Analysis of epithelial transport by measurement of K+, Cl- and pH gradients in extracellular unstirred layers: ion secretion and reabsorption by Malpighian tubules of Rhodnius prolixus

1997 ◽  
Vol 200 (11) ◽  
pp. 1627-1638 ◽  
Author(s):  
KA Collier ◽  
MJ O'Donnell
Keyword(s):  
Epithelial Transport ◽  
Malpighian Tubule ◽  
Fluid Secretion ◽  
Rhodnius Prolixus ◽  
Malpighian Tubules ◽  
Unstirred Layer ◽  
Ph Gradients ◽  
In Situ Preparation

Summary The pH and concentrations of K+ and Cl- in the unstirred layer (USL) associated with the basolateral surfaces of the upper and lower Malpighian tubules of Rhodnius prolixus were measured using extracellular ion-selective microelectrodes. When stimulated with 5-hydroxytryptamine (5-HT) in vitro, the upper Malpighian tubule secretes Na+, K+, Cl- and water at high rates; the lower Malpighian tubule reabsorbs K+ and Cl- but not water. Concentrations of K+ and Cl- in the unstirred layer of the lower Malpighian tubule ([K+]USL, [Cl-]USL) were greater than those in the bathing saline, consistent with the accumulation of K+ and Cl- in the USL during 5-HT-stimulated KCl reabsorption. [K+]USL exceeded [K+]Bath by as much as 5.3-fold. Calculations of K+ flux based on measurements of [K+]USL at various distances from the tubule surface agreed well with flux calculated from the rate of fluid secretion and the change in K+ concentration of the secreted fluid during passage through the lower tubule. Concentrations of K+ in the unstirred layer of the upper Malpighian tubule were reduced relative to those in the bathing saline, consistent with depletion of K+ from the USL during 5-HT-stimulated secretion of K+ from bath to lumen. Changes in [K+]USL during 5-HT-stimulated K+ secretion from single upper Malpighian tubule cells could be resolved. Although differences between [K+]USL and [K+]Bath were apparent for upper and lower tubules in an in situ preparation, they were reduced relative to the differences measured using isolated tubules. We suggest that convective mixing of the fluids around the tubules by contractions of the midgut and hindgut reduces, but does not eliminate, differences between [K+]USL and [K+]Bath in situ. The USL was slightly acidic relative to the bath in 5-HT-stimulated upper and lower tubules; contributions to USL acidification are discussed. The results also show that the techniques described in this paper can resolve rapid and localized changes in ion transport across different regions of Malpighian tubules in response to stimulants or inhibitors of specific membrane transporters.

Anti-diuresis in the blood-feeding insect Rhodnius prolixus Stål: the peptide CAP2b and cyclic GMP inhibit Malpighian tubule fluid secretion.

1997 ◽  
Vol 200 (17) ◽  
pp. 2363-2367 ◽  
Author(s):  
M C Quinlan ◽  
N J Tublitz ◽  
M J O'Donnell
Keyword(s):  
Cyclic Gmp ◽  
Physiological Role ◽  
Malpighian Tubule ◽  
Fluid Secretion ◽  
Blood Feeding ◽  
Rhodnius Prolixus ◽  
Malpighian Tubules ◽  
Tubule Fluid ◽  
Secretion Rates

Rhodnius prolixus eliminates NaCl-rich urine at high rates following its infrequent but massive blood meals. This diuresis involves stimulation of Malpighian tubule fluid secretion by diuretic hormones released in response to distention of the abdomen during feeding. The precipitous decline in urine flow that occurs several hours after feeding has been thought until now to result from a decline in diuretic hormone release. We suggest here that insect cardioacceleratory peptide 2b (CAP2b) and cyclic GMP are part of a novel mechanism of anti-diuresis. Secretion rates of 5-hydroxytryptamine-stimulated Malpighian tubules are reduced by low doses of CAP2b or cyclic GMP. Maximal secretion rates are restored by exposing tubules to 1 mmol l-1 cyclic AMP. Levels of cyclic GMP in isolated tubules increase in response to CAP2b, consistent with a role for cyclic GMP as an intracellular second messenger. Levels of cyclic GMP in tubules also increase as urine output rates decline in vivo, suggesting a physiological role for this nucleotide in the termination of diuresis.

Intracellular ion activities in Malpighian tubule cells ofRhodnius prolixus: evaluation of Na+-K+-2Cl-cotransport across the basolateral membrane

2002 ◽  
Vol 205 (11) ◽  
pp. 1645-1655 ◽  
Author(s):  
Juan P. Ianowski ◽  
Robert J. Christensen ◽  
Michael J. O'Donnell
Keyword(s):  
Basolateral Membrane ◽  
Malpighian Tubule ◽  
Fluid Secretion ◽  
Rhodnius Prolixus ◽  
Passive Movement ◽  
Malpighian Tubules ◽  
Intracellular Ion Activities ◽  
Ion Activities ◽  
Tubule Cells ◽  
Electrochemical Potentials

SUMMARYIntracellular ion activities (aion) and basolateral membrane potential (Vbl) were measured in Malpighian tubule cells of Rhodnius prolixus using double-barrelled ion-selective microelectrodes. In saline containing 103mmoll-1Na+, 6mmoll-1 K+ and 93mmoll-1Cl-, intracellular ion activities in unstimulated upper Malpighian tubules were 21, 86 and 32mmoll-1, respectively. In serotonin-stimulated tubules, aCl was unchanged, whereas aNa increased to 33mmoll-1 and aK declined to 71mmoll-1. Vbl was -59mV and -63mV for unstimulated and stimulated tubules, respectively. Calculated electrochemical potentials(Δμ/F) favour passive movement of Na+ into the cell and passive movement of Cl- out of the cell in both unstimulated and serotonin-stimulated tubules. Passive movement of K+ out of the cell is favoured in unstimulated tubules. In stimulated tubules, Δμ/F for K+ is close to 0 mV.The thermodynamic feasibilities of Na+-K+-2Cl-, Na+-Cl-and K+-Cl- cotransporters were evaluated by calculating the net electrochemical potential (Δμnet/F) for each transporter. Our results show that a Na+-K+-2Cl- or a Na+-Cl- cotransporter but not a K+-Cl- cotransporter would permit the movement of ions into the cell in stimulated tubules. The effects of Ba2+ and ouabain on Vbl and rates of fluid and ion secretion show that net entry of K+ through ion channels or the Na+/K+-ATPase can be ruled out in stimulated tubules. Maintenance of intracellular Cl- activity was dependent upon the presence of both Na+ and K+ in the bathing saline. Bumetanide reduced the fluxes of both Na+ and K+. Taken together, the results support the involvement of a basolateral Na+-K+-2Cl- cotransporter in serotonin-stimulated fluid secretion by Rhodnius prolixus Malpighian tubules.

Active transport of sodium by the malpighian tubules of the tsetse fly Glossian morsitans

1976 ◽  
Vol 64 (2) ◽  
pp. 357-368
Author(s):  
J. D. Gee
Keyword(s):  
Cell Function ◽  
Malpighian Tubule ◽  
Active Role ◽  
Fluid Secretion ◽  
Sodium Concentration ◽  
Tsetse Fly ◽  
Malpighian Tubules ◽  
Bathing Solution ◽  
Bathing Medium

Isolated Malpighian tubules of Glossina morsitans are able to transport sodium against its concentration gradient. Their rate of secretion is dependent on the sodium concentration of the bathing medium. Potassium must be present in the bathing solution for rapid secretion to be maintained, but it does not play an active role in fluid secretion. Lithium and ammonium ions are able to substitute partially for sodium, other monovalent cations cannot. Ouabain does not affect rapid secretion by Glossina tubules in vitro. Conclusions drawn from the results are incorporated into a model of Malpighian tubule cell function in this insect.

Measurements of osmotic permeability in the Malpighian tubules of an insect, Rhodnius prolixus Stål

1982 ◽  
Vol 216 (1204) ◽  
pp. 267-277 ◽  
Keyword(s):  
Ion Transport ◽  
Experimental Method ◽  
Malpighian Tubule ◽  
Rhodnius Prolixus ◽  
Malpighian Tubules ◽  
Unstirred Layer ◽  
Osmotic Permeability

(i) With use of a novel experimental method, an improved estimate has been obtained for the value of the osmotic permeability ( P os ) of Rhodnius Malpighian tubules, 4.3 x 10 -3 cm s -1 osmol -1 l. Corrections for unstirred layer effects permit a further 35% increase in the value of P os , to 5.8 x 10 -3 cm s -1 osmol -1 l. (ii) In the lower Malpighian tubule, P os decreases from 3.4 x 10 -3 cm s -1 osmol -1 l at the junction with the upper Malpighian tubule to 0.4 x 10 -3 cm s -1 osmol -1 l at the junction with the gut. In lower tubules stimulated with 5-HT, P os is reduced over most of their length by about 50% . It is suggested that reduced values of P os in the lower portions of the lower tubule prevent water movements accompanying 5-HT-stimulated KCl resorption over the same area of the tubule. (iii) Values of P os in the upper segment of the upper tubule are discussedin relation to proposed mechanisms for coupling water movements to ion transport during isosmotic secretion by the tubule.

scholarly journals Functional Plasticity of the gut and the Malpighian tubules underlies cold acclimation and mitigates cold-induced hyperkalemia in Drosophila melanogaster

2017 ◽  
Author(s):  
Gil Y. Yerushalmi ◽  
Lidiya Misyura ◽  
Heath A. MacMillan ◽  
Andrew Donini
Keyword(s):  
Water Balance ◽  
Cold Acclimation ◽  
Low Temperatures ◽  
Fluid Transport ◽  
Epithelial Transport ◽  
Ion Homeostasis ◽  
Malpighian Tubule ◽  
Adult Emergence ◽  
Fluid Secretion ◽  
Malpighian Tubules

AbstractAt low temperatures, Drosophila, like most insects, lose the ability to regulate ion and water balance across the gut epithelia, which can lead to a lethal increase of [K+] in the hemolymph (hyperkalemia). Cold-acclimation, the physiological response to low temperature exposure, can mitigate or entirely prevent these ion imbalances, but the physiological mechanisms that facilitate this process are not well understood. Here, we test whether plasticity in the ionoregulatory physiology of the gut and Malpighian tubules of Drosophila may aid in preserving ion homeostasis in the cold. Upon adult emergence, D. melanogaster females were subjected to seven days at warm (25°C) or cold (10°C) acclimation conditions. The cold acclimated flies had a lower critical thermal minimum (CTmin), recovered from chill coma more quickly, and better maintained hemolymph K+ balance in the cold. The improvements in chill tolerance coincided with increased Malpighian tubule fluid secretion and better maintenance of K+ secretion rates in the cold, as well as reduced rectal K+ reabsorption in cold-acclimated flies. To test whether modulation of ion-motive ATPases, the main drivers of epithelial transport in the alimentary canal, mediate these changes, we measured the activities of Na+-K+-ATPase and V-type H+-ATPase at the Malpighian tubules, midgut, and hindgut. Na+/K+-ATPase and V-type H+-ATPase activities were lower in the midgut and the Malpighian tubules of cold-acclimated flies, but unchanged in the hindgut of cold acclimated flies, and were not predictive of the observed alterations in K+ transport. Our results suggest that modification of Malpighian tubule and gut ion and water transport likely prevents cold-induced hyperkalemia in cold-acclimated flies and that this process is not directly related to the activities of the main drivers of ion transport in these organs, Na+/K+- and V-type H+-ATPases.Summary StatementAt low temperatures, insects lose the ability to regulate ion and water balance and can experience a lethal increase in hemolymph [K+]. Previous exposure to low temperatures can mitigate this effect and improve chill tolerance. Here, we show that plasticity of ion and fluid transport across the Malpighian tubule and rectal epithelia likely drive this response.

scholarly journals Role of an apical K,Cl cotransporter in urine formation by renal tubules of the yellow fever mosquito (Aedes aegypti)

2011 ◽  
Vol 301 (5) ◽  
pp. R1318-R1337 ◽  
Author(s):  
Peter M. Piermarini ◽  
Rebecca M. Hine ◽  
Matthew Schepel ◽  
Jeremy Miyauchi ◽  
Klaus W. Beyenbach
Keyword(s):  
Aedes Aegypti ◽  
Epithelial Transport ◽  
Input Resistance ◽  
Fluid Secretion ◽  
Malpighian Tubules ◽  
Cell Swelling ◽  
Renal Tubules ◽  
Transport Pathway ◽  
Principal Cells

The K,Cl cotransporters (KCCs) of the SLC12 superfamily play critical roles in the regulation of cell volume, concentrations of intracellular Cl−, and epithelial transport in vertebrate tissues. To date, the role(s) of KCCs in the renal functions of mosquitoes and other insects is less clear. In the present study, we sought molecular and functional evidence for the presence of a KCC in renal (Malpighian) tubules of the mosquito Aedes aegypti . Using RT-PCR on Aedes Malpighian tubules, we identified five alternatively spliced partial cDNAs that encode putative SLC12-like KCCs. The majority transcript is AeKCC1-A1; its full-length cDNA was cloned. After expression of the AeKCC1-A protein in Xenopus oocytes, the Cl−-dependent uptake of 86Rb+ is 1) activated by 1 mM N-ethylmaleimide and cell swelling, 2) blocked by 100 μM dihydroindenyloxyalkanoic acid (DIOA), and 3) dependent upon N-glycosylation of AeKCC1-A. In Aedes Malpighian tubules, AeKCC1 immunoreactivity localizes to the apical brush border of principal cells, which are the predominant cell type in the epithelium. In vitro physiological assays of Malpighian tubules show that peritubular DIOA (10 μM): 1) significantly reduces both the control and diuretic rates of transepithelial fluid secretion and 2) has negligible effects on the membrane voltage and input resistance of principal cells. Taken together, the above observations indicate the presence of a KCC in the apical membrane of principal cells where it participates in a major electroneutral transport pathway for the transepithelial secretion of fluid in this highly electrogenic epithelium.

scholarly journals 5-Hydroxytryptamine: a second diuretic hormone in Rhodnius prolixus

1991 ◽  
Vol 156 (1) ◽  
pp. 557-566 ◽  
Author(s):  
S. H. Maddrell ◽  
W. S. Herman ◽  
R. L. Mooney ◽  
J. A. Overton
Keyword(s):  
Salivary Glands ◽  
Malpighian Tubule ◽  
Peptide Hormone ◽  
Fluid Secretion ◽  
Rhodnius Prolixus ◽  
Malpighian Tubules ◽  
Biologically Active ◽  
Diuretic Hormone ◽  
Naturally Occurring ◽  
Tubule Fluid

Bioassays of 5-hydroxytryptamine (5-HT) in fifth-instar Rhodnius prolixus haemolymph using Calliphora salivary glands indicate that: (1) biologically active 5-HT is present, (2) in unfed animals there is not enough 5-HT to stimulate Malpighian tubule fluid secretion, and (3) there is enough 5-HT soon after the initiation of feeding to stimulate rapid tubule secretion. The 5-HT receptor antagonists ketanserin and spiperone reversibly and selectively inhibit 5-HT-induced fluid secretion, indicating the presence of specific 5-HT receptors on Rhodnius Malpighian tubules. The data provide evidence that 5-HT is a naturally occurring hormone acting with a previously described peptide hormone to regulate diuresis in this species.

scholarly journals Active transport of brilliant blue FCF across the Drosophila midgut and Malpighian tubule epithelia

2019 ◽  
Author(s):  
Dawson B.H. Livingston ◽  
Hirva Patel ◽  
Andrew Donini ◽  
Heath A. MacMillan
Keyword(s):  
Traumatic Injury ◽  
Malpighian Tubule ◽  
Malpighian Tubules ◽  
Brilliant Blue ◽  
Barrier Dysfunction ◽  
Concentration Gradients ◽  
Barrier Disruption ◽  
Brilliant Blue Fcf ◽  
Summary Statement

AbstractUnder conditions of stress, many animals suffer from epithelial barrier disruption that can cause molecules to leak down their concentration gradients, potentially causing a loss of organismal homeostasis, further injury or death. Drosophila is a common insect model, used to study barrier disruption related to aging, traumatic injury, or environmental stress. Net leak of a non-toxic dye (Brilliant blue FCF) from the gut lumen to the hemolymph is often used to identify barrier failure under these conditions, but Drosophila are capable of actively transporting structurally-similar compounds. Here, we examined whether cold stress (like other stresses) causes Brilliant blue FCF (BB-FCF) to appear in the hemolymph of flies fed the dye, and if so whether Drosophila are capable of clearing this dye from their body following chilling. Using in situ midgut leak and transport assays as well as Ramsay assays of Malpighian tubule transport, we tested whether these ionoregulatory epithelia can actively transport BB-FCF. In doing so, we found that the Drosophila midgut and Malpighian tubules can mobilize BB-FCF via an active transcellular pathway, suggesting that elevated concentrations of the dye in the hemolymph may occur from increased paracellular permeability, reduced transcellular clearance, or both.Summary StatementDrosophila are able to actively secrete Brilliant blue FCF, a commonly used marker of barrier dysfunction

Serotonin triggers cAMP and PKA-mediated intracellular calcium waves in Malpighian tubules of Rhodnius prolixus

2014 ◽  
Vol 307 (7) ◽  
pp. R828-R836 ◽  
Author(s):  
Paula Gioino ◽  
Brendan G. Murray ◽  
Juan P. Ianowski
Keyword(s):  
Second Messenger ◽  
Fluid Secretion ◽  
Rhodnius Prolixus ◽  
Malpighian Tubules ◽  
Insect Vector ◽  
Excess Water ◽  
Intracellular Ca ◽  
Single Meal ◽  
Hematophagous Insect

Rhodnius prolixus is a hematophagous insect vector of Chagas disease capable of ingesting up to 10 times its unfed body weight in blood in a single meal. The excess water and ions ingested with the meal are expelled through a rapid postprandial diuresis driven by the Malpighian tubules. Diuresis is triggered by at least two diuretic hormones, a CRF-related peptide and serotonin, which were traditionally believed to trigger cAMP as an intracellular second messenger. Recently, calcium has been suggested to act as a second messenger in serotonin-stimulated Malpighian tubules. Thus, we tested the role of calcium in serotonin-stimulated Malpighian tubules from R. prolixus. Our results show that serotonin triggers cAMP-mediated intracellular Ca2+ waves that were blocked by incubation in Ca2+-free saline containing the cell membrane-permeant Ca2+ chelator BAPTA-AM, or the PKA blocker H-89. Treatment with 8-Br-cAMP triggered Ca2+ waves that were blocked by H-89 and BAPTA-AM. Analysis of the secreted fluid in BAPTA-AM-treated tubules showed a 75% reduction in fluid secretion rate with increased K+ concentration, reduced Na+ concentration. Taken together, the results indicate that serotonin triggers cAMP and PKA-mediated Ca2+ waves that are required for maximal ion transport rate.

scholarly journals Action of activated 27,000 Mr toxin from Bacillus thuringiensis var. israelensis on Malpighian tubules of the insect, Rhodnius prolixus

1989 ◽  
Vol 94 (3) ◽  
pp. 601-608
Author(s):  
S.H. Maddrell ◽  
J.A. Overton ◽  
D.J. Ellar ◽  
B.H. Knowles
Keyword(s):  
Bacillus Thuringiensis ◽  
Cell Membrane ◽  
Basal Cell ◽  
Time Course ◽  
Fluid Secretion ◽  
Rhodnius Prolixus ◽  
Malpighian Tubules ◽  
Bathing Solution ◽  
Concentrated Solutions ◽  
Intracellular Milieu

The action of activated 27,000 Mr toxin from Bacillus thuringiensis var. israelensis (Bti toxin) on Malpighian tubules of Rhodnius prolixus has been investigated. Its binding to the tubules is slowed by low temperature but is not prevented even at 0 degree C. The binding is less effective at pH 10 than at pH7. Pretreatment of the tubules with 0.1 mmol l-1 ouabain or bumetanide or 1 mumol l-1 5-hydroxytryptamine did not affect the toxicity of the toxin. The toxin causes very large changes in the trans-epithelial potential difference; it changes from 40 mV, lumen negative, often to more than 100 mV, lumen positive. This reflects an initial collapse of the potential of the basal cell membrane, followed by a large positive-going potential change at the luminal cell membrane. Just prior to the effects of the toxin on rapid fluid secretion, the basal cell membrane becomes permeable to sucrose molecules. Raffinose at 170 mmol l-1 in the bathing solution does not protect the tubules from Bti toxin action but dextran, Mr5000, at 60 mmol l-1 significantly delayed failure of fluid secretion and, even more, the onset of staining of the tubule cells with Trypan Blue. Exposing tubules to saline that is calcium-free and/or magnesium-free, or has a composition adjusted to be similar to that of the intracellular milieu, does not affect the time course of failure of fluid secretion induced by the toxin. There is no evidence that effective aggregates of Bti toxin molecules are formed in concentrated solutions.(ABSTRACT TRUNCATED AT 250 WORDS)

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