Inhibition Of Fluid Secretion Research Articles

Piperine as potential therapy of post-weaning porcine diarrheas: an in vitro study using a porcine duodenal enteroid model

Post-weaning diarrhea in piglets is a major problem, resulting in a significant loss in pig production. This study aimed to investigate the effects of piperine, an alkaloid abundantly found in black peppers, on biological activities related to the pathogenesis of post-weaning diarrhea using a porcine duodenal enteroid model, a newly established intestinal stem cell-derived in vitro model recapitulating physiology of porcine small intestinal epithelia. Porcine duodenal enteroid models were treated with disease-relevant pathological inducers with or without piperine (8 μg/mL and/or 20 μg/mL) before measurements of oxidative stress, mRNA, and protein expression of proinflammatory cytokines, nuclear factor-kappa B (NF-κB) nuclear translocation, barrier leakage, and fluid secretion. We found that piperine (20 μg/mL) inhibited H2O2-induced oxidative stress, TNF-α-induced mRNA, and protein expression of proinflammatory cytokines without affecting NF-κB nuclear translocation, and prevented TNF-α-induced barrier leakage in porcine duodenal enteroid monolayers. Importantly, piperine inhibited fluid secretion induced by both forskolin and heat-stable toxins (STa) in a three-dimensional model of porcine duodenal enteroids. Collectively, piperine possesses both anti-inflammatory and anti-secretory effects in porcine enteroid models. Further research and development of piperine may provide novel interventions for the treatment of post-weaning porcine diarrhea.

CAPA Neuropeptides Inhibit VA Activity in Malpighian Tubules of the Mosquito, Aedes Aegypti

Abstract Haematophagus insects, such as the female Aedes aegypti mosquito, face the challenge of excess ion and water intake after engorgement on a blood meal. To cope with this, adult female A. aegypti have a specialized excretory system that includes the Malpighian tubules (MTs), which are under rigorous control by several neuroendocrine factors to regulate transepithelial movement of ions/water. Produced in the CNS, the mosquito anti-diuretic hormone is a member of the CAPA peptide family, which share homology to the vertebrate neuromedin U peptide. CAPA peptides inhibit fluid secretion of MTs stimulated by select diuretic factors, 5HT and DH31 through the NOS/cGMP/PKG pathway. However, the anti-diuretic signalling mechanism and downstream cellular targets, such as ion channels and transporters, remain unclear. To study whether the V-type H+-ATPase (VA) plays a role in CAPA inhibition, we performed fluid secretion assays in MTs treated with diuretics and bafilomycin, a known VA inhibitor. Bafilomycin significantly inhibited DH31-stimulated fluid secretion 30 min post treatment compared to diuretic controls (p<0.05, n=22). Similar, however, delayed responses were seen in 5HT-stimulated MTs, while no affect was observed in DH44-stimulated secretion. An indirect way to measure whether CAPA inhibits VA activity was to measure the pH of the secreted fluid from diuretic-stimulated MTs treated with CAPA. In DH31-stimulated MTs supplemented with CAPA, there was an immediate significantly higher pH at 40 min, increasing up to 7.73±0.038 compared to control, 7.56±0.038 (p=0.0007, n=20). The pH of 5HT-stimulated MTs treated with CAPA was seen to significantly increase up to 7.75±0.061 (p=0.03, n=10) at the 60 min mark, in agreement with the delayed response previously seen. Unlike the effects observed with DH31 and 5HT, CAPA did not alter the pH of the secreted fluid in DH44-stimulated MTs. Alkalization of the secreted fluid in response to CAPA suggests inhibition of the proton pump, which may lead to constrained cation entry across the apical membrane of the MTs. To understand Na+/K+-ATPase (NKA) and VA activity in response to CAPA, we performed activity assays in diuretic-stimulated MTs. Adult female MTs treated with DH31 resulted in an increase of both NKA and VA activity compared saline controls. As expected, MTs incubated with both DH31 and AedaeCAPA-1 had a lower NKA and VA activity (p<0.05) resulting in activity levels comparable to saline control levels. The results thus far could suggest a novel mechanism for CAPA inhibition, blocking the VA to hinder fluid secretion. Investigating the pathway of CAPA inhibition and its role in countering diuresis will help provide a deeper understanding of the critical process of diuresis and its signaling mechanism.

The role of the V‐type H + ‐ATPase in CAPA‐treated Malpighian tubules in the yellow fever mosquito, Aedes aegypti

Haematophagus insects, such as the female Aedes aegypti mosquito, face the challenge of excess ion and water intake after engorgement on a blood meal. To cope with this, adult female A. aegypti utilize their highly adapted excretory system that includes the Malpighian ‘renal’ tubules (MTs), which are under rigorous control by several hormones to regulate transepithelial movement of ions/water. Originating in the nervous system, the mosquito anti-diuretic hormone is a member of the CAPA peptide family and is a homolog of the vertebrate neuromedin U peptide. CAPA peptides inhibit fluid secretion of MTs stimulated by select diuretic factors, 5HT and DH31 through the NOS/cGMP/PKG pathway. However, the anti-diuretic signalling mechanism and downstream cellular targets, such as ion channels and transporters, remain unclear. Active ion transport in A. aegypti MTs is driven by the principal cells, whose brush-border membrane is densely packed by the V-type H+-ATPase (VA), serving as the primary energizer for transepithelial secretion of electrolytes and water. Due to its predominant role in fluid secretion, the VA is a likely target for diuretic and anti-diuretic factors. The effect of endocrine factors and the VA inhibitor, bafilomycin, on fluid secretion rate by MTs was measured using an in vitro bioassay. Bafilomycin was found to inhibit fluid secretion stimulated by serotonin (5HT) and the calcitonin-related diuretic hormone (DH31), while having no inhibitory action on MTs stimulated with the corticotropin-releasing factor (CRF)-related diuretic hormone. CAPA and bafilomycin treatment led to alkalization of the secreted fluid suggesting inhibition of the apical VA, which may lead to constrained entry of cations (Na+ and K+) across the apical membrane of MTs. VA activity was increased in adult female MTs treated with DH31, whereas CAPA treatment reduced VA activity, comparable to saline levels. Furthermore, a VA antibody was used to determine protein localization and expression in DH31 and CAPA-treated MTs. In DH31-incubated MTs, the VA was exclusively localized to the apical memrane of the MTs, whereas VA immunoreactivity was observed in both the apical membrane and cytosol of CAPA-treated tubules. These results suggest a novel mechanism for CAPA inhibition, blocking the VA to hinder fluid secretion. A. aegypti mosquitoes are vectors of a variety of pathogens leading to illnesses, such as Dengue fever, Yellow fever, and the Zika virus. The current results advance our knowledge of the anti-diuretic control of the A. aegypti MTs, suggesting that CAPA targets the VA to inhibit diuresis. Investigating the pathway of CAPA inhibition and its role in countering diuresis will help provide a deeper understanding of this critical physiological process. Studying the excretory system and discerning the underlying mechanisms behind ion and fluid transport may aid in developing novel pest management strategies to better control Aedes population and lessen its prominence as a disease vector.

Do Second Messengers of Neuroendocrine Factors Alter Aquaporin Expression in Malpighian tubules of the Mosquito, Aedes aegypti?

The mosquito Aedes aegypti is considered one of the most dangerous disease vectors because it can transmit several arboviral diseases including Dengue, Chikungunya, Zika virus, and yellow fever. The larvae are aquatic typically inhabiting hypotonic freshwater whereas the adults are terrestrial. Hence, the larvae are usually faced with the osmoregulatory challenge of eliminating excess water from body fluids while adults usually conserve water, that is, with the exception of females when they take a blood meal, at which time they too need to eliminate excess water. Osmoregulation is accomplished, in part by regulating water flux across epithelia of organs such as the midgut, Malpighian tubules (MTs), hindgut and, in the case of larvae, the anal papillae. The MTs with the hindgut are functionally analogous to the renal tubes of mammalian kidneys where urine is produced; however, the mechanisms of urine production are different. In MTs, urine production is accomplished by ion secretion driven by an apical V‐type H+‐ATPase and water is obliged to follow by osmosis. There is a plethora of literature which describes the regulation of MTs fluid secretion in insects but these studies mostly describe effects on ion transport. Little is known about the regulation of water transport across the epithelium of the MTs. The mosquito, A. aegypti, possesses six aquaporin (water transporter) genes designated AaAQP1 through 6, which are expressed in the MTs and transcript abundance is affected by blood feeding in adult females. Furthermore, AaAQP5 mediates water flux across larval MTs whereas AaAQP1 is expressed in tracheolar cells of adult MTs. In this study we have begun to examine if and how known second messengers of neuroendocrine factors influence AaAQP protein expression in the MTs of A. aegypti with western blots. Thus far we have found that in MTs of larvae incubated with cAMP, AaAQP1 protein expression is higher than unstimulated controls but AaAQP4 levels are unaffected while both AaAQP1 and AaAQP5 expression is not affected by cGMP also in comparison to unstimulated controls. In larvae and adult mosquitoes, cAMP is a second messenger of diuretic hormones that stimulate urine production whereas cGMP is the second messenger of anti‐diuretic hormones, including CAPA peptides, which inhibit fluid secretion at physiological doses. In heterologous expression systems, AaAQP1 and AaAQP5 transport water whereas AaAQP4 is an entomoglyceroporin which transports solutes like trehalose preferentially over water. We conclude that stimulatory hormones of MTs facilitate fluid secretion at least in part by increasing the basal expression of AaAQP1 and that inhibitory hormones may not affect basal AaAQP expression; however, effects of these factors on stimulated MTs and on the expression of the other AaAQP genes is the subject of continuing work.Support or Funding InformationNSERC‐Discovery RGPIN‐2018‐05841

Elucidating the Anti‐Diuretic Hormone Signalling System of CAPA Neuropeptides in the Disease Vector Mosquito, Aedes aegypti

Blood‐feeding insects, including female Aedes aegypti mosquitoes, face the challenge of excess water and ion intake after a blood meal. To cope with this, insects have a highly active excretory system that includes the Malpighian ‘renal’ tubules (MTs), which are under rigorous control by several neuroendocrine factors. While endocrine regulation has been extensively studied in insects, most research has focused on the effects of diuretic hormones with limited study on anti‐diuretic hormones. Insect CAPA neuropeptides are produced in the central nervous system and are evolutionarily related to the vertebrate neuromedinU peptides. While CAPA peptides elicit distinct responses in different insect species, their activity on stimulated MTs in the adult A. aegypti remained unknown. Given the CAPA receptor is highly enriched in insect MTs, the objectives of this study were to examine the effects of a CAPA peptide on adult female MTs stimulated with various diuretic factors.The effect of endocrine factors on fluid secretion rate was measured using an in vitro bioassay where CAPA was found to inhibit secretion of MTs stimulated by select diuretic factors, 5‐HT and DH31, while having no effect on other diuretic factors, including kinin‐related and corticotropin releasing factor‐related (CRF) peptides. Although CAPA elicits strong anti‐diuretic activity, it does not influence the relative proportions of cations transported by adult MTs, thus maintaining the kaliuretic activity of 5‐HT and the natriuretic activity of DH31. Notably, the inhibitory effect of CAPA was abolished by knocking down the receptor using RNAi, verifying its critical role in anti‐diuresis. To investigate the signalling cascade mediating this anti‐diuretic effect, the second messenger cGMP was tested on adult MTs revealing that both 5‐HT and DH31–stimulated secretion is strongly inhibited by cGMP, similar to effects seen with CAPA. Pharmacological inhibition of PKG/NOS signalling abolishes the anti‐diuretic activity of CAPA, confirming the role of cGMP/PKG/NOS in the CAPA signalling pathway. Furthermore, MTs treated with bafilomycin, a V‐type H+‐ATPase inhibitor, was found to inhibit fluid secretion stimulated by 5‐HT and DH31, while having no inhibitory action on CRF‐stimulated MTs. Additionally, alkalization of the secreted fluid in response to CAPA suggests inhibition of the apical V‐type H+‐ATPase, which may lead to constrained entry of cations (Na+and K+) across the apical membrane of the MTs.A. aegypti mosquitoes are vectors of a variety of pathogens leading to diseases including Dengue fever and Zika virus. In this study, the importance of hormonal regulation of the MTs is demonstrated, analyzing the importance of both diuretic and anti‐diuretic hormones. Given the central importance of the MTs in insect biology, these insights will be useful considering the need to develop novel strategies or compounds which more specifically and efficiently reduce the burden of insect disease vectors.Support or Funding InformationSupported by an NSERC Discovery Grant (JPP), Petro Canada Young Innovator Award (JPP), Ontario Ministry of Research & Innovation Early Researcher Award (JPP), NSERC Canada Graduate Scholarship (FS), Vernon Oliver Stong Graduate Scholarship in Science (FS), and The Carswell Scholarships in the Faculty of Science (FS)

Anti-diuretic action of a CAPA neuropeptide against a subset of diuretic hormones in the disease vector Aedes aegypti.

The mosquito Aedes aegypti is a vector responsible for transmitting various pathogens to humans, and their prominence as chief vectors of human disease is largely due to their anthropophilic blood feeding behaviour. Larval stage mosquitoes must deal with the potential dilution of their haemolymph in freshwater, whereas the haematophagus A. aegypti female faces the challenge of excess ion and water intake after a blood meal. The excretory system, composed of the Malpighian tubules (MTs) and hindgut, is strictly controlled by neuroendocrine factors, responsible for the regulation of diuresis across all developmental stages. The highly studied insect MTs are influenced by a variety of diuretic hormones and, in some insects, anti-diuretic factors. In the present study, we investigated the effects of AedaeCAPA-1 neuropeptide on larval and adult female A. aegypti MTs stimulated with various diuretic factors including serotonin (5-HT), a corticotropin-related factor (CRF) diuretic peptide, a calcitonin-related diuretic hormone (DH31) and a kinin-related diuretic peptide. Overall, our findings establish that AedaeCAPA-1 specifically inhibits secretion of larval and adult MTs stimulated by 5-HT and DH31, whilst having no activity on MTs stimulated by other diuretic factors. Furthermore, although AedaeCAPA-1 acts as an anti-diuretic, it does not influence the relative proportions of cations transported by adult MTs, thus maintaining the kaliuretic activity of 5-HT and natriuretic activity of DH31 In addition, we tested the effects of the second messenger cGMP in adult MTs. We established that cGMP has similar effects to AedaeCAPA-1, strongly inhibiting 5-HT- and DH31-stimulated fluid secretion, but with only minor effects on CRF-stimulated diuresis. Interestingly, although AedaeCAPA-1 has no inhibitory activity on kinin-stimulated fluid secretion, cGMP strongly inhibited fluid secretion by this diuretic hormone, which targets stellate cells specifically. Collectively, these results support that AedaeCAPA-1 inhibits select diuretic factors acting on the principal cells and this probably involves cGMP as a second messenger. Kinin-stimulated diuresis, which targets stellate cells, is also inhibited by cGMP, suggesting that another anti-diuretic factor in addition to AedaeCAPA-1 exists and may utilize cGMP as a second messenger.

Caffeine Inhibits Fluid Secretion by Interlobular Ducts From Guinea Pig Pancreas.

Caffeine is contained in coffee, tea, and numerous beverages and foods. We examined the direct effects of caffeine on the physiological function of pancreatic duct cells by using interlobular duct segments isolated from guinea pig pancreas. The rate of fluid secretion was continuously measured by monitoring the luminal volume of isolated duct segments. Changes in intracellular Ca concentration ([Ca]i) were estimated by microfluorometry in ducts loaded with Fura-2. Both secretin-stimulated and acetylcholine (ACh)-stimulated fluid secretions were substantially and reversibly inhibited by relatively low concentrations of caffeine as low as 0.03 mM relevant to blood levels after ingestion of caffeine-containing beverages. Caffeine inhibited ACh-induced elevation of [Ca]i and secretin-induced fluctuation of [Ca]i. Caffeine abolished thapsigargin-induced intracellular Ca release but did not affect the entry of extracellular Ca. Caffeine (0.05 mM) abolished ethanol (1 mM)-induced fluid hypersecretion in secretin-stimulated pancreatic duct. Low concentrations of caffeine directly inhibit pancreatic ductal fluid secretion stimulated by secretin or ACh and also ethanol-induced fluid hypersecretion. The inhibition by caffeine seems to be mediated by the blockade of intracellular Ca mobilization. Daily intake of caffeine may reduce the volume of pancreatic juice secretion.

Effects of Beverage Ingredients on Salivary Fluid Secretion with an <i>ex Vivo</i> Submandibular Gland Perfusion System: Tannic Acid as a Key Component for the Inhibition of Saliva Secretion

Tannic acid (TA) and TA containing beverage have been proved to inhibit Ca2+-activated Cl- channel located apical membrane of the secretory cells. However, their effect on salivary fluid secretion is not well investigated. We used mouse ex Vivo submandibular gland perfusion technique to identify the general effect of TA and related beverage samples on muscarinic agonist carbachol induced fluid secretion. Green tea inhibited fluid secretion by 64% from the control, where oolong tea was by 53%, and red wine by 43% which was linked with their TA concentration. On the other hand, though TA was contained at 4.7 μM in white wine sample and 33 μM in coffee extract, no adverse effect was observed. In addition, coffee induced salivation in the absence of carbachol. TA had a negative effect on fluid secretion with a concentration dependent manner. The effects of TA on carbachol induced calcium increase showed identical as fluid secretion, which was initially no effect, and then gradually decreased over the time. These results demonstrate that TA directly inhibits the salivary fluid secretion and it affects not only Ca2+-activated Cl- channel but also intracellular Ca2+ increasing mechanisms.

Cigarette smoke extract inhibits fluid secretion and CFTR activity in guinea pig pancreatic ductal cells

s / Pancreatology 14 (2014) S1eS129 S79 Results: Sequencing analysis of the discovery cohort revealed four common mutations: intronic mutations c.23+71_23+103del, c.183-4C>A and c.1134+32C>A; and exonic missense mutation p.V206M. . These four mutations were found in linkage disequilibrium indicating a conserved haplotype. We found this haplotype in 18 heterozygous and 2 homozygous cases, and in 24 heterozygous and 2 homozygous controls (allele frequency 11.4% and 14.1% respectively). A synonymous mutation p.P397P was also detected in a single case. Conclusion:We found a novel, commonhaplotype in the SLC26A6 gene, which did not show association with CP. Supported by T AMOP and OTKA.

Roles of PKC and phospho‑adducin in transepithelial fluid secretion by Malpighian tubules of the yellow fever mosquito

The diuretic hormone aedeskinin‑III is known to increase the paracellular Cl- conductance in Malpighian (renal) tubules of the mosquito Aedes aegypti via a G protein-coupled receptor. The increase serves the blood-meal-initiated diuresis and is associated with elevated levels of Ca2+ and phosphorylated adducin in the cytosol of tubule. In the present study we have cloned adducin in Aedes Malpighian tubules and investigated its physiological roles. Immunolabeling experiments are consistent with the association of adducin with the cortical cytoskeleton, especially near the apical brush border of the tubule. An antibody against phosphorylated adducin revealed the transient phosphorylation of adducin 2 min after stimulating tubules with aedeskinin‑III. The PKC inhibitor bisindolylmaleimide‑I blocked the phosphorylation of adducin as well as the electrophysiological and diuretic effects of aedeskinin‑III. Bisindolylmaleimide‑I also inhibited fluid secretion in control tubules. Phorbol 12‑myristate 13‑acetate increased phosphorylated adducin levels in Malpighian tubules, but it inhibited fluid secretion. Thus, the phosphorylation of adducin by PKC alone is insufficient to trigger diuretic rates of fluid secretion; elevated levels of intracellular Ca2+ may also be required. The above results suggest that the phosphorylation of adducin, which is known to destabilize the cytoskeleton, may (1) facilitate the traffic of transporters into the apical brush border supporting diuretic rates of cation secretion and (2) destabilize proteins in the septate junction thereby enabling paracellular anion (Cl‑) secretion at diuretic rates. Moreover, PKC and the phosphorylation of adducin play a central role in control and diuretic tubules, consistent with the dynamic behavior of both transcellular and paracellular transport pathways.

AedesCAPA-PVK-1 displays diuretic and dose dependent antidiuretic potential in the larval mosquito Aedes aegypti (Liverpool)

This study reveals that AedesCAPA-PVK-1 (GPTVGLFAFPRV-NH2) inhibits basal and serotonin stimulated fluid secretion in the Malpighian tubules of larval Aedes aegypti at femtomolar concentrations. Conversely 10−4moll−1 of the peptide stimulated fluid secretion rates. The diuretic effects of 10−4moll−1AedesCAPA-PVK-1 and antidiuretic effects of 10−15moll−1AedesCAPA-PVK-1 were abolished by protein kinase A (PKA) and protein kinase G (PKG) inhibition, respectively. Similar to the peptide, 10−3moll−1 cGMP stimulated fluid secretion but doses in the micromolar to nanomolar range inhibited fluid secretion of the Malpighian tubules. Stimulatory effects of cGMP were abolished by PKA inhibition and inhibitory effects of cGMP were abolished by PKG inhibition. Furthermore, the nitric oxide synthase inhibitor l-NAME attenuated the inhibitory effects of AedesCAPA-PVK-1 but did not affect inhibition by cGMP. Based on the results we propose that AedesCAPA-PVK-1 inhibits fluid secretion rates of larval Malpighian tubules via the NOS/cGMP/PKG pathway and that high doses of the peptide lead to diuresis through the cGMP mediated activation of PKA.

EFFECTS OF THE MICROBIAL METABOLITE DESTRUXIN A ON ION TRANSPORT BY THE GUT AND RENAL EPITHELIA OFDrosophila melanogaster

Destruxins have been implicated in the infection process by entomopathogenic fungi and have been also found to be highly toxic when applied topically or ingested by different insect species. To gain insight into the mechanism of action of this toxin on insect internal organs, we have evaluated the effects of destruxin A on Drosophila melanogaster Malpighian tubules and gut tissues. Destruxin A was toxic when injected into adults; the calculated EC(50) was 0.11 mM. Destruxin A significantly inhibited fluid secretion rate by Malpighian tubules as well; the calculated IC(50) was 0.25 μM. The Na(+) concentration in the secreted fluid increased significantly when tubules were exposed to 0.25 μM destruxin A, whereas pH and the concentrations of Ca(2+) and K(+) did not change. In gut, there was no effect of destruxin on H(+) flux, but there was a significant decrease in K(+) and Ca(2+) absorption. The concentration of Ca(2+) and K(+) in the hemolymph of destruxin A-injected flies was not significantly different from those of control flies after 3 h. Taken together, these results show that destruxin A produces differential effects on ion transport by renal and gut tissues. © 2012 Wiley Periodicals, Inc.

Translational Approaches for Pharmacotherapy Development for Acute Diarrhea

Areport is presented of a National Institute of Diabetes and Digestive and Kidney Diseases–sponsored single topic conference designed to determine scientific advances needed to encourage development of pharmacotherapy for diarrheal diseases. Acute diarrheal diseases are a global public health problem in developing and developed countries. They are a leading cause of mortality and morbidity for children under the age of 5 worldwide and have a major economic impact in developed countries. Although oral rehydration solution (ORS) along with oral zinc therapy has reduced mortality owing to acute diarrheal diseases, there are essentially no other approved, safe, and effective drugs to decrease stool volume and prevent fluid loss. On September 25–27, 2011, the National Institute of Diabetes and Digestive and Kidney Diseases convened a workshop on Translational Approaches for Pharmacotherapy Development for Acute Diarrhea to assess current treatments for acute diarrhea, to identify common host pathways targeted by infectious agents associated with acute diarrhea, to evaluate which pathophysiologic mechanisms are most clinically relevant as potential targets, and to identify promising areas for translational research.

Short-term effects of tolvaptan on renal function and volume in patients with autosomal dominant polycystic kidney disease

Tolvaptan and related V(2)-specific vasopressin receptor antagonists have been shown to delay disease progression in animal models of polycystic kidney disease. Slight elevations in serum creatinine, rapidly reversible after drug cessation, have been found in clinical trials involving tolvaptan. Here, we sought to clarify the potential renal mechanisms to see whether the antagonist effects were dependent on underlying renal function in 20 patients with autosomal dominant polycystic kidney disease (ADPKD) before and after 1 week of daily split-dose treatment. Tolvaptan induced aquaresis (excretion of solute-free water) and a significant reduction in glomerular filtration rate (GFR). The serum uric acid increased because of a decreased uric acid clearance, and the serum potassium fell, but there was no significant change in renal blood flow as measured by para-aminohippurate clearance or magnetic resonance imaging (MRI). No correlation was found between baseline GFR, measured by iothalmate clearance, and percent change in GFR induced by tolvaptan. Blinded post hoc analysis of renal MRIs showed that tolvaptan significantly reduced total kidney volume by 3.1% and individual cyst volume by 1.6%. Preliminary analysis of this small cohort suggested that these effects were more noticeable in patients with preserved renal function and with larger cysts. No correlation was found between changes of total kidney volume and body weight or estimated body water. Thus, functional and structural effects of tolvaptan on patients with ADPKD are likely due to inhibition of V(2)-driven adenosine cyclic 3',5'-monophosphate generation and its aquaretic, hemodynamic, and anti-secretory actions.

The effect of loperamide oxide on prostaglandin-stimulated fluid transport in rat small intestine

The effects of loperamide and loperamide oxide on basal and prostaglandin E2-stimulated fluid transport by rat small intestine have been investigated. In contrast to loperamide, loperamide oxide, when applied intraperitoneally, failed to inhibit either basal or prostaglandin E2-stimulated fluid transport. However, intraperitoneal administration of loperamide oxide following its incubation with the contents of the intestinal lumen under aerobic conditions resulted in an effective inhibition of fluid secretion. The activating material was present in the essentially non-particulate 3000 g supernatant fraction of the luminal contents and was heat-stable.

Effects of the cyclopeptide mycotoxin destruxin A on the Malpighian tubules of Rhodnius prolixus (Stål)

The production of peptide toxins by entomopathogenic fungi during the infection process plays critical roles in pathogenesis. To gain insight into the mechanism of action of these mycotoxins on insect internal organs, we have evaluated the effects of destruxin A, a cyclic peptide produced by Metarhizium anispliae, on Rhodnius prolixus Malpighian tubules measuring fluid secretion rate, transepithelial electrical potential (TEP), pH and ion composition of secreted fluid, and ATP content. Destruxin A dramatically inhibited fluid secretion rate on tubules stimulated by 5-Hydroxytryptamine (5-HT) or cAMP. The calculated IC 50 for destruxin A on 5-HT-stimulated tubules was 3 × 10 −7 M. Fluid secretion rate by Malpighian tubules exposed for 20 min to 10 −6 M destruxin A recovered completely when tubules were washed with saline; however, when tubules were exposed to 5 × 10 −6 M destruxin A the fluid secretion rate only partially recovered upon wash off. The use of Ca 2+-free saline or addition of the calcium channel blocker CoCl 2 to the bathing saline did not interfere with the effects of destruxin A, and neither did the modification of intracellular calcium by TMB-8. Measurement of TEP of tubules challenged with 5-HT after preincubation for 10 min in saline containing 10 −6 M destruxin A showed that the second and third phases of the typical triphasic response to 5-HT were disrupted. Likewise, the positive shift in TEP in response to 5-HT in chloride-free bathing saline was significantly reduced when tubules were preincubated for 10 min in 10 −6 M destruxin A. The pH of the secreted fluid, but not the Na + or K + concentration, increased significantly when 5-HT-stimulated tubules were exposed to 10 −6 M destruxin A. The ATP content was not significantly different when tubules stimulated with 5-HT were exposed to destruxin A. Taken together, these results show that destruxin A, without interfering with the intracellular ATP production, strongly inhibits fluid secretion rate by the Malpighian tubules of R. prolixus. Changes in properties of the TEP suggest that one of the target sites for this peptide toxin might be associated with inhibition of the apical V-type H + ATPase of tubule cells.

High glucose inhibits HCO 3 − and fluid secretion in rat pancreatic ducts

Cellular mechanisms underlying the impairment of pancreatic fluid and electrolyte secretion in diabetes were examined using interlobular ducts isolated from rat pancreas. Fluid secretion was assessed by monitoring changes in luminal volume. HCO3(-) uptake across the basolateral membrane was estimated from the recovery of intracellular pH following an acid load. Exposure to high glucose concentrations inhibited fluid secretion and reduced the rate of basolateral HCO3(-) uptake in secretin-stimulated ducts isolated from normal rats. In ducts isolated from streptozotocin-treated diabetic rats, fluid secretion and basolateral HCO3(-) uptake were also severely impaired but could be largely reversed by incubation in normal-glucose solutions. Sodium-dependent glucose cotransporter 1 (SGLT1), glucose transporter (GLUT)1, GLUT2, and GLUT8 transcripts were detected by reverse transcriptase polymerase chain reaction in isolated ducts. Raising the luminal glucose concentration in microperfused ducts caused a depolarization of the membrane potential, consistent with the presence of SGLT1 at the apical membrane. Unstimulated ducts filled with high-glucose solutions lost luminal fluid by a phlorizin-sensitive mechanism, indicating that pancreatic ducts are capable of active glucose reabsorption from the lumen via SGLT1. In ducts exposed to high glucose concentrations, continuous glucose diffusion to the lumen and active reabsorption via SGLT1 would lead to elevation of intracellular Na+ concentration and sustained depolarization of the apical membrane. These two factors would tend to inhibit the basolateral uptake and apical efflux of Cl(-) and HCO3(-) and could therefore account for the impaired fluid and electrolyte secretion that is observed in diabetes.

CRTH2 Is Not Involved in the Anti-Enteropooling Effect of PGD2 in the Small Intestine

The majority of prostaglandins (PGs) are known to induce intestinal fluid secretion (enteropooling). In contrast, PGD₂ has been demonstrated to inhibit fluid secretion induced by other PGs. This study was aimed to investigate, by the use of selective agonists/antagonists, which type of PGD₂ receptor mediates this inhibitory effect. The DP1 agonist BW245C dose-dependently inhibited the enteropooling effect of 16,16-dimethyl-PGE₂. This inhibition was counteracted by the DP1 antagonist BWA868C. In contrast, the CRTH2 receptor does not seem to be involved in the anti-enteropooling effect of PGD₂, since the selective agonists 13,14-dihydro-15-keto-PGD₂ and 15(R)-15-methyl-PGD₂ were without effect. Therefore, our results suggest that the inhibitory effect of PGD₂ in the small intestine is mediated via activation of the DP1 receptor.

Beetle diuretic peptides: The response of mealworm ( Tenebrio molitor) Malpighian tubules to synthetic peptides, and cross-reactivity studies with a dung beetle ( Onthophagus gazella)

This paper reports the effects of different diuretic factors on the Malpighian tubules of beetles. Calcitonin (CT)-like peptides from silkmoth and mosquito increase fluid secretion in a dose-dependent manner in the tubules of Tenebrio molitor, but the cockroach CT-like peptide, Dippu-DH 31, has no effect. Thapsigargin induces a small but significant increase in tubule secretion rates. The interactions between different factors in mealworm tubules were explored by testing CT-like peptides, thapsigargin and the mealworm CRF-related diuretic factor Tenmo-DH 37 in various combinations, but no synergistic effects were observed. C-terminal fragments of the CRF-related diuretic peptides Locmi-DH 46 and Dippu-DH 46 fail to increase fluid secretion in mealworm tubules, unlike their corresponding whole peptides. Cross-reactivity of factors between beetle species was investigated using the scarabaeid Onthophagus gazella. Tenmo-DH 37 increases fluid secretion in isolated tubules of O. gazella in a dose-dependent manner, revealing a high degree of cross-reactivity in this distantly related beetle species. However, homogenates of O. gazella brains inhibited fluid secretion in mealworm tubules.

Evaluation of the antidiarrhoeal activity of Byrsocarpus coccineus

Based on its use in traditional African medicine, the antidiarrhoeal activity of the aqueous leaf extract of Byrsocarpus coccineus, Connaraceae, was evaluated on normal and castor oil-induced intestinal transit, castor oil-induced diarrhoea, enteropooling and gastric emptying. The extract (50, 100, 200 and 400 mg/kg, p.o.) produced a significant ( P < 0.05) dose dependent decrease in propulsion in the castor oil-induced intestinal transit in mice. The mean peristaltic index (%) for these doses of extract, control (distilled water; 10 ml/kg, p.o.) and morphine (10 mg/kg, s.c.) were 55.27 ± 1.86, 53.12 ± 3.73, 38.60 ± 3.79, 30.25 ± 1.27, 89.33 ± 5.62 and 20.29 ± 3.38, respectively. The effect of the extract at the highest dose was significantly ( P < 0.05) lower than that of the standard drug. This effect was antagonised by yohimbine (1 mg/kg, s.c.) but not by isosorbide dinitrate (IDN, 150 mg/kg, p.o.). At 200 mg/kg, the extract produced a significant decrease in propulsion in normal intestinal transit. In a dose dependent manner, it delayed the onset of diarrhoea, produced a significant decrease in the frequency of defaecation, severity of diarrhoea and protected the mice treated with castor oil. Mean diarrhoea scores were 30.83 ± 1.72, 22.40 ± 1.71, 21.43 ± 1.32, 13.80 ± 0.33, 18.00 ± 3.94 and 7.67 ± 2.41 for control, extract (50, 100, 200 and 400 mg/kg) and morphine, respectively. This effect was not antagonized by IDN. The extract (400 mg/kg) significantly decreased the volume (ml) of intestinal fluid secretion induced by castor oil (0.60 ± 0.23) compared with 1.27 ± 0.12 for control. However, there was no significant effect on gastric emptying. The results obtained suggest that Byrsocarpus coccineus possesses antidiarrhoeal activity due to its inhibitory effect on gastrointestinal propulsion, mediated through α 2 adrenoceptors, and also inhibition of fluid secretion. Preliminary phytochemical analysis revealed the presence of alkaloids, tannins, saponins, reducing sugars, glycosides and anthraquinones.