Effect of somatostatin on electrogenic ion transport in the duodenum and colon of the mouse, Mus domesticus

In this study, we have used the mouse intestine and the Ussing short circuit technique to compare the effects and mechanism of action of somatostatin (SST, 0.1 μM) on cAMP- and Ca²⁺-mediated ion secretion in the duodenum and colon of the Swiss-Webster mouse. The cAMP-dependent secretagogues, prostaglandin E₂ (1 μM) and dibutyryl-cAMP (150 μM) increased short circuit current (I_sc) in both regions, but only the colonic response was inhibited by SST. This inhibition was independent of enteric nerves, suggesting a direct action on the epithelial cells. The Ca²⁺-dependent secretagogue carbachol (10 μM) stimulated a transient increase in I_sc in both intestinal segments. In the duodenum, SST partially inhibited this increase in I_sc and both the responses to carbachol and SST were independent of enteric nerves. In the colon, while SST inhibited the carbachol induced increase in I_sc, pre-treatment with tetrodotoxin (750 nM) profoundly inhibited the carbachol induced increase in I_sc, thus markedly reducing the inhibitory effect of SST. This indicates an involvement of the enteric nervous system in the response to carbachol and the action of SST in the colon. These data indicate marked regional differences within the mouse intestine of the effects of SST on ion secretion and demonstrate different mechanisms of action of SST in the duodenum and colon.     

Ca-sensitive sodium absorption in the colon of Xenopus laevis

Summary Transepithelial electrogenic Na transport (I_Na) was investigated in the colon of the frog Xenopus laevis with electrophysiological methods in vitro. The short circuit current (I_sc) of the voltage-clamped tissue was 24.2±1.8 A·cm^-2 (n=10). About 60% of this current was generated by electrogenic Na transport. Removal of Ca²⁺ from the mucosal Ringer solution stimulated I_Na by about 120%. I_Na was not blockable by amiloride (0.1 mmol·l^-1), a specific Na-channel blocker in epithelia, but a fully and reversible inhibition was achieved by mucosal application of 1 mmol·l^-1 lanthanum (La^3-). No Na-self-inhibition was found, because I_Na increased linearly with the mucosal Na concentration. A stimulation of I_Na by antidiuretic hormones was not possible. The analysis of fluctuations in the short circuit current (noise analysis) indicated that Na ions pass the apical cell membrane via a Ca-sensitive ion channel. The results clearly demonstrate that in the colon of Xenopus laevis Na ions are absorbed through Ca-sensitive apical ion channels. They differ considerably in their properties and regulation from the amiloride-sensitive Na channel which is typically found in the colon of vertebrates.Abbreviations G _T transepithelial conductance - I _sc short circuit current - I _Na transepithelial Na-current - m mucosal - s serosal - PDS power density spectrum - f frequency - f _c corner frequency of the Lorentzian component of the PDS - S(f) power density of the Lorentzian component of the PDS - S_o plateau value of the Lorentzian component of the PDS     

Na and Cl transport and short-circuit current in rabbit ileum

Summary Na and Cl fluxes and short-circuit current (I _sc) in rabbit ileum have been studied as a function of ionic concentrations in HCO₃-free solutions. Both net Na flux (J _net ^Na ) andI _sc show similar saturation functions of [Na] at fixed [Cl]. They show no significant difference between zero and 112mm Na but at 140mm NaI _sc is significantly greater than theJ _net ^Na . Net Cl transport, secretion, is observed only at 140mm Na and is approximately equivalent to the difference between theI _sc andJ _net ^Na . The transcellular mucosa-to-serosa Na fluxes measured at 140 and 70mm Na do not differ significantly from the correspondingI _sc. The net Cl flux varies with [Cl] at fixed [Na] whileI _sc is virtually not affected by [Cl]. These results suggest that the absorptive Na transport process is electrogenic and responsible for theI _sc and that the secretory fluxes of Na and Cl are coupled, require high [Na], vary with [Cl], and do not contribute toI _sc. K-free solution abolishes theI _sc after a prolonged lag. Finally, the effect of a low resistance shunt pathway on active Na absorption is examined with a four-compartment model.Deceased (October 16, 1974).     

Trimethyltin chloride induced chloride secretion across rat distal colon

TMT (trimethyltin chloride), an organotin, is ubiquitous in the environment. The consumption of contaminated food may cause exposure of the human diet to this toxic compound. The present study was to investigate the effects of TMT on the regulation of ion transport across the rat distal colon. The rat colonic mucosa was mounted in Ussing chambers. The effects of TMT were assessed using the I_sc (short‐circuit current). Both apical and basolateral TMT induced, dose‐dependently, an increase in I_sc, which was due to a stimulation of Cl^? secretion as measured using ion substitution experiments and pharmacological manoeuvres. The secretion was also inhibited by several K⁺ channel blockers administrated at the basolateral side. When the apical side was permeabilized by nystatin, the TMT‐induced K⁺ conductance was effectively blocked by tetrapentylammonium, a Ca²⁺‐sensitive K⁺ channel blocker. The response of TMT was sensitive to the basolateral Ca²⁺ and the intracellular Ca²⁺ store, which could be disclosed by applying the inhibitors of ryanodine receptors and inositol 1,4,5‐trisphosphate receptors. In conclusion, TMT led to Cl^? secretion, which was essentially regulated by basolateral Ca²⁺‐sensitive K⁺ channels. These results suggest the importance of K⁺ channels in the toxicity hazard of TMT.     

Different modes of electrogenic Na+ absorption in the coprodeum of the chicken embryo: role of extracellular Ca2+

Summary Transepithelial electrogenic Na⁺ transport (I_Na) was investigated in the coprodeum of 20-days-old chicken embryos in Ussing chambers. Short circuit current (I_sc) and transepithelial resistance (R_t) were 14.7±4.8 A · cm^-2 (n=12) and 0.53±0.09 k · cm^-2 (n=12), respectively. I_Na was calculated from changes in I_sc by substitution of mucosal Na⁺ by (N-methyl-d-glucamine) (NMDG). I_sc inversed during Na⁺ removal, and I_Na was found to be 27.8±4.7 A · cm^-2 (n=12). Amiloride (100 mol · l^-1) inhibited only about 60% of I_Na. Analysis of I_sc fluctuations revealed a Lorentzian component in the power density spectrum with a corner frequency of about 57 Hz. This component was not correlated to I_Na, and its origin is still unclear. Removal of mucosal Ca²⁺ increased I_Na about 2.5-fold due to an increase of the amiloride-insensitive component of I_Na in additionally investigated adult tissues. The results clearly show that this is due to a non-selective cation channel with an apparent order of selectivity Cs⁺>Na⁺=K⁺>Rb⁺>Li⁺. The Ca²⁺ concentration required to block 50% of the I_sc was about 18 mol · l^-1. The I _sc ^Ca could also be supressed by other divalent cations such as Mg²⁺ and Ba²⁺. Additionally, an I_Na-linked Lorentzian component occurred which dominated the control spectrum with a significantly higher corner frequency (about 88 Hz). The results indicate that Na⁺ absorption in the coprodeum of the chicken embryo is more complex than in adult hens. However, the Ca²⁺ sensitivity of I_Na is similar to comparable effects described for other epithelia. This possibly reflects the existence of two types of amiloride-insensitive apical cation channels as pathways for Na⁺ absorption, which may be involved to differing degrees in ontogenetic developments of nonselective channels to Na⁺-specific ion channels.Abbreviations DPL direct-linear-plot method - slope of the back-ground noise component - EGTA ethylene glycol-bi(2-amino-ethylether)-N,N,N,N-tetraacetic acid - f frequency - f _c corner frequency of the Lorentzian noise component - G _t transepithelial conductance - HEPES N-hydroxyethylpiperazine-N-ethanesulfonic acid - I _sc short-circuit current - I _Na transepithelial sodium current - I _sc ^Ca Ca²⁺-sensitive short-circuit current - K _m ^Ca Michaelis-Menten constant for Ca²⁺ - K _B power density of the background noise component at f=1Hz - m mucosal - NMDG N-methyl-D-glucamine - R _t transepithelial resistance - s serosal - SEM standard error of mean - S(f) power density of the Lorentzian noise component - S _o plateau value of the Lorentzian noise component     

Acetate transport by locust rectum in vitro

Everted rectal sacs of Schistocerca gregaria absorb ¹⁴C-acetate from the lumen side at high rates against large electrical and often small concentration differences. Most of the ¹⁴C-activity in the absorbed fluid remains as acetate, but small amounts serve as substrate for aerobic respiration within this tissue. When acetate is substituted for SO₄^?2 or Cl^? in external salines, both short-circuit current (I_sc) and the open-circuit transepithelial potential (PD) increase by as much as 2- to 3-fold. The stimulatory effect of acetate on I_sc and PD exhibits saturation kinetics. The ‘steady-state’ influx of ¹⁴C-acetate from lumen (L) to haemocoel (H) side greatly exceeds efflux (haemocoel to lumen) across short-circuited recta. Over the whole range of acetate concentrations tested, the resulting net flux of acetate is sufficient to explain all of the increase in I_sc caused by this organic anion. Acetate was detected in moderate concentrations in body fluids of locusts. The possible significance of acetate transport in vivo is discussed.     

Chloride-dependent transmural potential in the rectal wall of Schistocerca gregaria

Rectum transmural potential (PD) and short-circuit current (I_sc) of the desert locust, Schistocerca gregaria, have been studied in vitro, with everted rectal wall preparations in solutions of different ionic composition. Initially, a PD of about 35 mV (lumen positive) and a I_sc of about 300 μA cm^?2 were recorded. Omission of sodium or potassium (Tris as substitute), from the luminal side or from both sides led to an increase of 4 to 6 mV in PD (lumen more positive) together with an increase in I_sc. In the absence of chloride alone (sulphate as substitute) the PD quickly dropped to nearly zero. In each case the control values were recovered on replacing the corresponding ions. Neither the PD nor the I_sc changed when substitutions affected only the haemocoelic solution. These findings corroborate the assumption that active transport of chloride ions from lumen to haemolymph is the major factor for transmural PD and account for the short-circuit current in the rectal wall of desert locust. A working scheme is given to explain the influence of sodium, potassium, and chloride ions on the PD.     

Ion transport by rabbit colon: II. Unidirectional sodium influx and the effects of amphotericin B and amiloride

Summary The unidirectional influx of Na from the mucosal solution into the epithelium ofin vitro descending rabbit colon (J _me ^Na ) determined under short-circuit conditions, is comprised of two components: one represents entry of Na into transporting epithelial cells and is abolished by amiloride which also abolishes Na absorption (J _net ^Na ). The other represents diffusional Na entry into paracellular pathways traversing the epithelium. In all instances, exposure of the mucosal surface to amphotericin B increased tissue conductance andJ _me ^Na and elicited K secretion. Tissues showing a spontaneousI _sc of approximately 4 eq/cm²hr did not respond to amphotericin B with increasedI _sc andJ _net ^Na . However, in tissues characterized by a lowerI _sc under control conditions, amphotericin B increasedI _sc andJ _net ^Na to approximately 4eq/cm²hr. These findings suggest that amphotericin increasesJ _net ^Na and elicits K secretion by disrupting the normal permselectivity of the mucosal membrane. Under these conditions the extrusion of Na from cell-to-serosal solution becomes the rate limiting step in transepithelial Na transport. Finally, a close correlation betweenJ _me ^Na andJ _net ^Na was observed when the rate of Na absorption varied either spontaneously or experimentally with amiloride, suggesting that the backflux of Na from cell-to-mucosal solution is undetectably small.     

Modulation of Calcium-dependent Chloride Secretion by Basolateral SK4-like Channels in a Human Bronchial Cell Line

The human bronchial cell line16HBE14o– was used as a model of airway epithelial cells to study the Ca²⁺-dependent Cl^– secretion and the identity of K_Ca channels involved in the generation of a favorable driving force for Cl^– exit. After ionomycin application, a calcium-activated short-circuit current (I _sc) developed, presenting a transient peak followed by a plateau phase. Both phases were inhibited to different degrees by NFA, glybenclamide and NPPB but DIDS was only effective on the peak phase. ⁸⁶Rb effluxes through both apical and basolateral membranes were stimulated by calcium, blocked by charybdotoxin, clotrimazole and TPA. 1-EBIO, a SK-channel opener, stimulated ⁸⁶Rb effluxes. Block of basolateral K_Ca channels resulted in I _sc inhibition but, while reduced, I _sc was still observed if mucosal Cl^– was lowered. Among SK family members, only SK4 and SK1 mRNAs were detected by RT-PCR. KCNQ1 mRNAs were also identified, but involvement of K_cAMP channels in Cl^– secretion was unlikely, since cAMP application had no effect on ⁸⁶Rb effluxes. Moreover, chromanol 293B or clofilium, specific inhibitors of KCNQ1 channels, had no effect on cAMP-dependent I _sc. In conclusion, two distinct components of Cl^– secretion were identified by a pharmacological approach after a Ca _i ²⁺ rise. K_Ca channels presenting the pharmacology of SK4 channels are present on both apical and basolateral membranes, but it is the basolateral SK4-like channels that play a major role in calcium-dependent chloride secretion in 16HBE14o– cells.     

Aldosterone regulation of active sodium chloride transport in the lizard colon (Gallotia galloti)

Summary Bioelectrical parameters and unidirectional sodium and chloride fluxes were measured under voltageclamp conditions in groups of lizards submitted to single or chronic aldosterone treatment. Both acute (AT) and chronic (CT) treatment induced significant increases in the short-circuit current (I _sc), as well as in the mucosa-to-serosa (J _m-s ^Na ) and net sodium flux (J _net ^Na ). In AT tissues, aldosterone did not change net chloride flux (J _net ^Cl ) but did so in CT tissues. Amiloride reduced the aldosterone-increased I _sc in AT and CT tissues, inhibited J _net ^Na in AT tissues and abolished it in CT colons. J _net ^Cl was also reduced by the diuretic in the group of AT colons, whereas no changes were observed in the CT tissues. Addition of luminal DIDS reduced Na⁺ absorption and totally inhibited Cl^- absorption in the AT tissues, but did not change I _sc. However, in CT tissues neither Na⁺ nor Cl^- transport were affected by DIDS. A good relationship between I _sc and J _m-s ^Na was apparent after DIDS treatment in AT tissues. In this group, simultaneous addition of DIDS and amiloride totally abolished J _net ^Na and reduced I _sc to untreated control values. Addition of serosal ouabain abolished I _sc and Na⁺ absorption in AT and CT colons, but Cl^- absorption was only altered in AT tissues. These results support the hypothesis that aldosterone induces an electrogenic, amiloride-sensitive sodium absorption, and in a dose-dependent fashion suppresses electroneutral NaCl absorption in the lizard colon.Abbreviations AT acutely treated - CT chronically treated animals - DIDS 4-4-diisothiocyanatostibene-2-2-disulfonic acid - DMSO dimethylsulphoxide - G _t tissue conductance - I _sc short circuit current - PD transepithelial potential difference - SITS 4-acetamido-4-isothiocyanatostilbene-2-2-disulfonic acid - UC untreated controls Preliminary results of this paper were presented at the X ^th meeting of the European Intestinal Transport Group (EITG), Askov Hojskole, Denmark, 16–19 September 1990     

Relationship of transmural electrical parameters to the luminal Na concentration in the colon of the fowl (Gallus domesticus)

Summary Electrical parameters: PD, resistance,I _sc and amiloride-sensitive-I _sc across the fowl colon (in vitro) change in response to the Na content of the diet. On a low-Na diet these changes appear to reflect increases in ion transport, especially amiloride-inhibitable Na transport. In vitro the magnitudes of the changes are related to the Na concentration in the luminal (mucosal) fluid and in birds on a low-Na diet peak at a concentration of about 12.5 mM. Such Na concentrations are similar to those in the colonic fluid of Na-deprived birds. Typical Michaelis-Menten kinetics do not appear to apply, possibly reflecting a local adaptation of the ion transport process in response to its external Na concentration.     

Effect of Adenosine on Na+ and Cl− Currents in A6 Monolayers. Receptor Localization and Messenger Involvement

The effect of adenosine regulation on sodium and chloride transport was examined in cultured A₆ renal epithelial cells. Adenosine and its analogue N⁶-cyclopentyladenosine (CPA) had different effects on short-circuit current (I _sc) depending on the side of addition. Basolateral CPA addition induced an approximately threefold increase of the I _sc that reached a maximum effect 20 min after addition and was completely inhibited by preincubation with either an A₂ selective antagonist, CSC, or the sodium channel blocker, amiloride. Apical CPA addition induced a biphasic I _sc response characterized by a rapid fourfold transient increase over its baseline followed by a decline and a plateau phase that were amiloride insensitive. The A₁ adenosine antagonist, CPX, completely prevented this response. This I _sc response to apical CPA was also strongly reduced in Cl⁻-free media and was significantly inhibited either by basolateral bumetanide or apical DPC preincubation. Only basolateral CPA addition was able to induce an increase in cAMP level. CPA, added to cells in suspension, caused a rapid rise in [Ca²⁺] _i that was antagonized by CPX, not affected by CSC and prevented by thapsigargin preincubation. These data suggest that basolateral CPA regulates active sodium transport via A₂ adenosine receptors stimulating adenylate cyclase while apical CPA regulates Cl⁻ secretion via A₁ receptor-mediated changes in [Ca²⁺] _i .     

Capsaicin inhibits intestinal Cl- secretion and promotes Na+ absorption by blocking TRPV4 channels in healthy and colitic mice

Although capsaicin has been studied extensively as an activator of the transient receptor potential vanilloid cation channel subtype 1 (TRPV1) channels in sensory neurons, little is known about its TRPV1-independent actions in gastrointestinal health and disease. Here, we aimed to investigate the pharmacological actions of capsaicin as a food additive and medication on intestinal ion transporters in mouse models of ulcerative colitis (UC). The short-circuit current (I_sc) of the intestine from WT, TRPV1-, and TRPV4-KO mice were measured in Ussing chambers, and Ca²⁺ imaging was performed on small intestinal epithelial cells. We also performed Western blots, immunohistochemistry, and immunofluorescence on intestinal epithelial cells and on intestinal tissues following UC induction with dextran sodium sulfate. We found that capsaicin did not affect basal intestinal I_sc but significantly inhibited carbachol- and caffeine-induced intestinal I_sc in WT mice. Capsaicin similarly inhibited the intestinal I_sc in TRPV1 KO mice, but this inhibition was absent in TRPV4 KO mice. We also determined that Ca²⁺ influx via TRPV4 was required for cholinergic signaling–mediated intestinal anion secretion, which was inhibited by capsaicin. Moreover, the glucose-induced jejunal I_scvia Na⁺/glucose cotransporter was suppressed by TRPV4 activation, which could be relieved by capsaicin. Capsaicin also stimulated ouabain- and amiloride-sensitive colonic I_sc. Finally, we found that dietary capsaicin ameliorated the UC phenotype, suppressed hyperaction of TRPV4 channels, and rescued the reduced ouabain- and amiloride-sensitive I_sc. We therefore conclude that capsaicin inhibits intestinal Cl^- secretion and promotes Na⁺ absorption predominantly by blocking TRPV4 channels to exert its beneficial anti-colitic action.     

Correlation Between Transepithelial Na+ Transport and Transepithelial Water Movement Across Isolated Frog Skin (Rana esculenta)

In the present work the coupling under short-circuited conditions between the net Na⁺-influx across isolated frog skin and the transepithelial transport of water was examined i.e., the short-circuit current (I _sc ) and the transepithelial water movement (TEWM) were measured simultaneously. It has been shown repeatedly that the I _sc across isolated frog skin is equal to the net transepithelial Na⁺ transport. Furthermore the coupling between transepithelial uptake of NaCl under open-circuit conditions and TEWM was also measured. The addition of antidiuretic hormone (AVT) to skins incubated under short-circuited conditions resulted in an increase in the I _sc and TEWM. Under control conditions I _sc was 9.14 ± 2.43 and in the presence of AVT 45.9 ± 7.3 neq cm⁻² min⁻¹ (n= 9) and TEWM changed from 12.45 ± 4.46 to 132.8 ± 15.8 nL cm⁻² min⁻¹. The addition of the Na⁺ channel blocking agent amiloride resulted in a reduction both in I _sc and TEWM, and a linear correlation between I _sc and TEWM was found. The correlation corresponds to that 160 ± 15 (n= 7) molecules of water follow each Na⁺ across the skin. In another series of experiments it was found that there was a linear correlation between I _sc and the increase in apical osmolarity needed to stop the TEWM. The data presented indicate that the observed coupling between the net transepithelial Na⁺ transport and TEWM is caused by local osmosis. Received: 16 October 1996/Revised: 6 March 1997     

KCl transport across and insect epithelium: I. Tracer fluxes and the effects of ion substitutions

Summary Models for active Cl transport across epithelia are often assumed to be universal although they are based on detailed studies of a relatively small number of epithelia from vertebrate animals. Epithelial Cl transport is also important in many invertebrates, but little is known regarding its cellular mechanisms. We used short-circuit current, tracer fluxes and ion substitutions to investigate the basic properties of Cl absorption by locust hindgut, an epithelium which is ideally suited for transport studies. Serosal addition of 1mm adenosine 35-cyclic monophosphate (cAMP), a known stimulant of Cl transport in this tissue, increased short-circuit current (I _sc) and net reabsorptive³⁶Cl flux (J _net ^Cl ) by 1000%. Cl absorption did not exhibit an exchange diffusion component and was highly selective over all anions tested except Br. Several predictions of Na- and HCO₃-coupled models for Cl transport were tested: Cl-dependentI _sc was not affected by sodium removal (<0.05mm) during the first 75 min. Also, a large stimulation ofJ _net ^Cl was elicited by cAMP when recta were bathed for 6 hr in nominally Na-free saline (<0.001 to 0.2mm) and there was no correlation between Cl transport rate and the presence of micromolar quantities of Na contamination. Increased unidirectional influx of³⁶Cl into rectal tissue during cAMP-stimulation was not accompanied by a comparable uptake of²²Na.J _net ^Cl was independent of exogenous CO₂ and HCO₃, but was strongly dependent on the presence of K. These results suggest that the major fraction of Cl transport across this insect epithelium occurs by an unusual K-dependent mechanism that does not directly require Na or HCO₃.     

Expression of an Artificial Cl<Superscript>−</Superscript> Channel in Microperfused Renal Proximal Tubules

To better understand the process of fluid movement driven by Cl^– conductance, a Cl^– channel-forming peptide was delivered to the luminal membrane of microperfused rabbit renal proximal tubules. When the peptide (NK₄-M2GlyR) was perfused, a significant new conductance was observed within 3 min and stabilized at 10 min. Alteration of the ion composition revealed it to be a Cl^–-specific conductance. Reabsorption of Cl^– (J _Cl) was increased by NK₄-M2GlyR, but not by a scramble NK₄-M2GlyR sequence, suggesting that the active peptide formed de novo Cl^– channels in the luminal membrane of the perfused tubules. In the presence of the peptide, reabsorption of fluid (J _v) was dramatically increased and J _Na and J _Ca were concomitantly increased. We propose that introduction of the new Cl^– conductance in the luminal membrane leads to a coordinated efflux of water across the membrane and an increase in cation translocation via the paracellular pathway, resulting in an increase in J _v. This novel method could prove useful in characterizing mechanisms of fluid transport driven by Cl^– gradients.     

House dust mite extract activates apical Cl− channels through protease‐activated receptor 2 in human airway epithelia

Adequate fluid secretion from airway mucosa is essential for maintaining mucociliary clearance, and fluid hypersecretion is a prominent feature of inflammatory airway diseases such as allergic rhinitis. House dust mite extract (HDM) has been reported to activate protease‐activated receptors (PARs), which play various roles in airway epithelia. However, the role of HDM in regulating ion transporters and fluid secretion has not been investigated. We examined the effect of HDM on ion transport in human primary nasal epithelial cells. The Ca²⁺‐sensitive dye Fura2‐AM was used to determine intracellular Ca²⁺ concentration ([Ca²⁺]_i) by means of spectrofluorometry in human normal nasal epithelial cells (NHNE). Short‐circuit current (Isc) was measured using Ussing chambers. Fluid secretion from porcine airway mucosa was observed by optical measurement. HDM extract (10 µg/Ml) effectively cleaved the PAR‐2 peptide and induced an increase of [Ca²⁺]_i that was abolished by desensitization with trypsin, but not with thrombin. Apical application of HDM‐induced Isc sensitive to both a cystic fibrosis transmembrane conductance regulator (CFTR) inhibitor and a Ca²⁺‐activated Cl^? channel (CaCC) inhibitor. HDM extract also stimulated fluid secretion from porcine airway mucosa. HDM extract activated PAR‐2 and apical Cl^? secretion via CaCC and CFTR, and HDM‐induced fluid secretion in porcine airway mucosa. Our results suggest a role for PAR‐2 in mucociliary clearance and fluid hypersecretion of airway mucosa in response to air‐borne allergens such as HDM. J. Cell. Biochem. 109: 1254–1263, 2010. © 2010 Wiley‐Liss, Inc.     

Effects of aldosterone on Na transport in the toad bladder II. The anaerobic response

The action of aldosterone on active Na⁺ transport was assessed under aerobic and anaerobic conditions in the isolated urinary bladder of the toad, BUfo marinus. Aldesterone augmented the short-circuit current (I_sc) under rigorous anaerobiosis. Four lines of evidence indicate that the increase in anaerobic I_sc does not represent an equivalent increase in active Na⁺ transport: 1. Net Na⁺ transport, determined by isotopic fluxes, was the same in the aldosterone-treated and control quarter-bladders, and significantly greater than the simultaneously measured I_sc. 2. Amiloride, an inhibitor of the apiral entry of Na⁺, did not reduce the steroid-dependent increase in the anaerobic I_sc. 3. Substitution of choline for Na⁺ in the mucosal medium reduced the magnitude of the anaerobic I_sc values did not eliminate the effect of aldosterone. 4. Addition of ouabain, a potent inhibitor of the Na⁺ pump, partially inhibited the effect of aldosterone on the anerobic I_sc but a significant hormonal increment remained. The source of the anaerobic I_sc was not identified; an effort was made, however, to determine the dependence of this current on glycolysis. During anaerobics, aldosterone increased the integral I_sc by 42% but did not alter lactate production. These results suggest that the steroid-dependent increase in the anaerobic I_sc may involve effects on permeability properties of the epithelium rather than on active tranport systems.     

Signal transduction for Schistocerca gregaria ion transport peptide is mediated via both cyclic AMP and cyclic GMP

The second messengers involved in the signal transduction for Schistocerca gregaria, ion transport peptide (Schgr-ITP) that regulates ion and fluid transport across the ileum of the desert locust S. gregaria, were measured using competitive enzyme-linked immunosorbent assays (ELISAs). Synthetic Schgr-ITP elevates intracellular levels of both cyclic AMP and cyclic GMP, measured over a 15 min period in the presence of 3-isobutyl-1-methylxanthine, in a dose-dependent manner. Furthermore, crude corpora cardiaca (CC) extracts elevate intracellular cyclic AMP levels 2-fold greater than Schgr-ITP, suggesting that factors present in the CC, other than Schgr-ITP, also act via this second messenger. These results suggest that the interaction of Schgr-ITP with two separate receptors, most likely a G-protein coupled receptor and a membrane bound guanylate cyclase, elevates intracellular levels of cyclic AMP and cyclic GMP to regulate ion and fluid transport across the locust ileum. Cyclic AMP stimulates Cl⁻, K⁺ and Na⁺ reabsorption, whereas secretion of H⁺ into the lumen of the ileum is most likely mediated via cyclic GMP. Cyclic GMP also stimulates Cl⁻ uptake in a similar manner to cyclic AMP. The measurement of tissue (central nervous system) levels of Schgr-ITP using an indirect ELISA confirms that the peptide is only present in the locust brain and the CC. The amounts present are greatest in the CC, where the peptide is presumably stored for release into the hemolymph when locusts feed.     

The effect of short-term low-temperature treatments on gene expression in Arabidopsis correlates with changes in intracellular Ca2+ levels

The role of changes in intracellular calcium ion concentration ([Ca²⁺]_i) in low‐temperature signal transduction in plants has lately been supported by several studies. An analysis to determine whether the low‐temperature‐induced increase in cytosolic Ca²⁺ concentration ([Ca²⁺]_cyt) could be correlated with a downstream response such as gene expression was carried out. The induction of the low‐temperature‐regulated gene LTI78 was used as an end point marker of the signal transduction pathway. It was found that this gene is induced by very brief low‐temperature exposures and that the induction does not depend on a continuous exposure to low temperature. By altering the cooling rate, different patterns of the Ca²⁺ response were obtained which could be correlated with different patterns of LTI78 induction. Furthermore, reducing the Ca²⁺ transients by pre‐treatment with the Ca²⁺ channel blocker La³⁺ also led to a reduced level of gene induction. The results show that brief exposures to low temperature results in the onset of a signalling pathway that leads to the induction of gene expression. This indicates the involvement of changes in [Ca²⁺]_cyt in low‐temperature signalling leading to LTI78 expression but the presence of multiple signalling pathways is suggested.