Effects of potassium levels over the range occurring in host plants on the growth of an insect herbivore (Manduca sexta)

Third instar tobacco hornworms (Manduca sexta L.: Sphingidae) on low dietary potassium had a lower relative growth rate than individuals on diets with potassium concentrations reflecting those in host-plants, due to decreased consumption rate, lower efficiencies of conversion of ingested and digested food (ECI and ECD), and a prolonged growth/feeding phase. Furthermore, these larvae, when placed on a diet with a moderate potassium concentration through the fourth stadium, ended up being smaller due to lower ECI and less biomass gained, and had a prolonged growth phase, which suggest an irreversible cost of the previous low potassium diet. Third instar hornworms on high potassium diets had lower ECI and ECD, and they had a prolonged growth phase. These individuals, when placed on a moderate potassium diet in the fourth stadium, gained less biomass, than those previously offered hostplant-like-potassium diets. Body potassium concentrations (% dw) at the end of the third stadium were similar among treatment groups. With increasing potassium concentrations in the diet, utilization efficiencies of potassium decreased and potassium concentrations in the frass increased. Correspondingly, water content (% fw) of the newly-molted fourth instar larvae declined with increasing potassium, indicating a passive loss of water during potassium excretion. Low and high dietary potassium reduced survivorship of third instar larvae; fourth instar caterpillars previously fed the low potassium diet also had poor survivorship. We conclude that, within the normal range of potassium concentrations in the hostplants, caterpillar performance is largely unaffected by potassium concentration, but that potassium-poor and potassium-rich diets, such as those hornworms may sometimes experience, can reduce growth and survivorship.     

The spatial distribution of cations in nematocytes of Hydra vulgaris

The spatial distribution of cations was assayed qualitatively and quantitatively in tentacular nematocytes of Hydra vulgaris in a scanning transmission electron microscope by means of x-ray microanalysis performed on 100 nm thick freeze-dried cryosections. The matrix of undischarged cysts (stenoteles, desmonemes and isorhizas) was found to contain mainly K⁺. In isolated nematocysts of Hydra the intracapsular potassium can be readily substituted by practically any other mono- and divalent cation (Na⁺, NH₄ ⁺, Mn²⁺, Co²⁺, Mg²⁺, Ca²⁺, Fe²⁺) all, except Fe²⁺, without impairing the ability of the cyst to respond to the chemical triggering with dithioerythritol or proteases. Monovalent cations increase the osmotically generated intracapsular pressure compared to divalent ions.     

Intracellular ion concentrations and cell volume during cholinergic stimulation of eccrine secretory coil cells

Summary Methacholine (MCh)-induced changes in intracellular concentrations of Na, K, and Cl ([Na]_i, [K]_i, and [Cl]_i, respectively) and in cellular dry mass (a measure of cell shrinkage) were examined in isolated monkey eccrine sweat secretory coils by electron probe X-ray microanalysis using the peripheral standard method. To further confirm the occurrence of cell shrinkage during MCh stimulation, the change in cell volume of dissociated clear and dark cells were directly determined under a light microscope equipped with differential interference contrast (DIC) optics. X-ray microanalysis revealed a biphasic increase in cellular dry mass in clear cells during continuous MCh stimulation; an initial increase of dry mass to 158% (of control) followed by a plateau at 140%, which correspond to the decrease in cell volume of 37 and 29%, respectively. The latter agrees with the MCh-induced cell shrinkage of 29% in dissociated clear cells. The MCh-induced increase in dry mass in myoepithelial cells was less than half that of clear cells. During the steady state of MCh stimulation, both [K]_i and [Cl]_i of clear cells decreased by about 45%, whereas [Na]_i increased in such a way as to maintain the sum of [Na]_i+[K]_i constant. There was a small (12–15mm) increse in [Na]_i and a decrease in [K]_i in myoepithelial cells during stimulation with MCh. Dissociated dark cells failed to significantly shrink during MCh stimulation. The decrease in [Cl]_i in the face of constant [Na]_i+[K]_i suggests the accumulation of unknown anion(s) inside the clear cell during MCh stimulation. While the decrease in [K]_i and [Cl]_i may be instrumental in facilitating influx of ions via Na–K–2Cl cotransporters, the functional significance of MCh-induced cell shrinkage remains unknown.     

Regulation of basolateral membrane potential after stimulation of Na+ transport in proximal tubules

Summary We have previously shown that stimulation of apical Na-coupled glucose and alanine transport produces a transient depolarization of basolateral membrane potential (V _bl) in rabbit proximal convoluted tubule (PCT. Sl segment). The present study is aimed at understanding the origin of the membrane repolarization following the intial effect of addition of luminal cotransported solutes. Luminal addition of 10–15mMl-alanine produced a rapid and highly significant depolarization ofV _bl (20.3±1.1 mV,n=15) which was transient and associated with an increase in the fractional K⁺ conductance of the basolateral membrane (t _K) from 8 to 29% (P<0.01,n=6). Despite the significant increase int _K, the repolarization was only slightly reduced by the presence of basolateral Ba²⁺ (2mM,n=6) or quinine (0.5 mM,n=5). The repolarization was greatly reduced in the presence of 0.1 mM 4-acetamino-4isothiocyamostilbene-2,2-disulfonic acid (SITS) and blunted by bicarbonate-free solutions. Intracellular pH (pH _i ) determined with the fluorescent dye 2, 7-bis-2-carboxyethyl-5(and-6)-carboxyfluorescein (BCECF), averaged 7.39±0.02 in control solution (n=9) and increased to 7.50±0.03 in the first 15 sec after the luminal application of alanine. This was followed by a significant acidification averaging 0.16±0.01 pH unit in the next 3 min. In conclusion, we believe that, contrary to other leaky epithelia, rabbit PCT can regulate its basolateral membrane potential not only through an increase in K⁺ conductance but also through a cellular acidification reducing the basolateral HCO ₃ ^– exit through the electrogenic Na-3(HCO₃) cotransport mechanism.     

A TEA-insensitive flickering potassium channel active around the resting potential in myelinated nerve

A novel potassium-selective channel which is active at membrane potentials between -100 mV and +40 mV has been identified in peripheral myelinated axons of Xenopus laevis using the patch-clamp technique. At negative potentials with 105 mM-K on both sides of the membrane, the channel at 1 kHz resolution showed a series of brief openings and closings interrupted by longer closings, resulting in a flickery bursting activity. Measurements with resolution up to 10 kHz revealed a single-channel conductance of 49 pS with 105 mM-K and 17 pS with 2.5 mM-K on the outer side of the membrane. The channel was selective for K ions over Na ions (PNa/PK = 0.033). The probability of being within a burst in outside-out patches varied from patch to patch (> 0.2, but often > 0.9), and was independent of membrane potential. Open-time histograms were satisfactorily described with a single exponential (tau o = 0.09 msec), closed times with the sum of three exponentials (tau c = 0.13, 5.9, and 36.6 msec). Sensitivity to external tetraethylammonium was comparatively low (IC50 = 19.0 mM). External Cs ions reduced the apparent unitary conductance for inward currents at Em = -90 mV (IC50 = 1.1 mM). Ba and, more potently, Zn ions lowered not only the apparent single-channel conductance but also open probability. The local anesthetic bupivacaine with high potency reduced probability of being within a burst (IC50 = 165 nM). The flickering K channel is clearly different from the other five types of K channels identified so far in the same preparation. We suggest that this channel may form the molecular basis of the resting potential in vertebrate myelinated axons.     

Regulation of a potassium-selective current in rabbit corneal epithelium by cyclic GMP,carbachol and diltiazem

Summary The effects of cyclic GMP (cGMP), carbachol and diltiazem on a potassium-selective, delayed-rectifier current in freshly dissociated rabbit corneal epithelial cells were studied using a modified perforated-patch-clamp technique. The current was stimulated by both 500 m cGMP (2.3–4.5-fold, mean = 2.9) and 250 nm carbachol, a muscarinic agonist (1.12–7.04-fold, mean = 3.8), and the stimulated current was totally blocked by diltiazem (10 m). The effects of cGMP appeared to be, at least in part, different from those of carbachol as they required the presence of external calcium. Single-channel data suggest that cGMP and carbachol activate the potassium current by increasing the open probability of the channel via a second-messenger system and that the action of diltiazem is probably through a direct blocking effect on the open channel.We are grateful to Erika Wohlfiel for secretarial help, Helen Hendrickson for cell preparation, and Joan Rae for software development. The work was supported by NIH grants EY06005 and EY03282 and an unrestricted award from Research to Prevent Blindness.     

Permeation of Ca2+ through K+ channels in the plasma membrane of Vicia faba guard cells

Summary The whole-cell patch-clamp method has been used to measure Ca²⁺ influx through otherwise K⁺-selective channels in the plasma membrane surrounding protoplasts from guard cells of Vicia faba. These channels are activated by membrane hyperpolarization. The resulting K⁺ influx contributes to the increase in guard cell turgor which causes stomatal opening during the regulation of leaf-air gas exchange. We find that after opening the K⁺ channels by hyperpolarization, depolarization of the membrane results in tail current at voltages where there is no electrochemical force to drive K⁺ inward through the channels. Tail current remains when the reversal potential for permeant ions other than Ca²⁺ is more negative than or equal to the K⁺ equilibrium potential (–47 mV), indicating that the current is due to Ca²⁺ influx through the K⁺ channels prior to their closure. Decreasing internal [Ca²⁺] (Ca _i ) from 200 to 2 nm or increasing the external [Ca²⁺] (Ca _o ) from 1 to 10 mm increases the amplitude of tail current and shifts the observed reversal potential to more positive values. Such increases in the electrochemical force driving Ca²⁺ influx also decrease the amplitude of time-activated current, indicating that Ca²⁺ permeation is slower than K⁺ permeation, and so causes a partial block. Increasing Ca _o also (i) causes a positive shift in the voltage dependence of current, presumably by decreasing the membrane surface potential, and (ii) results in a U-shaped current-voltage relationship with peak inward current ca. –160 mV, indicating that the Ca^2– block is voltage dependent and suggesting that the cation binding site is within the electric field of the membrane. K⁺ channels in Zea mays guard cells also appear to have a Ca _i -, and Ca _o -dependent ability to mediate Ca²⁺ influx. We suggest that the inwardly rectiying K⁺ channels are part of a regulatory mechanism for Ca _i . Changes in Ca _o and (associated) changes in Ca _i regulate a variety of intracellular processes and ion fluxes, including the K⁺ and anion fluxes associated with stomatal aperture change.This work was supported by grants to S.M.A. from NSF (DCB-8904041) and from the McKnight Foundation. K.F.-G. is a Charles Gilbert Heydon Travelling Fellow. The authors thank Dr. R. MacKinnon (Harvard Medical School) and two anonymous reviewers for helpful comments.     

Lipopolysaccharide induction of outward potassium current expression in human monocyte-derived macrophages: Lack of correlation with secretion

Summary Although an outwardly rectifying K⁺ conductance (I _K, A) is prominently expressed in human alveolar macrophages, the expression of this conductance in human monocyte-derived macrophages (HMDMs) is rare. We have analyzed the induction of the expression of I _K, A in voltage-clamped, in vitro differentiated HMDMs by a number of stimuli which produce either priming or activation of macrophages. Cultures were stimulated with lipopolysaccharide (LPS, 2 g/ml), interleukin 2 (IL-2, 100 U/ml), or combinations of LPS and either recombinant interferon-gamma (-IFN, 10 U/ml), phorbol myristate acetate (PMA, 0.01 or 1 g/ ml) and platelet activating factor (PAF, 20 ng/ml) for periods of up to 24 hr. Treatment of the cells with either LPS or IL-2 greatly enhanced the frequency of current expression. Treatment with either PMA or -IFN alone did not induce current expression; treatment of the cells with a combination of LPS and either PMA, -IFN, or PAF did not enhance current expression over that observed with LPS alone. The expression of the outwardly rectifying K⁺ current was observed in 36% (n=321) of the cells for cultures treated with LPS and 33% (n=55) of the cells for cultures treated with IL-2. The inactivating outward K⁺ current was absent in cells which were not treated with either LPS or IL-2. The kinetics of current activation and inactivation appeared identical to that previously described for the transient-inactivating outward current of the human alveolar macrophage. Cycloheximide (1 g/ml), an inhibitor of protein synthesis, completely suppressed LPS-induced current expression. No correlation was found between peak current amplitude and cell size in LPS-activated cells expressing the outwardly rectifying K⁺ current, indicating that current density was not held constant from cell to cell. The coupling of ion channel expression and secretion in individual HMDMs was studied using the reverse hemolytic plaque assay. Although an enhancement of K ⁺ current expression was observed following either LPS or IL-2 treatment, a quantitatively similar and uniform increase in the percentage of either IL-1 or lysozyme-secreting cells was not observed. The frequency of current expression in cells identified as secreting tumor necrosis factor- (TNF-), interleukin 1 (IL-1), or lysozyme was the same or decreased over that observed for nonsecreting cells. Thus, LPS treatment increases the number of K⁺ channels on HMDM membranes; however, K⁺ channel expression alone was not sufficient to give rise to enhanced secretion in LPS-activated macrophages. Enhanced K⁺ channel expression appears to be a part of the primary activation signal. K⁺-channel activation would hyperpolarize the membrane potential, potentially providing the driving force for calcium entry through voltage-independent pathways activated by the subsequent binding of soluble substances to membrane surface receptors, the secondary signal linked to secretion.This work was supported by NIH grant RO 1 GM36823.     

Leaf water relations characteristics of differently potassium fertilized and watered field grown barley plants

Seasonal leaf water relations characteristics were studied in fully irrigated spring barley (Hordeum distichum L. cv. Gunnar) fertilized at low (50 kg K ha⁻¹) or high (200 kg K ha⁻¹) levels of potassium applied as KCl. The investigation was undertaken from about 14 days before anthesis until the milk ripe stage in leaves of different position and age. Additionally, the effects of severe water stress on leaf water relations were studied in the middle of the grain filling period in spring barley (cv. Alis). The leaf water relations characteristics were determined by the pressure volume (PV) technique. Water relations of fully irrigated plants were compared in leaf No 7 with the water relations of slowly droughted plants (cv. Alis). Leaf osmotic potential at full turgor (ψ _π ¹⁰⁰ ) decreased 0.1 to 0.3 MPa in droughted leaves indicating a limited osmotic adjustment due to solute accumulation. The leaf osmotic potential at zero turgor (ψ _π ⁰ ) was about −2.2 MPa in fully irrigated plants and −2.6 MPa in droughted plants. The relative water content at zero turgor (R⁰) decreased 0.1 unit in severely droughted leaves. The ratio of turgid leaf weight to dry weight (TW/DW) tended to be increased by drought. The tissue modulus of elasticity (ε) decreased in droughted plants and together with osmotic adjustment mediated turgor maintenance during drought. A similar response to drought was found in low and high K plants except that the R⁰ and ε values tended to be higher in the high K plants. Conclusively, during drought limited osmotic adjustment and increase in elasticity of the leaf tissue mediated turgor maintenance. These effects were only slightly modified by high potassium application. The seasonal analysis in fully irrigated plants (cv. Gunnar) showed that within about 14 days from leaf emergence ψ _π ¹⁰⁰ decreased from about −0.9 to −1.6 MPa in leaf No 7 (counting the first leaf to emerge as number one) and from about −1.1 to −1.9 MPa in leaf No 8 (the flag leaf) due to solute accumulation. A similar decrease took place in ψ _π ⁰ except that the level of ψ _π ⁰ was displaced to a lower level of about 0.2 to 0.3 MPa. Both ψ _π ¹⁰⁰ and ψ _π ⁰ tended to be 0.05 to 0.10 MPa lower in high K than in low K plants. R⁰ was about 0.8 to 0.9 and was independent of leaf position and age, but tended to be highest in high K plants. The TW/DW ratio decreased from about 5.5 in leaf No 6 to 4.5 in leaf No 7 and 3.8 in leaf No 8. The TW/DW ratio was 4 to 10% higher in high K than in low K plants indicating larger leaf cell size in the former. The apoplastic water content (V_a) at full turgor constituted about 15% in leaf No 7. ε was maximum at full turgor and varied from about 11 to 34 MPa. ε tended to be higher in high K plants. Conclusively, in fully watered plants an ontogenetically determined accumulation of solutes (probably organic as discussed) occurred in the leaves independent of K application. The main effect of high K application on water relations was an increase in leaf water content and a slight decrease in leaf ψ_π. The effect of K status on growth and drought resistance is discussed.