Effect of extracellular pH on sodium current in isolated,single rat ventricular cells

Summary Effects of extracellular pH on the sodium current (I _Na) of single rat ventricular cells were examined under conditions of voltage clamp and internal perfusion. In this way, pH _i was controlled while pH _o was changed. The combined suction pipette-microelectrode method was used. The suction pipette passed current and perfused the cell's interior; the microelectrode measured membrane potential. Increasing extracellular H⁺ depressedI _Na and slowed inactivation. The current-voltage curves forI _Na and Slowed inactivation. The current-voltage curves forI _Na were shifted to positive and negative potentials at low and high pH _o , respectively. Similar potential shifts were observed in both the conductance voltage curve and the steadystate inactivation voltage curve (h ). Conduction was also depressed at low pH _o . The shifts were probably due to surface charge effects, while the impaired conduction was probably due to protonation of a site in the Na channel.     

Modulation of inwardly rectifying Na+-K+ channels by serotonin and cyclic nucleotides in salivary gland cells of the leech,Haementeria

Summary The electrically excitable salivary cells of the giant Amazon leech, Haementeria, display a time-dependent inward rectification. Under voltage clamp, hyperpolarizing steps to membrane potentials negative to about –70 mV were associated with the activation of a slow inward current (I _h) which showed no inactivation with time. The time course of activation of I _hwas described by a single-exponential function and was strongly voltage dependent. The activation curve of_hranged from –72 to –118 mV, with half-activation occurring at –100 mV. Ion-substitution experiments indicated that I _his carried by both Na⁺ and K⁺ ions. 5-Hydroxytryptamine (5-HT) increased the amplitude of I _hand its rale of activation. It also produced a positive shift of the activation curve of the conductance underlying I _h G_hwithout altering the slope factor, thus indicating that the voltage dependence of I _hwas modulated by 5-HT. Cs⁺ blocked both I _hand the 5-HT-polentiated current in a voltage-independent manner, whereas Ba²⁺ had little effect. It is concluded that 5-HT increases I _hby modulating the inwardly rectifying Na⁺-K⁺ channels in the salivary cells. The effect of 5-HT may be mediated by an increase in adenylate cyclase activity since I _hwas increased by 8-bromocyclic AMP and by the phosphodiesterase inhibitor, 3-isobutyl-l-methylxanthine. In contrast, I _hwas reduced by 8-bromo-cyclic GMPand by zaprinast (an inhibitor of cyclic GMP-scnsitive phosphodieslerase). Cyclic GMP itself also reduced I _h, and the effect was specific to the 3,5 form; 2,3-cyclic GMP was inactive. The results suggest that the inward-rectifier channel may be modulated in opposite directions by cyclic AMP and cyclic GMPThis work was supported by a grant from the Science and Engineering Research Council (no. GR/F/17087). We are grateful to the SmithKline (1982) Foundation for provision of a pulse generator     

Ca-sensitive sodium absorption in the colon of Xenopus laevis

Summary Transepithelial electrogenic Na transport (I_Na) was investigated in the colon of the frog Xenopus laevis with electrophysiological methods in vitro. The short circuit current (I_sc) of the voltage-clamped tissue was 24.2±1.8 A·cm^-2 (n=10). About 60% of this current was generated by electrogenic Na transport. Removal of Ca²⁺ from the mucosal Ringer solution stimulated I_Na by about 120%. I_Na was not blockable by amiloride (0.1 mmol·l^-1), a specific Na-channel blocker in epithelia, but a fully and reversible inhibition was achieved by mucosal application of 1 mmol·l^-1 lanthanum (La^3-). No Na-self-inhibition was found, because I_Na increased linearly with the mucosal Na concentration. A stimulation of I_Na by antidiuretic hormones was not possible. The analysis of fluctuations in the short circuit current (noise analysis) indicated that Na ions pass the apical cell membrane via a Ca-sensitive ion channel. The results clearly demonstrate that in the colon of Xenopus laevis Na ions are absorbed through Ca-sensitive apical ion channels. They differ considerably in their properties and regulation from the amiloride-sensitive Na channel which is typically found in the colon of vertebrates.Abbreviations G _T transepithelial conductance - I _sc short circuit current - I _Na transepithelial Na-current - m mucosal - s serosal - PDS power density spectrum - f frequency - f _c corner frequency of the Lorentzian component of the PDS - S(f) power density of the Lorentzian component of the PDS - S_o plateau value of the Lorentzian component of the PDS     

Effect of dilution rate on the release of pertussis toxin and lipopolysaccharide ofBordetella pertussis

Summary The kinetics ofBordetella pertussis growth was studied in a glutamate-limited continuous culture. Growth kinetics corresponded to Monod's model. The saturation constant and maximum specific growth rate were estimated as well as the energetic parameters, theoretical yield of cells and maintenance coefficient. Release of pertussis toxin (PT) and lipopolysaccharide (LPS) were growth-associated. In addition, they showed a linear relationship between them. Growth rate affected neither outer membrane proteins nor the cell-bound LPS pattern.Nomenclature X cell concentration (g L^–1) - specific growth rate (h^–1) - _m maximum specific growth rate (h^–1) - D dilution rate (h^–1) - S concentration of growth rate-limiting nutrient (glutamate) (mmol L^–1 or g L^–1) - Ks substrate saturation constant (mol L^–1) - m_s maintenance coefficient (g g^–1 h^–1) - Y_x/s theoretical yield of cells from glutamate (g g^–1) - Y_x/s yield of cells from glutamate (g g^–1) - Y_PT/s yield of soluble PT from glutamate (mg g^–1) - Y_KDO/s yield of cell-free KDO from glutamate (g g^–1) - Y_PT/x specific yield of soluble PT (mg g^–1) - Y_KDO/x specific yield of cell-free KDO (g g^–1) - q_PT specific soluble PT production rate (mg g^–1 h^–1) - q_KDO specific cell-free KDO production rate (g g^–1 h^–1)     

Different modes of electrogenic Na+ absorption in the coprodeum of the chicken embryo: role of extracellular Ca2+

Summary Transepithelial electrogenic Na⁺ transport (I_Na) was investigated in the coprodeum of 20-days-old chicken embryos in Ussing chambers. Short circuit current (I_sc) and transepithelial resistance (R_t) were 14.7±4.8 A · cm^-2 (n=12) and 0.53±0.09 k · cm^-2 (n=12), respectively. I_Na was calculated from changes in I_sc by substitution of mucosal Na⁺ by (N-methyl-d-glucamine) (NMDG). I_sc inversed during Na⁺ removal, and I_Na was found to be 27.8±4.7 A · cm^-2 (n=12). Amiloride (100 mol · l^-1) inhibited only about 60% of I_Na. Analysis of I_sc fluctuations revealed a Lorentzian component in the power density spectrum with a corner frequency of about 57 Hz. This component was not correlated to I_Na, and its origin is still unclear. Removal of mucosal Ca²⁺ increased I_Na about 2.5-fold due to an increase of the amiloride-insensitive component of I_Na in additionally investigated adult tissues. The results clearly show that this is due to a non-selective cation channel with an apparent order of selectivity Cs⁺>Na⁺=K⁺>Rb⁺>Li⁺. The Ca²⁺ concentration required to block 50% of the I_sc was about 18 mol · l^-1. The I _sc ^Ca could also be supressed by other divalent cations such as Mg²⁺ and Ba²⁺. Additionally, an I_Na-linked Lorentzian component occurred which dominated the control spectrum with a significantly higher corner frequency (about 88 Hz). The results indicate that Na⁺ absorption in the coprodeum of the chicken embryo is more complex than in adult hens. However, the Ca²⁺ sensitivity of I_Na is similar to comparable effects described for other epithelia. This possibly reflects the existence of two types of amiloride-insensitive apical cation channels as pathways for Na⁺ absorption, which may be involved to differing degrees in ontogenetic developments of nonselective channels to Na⁺-specific ion channels.Abbreviations DPL direct-linear-plot method - slope of the back-ground noise component - EGTA ethylene glycol-bi(2-amino-ethylether)-N,N,N,N-tetraacetic acid - f frequency - f _c corner frequency of the Lorentzian noise component - G _t transepithelial conductance - HEPES N-hydroxyethylpiperazine-N-ethanesulfonic acid - I _sc short-circuit current - I _Na transepithelial sodium current - I _sc ^Ca Ca²⁺-sensitive short-circuit current - K _m ^Ca Michaelis-Menten constant for Ca²⁺ - K _B power density of the background noise component at f=1Hz - m mucosal - NMDG N-methyl-D-glucamine - R _t transepithelial resistance - s serosal - SEM standard error of mean - S(f) power density of the Lorentzian noise component - S _o plateau value of the Lorentzian noise component     

Ion transport across leech integument

The dorsal skin of the leech Hirudo medicinalis was used for electrophysiological measurements performed in Ussing chambers. The leech skin is a tight epithelium (transepithelial resistance = 10.5±0.5 k· cm^-2) with an initial short-circuit current of 29.0±2.9 A·cm^-2. Removal of Na⁺ from the apical bath medium reduced short-circuit current about 55%. Ouabain (50mol·l^-1) added to the basolateral solution, depressed the short-circuit current completely. The Na⁺ current saturated at a concentration of 90 mmol Na⁺·l^-1 in the apical solution (K _M=11.2±1.8 mmol·l^-1). Amiloride (100 mol·l^-1) on the apical side inhibited ca. 40% of the Na⁺ current and indicated the presence of Na⁺ channels. The dependence of Na⁺ current on the amiloride concentration followed Michaclis-Menten kinetics (K _i=2.9±0.4 mol·l^-1). The amiloride analogue benzamil had a higher affinity to the Na⁺ channel (K _i=0.7±0.2 mol·l^-1). Thus, Na⁺ channels in leech integument are less sensitive to amiloride than channels known from vertebrate epithelia. With 20 mmol Na⁺·l^-1 in the mucosal solution the tissue showed an optimum amiloride-inhibitable current, and the amiloride-sensitive current under this condition was 86.8±2.3% of total short-circuit current. Higher Na⁺ concentrations lead to a decrease in amiloride-blockade short-circuit current. Sitmulation of the tissue with cyclic adenosine monophosphate (100 mol·l^-1) and isobutylmethylxanthine (1 mmol·l^-1) nearly doubled short-circuit current and increased amiloride-sensitive Na⁺ currents by 50%. By current fluctuation analysis we estimated single Na⁺ channel current (2.7±0.9 pA) and Na⁺ channel density (3.6±0.6 channels·m^-2) under control conditions. After cyclic adenosine monophosphate stimulation Na⁺ channel density increased to 5.4±1.1 channels·m^-2, whereas single Na⁺ channel current showed no significant change (1.9±0.2 pA). These data present a detailed investigation of an invertebrate epithelial Na⁺ channel, and show the similarities and differences to vertebrate Na⁺ channels. Whereas the channel properties are different from the classical vertebrate Na⁺ channel, the regulation by cyclic adenosine monophosphate seems similar. Stimulation of Na⁺ uptake by cyclic adenosine monophosphate is mediated by an increasing number of Na⁺ channels.Abbreviations slope of the background noise component - ADH antidiuretic hormone - cAMP cyclic adenosine monophosphate - f frequency - f _c coner frequency of the Lorentzian noise component - Hepes N-hydroxyethylpiperazine-N-ethanesulphonic acid - BMX isobutyl-methylxanthine - i _Na single Na⁺ channel current - I _Na max, maximal inhibitable Na⁺ current - I _SC short circuit current - K _i half maximal blocker concentration - K _M Michaelis constandard error of the mean - S _(f) power density of the Lorentzian noise component - S ₀ plateau value of the Lorentzian noise component - TMA tetramethylammonium - Trizma TRIS-hydroxymethyl-amino-methane - V _max maximal reaction velocity - V _T transepithelial potential - K half maximal blocker concentration     

Single point mutations of the sodium channel drastically reduce the pore permeability without preventing its gating

1. Two mutants of the sodium channel II have been expressed inXenopus oocytes and have been investigated using the patch-clamp technique. In mutant E387Q the glutamic acid at position 387 has been replaced by glutamine, and in mutant D384N the aspartic acid at position 384 has been replaced by asparagine.2. Mutant E387Q, previously shown to be resistant to block by tetrodotoxin (Noda et al. 1989), has a single-channel conductance of 4 pS, that can be easily measured only using noise analysis. At variance with the wild-type, the openchannel current-voltage relationship of mutant E387Q is linear over a wide voltage range even under asymmetrical ionic conditions.3. Mutant D384N has a very low permeability for any of the following ions: Cl^–, Na⁺, K⁺, Li⁺, Rb⁺, Ca²⁺, Mg²⁺, NH₄ ⁺ , TMA⁺, TEA⁺. However, asymmetric charge movements similar to the gating currents of the Na⁺-selective wild-type are still observed.4. These results suggest that residues E387 and D384 interact directly with the pathway of the ions permeating the open channel.Abbreviations TTX tetrodotoxin; Na⁺, sodium; K⁺, potassium; - NFR normal frog Ringer - HEPES N-2-hydroxylethyl piperazine-N-2-ethanesulfonic acid - EGTA ethyleneglycol-bis(-amino-ethyl ether) N,N,N',N'-tetra acetic acid - TEA tetraethylammonium - TMA tetramethylammonium;I _g , gating current; , single-channel conductance     

Electrogenic properties of the sodium-alanine cotransporter in pancreatic acinar cells: II. Comparison with transport models

Summary In this paper, the results of the preceding electrophysiological study of sodium-alanine cotransport in pancreatic acinar cells are compared with kinetic models. Two different types of transport mechanisms are considered. In the simultaneous mechanism the cotransporterC forms a ternary complexNCS with Na⁺ and the substrateS; coupled transport of Na⁺ andS involves a conformational transition between statesNCS andNCS with inward- and outward-facing binding sites. In the consecutive (or ping-pong) mechanism, formation of a ternary complex is not required; coupled transport occurs by an alternating sequence of association-dissociation steps and conformational transitions. It is shown that the experimentally observed alanine- and sodium-concentration dependence of transport rates is consistent with the predictions of the simultaneous model, but incompatible with the consecutive mechanism. Assuming that the association-dissociation reactions are not rate-limiting, a number of kinetic parameters of the simultaneous model can be estimated from the experimental results. The equilibrium dissociation constants of Na⁺ and alanine at the extracellular side are determined to beK _N <-64mm andK _S <-18mm. Furthermore, the ratioK _N /K _N ^S of the dissociation constants of Na⁺ from the binary (NC) and the ternary complex (NCS) at the extracellular side is estimated to be <-6. This indicates that the binding sequence of Na⁺ andS to the transporter is not ordered. The current-voltage behavior of the transporter is analyzed in terms of charge translocations associated with the single-reaction steps. The observed voltage-dependence of the half-saturation concentration of sodium is consistent with the assumption that a Na⁺ ion that migrates from the extracellular medium to the binding site has to traverse part of the transmembrane voltage.     

Use-dependent block with tetrodotoxin and saxitoxin at frog Ranvier nodes I. Intrinsic channel and toxin parameters

The use-dependent phasic blockage of sodium channels by tetrodotoxin (TTX) and saxitoxin (STX) was examined in frog nodes of Ranvier using trains of depolarizing pulses. The decline of the peak Na⁺ current from its initial value (I ₀) before the train to a stationary value (I ) after the train was more pronounced at more negative holding potentials. The relationship betweenI /I ₀ and holding potential was fitted by a sigmoid function which yielded values for the steepness of the voltage dependencies of around –15 mV for TTX and – 8 mV for STX. Similar values were obtained at toxin concentrations of 4 and 8 nM. The higher voltage sensitivity of STX versus TTX is interpreted in terms of the higher charge and the faster binding kinetics of STX. These differences also explain the frequency dependence of the decline of Na⁺ currents with STX (between 0.5 and 2 Hz) and the frequency independence with TTX. Variation of the pulse amplitude in a train of conditioning pulses revealed that the magnitude of the use-dependent actions of STX parallels the steady-state Na⁺ inactivation curveh . Inhibition of inactivation, by pre-treatment with chloramine-T, did not, however, abolish the use dependence. Instead, it introduced a change in the time constants of the decline of the Na⁺ currents and the magnitude became independent of the holding potential.     

Sodium Conductance Activation without Inactivation in Pronase-perfused Axons

A controversy of long standing in membrane electrophysio-logy is whether the sodium ion current (I_Na) and potassium ion current (I_K) pass through the membrane in separate channels, or through a single set of channels which conduct first I_Na and then I_K. In support of the latter hypothesis it has been noted that the sodium conductance (g_Na) decline, called inactivation, proceeds with about the same time course as the potassium conductance (g_K) increase. This could mean that Na⁺ selective channels are being converted into K⁺ selective channels. The hypothesis is especially interesting because of the possibility that the carrier postulated in active transport is convertible from Na⁺ to K⁺ selectivity¹. An explicit statement of the single channel hypothesis and the means for disproving it were given by Mullins². Because a single channel could not simultaneously conduct I_Na and I_K, disproof requires that membrane conductance (g_m) be made somehow to exceed the maximum value of g_Na or g_K. We report here that inactivation of g_Na can be destroyed fairly selectively by the action from inside the axon of the unspecific proteolytic enzymes of pronase. In many cases g_m after pronase treatment is greater than maximum g_K before treatment, making untenable the single channel hypothesis.     

Some regulatory aspects of [14C]methylamine influx intoPisum sativum L. cv. Feltham First seedlings

[¹⁴C]Methylamine influx intoPisum sativum L. cv. Feltham First seedlings showed Michaelis-Menten-type kinetics with apparentV _max=49.2 mol·g^-1 FW·h^-1 and apparentK _m=0.51 mM. The competitive interactions between ammonium and methylamine were most obvious when biphasic kinetics were assumed with saturation of the first phase at 0.05 mM. The inhibitor constant for ammonium (K _i)=0.027 mM. When [¹⁴C]methylamine was used in trace amounts with ammonium added as substrate, the influx of tracer showed Michaelis-Menten-type kinetics with apparentV _max=3.46 mol·g^-1 FW·h^-1 and apparentK _m=0.15 mM. The initial rate of net ammonium uptake corresponded with that found when [¹⁴C]methylamine was used to trace ammonium influx. The latter was also stimulated by high pH_o and inhibited by nitrate. Ammonium pretreatment±methionine sulphoximine or glutamine pretreatment of the seedlings inhibited subsequent [¹⁴C]methylamine influx, while methylamine or asparagine pretreatment stimulated [¹⁴C]methylamine influx. There was also a stimulatory effect of prior inoculation withRhizobium. The results are discussed in terms of current models for the regulation of ammonium uptake in plants.     

Properties of placental alkaline phosphatase. I. Molecular size and electrical charge of the various electrophoretic components of the six common phenotypes

The properties of the six common phenotypes (F₁, S₁, I₁, F₁S₁, S₁I₁, F₁I₁) of placental alkaline phosphatase have been studied by DEAE-Sephadex chromatography, gel filtration, two-dimensional paper-starch gel and starch block electrophoresis. The rapidly migrating component (component I) had the same molecular weight (118,000) in all phenotypes studied but different negative charge (F₁>I₁>S₁). The fastest of the slow-moving components (component II) had a molecular weight of 130,000 and a higher absolute negative charge, but slightly lower net negative charge than I. The very slow-moving components (II) were usually composed of two major and several minor components. The major components had a higher absolute negative charge but a lower net negative charge than II. All II components had a molecular weight of over 200,000. It is suggested that II is an enzyme bound to a small molecule, while II components are aggregates of enzyme with various substances or with itself.This work was supported in part by U.S. Public Health Service Grant HD02552. K.H. is a Career Scientist of the Health Research Council of the City of New York (I-513).     

Calculation of leaf photosynthetic parameters from light-response curves for ecophysiological applications

Measurement of the light response of photosynthetic CO₂ uptake is often used as an implement in ecophysiological studies. A method is described to calculate photosynthetic parameters, such as the maximum rate of whole electron transport and dissimilative respiration in the light, from the light response of CO₂ uptake. Examples of the light-response curves of flag leaves and ears of wheat (Triticum aestivum cv. ARKAS) are shown.Abbreviations and symbols A net photosynthesis rate - D ¹ rate of dissimilative respiration occurring in the light - f loss factor - I incident PPFD - I effective absorbed PPFD - J rate of whole electron transport - J _m maximum rate of whole electron transport - p ^c intercellular CO₂ partial pressure - PPFD photosynthetic photon flux density - q effectivity factor for the use of light (electrons/quanta) - absorption coefficient - I ^* CO₂ compensation point in the absence of dissimilative respiration (bar) - II conversion factor for calculation of CO₂ uptake from the rate of whole electron transport - convexity factor Gas-exchange rates relate to the projective area and are given in mol·m^-2·s^-1. Electron-transport rates are given in mol electrons·m^-2·s^-1; PPFD is given in mol quanta·m^-2·s^-1.     

Model predictions of the ionic mechanisms underlying the beating and bursting pacemaker characteristics of molluscan neurons

The general properties of the excitable membrane on molluscan pacemaker neurons can be described on the basis of a fair amount of experimental evidence available in the literature. The neuronal membrane exhibits under voltage clamp an initial inward current carried by both Na⁺ and Ca²⁺ ions, the time- and voltage-dependent characteristics of which are similar to that of other excitable structures. The conductance mechanism for the two ion species and the transport kinetics appear to be closely similar. The time course and amplitude of the delayed outward current carried by K⁺ ions shows a marked dependence on the membrane potential. Characteristic for the molluscan neurons is the existence of an additional fast transient outward current which is only activated by hyperpolarizing shifts from the membrane potential. A regular beating discharge over a wide range of frequencies can be predicted by making the assumption of a metabolically controlled driving of the Na⁺ conductance. Bursting pacemaker characteristics can be correctly simulated by the model if sinusoidal variations of an additional Na⁺ and Ca²⁺ conductances g _Na and g _Ca, and periodic variations of the K⁺ conductance g _K, governed by the known operation of a metabolic substrate cycle are introduced. The close approximation of experimentally observed impulse bursts requires that the actual inpulse-frequency and the amplitude of the after-spike hyperpolarization are determined by the temporal pattern of g _Na, while the spike amplitude is controlled by g _Na which (although of similar time course) is lagging in phase behing g _Na. The periodic changes in additional K⁺ conductance g _K, are responsible for burst termination and the changes in inter-burst interval, to the effect that spike doublets, triplets and multi-spike bursts can be simulated by a suitable choice for the time characteristics of g _K. The model makes use of the finding that the Ca²⁺ inflow associated with a spike discharge actually activates g _K, so that large postburst hyperpolarizations can be obtained in high-frequency bursts.Supported by the Deutsche Forschungsgemeinschaft (Grant Ch 25/1)     

The mechanism of Na+ transport by rabbit urinary bladder

Summary The mechanism of Na⁺ transport in rabbit urinary bladder has been studied by microelectrode techniques. Of the three layers of epithelium, the apical layer contains virtually all the transepithelial resistance. There is radial cell-to-cell coupling within this layer, but there is no detectable transverse coupling between layers. Cell coupling is apparently interrupted by intracellular injection of depolarizing current. The cell interiors are electrically negative to the bathing solutions, but the apical membrane of the apical layer depolarizes with increasingI _sc. Voltage scanning detects no current sinks at the cell junctions or elsewhere. The voltage-divider ratio, , (ratio of resistance of apical cell membrane,R _a, to basolateral cell membrane,R _b) decreases from 30 to 0.5 with increasingI _sc, because of the transportrelated conductance pathway in the apical membrane. Changes in effective transepithelial capacitance withI _sc are predicted and possibly observed. The transepithelial resistance,R _t, has been resolved intoR _a, R_b, and the junctional resistance,R _j, by four different methods: cable analysis, resistance of uncoupled cells, measurements of pairs of (R _t, ) values in the same bladder at different transport rates, and the relation betweenR _t andI _sc and between andI _sc.R _j proves to be effectively infinite (nominally 300 k F) and independent ofI _sc, andR _a decreases from 154 to 4 k F with increasingI _sc. In the resulting model of Na⁺ transport in tight epithelia, the apical membrane contains an amiloride-inhibited and Ca⁺⁺-inhibited conductance pathway for Na⁺ entry; the basolateral membrane contains a Na⁺–K⁺-activated ATPase that extrudes Na⁺; intracellular (Na⁺) may exert negative feedback on apical membrane conductance; and aldosterone acts to stimulate Na⁺ entry at the apical membrane via the amiloride-sensitive pathway.     

Re-sorption of organic components by roots of Zea mays L. and its consequences in the rhizosphere

The re-sorption of carbon compounds from the rhizosphere was investigated using ¹⁴C-labelled glucose, mannose and citric acid. Uptake in roots of 5-day-old, intact Zea mays plants in sterile solution culture was determined over a period of 48 hours. Under optimal growth conditions significant re-absorption of glucose and mannose occurred with the uptake rates being 70.5 and 40.2 g compound g^-1 root DW h^-1, respectively. For glucose and mannose approximately 25% of the ¹⁴C label taken up by the root was recovered inside the plant as low-MW compounds and 33% polymerized into high MW compounds. 42% was respired as ¹⁴C-CO₂. Citric acid by comparison showed little accumulation within plant tissues (11.4%) with most being respired and recovered as ¹⁴C-CO₂ in KOH traps (88%). The uptake rate for citric acid was 34.8 g g^-1 root DW h^-1. Over the 48-hour period a net efflux (i.e. exudation) of labelled plus unlabelled C was observed at a rate of 608 g C g^-1 root DW h^-1 (equivalent to 1520 g glucose/mannose). Of the C released as root exudates, a minimum estimate of the amount of C taken back into the plant was therefore 9.5%. The two main C fluxes within the rhizosphere, namely release of C by the root and uptake by the microorganisms, have been well documented in recent years. It is now apparent however that a third flux term, re-sorption of C by roots, can also be identified. This may play an important but previously overlooked role within the rhizosphere, and further work is needed to determine its significance.A comparison between exudate release in static (permitting accumulation of C) and flowing culture (C removed as it is released) was also made with the respective rates being 15.36 and 45.18 mg C g^-1 root DW in 2 days. The relative important of re-sorption in natural environments and laboratory experiments is discussed.     

Na/H exchange in cultured epithelial cells from fish gills

Using primary cultures of gill pavement cells from freshwater rainbow trout, a method is described for achieving confluent monolayers of the cells on glass coverslips. A continuous record of intracellular pH was obtained by loading the cells with the pH-sensitive flourescent dye 2,7-bis(2-carboxyethyl)-5(6)-carboxyfluorescein and mounting the coverslips in the flowthrough cuvette of a spectrofluorimeter. Experiments were performed in HEPES-buffered media nominally free of HCO₃. Resting intracellular pH (7.43 at extracellular pH=7.70) was insensitive to the removal of Cl^– or the application of 4-acetamido-4-isothiocyanatostilbene-2,2-disulfonic acid (0.1 mmol·l^–1), but fell by about 0.3 units when Na⁺ was removed or in the presence of amiloride (0.2 mmol·l^–1). Exposure to elevated ammonia (ammonia prepulse; 30 mmol·l^–1 as NH₄Cl for 6–9 min) produced an increase in intracellular pH (to about 8.1) followed by a slow decay, and washout of the pulse caused intracellular pH to fall to about 6.5. Intracellular non-HCO ₃ ^– buffer capacity was about 13.4 slykes. Rapid recovery of intracellular pH from intracellular acidosis induced by ammonia prepulse was inhibited more than 80% in Na⁺-free conditions or in the presence of amiloride (0.2 mmol·l^–1). Neither bafilomycin A₁ (3 mol·l^–1) nor Cl removal altered the intracellular pH recovery rate. The K _m for Na⁺ of the intracellular pH recovery mechanism was 8.3 mmol·l^–1, and the rate constant at V _max was 0.008·s^–1 (equivalent to 5.60 mmol H⁺·l^–1 cell water·min^–1), which was achieved at external Na⁺ levels from 25 to 140 mmol·l^–1. We conclude that intracellular pH in cultured gill pavement cells in HEPES-buffered, HCO ₃ ^– -free media, both at rest and during acidosis, is regulated by a Na⁺/H⁺ antiport and not by anion-dependent mechanisms or a vacuolar H⁺-ATPase.Abbreviations BCECF 2,7-bis(2-carboxyethyl)-5(6)-carboxy-fluorescein - BCECF/AM 2,7-bis(2-carboxyethyl)-5(6)-carboxy-fluorescein, acetoxymethylester - Cholin-Cl choline chloride - DMSO dimethyl sulfoxide - EDTA ethylene diamine tetra-acetic acid - FBS foetal bovine serum - H ⁺ -ATPase Proton-dependent adenosine triphosphatase - HEPES N-[2-hydroxyethyl]piperazine-N[2-ethanesulfonic acid] - pH _i intracellular pH - pH _e extracellular pH - PBS phosphate-buffered saline - SITS 4-acetamido-4-isothiocyanatostilbene-2,2-disulfonic acid     

Sodium channel opening as a precursor to inactivation

The time constant of the process producing the delay in Na inactivation development as determined by the two pulse method (_delay) was extracted and compared to that of the slowest Na activation process ₃ for the I _Na during the conditioning pulse of that same determination. _delay and two pulse inactivation _c values were computer generated using a nonlinear least squares algorithm. _h and single pulse inactivation _h values were independently generated for each determination also with the aid of the computer using the same non-linear least squares algorithm. In one determination at 2 mV, _c was 4.68 and _delay 0.494 ms while _h was 4.70 and ₃ 0.491 ms for a _c/_h of 0.996 and a _delay/₃ of 1.006. Mean _delay/₃ from five determinations in four axons, both Cs and K perfused, and spanning a potential range of-27 to 2mV was 1.068. The precursor process to inactivation is channel opening. Some fraction of channels presumably inactivate via another route where prior channel opening is not required.     

Optimal control of continuous fermentation processes

Three layer control structure is proposed for optimal control of continuous fermentation processes. The start-up optimization problems are solved as a first step for optimization layer building. A steady state optimization problem is solved by a decomposition method using prediction principle. A discrete minimum time optimal control problem with state delay is formulated and a decomposition method, based on an augmented Lagrange's function is proposed to solve it. The problem is decomposed in time domain by a new coordinating vector. The obtained algorithms are used for minimum time optimal control calculation of Baker's Yeast fermentation process.List of Symbols x(t) g/l biomass concentration - s(t) g/l limiting substrate concentration - x ⁰ g/l inlet biomass concentration - s ⁰(t) g/l inlet substrate concentration - D(t) h^–1 dilution rate - (t) h^–1 specific growth rate - Y g/g yield coefficient - (t) h^–1 specific limiting substrate consumption rate - k _D h^–1 disappearing constant - w ₁, w ₂ known constant or piece-wise disturbances - _m h^–1 maximum specific growth rate - k _s g/l Michaelis-Menten's parameter - h time delay - x ₀, s ₀ g/l initial concentrations - ¯x, ¯s, ¯D optimal steady state value - V _min , V _max , v=x,s,d,t bounds of variables - t h sampling period - K number of steps in the optimization horison - Js, J _d performance indexes - L _s Lagrange's function - L _d Lagrange's functional - ₀ weighting coefficient for the amount of the limiting substrate throwing out of the fermentor - ₁, ₂ dual variables of Lagrange's function - steps in steady state coordination procedure - errors values for steady state coordination process - _v , v=x, s conjugate variables of Lagrange's functional - _v , v=x,s penalty coefficients of augmented Lagrange's functional - _v , v=x, s interconnections of the time - e _v , v=x,s, D, _x , _s gradients of Lagrange's functional - j, l indexes of calculation procedures - values of errors in calculations The researches was supported by National Scientific Research Foundation under grants No NITN428/94 and No NITN440/94     

Kinetics of sodium tail current during repolarization of axon membrane normally and in the presence of scorpion toxin

The kinetics of the sodium tail current (tail I_Na) in myelinated frog nerve fibers in the region of repolarization potentials (V_r) from –40 to –70 mV has two exponential components: fast and slow. The component composition of the tail I_Na depends on V_r: with an increase in negative values of V_r the contribution of the slow component of the tail (_s) decreases, and at V_r values higher than –80 mV, the tail I_Na follows virtually one fast exponential curve. The component composition of the tail I_Na at a fixed level of V_r depends on the initial conditions: with an increase in the duration of the test pulse (V_t) the amplitude of the fast component of the tail falls much faster than the amplitude of the slow component. In that case the kinetics of the fall in amplitude of the fast component corresponds to the kinetics of inactivation of I_Na. Scorpion toxin causes slowing of the kinetics of the tail I_Na at all values of V_r, mainly on account of an increase in _s. For qualitative interpretation of the results a kinetic scheme assuming the presence of two open states of the sodium channel of the axon membrane is suggested. The hypothesis is put forward that scorpion toxin interacts with the same site of the gating mechanism of the channel as DDT and trinitrophenol.Institute of Cytology, Academy of Sciences of the USSR, Leningrad. Translated from Neirofiziologiya, Vol. 12, No. 5, pp. 541–549, September–October, 1980.