scholarly journals Barium and calcium block Bacillus thuringiensis subspecies Kurstaki delta-endotoxin inhibition of potassium current across isolated midgut of larval Manduca sexta

1988 ◽  
Vol 137 (1) ◽  
pp. 277-286 ◽  
Author(s):  
D. N. Crawford ◽  
W. R. Harvey
Keyword(s):  
Steady State ◽  
Manduca Sexta ◽  
Potassium Current ◽  
Short Circuit ◽  
Short Circuit Current ◽  
K Channel ◽  
Midgut Cell ◽  
Bt Toxin ◽  
Steady State Model ◽  
Delta Endotoxin

Ba2+ and Ca2+ prevent and reverse the Btk delta-endotoxin inhibition of the short-circuit current in isolated lepidopteran midgut. These findings support the K+ pump-leak steady-state model for midgut K+ homeostasis and the K+ channel mechanism of Bt toxin action. They provide a new tool with which to study the interactions between Bt toxin and midgut cell membranes.

scholarly journals K+ CHANNEL PERMEATION AND BLOCK IN THE MIDGUT EPITHELIUM OF THE TOBACCO HORNWORM MANDUCA SEXTA

1994 ◽  
Vol 197 (1) ◽  
pp. 179-200
Author(s):  
K Schirmanns ◽  
W Zeiske
Keyword(s):  
Manduca Sexta ◽  
Short Circuit ◽  
Short Circuit Current ◽  
K Channel ◽  
K Channels ◽  
Zero Current ◽  
Channel Opening ◽  
Voltage Dependent ◽  
K Current

The K+-secreting larval midgut of Manduca sexta in vitro was voltage- or current-clamped. In contrast to Tl+, NH4+ and Na+, both Rb+ and K+ generated a short-circuit current, although with different saturation kinetics. The dependence of the short-circuit current on Rb+/K+ mole fraction gave no evidence for multi-ion occupation of the basolateral K+ channels. After 'functionally' eliminating the apical membranes using the ionophore amphotericin B and the 'apical K+ pump' blockers trimethyltin chloride or Tl+, the K+ channels could be more closely investigated. By measuring zero-current potentials, permeability ratios PX/PK were estimated using an adapted version of the Goldman­Hodgkin­Katz voltage equation. Their sequence was K+ (1) = Tl+ > Rb+ (0.38) > NH4+ (~0.3) > Cs+ (0.03) > Na+ (~0). The K+ channels could not be blocked by basally applied Cs+, Na+ or tetraethylammonium. Blockade of K+ current by Ba2+ was typically voltage-dependent, but only at moderate transbasal voltages. The relative electrical distance delta of the Ba2+ binding site from the basal channel opening was determined to be 0.2. At zero transbasal voltage, the apparent inhibition constant for barium KBa* was 1.7 mmol l-1.

Active transport of magnesium across the isolated midgut of Hyalophora cecropia

1975 ◽  
Vol 63 (2) ◽  
pp. 313-320
Author(s):  
J. L. Wood ◽  
A. M. Jungreis ◽  
W. R. Harvey
Keyword(s):  
Steady State ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Potassium Transport ◽  
Magnesium Concentration ◽  
Magnesium Transport ◽  
Wet Weight ◽  
Net Flux ◽  
Burk Plot

1. The 28Mg-measured net flux of magnesium from lumen-side to haemolymph-side of the isolated and short-circuited midgut was 1.97 +/− 0.28 mu-equiv cm(−2) /(−1) in 8 mM-Mg2+. 2. The magnesium-influx shows a delay before the tracer steady-state is attained, indicating the existence of a magnesium-transport pool equivalent to 6.7 mu-equiv/g wet weight of midgut tissue. 3. Magnesium depresses the short-circuit current produced the midgut but not the potassium transport, the depression being equal to the rate of magnesium transport. 4. Magnesium transport yields a linear Lineweaver-Burk plot with an apparent Km of 34 mM-Mg2+ and an apparent Vmax of 14.9 mu-equiv cm(−1) /(−1). 5. Magnesium is actively transported across the midgut and contributes to the regulation of the haemolymph magnesium concentration in vivo.

Bathing Solution Tonicity and Potassium Transport by the Midgut of the Tobacco Hornworm Manduca Sexta

1979 ◽  
Vol 78 (1) ◽  
pp. 213-223
Author(s):  
DAVID F. MOFFETT
Keyword(s):  
Manduca Sexta ◽  
Short Circuit ◽  
Weight Ratio ◽  
Short Circuit Current ◽  
Dry Weight ◽  
Potassium Transport ◽  
Bathing Solution ◽  
Histological Studies ◽  
Wet Weight ◽  
Small Solute

Potassium transport by the isolated midgut of Manduca larvae, as measured by the short circuit current, is inhibited by substitution of small organic solutes (M.W. < 340) for the sucrose normally included in bathing solution formulated for this tissue. Other solutes of molecular weight equal to or greater than sucrose are essentially as effective as sucrose in promoting the short circuit current. Equilibration of midgut in solutions containing the small solute mannitol results in a decrease in the dry weight/wet weight ratio of the tissue, suggesting that the small solutes can penetrate into areas of the tissue which are not accessible to sucrose. Histological studies suggest that sites of swelling in the presence of mannitol include both cytoplasm and goblet cell lumen. The inhibition of the short circuit current is rapidly reversible on return to bathing solution containing sucrose or another large solute. The effect of small solutes probably does not involve compromise of the energy source for potassium transport since oxygen uptake is unchanged in the presence of a small solute.

scholarly journals Zinc inhibits cAMP-stimulated Cl secretion via basolateral K-channel blockade in rat ileum

2005 ◽  
Vol 288 (5) ◽  
pp. G956-G963 ◽  
Author(s):  
Kazi Mirajul Hoque ◽  
Vazhaikkurichi M. Rajendran ◽  
Henry J. Binder
Keyword(s):  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
K Channel ◽  
Isolated Cells ◽  
Rat Ileum ◽  
Anion Secretion ◽  
Cl Secretion ◽  
Ion Absorption ◽  
Isotonic Buffer

Zn, an essential micronutrient and second most abundant trace element in cell and tissues, reduces stool output when administered to children with acute diarrhea. The mechanism by which Zn improves diarrhea is not known but could result from stimulating Na absorption and/or inhibiting anion secretion. The aim of this study was to investigate the direct effect of Zn on intestinal epithelial ion absorption and secretion. Rat ileum was partially stripped of serosal and muscle layers, and the mucosa was mounted in lucite chambers. Potential difference and short-circuit current were measured by conventional current-voltage clamp method.86Rb efflux and uptake were assessed for serosal K channel and Na-K-2Cl cotransport activity, respectively. Efflux experiments were performed in isolated cells preloaded with86Rb in the presence of ouabain and bumetanide, whereas uptake experiments were performed in low-Cl isotonic buffer containing Ba and ouabain. Neither mucosal nor serosal Zn affected glucose-stimulated Na absorption. In contrast, forskolin-induced Cl secretion was markedly reduced by serosal but not mucosal addition of Zn. Zn also substantially reversed the increase in Cl secretion induced by 8-bromoadenosine 3′,5′-cyclic monophosphate (8-BrcAMP) with half-maximal inhibitory concentration of 0.43 mM. In contrast, serosal Zn did not alter Cl secretion stimulated by carbachol, a Ca-dependent agonist. Zn inhibited 8-BrcAMP-stimulated86Rb efflux but not carbachol-stimulated86Rb efflux. Zn had no effect on bumetanide-sensitive86Rb uptake, Na-K-ATPase, or CFTR. We conclude from these studies that Zn inhibits cAMP-induced Cl secretion by blocking basolateral membrane K channels.

Electron probe X-ray microanalysis of the effects of Bacillus thuringiensis var kurstaki crystal protein insecticide on ions in an electrogenic K+-transporting epithelium of the larval midgut in the lepidopteran, Manduca sexta, in vitro

1985 ◽  
Vol 74 (1) ◽  
pp. 137-152
Author(s):  
B.L. Gupta ◽  
J.A. Dow ◽  
T.A. Hall ◽  
W.R. Harvey
Keyword(s):  
Bacillus Thuringiensis ◽  
Manduca Sexta ◽  
Goblet Cell ◽  
Electron Probe ◽  
Apical Membrane ◽  
Short Circuit ◽  
Short Circuit Current ◽  
X Ray ◽  
Elemental Concentrations

An alkaline hydrolysate of Bacillus thuringiensis var kurstaki HD1 (Btk) parasporal crystals was administered at 25 micrograms ml-1 (f.c.) to isolated, short-circuited, midguts of tobacco hornworm (Manduca sexta) larvae. The short-circuit current (s.c.c.), a precise measure of K+ active transport, was inhibited by 78% in 10 min in Btk-treated midguts as compared to controls. The elemental concentrations of K, together with Na, Mg, P, S, Cl and Ca, as well as the water content, were determined by electron probe X-ray microanalysis (EPXMA) in the muscle cells, columnar cells and goblet cells, as well as in the extracellular goblet cavity and the bathing media. The average K concentration in the goblet cell cavity was 129 mmol/kg wet wt in control midguts but only 37 mmol/kg wet wt in Btk-treated midguts. The elemental concentrations, including that of K, in other cell compartments were much less affected by Btk, but a rise in total cell calcium is suggested. It has been previously suggested that in vitro Btk acts specifically on limited regions of the apical membrane of the midgut epithelial cells. The simplest interpretation of the EPXMA results would be that initially Btk interacts specifically with the goblet cell apical membrane, which bounds the goblet cavity and contains the K+ pump responsible for the s.c.c. and high transepithelial potential difference (p.d.). Such interaction results in a rapid disruption of K+ transport across the goblet cell apical membrane, leading to dissipation of the K+ gradient and loss of p.d. The histopathological changes previously reported by other workers would then be a consequence of K+ pump inhibition causing changes in the intracellular pH, Ca2+ etc. Some possible molecular bases for these specific interactions between Btk and cell membrane are discussed.

Potassium conductance in rabbit esophageal epithelium

1993 ◽  
Vol 265 (1) ◽  
pp. G28-G34 ◽  
Author(s):  
W. E. Khalbuss ◽  
R. Alkiek ◽  
C. G. Marousis ◽  
R. C. Orlando
Keyword(s):  
Steady State ◽  
Epithelial Cells ◽  
Short Circuit ◽  
Basolateral Membrane ◽  
Potassium Conductance ◽  
Short Circuit Current ◽  
Sodium Absorption ◽  
High K ◽  
Esophageal Epithelial Cells ◽  
Apical Membranes

K+ conductance in apical and basolateral cell membranes of rabbit esophageal epithelial cells was investigated within intact epithelium by impalement with conventional microelectrodes from luminal or serosal sides. Under steady-state conditions, K+ conductance was demonstrated in basolateral, but not apical, membranes by showing 1) membrane depolarization upon exposure to either solutions high in K+ (20-65 mM) or containing Ba2+, tetraethylammonium, or quinine, and 2) a resistance ratio that increased on exposure to high K+ solution and decreased on exposure to Ba2+, quinine, and tetraethylammonium. From exposures to high K+, the apparent K+ transference number and electromotive force generated at the basolateral membrane were calculated and found to be 0.42 +/- 0.01 and -83 +/- 3 mV, respectively. Furthermore, basolateral K+ conductance was shown to be important for maintaining resting net transepithelial Na+ absorption in that high K+ or barium inhibited the transepithelial potential difference and short-circuit current of Ussing-chambered epithelia. We conclude that under steady-state conditions the basolateral, but not apical, membranes of esophageal epithelial cells contain a K(+)-conductive pathway and that this pathway is important for active sodium absorption.

Basal membrane uptake in potassium-secreting cells of midgut of tobacco hornworm (Manduca sexta)

1990 ◽  
Vol 258 (1) ◽  
pp. R112-R119
Author(s):  
A. C. Chao ◽  
A. R. Koch ◽  
D. F. Moffett
Keyword(s):  
Manduca Sexta ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Basal Membrane ◽  
Bathing Solution ◽  
K Uptake ◽  
Active Secretion ◽  
Uptake Process ◽  
Intracellular K Activity ◽  
K Activity

Basal membrane voltage (Vb), intracellular K+ activity [(K+)i], and short-circuit current (Isc) were measured in isolated posterior midguts of Manduca sexta wherein Isc is a measured of active secretion of K+ from blood into lumen. When bathed in 32 mM K+ and exposed to 100% O2, average values were Isc = 244 microAmp/cm2, Vb = -33.1 mV, and (K+)i = 88.6 mM. The electrochemical gradient across the basal membrane (d mu) averaged +5.8 mV (a gradient favorable for K+ entry). Exposure to 5% O2 led to a new steady state in which Isc = 71 microAmp/cm2, Vb = -18.7 mV, and (K+)i = 99.4 mM. During hypoxia, d mu averaged -9.9 mV (a gradient unfavorable for K+ entry). When the external bathing solution was 10 mM K+, comparable values were, for 100% O2, Isc = 139 microAmp/cm2, Vb = -56.1 mV, (K+)i = 72.2 mM, and d mu = +3.6 mV, and in 5% O2 the values were Isc = 28.3 microAmp/cm2, Vb = -43.7 mV, (K+)i = 76.1 mM, and d mu = -10.2 mV. The failure of cellular K+ to fall during prolonged hypoxia is evidence for a thermodynamically active basal K+ uptake process.

Changes in intracellular sodium with chloride secretion in dog tracheal epithelium

1986 ◽  
Vol 250 (4) ◽  
pp. C646-C650 ◽  
Author(s):  
S. R. Shorofsky ◽  
M. Field ◽  
H. A. Fozzard
Keyword(s):  
Steady State ◽  
Kinetic Models ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Tracheal Epithelium ◽  
Cl Secretion ◽  
Intracellular Na Activity ◽  
Sufficient Energy ◽  
Stimulation Of

Na-selective microelectrodes were employed to investigate the mechanism of Cl secretion by canine tracheal epithelium. In control tissues with a mean short-circuit current (Isc) of 30.1 microA/cm2, the intracellular Na activity (aiNa) was 10.7 mM. Following steady-state stimulation of Cl secretion with epinephrine (Isc = 126.4 microA/cm2), aiNa was 21.3 mM. These data indicate that there is sufficient energy in the Na gradient to drive Cl secretion by this tissue. When analyzed with simple kinetic models for the Na-K pump, they also suggest that the basolateral entry step involves the Na-K-2Cl cotransporter.

scholarly journals On the mechanism of the amiloride-sodium entry site interaction in anuran skin epithelia.

1979 ◽  
Vol 73 (3) ◽  
pp. 307-326 ◽  
Author(s):  
D J Benos ◽  
L J Mandel ◽  
R S Balaban
Keyword(s):  
Steady State ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Inhibitory Effect ◽  
Transport Kinetics ◽  
Entry Site ◽  
Interaction Sites ◽  
Noncompetitive Inhibitor ◽  
Kinetics Of ◽  
Entry Mechanism

The steady-state transport kinetics of the interaction between external sodium and the diuretic drug, amiloride, was studied in isolated anuran skin epithelia. We also investigated the effect of calcium on the amiloride-induced inhibition of short-circuit current (Isc) in these epithelial preparations. The major conclusions of this study are: (a) amiloride is a noncompetitive inhibitor of Na entry in bullfrog and grassfrog skin, but displays mixed inhibition in R. temporaria and the toad. A hypothesis which states that the interaction sites for amiloride and Na on the putative entry protein are spatially distinct in all of these species is proposed. (b) The stoichiometry of interaction between amiloride and the Na entry mechanism is not necessarily one-to-one. (c) The external Ca requirement for the inhibitory effect of amiloride is not absolute. Amiloride, at all concentrations, is equally effective in inhibiting Isc of bullfrog skin independently from the presence or absence of external Ca.

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