Mechanism of Inhibition of Active Potassium Transport in Isolated Midgut of Manduca Sexta by Bacillus Thurjngiensis Endotoxin

1979 ◽  
Vol 83 (1) ◽  
pp. 293-304
Author(s):  
WILLIAM R. HARVEY ◽  
MICHAEL G. WOLFERSBERGER
Keyword(s):  
Manduca Sexta ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Potassium Transport ◽  
Reverse Direction ◽  
Transport Parameters ◽  
Toxic Activity ◽  
Mechanism Of Inhibition ◽  
Manduca Sexta Larvae ◽  
Hydrolysis Of

After incubation at pH 10 or higher, Bacillus thuringiensis spores and endotoxin, at concentrations above 0.1 IU/ml, affected transport parameters in the isolated midgut of Manduca sexta larvae. (Toxic activity was lost during roughly 1 week at pH 11.) About 60% of the short-circuit current was in-hibited, and the remainder was reversibly inhibited by anoxia. Electrical resistance was reduced by about 55% and oxygen uptake stimulated by about 30%. Influx of potassium from blood-side to lumen-side (‘active’ flux) was unaffected but flux in the reverse direction was nearly tripled. These results suggest that hydrolysis of the toxin yields an inhibitor of potassium transport, presumably a polypeptide. It is argued that inhibition is not primarily by uncoupling of oxidative phosporylation, but instead by inter-ference with an active depression of the efflux of potassium from lumen-side to blood-side.

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.

Comparison of Potassium Transport in Three Structurally Distinct Regions of The Insect Midgut

1981 ◽  
Vol 91 (1) ◽  
pp. 103-116
Author(s):  
MOIRA CIOFFI ◽  
WILLIAM R. HARVEY
Keyword(s):  
Goblet Cell ◽  
Apical Membrane ◽  
Close Association ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Potassium Transport ◽  
Structural Requirement ◽  
Posterior Midgut ◽  
New Information ◽  
Structural And Functional Properties

1. Active potassium transport across the isolated midgut of the Tobacco Hornworm larva, Manduca sexta, was studied by measuring the short circuit current (ISC) and unidirectional 42-potassium fluxes. 2. The midgut is composed of structurally distinct anterior, middle and posterior regions, all of which are shown to transport potassium, so that by comparing and contrasting their structural and functional properties new information on the mechanism of midgut potassium transport was obtained. 3. It has previously been shown that the potassium pump is located on the apical membrane of the goblet cell. In the anterior and middle regions of the midgut the goblet cell has a large cavity and mitochondria are closely associated with the apical membrane while in the posterior midgut the goblet cavity is much smaller, and mitochondria are not associated with the apical membrane. However, the apical membrane particles which have been implicated in active potassium transport in a number of other insect epithelia are present in all three regions. This observation suggests that the particles are a structural requirement for active transport, and that close association between mitochondria and the transporting membrane is not essential. 4. Comparison of the kinetic influx pool size and the differences in the ISC decay profiles between the three midgut regions suggest that part of the influx pool is a transported pool located in the goblet cavity. 5. A new model to explain the driving force for potassium transport in the midgut is proposed, in which the rate of potassium transport controls the entrance of potassium into the cell, rather than the opposite, currently accepted view.

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.

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.

Active Transport by the Cecropia Midgut

1968 ◽  
Vol 48 (1) ◽  
pp. 1-12
Author(s):  
W. R. HARVEY ◽  
J. A. HASKELL ◽  
S. NEDERGAARD
Keyword(s):  
Transport Mechanism ◽  
Potassium Concentration ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Potassium Transport ◽  
Nernst Equation ◽  
Tissue Concentrations ◽  
Sodium Magnesium ◽  
Lines Of Evidence ◽  
Stimulation Of

1. From two lines of evidence, we conclude that the potassium transport gives rise directly to the midgut potential, i.e. that the active potassium transport mechanism is electrogenic. 2. First, diffusion potentials of neither potassium, sodium, magnesium, calcium, nor chloride could give rise to the large midgut potential if values for tissue concentrations are accepted for their respective activities in the epithelium. 3. Secondly, no externally added cation other than potassium is required to sustain either the potential or short circuit current, no specific anion is required, and no metabolic ion is known to be produced in sufficient amount to act as a counter ion for potassium in a non-electrogenic process. 4. Changes in the concentration of potassium on the blood-side of the midgut always lead to changes in potential in the direction predicted by the Nernst equation. Moreover, a tenfold change in potassium concentration leads to the expected 59 mV. potential change provided that the prior midgut potential is at least 130 mV. This effect could be attributed either to the stimulation of an electrogenic potassium pump or to a potassium diffusion potential across the blood-side barrier.

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.

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.

Potassium transport by rabbit descending colon

1982 ◽  
Vol 242 (1) ◽  
pp. C81-C86 ◽  
Author(s):  
R. McCabe ◽  
H. J. Cooke ◽  
L. P. Sullivan
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Potassium Transport ◽  
Paracellular Pathway ◽  
Potential Difference ◽  
Metabolic Energy ◽  
Sodium Absorption ◽  
Potassium Absorption ◽  
Potassium Secretion ◽  
Descending Colon

Unidirectional mucosal-to-serosal (Jm leads to s) and serosal-to-mucosal (Js leads to m) fluxes of potassium and sodium were determined simultaneously on paired sections of descending colon from the same rabbit under short-circuit conditions. In 13-16 pairs of tissues, net potassium secretion and sodium absorption averaged 0.49 +/- 0.08 and 4.0 +/- 0.8 mueq.cm-2.h-1, respectively. Short-circuit current (Jsc) averaged 3.7 +/- 0.4 mueq.cm-2.h-1 and was approximately equal to the algebraic sum of net potassium and sodium fluxes. Treatment of both sides of the colon with 10(-4) M ouabain reduced the Jsc and transmural potential difference to near zero. Ouabain abolished net potassium secretion by reducing JKs leads to m and abolished net sodium absorption by inhibiting JNam leads to s. In the presence of ouabain, net potassium absorption averaging 0.15 +/- 0.07 mueq.cm-2.h-1 (n = 11) was observed. In the presence of 10(-3) M 2,4-dinitrophenol, both net potassium and net sodium fluxes were abolished, primarily as a result of a reduction in JKs leads to m and JNam leads to s without altering JKm leads to s and JNas leads to m. These results suggest that the rabbit descending colon has the capacity to secrete and possibly to absorb potassium by active mechanisms requiring metabolic energy. Comparison of potassium and sodium fluxes suggest that the paracellular pathway in the rabbit colon is not potassium selective.

Glucose flux from dietary disaccharides: all sugars are not absorbed at equal rates

1991 ◽  
Vol 261 (5) ◽  
pp. G818-G822 ◽  
Author(s):  
L. A. Heitlinger ◽  
B. U. Li ◽  
R. D. Murray ◽  
H. J. McClung ◽  
H. R. Sloan ◽  
...  
Keyword(s):  
Glucose Uptake ◽  
Glucose Transporter ◽  
Short Circuit ◽  
Hydrolytic Enzyme ◽  
Short Circuit Current ◽  
Glucose Absorption ◽  
Glucose Flux ◽  
Free Glucose ◽  
Hydrolysis Of

Considerable discrepancies exist in the literature regarding the rates of glucose absorption from the common dietary disaccharides, lactose, maltose, and sucrose. This study compared the unidirectional flux of glucose derived from dietary disaccharides with that of their constituent monosaccharides in vitro. Lactose-stimulated short-circuit current (Isc) and mucosal-to-serosal flux (Jm----s) were lower than that of an equimolar glucose-galactose mixture and were phlorizin inhibitable. Maltose- and glucose-stimulated Isc were similar, but Jm----s of glucose derived from the hydrolysis of maltose was lower than that of free glucose. Sucrose-stimulated Isc and Jm----s were similar to that of an equimolar glucose-fructose mixture. Isc and Jm----s of glucose from both maltose and sucrose were phlorizin and acarbose inhibitable. We conclude that the rate of glucose uptake from disaccharides is less than or equal to that of free glucose and is dependent on the glucose source. We speculate that regulation of glucose uptake from disaccharides can occur at three sites: the hydrolytic enzyme, the glucose transporter, and the tight junctions.

Altered regulation of intestinal ion transport by enteric nerves in diabetic rats

1988 ◽  
Vol 254 (3) ◽  
pp. G444-G449 ◽  
Author(s):  
M. H. Perdue ◽  
J. S. Davison
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Cholinergic Receptor ◽  
Diabetic Rats ◽  
Muscarinic Agonist ◽  
Transport Parameters ◽  
Ileal Mucosa ◽  
Enteric Nerves ◽  
Muscarinic Cholinergic

We compared ion transport parameters in isolated ileal mucosa from diabetic rats (8 wk after streptozotocin injection) and littermate controls under basal conditions and in response to electrical transmural stimulation (TS). Stripped ileal mucosa (submucosal plexus intact) was mounted in Ussing flux chambers modified to include stimulating electrodes on opposite sides of the tissue. Under basal conditions unidirectional fluxes of Na+ and Cl- were decreased across mucosa from diabetic rats compared with controls, whereas net fluxes were not significantly different. TS caused a tetrodotoxin (TTX)-sensitive transient increase in short-circuit current (Isc) that was significantly less in tissue from diabetic than control rats. The muscarinic cholinergic receptor antagonist, atropine, significantly reduced the Isc response to TS in ileum from control but not diabetic rats. In addition, the noncholinergic component of the response was smaller. The muscarinic agonist, Urecholine chloride (bethanechol chloride), caused an increase in Isc that was unaffected by pretreatment with TTX and was the same in tissue from control and diabetic rats. Our results suggest that the intestinal abnormalities that occur in diabetes may include a defect in the regulation of ion transport by enteric nerves resulting in an abnormal ability to respond to luminal and other stimuli.

Sign in / Sign up

Export Citation Format

Share Document