Single-cell, real-time measurements of extracellular oxygen and proton fluxes fromSpirogyra grevilleana

Summary We have adapted the self-referencing microelectrode technique to allow sensitive and noninvasive measurement of oxygen fluxes around single cells. The self-referencing technique is based on the translational movement of a selective microelectrode through the gradient next to the cell wall or membrane. The electrode is moved at a known frequency and between known points. The differential electrode output values are converted into a directional measurement of flux by the Fick equation. By coupling the newly developed oxygen-selective self-referencing electrochemical microelectrode (SREM-O₂) system with self-referencing ionselective proton measurements (SRIS-H⁺) we have characterized oxygen and proton fluxes from a single cell of the filamentous green algaSpirogyra gre illeana (Hass.). Oxygen showed a net efflux and protons showed a net influx when the cell was illuminated. These photosynthesis-dependent fluxes were found to be spatially associated with the chloroplasts and were sensitive to treatment with dichlorophenyldimethylurea. In the dark the directions of oxygen and proton fluxes were reversed. This oxygen influx was associated with mitochondrial respiration and was reduced by 78% when the cells was treated with 0.5 mM KCN. The residual cyanide-resistant respiration was inhibited by the application of 5 mM salicylhydroxamic acid, an inhibitor of the alternative oxidase. Similarly the cytochrome pathway was also inhibited by the presence of 20 M NO, while the cyanide-resistant alternative oxidase was not. These results demonstrate the use of the newly developed SREM-O₂ system to measure and characterize metabolic fluxes at a level of sensitivity that allows for subcellular resolution. These measurements, in conjunction with SERIS-H⁺ measurements, have led to new insights in our understanding of basic cellular physiology in plant cells.Abbreviations SRIS self-referencing ion selective - SREM self-referencing electrochemical microelectrode - ICP inductive coupled plasma spectroscopy     

An analytical model of ionic movements in airway epithelial cells.

A new mathematical model of ion movements in airway epithelia is presented, which allows predictions of ion fluxes, membrane potentials and ion concentrations. The model includes sodium and chloride channels in the apical membrane, a Na/K pump and a cotransport system for Cl- with stoichiometry Na+:K+:2Cl- in the basolateral membrane. Potassium channels in the basolateral membrane are used to regulate cell volume. Membrane potentials, ion fluxes and intracellular ion concentration are calculated as functions of apical ion permeabilities, the maximum pump current and the cotransport parameters. The major predictions of the model are: (1) Cl- concentration in the cell is determined entirely by the intracellular concentration of negatively charged impermeable ions and the osmotic conditions; (2) changes in intracellular Na+ and K+ concentrations are inversely related; (3) cotransport provides the major driving force for Cl- flux, increases intracellular Na+ concentration, decreases intracellular K+ concentration and hyperpolarizes the cell interior; (4) the maximum rate of the Na/K pump, by contrast, has little effect on Na+ or Cl- transepithelial fluxes and a much less pronounced effect on cell membrane polarization; (5) an increase in apical Na+ permeability causes an increase in intracellular Na+ concentration and a significant increase in Na+ flux; (6) an increase in apical Cl- permeability decreases intracellular Na+ concentration and Na+ flux; (7) assuming Na+ and Cl- permeabilities equal to those measured in human nasal epithelia, the model predicts that under short circuit conditions, Na+ absorption is much higher than Cl- secretion, in agreement with experimental measurements.     

Measurements of net fluxes and extracellular changes of H+, Ca2+, K+, and NH4+ in Escherichia coli using ion-selective microelectrodes.

This study introduced the use of a non-invasive ion-selective microelectrode (MIFE) technique to study membrane-transport processes in bacteria. Net ion fluxes and changes in the extracellular concentrations of H+, Ca2+, K+ and NH4+ in adherent bacteria, isolated from cultures at different growth stages (exponential, late exponential, and stationary phases), were monitored. With the exception of Ca2+, a significant (P=0.05) difference was found in the magnitude of net fluxes of the ions measured from bacterial cells at different stages of the population growth curve. The magnitude of the H+ response was glucose-dependent with maximum changes occurring at the highest concentration. There was a progressive increase in H+ extrusion followed by a gradual return to zero at late stationary phase. Measurements of net ion fluxes crossing the bacterial cytoplasmic membrane, demonstrated here for the first time, may offer insight into underlying mechanisms of ion transport kinetics. Applications of the non-invasive ion-selective microelectrode technique in microbiology are discussed.     

Changes in ion fluxes during phototropic bending of etiolated oat coleoptiles

BACKGROUND AND AIMS: This work has been conducted to assist theoretical modelling of the different stages of the blue light (BL)-induced phototropic signalling pathway and ion transport activity across plant membranes. Ion fluxes (Ca(2+), H(+), K(+) and Cl(-)) in etiolated oat coleoptiles have been measured continuously before and during unilateral BL exposure. METHODS: Changes in ion fluxes at the illuminated (light) and shadowed (dark) sides of etiolated oat coleoptiles (Avena sativa) were studied using a non-invasive ion-selective microelectrode technique (MIFE). The bending response was also measured continuously, and correlations between the changes in various ion fluxes and bending response have been investigated. For each ion the difference (Delta) between the magnitudes of flux at the light and dark sides of the coleoptile was calculated. KEY RESULTS: Plants that demonstrated a phototropic bending response also demonstrated Ca(2+) influx into the light side approximately 20 min after the start of BL exposure. This is regarded as part of the perception and transduction stages of the BL-induced signal cascade. The first 10 min of bending were associated with substantial influx of H(+), K(+) and Cl(-) into the light (concave) side of the coleoptiles. CONCLUSIONS: The data suggest that Ca(2+) participates in the signalling stage of the BL-induced phototropism, whereas the phototropic bending response is linked to changes in the transport of H(+), K(+) and Cl(-).     

Ionic currents and ion fluxes in Neurospora crassa hyphae

Voltage dependence of ionic currents and ion fluxes in a walled, turgor-regulating cell were measured in Neurospora crassa. The hyphal morphology of the model organism Neurospora simplifies cable analysis of ionic currents to determine current density for quantitative comparisons with ion fluxes. The ion fluxes were measured directly and non-invasively with self-referencing ion-selective microelectrodes. Four ions (H(+), Ca(2+), K(+), and Cl(-)) were examined. H(+) net uptake and Ca(2+) net release were small (10.2 nmol m(-2) s(-1) and 1.1 nmol m(-2) s(-1), respectively) and voltage independent. K(+) and Cl(-) fluxes were larger and voltage dependent. Maximal K(+) net release ( approximately 1440 nmol m(-2) s(-1)) was observed at positive voltages (+15 mV), while maximal Cl(-) net release ( approximately 905 nmol m(-2) s(-1)) was observed at negative voltage (-210 mV). A possible function of the net outward K(+) and Cl(-) fluxes is regulation of the plasma membrane potential. Total ion fluxes were 37-58% of the total ionic current density (about +/-244 mA m(-2), equivalent to +/-2500 nmol m(-2) s(-1), at 0 mV and -200 mV) so other ions must contribute significantly to the ionic currents.     

The Double Fixed Charge Membrane: Solution-Membrane Ion Partition Effects and Membrane Potentials

An analysis is made of the effect of solution-membrane partition of ions on the electrostatic potential and ion concentration profiles in fixed charge membranes. It is shown that the inclusion of partition effects gives rise to large solution-membrane “Donnan” potentials even when the concentration of fixed charges is of the same order as the concentration of the external solution. This effect renders the system and the simplified analysis of the double fixed charge membrane (FCM) previously given more applicable to biological membranes. An analysis is also given of the voltage dependence of the fluxes of individual ion species in the double FCM when it separates different ionic solutions and an expression is deduced for the membrane resting potential. Although the latter is similar in form to the Goldman-Hodgkin-Katz (GHK) equation the corresponding value of the permeability ratio P_C1/P_K is under certain specified conditions both concentration and potential dependent.     

Diuretic factors and second messengers stimulate secretion of the organic cation TEA by the Malpighian tubules of Drosophila melanogaster

This study showed that four factors which stimulate transepithelial fluid secretion and inorganic ion transport across the main segment of the Malpighian tubules of Drosophila melanogaster also stimulate transepithelial secretion of the prototypical organic cation tetraethylammonium (TEA). TEA fluxes across the Malpighian tubules and gut were measured using a TEA-selective self-referencing (TEA-SeR) microelectrode. TEA flux across isolated Malpighian tubules was also measured using a TEA-selective microelectrode positioned in droplets of fluid secreted by tubules set up in a modified Ramsay assay. TEA flux was stimulated by the intracellular second messengers cAMP and cGMP, which increase the lumen-positive transepithelial potential (TEP), and also by tyramine and leucokinin-I (LK-I), which decrease TEP. The largest increase was measured in response to 1 micromol l-1 LK-I which increased transepithelial TEA flux by 72%. TEA flux in the lower tubule was stimulated slightly (13%) by 1 micromol l-1 tyramine but not by any of the other factors. TEA flux across the midgut was unaffected by cAMP, cGMP or tyramine. This is the first study to demonstrate the effects of insect diuretic factors and second messengers on excretion of organic cations.     

Comparative analysis of Ca2+ and H+ flux magnitude and location along growing hyphae of Saprolegnia ferax and Neurospora crassa

Calcium and proton ion fluxes were mapped at the growing apices of two hyphal organisms, the oomycete Saprolegnia ferax and the ascomycete Neurospora crassa and pseudohyphal Saccharomyces cerevisiae using self-referencing ion-selective probes. S. ferax exhibited well-defined transport zones absent in N. crassa. Ca2+ fluxes were located within 8 microm of the growing hyphal tip; the net Ca2+ flux was either inward (75% of all experiments) or outward. The inward component of the net flux was inhibited by Gd3+, known to inhibit Ca2+ permeable stretch-activated channels. Because the Ca2+ flux is located at the region of maximal hyphal expansion, exocytosis may contribute to Ca2+ efflux, in addition to the stretch-activated channel mediated influx. Maximal inward H+ flux was observed 10-30 microm behind the hyphal tip where peak mitochondria densities taper off at the onset of a vacuolation zone, presumably due to highly localized H+ cotransporter activity. By contrast, N. crassa exhibited no net Ca2+ flux and a consistently inward H+ flux (93% of all experiments) that was homogeneously distributed up to 60 microm behind the hyphal apex. Both hyphal organisms have similar tip morphology and growth rates, and are reported to have tip-high cytosolic Ca2+ gradients associated with growth. Only S. ferax exhibited tip-localized Ca2+ fluxes and a well defined H+ influx zone just behind the tip. Differences in ecological habitats and cytology--S. ferax is an aquatic organism that grows as a migrating plug of cytoplasm while N. crassa is normally terrestrial with a cytoplasm-rich mycelium and highly active cytoplasmic streaming behind the growing margin--may account for the differences in the 'architecture' of ion transport occurring during the process of tip growth. Net Ca2+ efflux and H+ influx of growing S. cerevisiae pseudohyphae were also measured but localization was not possible due to small cell size.     

Use of an extracellular,ion-selective,vibrating microelectrode system for the quantification of K+, H+, and Ca2+ fluxes in maize roots and maize suspension cells

An ion-selective vibrating-microelectrode system, which was originally used to measure extracellular Ca²⁺ gradients generated by Ca²⁺ currents, was used to study K⁺, H⁺ and Ca²⁺ transport in intact maize (Zea mays L.) roots and individual maize suspension cells. Comparisons were made between the vibrating ion-selective microelectrode, and a technique using stationary ion-selective microelectrodes to measure ionic gradients in the unstirred layer at the surface of plant roots. The vibrating-microelectrode system was shown to be a major improvement over stationary ion-selective microelectrodes, in terms of sensitivity and temporal resolution. With the vibrating ion microelectrode, it was easy to monitor K⁺ influxes into maize roots in a background K⁺ concentration of 10 mM or more, while stationary K⁺ electrodes were limited to measurements in a background K⁺ concentration of 0.3 mM or less. Also, with this system it was possible to conduct a detailed study of root Ca²⁺ transport, which was previously not possible because of the small fluxes involved. For example, we were able to investigate the effect of the excision of maize roots on Ca²⁺ influx. When an intact maize root was excised from the seedling at a position 3 cm from the site of measurement of Ca²⁺ transport, a rapid fourfold stimulation of Ca²⁺ influx was observed followed by dramatic oscillations in Ca²⁺ flux, oscillating between Ca²⁺ influx and efflux. These results clearly demonstrate that wound or perturbation responses of plant organs involve transient alterations in Ca²⁺ transport, which had previously been inferred by demonstrations of touch-induced changes in cytoplasmic calcium. The sensitivity of this system allows for the measurement of ion fluxes in individual plant cells. Using vibrating K⁺ and H⁺electrodes, it was possible to measure H⁺efflux and both K⁺ influx and efflux in individual maize suspension cells under different conditions. The availability of this technique will greatly improve our ability to study ion transport at the cellular level, in intact plant tissues and organs, and in specialized cells, such as root hairs or guard cells.Symbol X amplitude of vibration The authors would like to thank Richard Sanger for his invaluable work on the design and improvement of the ion-selective vibratingmicroelectrode system. The research presented here was supported in part by U.S. Department of Agriculture Competitive Grant No. 90-37261-5411 to Leon Kochian and William Lucas.     

选择性微电极在植物生理学研究中的应用

选择性微电极技术是一种不仅能直接测定活的生物细胞或细胞器内的离子或分子活度,而且能对活的生物相邻的位置、功能和代谢速率可能不同的特定微区细胞表面的离子或分子流（flux）分别测定的电生理方法。具有操作简便、实时、非损伤性（测定离子或分子流）、灵敏度高（可达10^-12moles cm^-2s^-1）等优点。因为它是用微型化（尖端直径为0.5-5μm）的离子或分子选择性电极直接对准样品测定,不同于其他化学测定需取样品,所以能连续测定和自动监测,具有广阔的应用前景。该文阐述了选择性微电极测定原理,总结了选择性微电极技术在植物生理学研究中的应用进展,并展望了其发展前景。     

Double-barreled and Concentric Microelectrodes for Measurement of Extracellular Ion Signals in Brain Tissue

Electrical activity in the brain is accompanied by significant ion fluxes across membranes, resulting in complex changes in the extracellular concentration of all major ions. As these ion shifts bear significant functional consequences, their quantitative determination is often required to understand the function and dysfunction of neural networks under physiological and pathophysiological conditions. In the present study, we demonstrate the fabrication and calibration of double-barreled ion-selective microelectrodes, which have proven to be excellent tools for such measurements in brain tissue. Moreover, so-called “concentric” ion-selective microelectrodes are also described, which, based on their different design, offer a far better temporal resolution of fast ion changes. We then show how these electrodes can be employed in acute brain slice preparations of the mouse hippocampus. Using double-barreled, potassium-selective microelectrodes, changes in the extracellular potassium concentration ([K⁺]_o) in response to exogenous application of glutamate receptor agonists or during epileptiform activity are demonstrated. Furthermore, we illustrate the response characteristics of sodium-sensitive, double-barreled and concentric electrodes and compare their detection of changes in the extracellular sodium concentration ([Na⁺]_o) evoked by bath or pressure application of drugs. These measurements show that while response amplitudes are similar, the concentric sodium microelectrodes display a superior signal-to-noise ratio and response time as compared to the double-barreled design. Generally, the demonstrated procedures will be easily transferable to measurement of other ions species, including pH or calcium, and will also be applicable to other preparations.     

Ion transport in the simplest single file pore

A kinetic scheme is developed to describe single-file transport through pores containing up to two ions which may be of different species. The solution for the fluxes in terms of rate constants for entry, exit, and transfer is derived without specific assumptions about symmetry or the voltage and activity dependence of the constants. For a symmetrical pore the relation between the slope conductance at zero applied potential and ion activity can have two distinct regions in which the conductance increases linearly. Zero current or reversal potentials depend on the absolute values of the activities as well as their ratios. The use of this theory to describe the cation fluxes through the pores formed by gramicidin A will be considered in a subsequent paper. Here the model is discussed for a number of more specific assumptions, most extensively the following combination: (1) while entry to a pore is less likely when the pore is already occupied at the opposite end, this entry is still rapid; (2) exit is much more rapid when the pore is occupied by two ions; and (3) transfer from one end to the other of a singly occupied pore is rapid. With these assumptions and for a range of concentrations over which the fluxes are proportional to ion activities, the model predicts a flux ratio exponent nearly equal to 2, blocking by impermeant ions, rectification due to blocking particles on one side only, relief of block by increase in the permeant ion concentration on the opposite side, and anomalous variations of the conductance and zero current potential with mole ratio when the total concentration of the two permeants is held constant.     

Ion transport in the simplest single file pore.

A kinetic scheme is developed to describe single-file transport through pores containing up to two ions which may be of different species. The solution for the fluxes in terms of rate constants for entry, exit, and transfer is derived without specific assumptions about symmetry or the voltage and activity dependence of the constants. For a symmetrical pore the relation between the slope conductance at zero applied potential and ion activity can have two distinct regions in which the conductance increases linearly. Zero current or reversal potentials depend on the absolute values of the activities as well as their ratios. The use of this theory to describe the cation fluxes through the pores formed by gramicidin A will be considered in a subsequent paper. Here the model is discussed for a number of more specific assumptions, most extensively the following combination: (1) while entry to a pore is less likely when the pore is already occupied at the opposite end, this entry is still rapid; (2) exit is much more rapid when the pore is occupied by two ions; and (3) transfer from one end to the other of a singly occupied pore is rapid. With these assumptions and for a range of concentrations over which the fluxes are proportional to ion activities, the model predicts a flux ratio exponent nearly equal to 2, blocking by impermeant ions, rectification due to blocking particles on one side only, relief of block by increase in the permeant ion concentration on the opposite side, and anomalous variations of the conductance and zero current potential with mole ratio when the total concentration of the two permeants is held constant.     

Cation fluxes cause plasma membrane depolarization involved in beta-glucan elicitor-signaling in soybean roots

Inducible and specific ion fluxes on plasma membranes represent very early events during elicitation of plant cells. The hierarchy of such ion fluxes involved is still unknown. The effect of Phytophthora sojae-derived beta-glucan elicitors on the plasma membrane potential as well as on surface K+, Ca2+, and H+ fluxes has been investigated on soybean roots using ion-selective microelectrodes. Beta-Glucans with different degrees of polymerization transiently depolarized the plasma membrane. The elicitor concentration necessary for half-maximal depolarization closely resembled the corresponding binding affinities of soybean root membranes toward the respective beta-glucans. Upon repeated elicitor treatment, the root cells responded partially refractory, suggesting a complex responsiveness of the system. Within the root hair space, characteristic decreasing K(+)- and Ca(2+)-free concentrations were induced by the elicitors, probably causing depolarization through the influx of positive charges. Whereas K+ fluxes were inverted after passing the K+ equilibrium (Nernst-) potential, Ca2+ influx continued. No anion fluxes sufficient to account for charge compensation were observed under the same experimental conditions. K+ and Ca2+ fluxes as well as depolarization were inhibited by 100 microM or less of the Ca2+ antagonist La3+. Contrasting other systems, in soybean the main cause for elicitor-induced plasma membrane depolarization is the activation of cation instead of anion fluxes.     

Sustaining olfaction at low salinities: evidence for a paracellular route of ion movement from the hemolymph to the sensillar lymph in the olfactory sensilla of the blue crab Callinectes sapidus

Evidence reported previously suggests that in low-salinity conditions the integrity of the olfactory dendrites of the blue crab is sustained by a diffusion-generated ionic microenvironment within the aesthetascs. Diffusion of ions from the hemolymph to the sensillar lymph is proposed to maintain this microenvironment. In this study, using lanthanum as an electron-dense marker of extracellular fluid space, we find morphological evidence for paracellular continuity between the hemolymph and the sensillar lymph. Lanthanum penetrates extracellular fluid spaces within the aesthetascs when antennules are either perfused or bathed externally with solutions containing lanthanum nitrate. This was found in both freshwater- and seawater-acclimated animals. Evidence for ion diffusion from the aesthetascs was obtained using self-referencing, ion-selective microelectrodes. Both Ca2+ and K+ exhibit outwardly directed flux gradients associated with the aesthetasc tuft in low-salinity conditions. These findings are consistent with the concept that ion diffusion from the hemolymph to the sensillar lymph generates an ionic/osmotic microenvironment within the aesthetascs at low salinities.     

Non-invasive self-referencing electrochemical sensors for quantifying real-time biofilm analyte flux

Current techniques for characterizing biofilm physiology lack the signal filtering capability required for quantifying signals associated with real time biologically active transport. Though a great deal was learned from previous investigations, no results have been reported on the characterization of in vivo, real time biofilm flux using non-invasive (non-destructive) techniques. This article introduces the self-referencing technique for applications in biofilm physiology. Self-referencing is a non-invasive sensing modality which is capable of sensing changes in biologically active analyte flux as small as 10 fmol cm(-2) s(-1). Studies directly characterizing flux, as opposed to concentration, have the advantage of quantifying real time changes in biologically active transport which are otherwise lost to background noise. The use of this modality for characterizing biofilm physiology is validated with a reversible enzyme inhibition study. The experiment used self-referencing potentiometric sensors for quantifying real time ammonium and nitrite flux. Amperometric and optical sensing methods, though not presented herein, are also powerful sensing tools which benefit from operation in self-referencing mode. Reversible ammonia monooxygenase inhibition by a copper chelator (thiourea), and subsequent relief by excess copper addition was successfully demonstrated using self-referencing ion-selective microelectrodes for a mature Nitrosomonas europaea biofilm.     

Simultaneous flux and current measurement from single plant protoplasts reveals a strong link between K+ fluxes and current, but no link between Ca2+ fluxes and current

We present a thorough calibration and verification of a combined non-invasive self-referencing microelectrode-based ion-flux measurement and whole-cell patch clamp system as a novel and powerful tool for the study of ion transport. The system is shown to be capable of revealing the movement of multiple ions across the plasma membrane of a single protoplast at multiple voltages and in complex physiologically relevant solutions. Wheat root protoplasts are patch clamped in the whole-cell configuration and current-voltage relations obtained whilst monitoring net K+ and Ca2+ flux adjacent to the membrane with ion-selective electrodes. At each voltage, net ion flux (nmol m(-2) sec(-1)) is converted to an equivalent current density (mA m(-2)) taking into account geometry and electrode efficiency, and compared with the net current density measured with the patch clamp system. Using this technique, it is demonstrated that the K+-permeable outwardly rectifying conductance (KORC) is responsible for net outward K+ movement across the plasma membrane [1:1 flux-to-current ratio (1.21 +/- 0.14 SEM, n = 15)]. Variation in the K+ flux-to-current ratio among single protoplasts suggests a heterogeneous distribution of KORC channels on the membrane surface. As a demonstration of the power of the technique we show that despite a significant Ca2+ permeability being associated with KORC (analysis of tail current reversal potentials), there is no correlation between Ca2+ flux and KORC activity. A very significant observation is that large Ca2+ fluxes are electrically silent and probably tightly coupled to compensatory charge movements. This analysis demonstrates that it is mandatory to measure flux and currents simultaneously to investigate properly Ca2+ transport mechanisms and selectivity of ion channels in general.     

Time resolved secretion of chloride from a monolayer of mucin-secreting epithelial cells

Short-circuit current (Isc) measurement is used to quantify transepithelial ion flux. This technique provides a direct measure of net charge transport across a cell monolayer. Isc however, lacks chemical selectivity. Chemically resolved ion fluxes may be much greater than Isc, and differ in different biological processes. This work describes a novel experimental approach and deconvolution method to obtain temporally resolved ion fluxes at epithelial cell monolayers. HT29-Cl.16E cells, a sub clone of the human colonic cancer cell line HT29 was used as a model cell line to validate this approach in the context of epithelial transport studies. This cell line is known to secrete chloride in response to purinergic stimulation. Changes in chloride concentration after stimulation with 1 mM ATP plus 50 nM phorbol-myristate acetate (PMA) are recorded with a chloride ion-selective electrode (ISE) at a short distance (∼50 μm) from the monolayer. The recorded concentrations are transformed to corresponding chloride flux across the monolayer using a deconvolution algorithm for extracellular mass transport based on minimization of the shape error function (Nair and Gratzl in Anal Chem 77:2875–2888, 2005). Simultaneous voltage clamp yields the associated net electrical charge flux (Isc). The dynamics of Cl⁻ flux did correlate with that of the electrical flux, but was found to be greater in amplitude. This suggests that Cl⁻ may not be the only ion secreted. The method of simultaneously assessing ionic and electrical fluxes with a temporal resolution of seconds provides unique information about the dynamics of solute fluxes across the apical membrane. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Electrolyte composition of pulmonary alveolar subphase in anesthetized rabbits

We measured the concentration of Na+, K+, Ca2+, and Cl- in the aqueous subphase of the alveolar lining by puncturing the most superficial alveoli of the exposed lungs of anesthetized rabbits with ion-selective microelectrodes and a nonselective KCl microelectrode. A buffered electrolyte solution bathed the lung surface to keep it moist and warm (38 +/- 1 degrees C) and to serve as a reference for each measurement of ionic concentration. The serum and alveolar concentrations (meq/l) were Na+ 134 +/- 6 and 135 +/- 5, K+ 3.4 +/- 0.2 and 7.3 +/- 0.7, Ca2+ 3.1 +/- 0.2 and 3.2 +/- 0.4, and Cl- 106 +/- 7 and 103 +/- 5 (mean +/- SD). Only K+ was significantly different (P less than 0.001). There was a small electrical potential difference between the alveolar lumen and the pleural surface (-3.5 +/- 0.8 mV, lumen negative) that was significantly different from zero (P less than 0.001). Although it is not possible to measure ion fluxes with these techniques, the results are consistent with active transport of one or more of the ions studied.     

Electrical correlates of secretion in endocrine and exocrine cells

Many types of secretory cells including neurons and cells of endocrine and exocrine glands show changes in electrical potential and resistance when secretion is stimulated. These electrical correlates result from the movement of ions across the cell membrane through specific ion-selective channels. In neurons and certain endocrine cells (such as pancreatic beta cells and certain cells of the anterior pituitary), these channels are voltage dependent and open transiently upon depolarization leading to action potentials. Thus some endocrine cells are electrically excitable, a property previously held to occur only in nerve and muscle. In other nonexcitable endocrine and exocrine cells (such as the pancreas and parotid), ion channels are responsive to either occupancy of specific membrane receptors or changes in intracellular metabolites and second messengers. Ion fluxes through these latter channels also lead to changes in the electrical potential and resistance, but these changes are generally more sustained and action potentials are not seen. The entry of Ca2+ through both voltage-dependent and voltage-independent ion channels plays a major role in the activation of secretion via exocytosis.