium from 7 to 9 led to a decrease in amplitude of the action potential and of its derivative (V_max). During hyperpolarization of the membrane these parameters were fully restored. Lowering the pH of the solution led to an increase in the action potential and a decrease in V_max. During hyperpolarization of the membrane the action potential and its derivative were not completely restored. Under the influence of solutions with low and high pH values the duration of the action potential was increased. Changes in the action potential and in V_max with an increase in pH can be attributed to increased inactivation of the sodium permeability of the membrane, and in solutions with low pH to a decrease in the maximal sodium permeability and to weakening of its inactivation.A. V. Vishnevskii Institute of Surgery, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 6, No. 2, pp. 205–210, March–April, 1974.     

Electrogenic H<Superscript>+</Superscript> Transport and pH Gradients Generated by a V-H<Superscript>+</Superscript>-ATPase in the Isolated Perfused Larval <Emphasis Type="Italic">Drosophila</Emphasis> Midgut

A method for microperfusion of isolated segments of the midgut epithelium of Drosophila larvae has been developed to characterize cellular transport pathways and membrane transporters. Stereological ultrastructural morphometry shows that this epithelium has unusually long tight junctions, with little or no lateral intercellular volume normally found in most epithelia. Amplification of the apical and basal aspects of the cells, by ≈ 17-fold and ≈ 7-fold, respectively, predicts an almost exclusively transcellular transport system for solutes. This correlates with the high lumen-negative transepithelial potential (V_t) of 38 to 45 mV and high resistance (R_t) of 800 to 1400 Ω • cm² measured by terminated cable analysis, in contrast to other microperfused epithelia like the renal proximal tubule. Several blockers (amiloride 10⁻⁴ M, ouabain 10⁻⁴ M, bumetanide 10⁻⁴ M), K⁺-free solutions, or organic solutes such as D-glucose 10 mM or DL-alanine 0.5 mM failed to affect V_t or R_t. Bafilomycin-A₁ (3 to 5 μM) decreased V_t by ≈ 40% and short-circuit current (I_sc) by ≈ 50%, and decreased intracellular pH when applied from the basal side only, consistent with an inhibition of an electrogenic V-H⁺-ATPase located in the basal membrane. Gradients of H⁺ were detected by pH microelectrodes close to the basal aspect of the cells or within the basal extracellular labyrinth. The apical membrane is more conductive than the basal membrane, facilitating secretion of base (presumably HCO₃⁻), driven by the basal V-H⁺-ATPase.     

Fused cells of frog proximal tubule: II. Voltage-dependent intracellular pH

Summary Experiments were performed in intact proximal tubules of the doubly perfused kidney and in fused proximal tubule cells ofRaha esculenta to evaluate the dependence of intracellular pH (pH_i) on cell membrane potential applying pH-sensitive and conventional microelectrodes. In proximal tubules an increase of the K^– concentration in the peritubular perfusate from 3 to 15 mmol/liter decreased the peritubular cell membrane potential from –55±2 to –38±1 mV paralleled by an increase of pH _i , from 7.54±0.02 to 7.66±0.02. The stilbene derivative DIDS hyperpolarized the cell membrane potential from –57 ± 2 to –71 ±4 mV and led to a significant increase of the K^–-induced cell membrane depolarization, but prevented the K^–-induced intracellular alkalinization. Fused proximal tubule cells were impaled by three microelectrodes simultaneously and cell voltage was clamped stepwise while pH _i changes were monitored. Cell membrane hyperpolarization acidified the cell cytoplasm in a linear relationship. This voltage-induced intracellular acidification was reduced to about one-third when HCO₃ ions were omitted from the extracellular medium. We conclude that in proximal tubule cells pH _i depends on cell voltage due to the rheogenicity of the HCO ₃ ^– transport system.     

A kinetic analysis of the electrogenic pump ofChara corallina: I. Inhibition of the pump by DCCD

Summary The current-voltage curve of theChara membrane was obtained by applying a slow ramp depo- and hyperpolarization by use of voltage clamp. With the progress of poisoning by DCCD (dicyclohexylcarbodiimide) theI–V curve moved by about 50 mV (depolarization) along the voltage axis, reducing its slope, and finally converged to thei _d -V curve of the passive diffusion channel. Changes ofi _p -V curve of the electrogenic pump channel could be obtained by subtracting the latter from the former.The sigmoidali _p -V curve could be simulated satisfactorily by adopting a simple reaction kinetic model. Kinetic parameters of the successive changes of state of the H⁺ ATPase could be evaluated. Changes of these kinetic parameters during inhibition gave useful information about the molecular mechanism of the electrogenic pump.Depolarization of the membrane potential, decrease of membrane conductance, and decrease of pump current during inhibition of the pump with DCCD are caused mainly by the decrease of conductance of the pump channel. The decrease of this pump conductance is caused principally by a marked decrease of the rate constant for releasing H⁺ to the outside.     

Aluminium effects on transmembrane potential in cells of fibrous roots of sugar beet

The effect of aluminium (Al) on the electrical transmembrane potential of epidermal and outer cortical root cells of intact seedlings of sugar beet (Beta vulgaris L. cv. Monohill) was studied. The potential difference to the surrounding medium was recorded with microelectrodes inserted into the vacuoles (PD_v) and cytoplasm (PD_c) of intact roots. Both long-term effects of AlCl₃ (100, μM present during cultivation) and immediate effects of AlCl₃ (10, 50, or 100 μM present in the assay medium), were measured. The effect of Al was measured at pH 4.0, 5.0 and 6.5 in order to obtain information on the toxicity of different Al forms existing at different pH values. Low pH and/or the presence of AlCl₃ during cultivation caused large depolarizations of the PD_v. Since the immediate effect of 2,4-dinitrophenol (DNP) on the resting potential of cells from Al-cultivated plants was negligible, it is likely that Al affects the metabolic component of the transmembrane potential. Aluminium also had an immediate effect on the PD in root cells of plants cultivated without Al. Addition of 10 or 50 μM Al to the assay medium caused hyperpolarization of PD_v in the presence of 0.5 mM Ca²⁺ at all pH values studied, depolarization of PD_c at pH 6.5, and hyperpolarization of PD_c at lower pH. At 1 mM Ca²⁺, or in the presence of K⁺ (≥ 2 mM), however, the same Al concentrations had little effect on PD_c. The strongest depolarizing effects of 10 or 50 μM Al in short-term treatments were obtained at pH 6.5, and were probably due to the soluble species Al(OH)₃, which is more frequent at pH 6.5 than at a lower pH. Addition of 50 μM Al caused alkalinization of the root medium at pH 6.5, but not at pH 4.0. Therefore, it is possible that Al at pH 6.5 is bound to, or translocated across, the membrane without the accompanying hydroxide ions. It is likely that most of the Al is bound to the root cells, since removal of Al from the buffer surrounding the roots did not cause the changed PD values to return to the original values. Aluminium also interacts with effects of Ca²⁺ and K⁺ on the membrane potential, since changes in PD, induced by changes in concentrations of Ca²⁺ and K⁺ are different in the absence and presence of Al.     

Relationship of transepithelial electrical potential to membrane potentials and conductance ratios in frog skin

Summary Previous studies in anuran epithelia have shown that, after clamping the transepithelial voltage in symmetrical sequences for 4–6 min there is near-constancy of the rate of active Na transport and the associated oxidative metabolism, with a near-linear potential dependence of both. Here we have investigated in frog skin the cellular electrophysiological events associated with voltage clamping (V _t =inside-outside potential). Increase and decrease ofV _t produced converse effects, related directly to the magnitude ofV _t .Hyperpolarization resulted in prompt decrease in inward transepithelial currentI _t and increase in fractional outer membrane resistancefR ₀ (as evaluated from small transient voltage perturbations) and in outer membrane potentialV ₀. Overshoot ofV ₀ was followed by relaxation to a quasi-steady state in minutes. Changes infR ₀ were progressive, with half times of some 1–5 sec. Changes in transepithelial slope conductanceg _t were more variable, usually preventing precise evaluation of the outer and inner cell membrane conductancesg ₀ andg _i . Nevertheless, it was shown thatg ₀ is related inversely toV _t andV ₀. Presuming insensitivity ofV _i toV _t , the dependence ofg ₀ onV ₀ in the steady state much exceeds that predicted by the constant field equation. Apparent inconsistencies with earlier results of others may be attributable to differences in protocol and the complex dependence ofg ₀ onV ₀ and/or cellular current. In contrast to previous findings in tight epithelia at open circuit, differences inV _t were associated with substantial differences infR ₀ and inner membrane potentialV _i . Hyperpolarization ofV _t over ranges commonly employed in studies of active transport and metabolism appears to increase significantly the electrochemical work per Na ion transported.     

Photosynthetic inorganic carbon utilization and growth of Porphyra linearis (Rhodophyta)

Photosynthetic (oxygen evolution) and growth (biomass increase) responses to ambient pH and inorganic carbon (Ci) supply were determined for Porphyralinearis grown in 0.5 L glass cylinders in the laboratory, or in 40 L fibreglass outdoor tanks with running seawater. While net photosynthetic rates were uniform at pH 6.0–8.0, dropping only at pH 8.7, growth rates were significantly affected by pH levels other than that of seawater (c. pH 8.3). In glass cylinders, weekly growth rates averaged 76% at external pH 8.0, 13% at pH 8.7 and 26% at pH 7.0. Photosynthetic O₂ evolution on a daily basis(i.e. total O₂ evolved during day time less total O₂ consumed during night time) was similar to the growth responses at all experimental pH levels, apparently due to high dark respiration rates measured at acidic pH. Weekly growth rates averaged 53% in algae grown in fibreglass tanks aerated with regular air (360 mg L_-1 CO₂) and 28% in algae grown in tanks aerated with CO₂-enriched air (750 mg L^-1 CO₂). The pH of the seawater medium in which P. linear is was grown increased slightly during the day and only rarely reached 9.0. The pH at the boundary layer of algae submerged in seawater increased in response to light reaching, about pH 8.9 within minutes, or remained unchanged for algae submerged in a CO₂-free artificial sea water medium. Photosynthesis of P. linearissaturated at Ci concentrations of seawater (K_0.5560 μM at pH 8.2) and showed low photosynthetic affinity for CO₂(K_0.5 61 μM) at pH 6.0. It is therefore concluded that P. linearisuses primarily CO₂ with HCO₃ ^- being an alternative source of Ci for photosynthesis. Its fast growth could be related to the enzyme carbonic anhydrase whose activity was detected intra- and extracellularly. This revised version was published online in August 2006 with corrections to the Cover Date.     

Mechanism of Cl- sensitivity in internal ion receptors of the leech; an inward current gated off by Cl- in the nephridial nerve cells

Summary The nephridial nerve cells of the leech, Hirudo medicinalis, 34 sensory cells, each associated with one nephridium, are sensitive to changes in extracellular Cl^- concentration, an important factor in ion homeostasis. Using single-electrode current- and voltage clamp and ion substitution techniques, the specificity and mechanism of Cl^- sensitivity of the nephridial nerve cell was studied in isolated preparations. Increase of the normally low external Cl^- concentration leads to immediate and sustained hyperpolarization, decrease of the frequency of bursts and decrease of membrane conductance. The response is halogen specific: Cl^- can be replaced by Br^–, but not by organic mono- or divalent anions or inorganic divalent anions.At physiological Cl^- concentrations (36mM extra-cellular Cl^-), the nephridial nerve cell has a high resting conductance for Cl^- and the membrane potential is governed by Cl^-. In high extracellular Cl^- concentrations (110–130 mM), membrane conductance is low, most likely due to the gating off of Cl^- channels. Under these conditions, membrane potential is dominated by the K⁺ distribution and the nephridial nerve cell hyperpolarizes towards E_K.Abbreviations NNC nephridial nerve cell - V _m membrane potential - E _Cl(k) equilibrium potential for Cl (K) - IV-curve current-voltage relationship     

Two steady states in membrane potential deflection in relation to chemoreception and chemotaxis byPhysarum polycephalum

Summary In the plasmodium ofPhysarum polycephalum application of various monovalent cation salts elicited either a depolarization or a hyperpolarization of the membrane potential. The hyperpolarization was restricted to phosphates and bicarbonates of large cations (Na⁺, Li⁺, NMe₄ ⁺, NEt₄ ⁺). More than 50 other combinations of cations (K⁺, Rb⁺, Cs⁺, NH₄ ⁺) and anions (Cl^–, NO₃ ^–, SO₄ ^2–, acetate, lactate, citrate, etc.) induced the depolarization. In both cases the magnitude of the deflection in membrane potential () varied linearly with logarithm of concentration above the threshold C_th (10^–4 M for all monovalent cation salts examined) according to the following equation: =± R log (C/C_th).The value of R was 10–15 mV, and plus and minus signs correspond to depolarization and hyperpolarization, respectively. Depolarizing and hyperpolarizing agents competed with each other and exhibited a sharp transition between the two states of the membrane which were characterized by — R and R in the above equation or displayed a strong hysteresis, depending on which agents had first been applied to the plasmodial membrane. This transition in the membrane potential corresponded to the transition between positive and negative taxis at the behavioral level.