Actions of hydrogen sulphide on ion transport across rat distal colon

Background and purpose:

The aim of this study was to identify the actions of H₂S on ion transport across rat distal colon.

Experimental approach:

Changes in short-circuit current (Isc) induced by the H₂S-donor, NaHS, were measured in Ussing chambers. Cytosolic Ca²⁺ concentration was evaluated using fura-2.

Key results:

NaHS concentration-dependently induced a change in Isc, that was only partially inhibited by the neurotoxin, tetrodotoxin. Lower concentrations (≤10⁻³ mol·L⁻¹) of NaHS induced a monophasic increase in Isc, whereas higher concentrations induced an additional, secondary fall of Isc, before a third phase when Isc rose again. Blockers of H₂S-producing enzymes (expression demonstrated immunohistochemically) decreased basal Isc, suggesting that endogenous production of H₂S contributes to spontaneous anion secretion. The positive Isc phases induced by NaHS were due to Cl⁻ secretion as shown by anion substitution and transport inhibitor experiments, whereas the transient negative Isc induced by higher concentrations of the H₂S-donor was inhibited by mucosal tetrapentylammonium suggesting a transient K⁺ secretion. When applied from the serosal side, glibenclamide, an inhibitor of ATP-sensitive K⁺ channels, and tetrapentylammonium, a blocker of Ca²⁺-dependent K⁺ channels, suppressed NaHS-induced Cl⁻ secretion suggesting different types of K⁺ channels are stimulated by the H₂S-donor. NaHS-induced increase in cytosolic Ca²⁺ concentration was confirmed in isolated, fura-2-loaded colonic crypts. This response was not dependent on extracellular Ca²⁺, but was inhibited by blockers of intracellular Ca²⁺ channels present on Ca²⁺ storage organelles.

Conclusions and implications:

H₂S induces colonic ion secretion by stimulation of apical as well as basolateral epithelial K⁺ channels.     

Multiple action sites of flufenamate on ion transport across the rat distal colon

The antisecretory effects of flufenamate in the rat distal colon were investigated with the Ussing-chamber and the patch-clamp method as well as by measurements of the intracellular Ca(2+) concentration using fura-2-loaded isolated crypts. Flufenamate (5.10(-4) mol l(-1)) suppressed the short-circuit current (Isc) induced by carbachol (5.10(-5) mol l(-1)), forskolin (5.10(-6) mol l(-1)) and the Isc induced by the membrane-permeable analogue of cyclic AMP, CPT - cyclic AMP (10(-4) mol l(-1)). Indomethacin (10(-6) - 10(-4) mol l(-1)) did not mimic the effect of flufenamate, indicating that the antisecretory effect of flufenamate is not related to the inhibition of the cyclo-oxygenase. When the basolateral membrane was depolarized by a high K(+) concentration and a Cl(-) current was induced by a mucosally directed Cl(-) gradient, the forskolin-stimulated Cl(-) current was blocked by flufenamate, indicating an inhibition of the cyclic AMP-stimulated apical Cl(-) conductance. When the apical membrane was permeabilized by the ionophore, nystatin, flufenamate decreased the basolateral K(+) conductance and inhibited the Na(+) - K(+)-ATPase. Patch-clamp experiments revealed a variable effect of flufenamate on membrane currents. In seven out of 11 crypt cells the drug induced an increase of the K(+) current, whereas in the remaining four cells an inhibition was observed. Experiments with fura-2-loaded isolated crypts indicated that flufenamate increased the basal as well as the carbachol-stimulated intracellular Ca(2+) concentration. These results demonstrate that flufenamate possesses multiple action sites in the rat colon: The apical Cl(-) conductance, basolateral K(+) conductances and the Na(+) - K(+)-ATPase.     

Effects of higenamine on regulation of ion transport in guinea pig distal colon

Effects of higenamine on Na+, K+ and Cl- transport were studied on stripped guinea pig distal colonic mucosa in vitro using Ussing chambers. Addition of 10(-5) M higenamine induced a biphasic change in short circuit current (Isc): a transient increase followed by a long-lasting decrease that was accompanied by an increase in transepithelial conductance (Gt). The initial phase with an increase in Isc was partially inhibited by serosal bumetanide and abolished by mucosal diphenylamine-2-carboxylate, a chloride channel blocker, indicating transient induction of Cl- secretion. The second phase with a decrease in Isc was composed of two effects: the inhibition of the amiloride-sensitive electrogenic Na+ absorption and the stimulation of the bumetanide-sensitive K+ secretion. However, the initial transient increase was not observed at the lower concentration of higenamine (10(-8)-10(-6) M). All the changes in Isc and Gt induced by higenamine were suppressed by the non-selective beta-adrenergic receptor antagonist propranolol and by the beta2-adrenergic receptor antagonist ICI-118,551, but not by the beta1-adrenergic-receptor-selective antagonist atenolol or by the alpha-antagonists phentolamine, prazosin and yohimbine. These results suggest that higenamine inhibits electrogenic Na+ absorption and stimulates electrogenic K+ and Cl- secretion through beta2-adrenergic receptors in guinea pig distal colon.     

Segmental heterogeneity of epithelial ion transport induced by stimulants in rat distal colon

Mammalian colon plays an important role in electrolyte and water balance, and exhibits significant segmental heterogeneity. The different responses to stimulants even exist in different segments of rat distal colon. The present study focus on the segmental heterogeneity of epithelial responses to different stimulants and investigates the possible mechanism by using short circuit current recording technique. Baseline I(SC) in the segment 4 of distal colon (DC(4), 20.8+/-2.8 microA.cm(-2)) was significantly lower than that in the segment 1 of distal colon (DC(1), 40.5+/-1.9 microA.cm(-2)). Basolateral application of indomethacin induced a larger reduction of the baseline I(SC) in DC(4) (-28.2+/-3.9 microA.cm(-2)) than that in DC(1) (-10.1+/-3.9 microA.cm(-2)). Moreover DC(4) are more sensitive to foskolin (a cAMP activator, apical side), Ach (basolateral side) and 5-HT (basolateral side) than DC(1), which was not affected by pretreatment with amiloride, a blocker of epithelial Na(+) channel. Basolateral pretreatment with atropine (muscarinic cholinergic receptor antagonist) for 10 min, Ach-induced DeltaI(SC) increases in both DC(1) and DC(4 )were totally blocked. Otherwise, 5-HT(4) receptor antagonist GR113808 (basolateral side) and SB-204070 (basolateral side) completely inhibited 5-HT-induced I(SC) increases in both DC(1) and DC(4). Taken together, the results suggested that the segmental heterogeneity of epithelial responses to different stimulants exists in rat distal colon. And it is more likely related to the dissimilar distribution of membrane proteins involved in the ion transports within different segments of rat distal colon.     

Sites of action of hydrogen peroxide on ion transport across rat distal colon

BACKGROUND AND PURPOSE: The aim of this study was the identification of the mechanism of oxidant-induced intestinal secretion. EXPERIMENTAL APPROACH: The action of H2O2 on ion transport across rat distal colon was evaluated in Ussing chambers. Changes in cytosolic Ca2+ concentration were measured using fura-2. KEY RESULTS: H2O2 concentration-dependently induced an increase in short-circuit current (Isc), which was due to a stimulation of Cl(-) secretion. The effect of H2O2 was dependent on the presence of serosal Ca2+. It was inhibited after emptying of intracellular Ca2+ stores by cyclopiazonic acid or blockade of ryanodine receptors by ruthenium red, whereas a blocker of inositol 1,4,5-trisphosphate receptors was less effective. Fura 2-experiments confirmed an increase in the cytosolic Ca2+ concentration in the presence of H2O2. Measurements of Cl- currents across the apical membrane at basolaterally depolarized epithelia revealed the activation of a glibenclamide-sensitive, SITS-resistant Cl- conductance by the oxidant. The activation of this conductance was inhibited after blockade of protein kinases with staurosporine. When the apical membrane was permeabilized with nystatin, two sites of action of H2O2 were identified at the basolateral membrane. The oxidant stimulated a basolateral tetrapentylammonium-sensitive K+ conductance and increased the current generated by the Na+-K+ pump. Pretreatment of the tissues with H2O2 reduced the action of subsequently administered Ca2+-, cAMP- and cGMP-dependent secretagogues demonstrating a long-term downregulation after the initial secretory response evoked by the oxidant. CONCLUSIONS AND IMPLICATIONS: H2O2 affects colonic anion secretion by action sites at both the apical, as well as the basolateral membrane.     

Melatonin inhibits prostaglandin E2- and sodium nitroprusside-induced ion secretion in rat distal colon

Although the gastrointestinal tract is a rich source of melatonin and possesses numerous melatonin-binding sites, the role of melatonin in this tissue has not yet been fully elucidated. In this work we focused on the role of melatonin in the modulation of ion transport in rat distal colon. Whereas melatonin had no effect on colonic secretion or caused only infrequent and small changes in the short circuit current (Isc) due to its solvent ethanol, this mediator significantly modulated the secretion elicited by some secretagogues. Out of the five substances tested (prostaglandin E(2); 5-hydroxytryptamine; bethanechol; histamine; sodium nitroprusside) melatonin inhibited the effect of prostaglandin E(2) (PGE(2)) and sodium nitroprusside (SNP). Melatonin concentration-dependently decreased PGE(2)-evoked Isc and this inhibitory effect was more obvious from the mucosal side. The basal level of cAMP in colonic mucosa was not influenced by melatonin, but this drug prevented a PGE(2)-induced increase in the level of cAMP. The neurotoxin tetrodotoxin blocked the inhibitory effect of melatonin on SNP-induced Isc. Our data suggests that melatonin takes part in the modulation of colonic ion transport. The modulatory effect of melatonin on PGE(2)-induced Isc occurs directly at the level of the epithelium, whereas the effect on SNP-induced Isc is indirect and located in tetrodotoxin-sensitive enteric neurons.     

A dual role of 5-hydroxytryptamine receptor 3 in serotonin induced ion transport in rat distal colon

5-hydroxytryptamine (5-HT)-evoked intestinal secretion can be divided into neural and non-neural pathway. Recently, 5-HT(3) receptor in neural pathway received much attention as a possible target in bowel diseases. The present study aims to investigate the effects of 5-HT(3) receptor in different enteric neural plexus (myenteric plexus and submucosal plexus) on rat colonic ion transport by using rat intact colon and mucosa/submucosa preparations. Ussing chamber and real-time PCR techniques were performed in our present study. Surprisingly, we found that in mucosa/submucosa preparations, 5-HT-induced DeltaI(SC) (change in short-circuit current) was not inhibited, but further increased by pretreatment with 5-HT(3) receptor antagonists, MDL72222 and Tropanyl-3, 5-dimethylbenzoate. And this response was significantly attenuated in the presence of tetrodotoxin (TTX). Conversely, in rat intact colon, 5-HT(3) receptor antagonists significantly inhibited 5-HT-induced DeltaI(SC). The results from real-time PCR proved the existence of 5-HT(3) receptor in muscularis externa and submucosa. Taken together, 5-HT(3) receptors possess a role of dual regulation on electrolyte secretion in rat distal colon, the neural stimulatory effect of 5-HT(3) receptor in myenteric plexus and the inhibitory effect of 5-HT(3) receptor in submucosal plexus.     

The effect of neuropeptide Y on sodium, chloride and potassium transport across the rat distal colon.

1. Neuropeptide Y (NPY; 10(-10)-10(-7) mol l-1) reduced basal short-circuit current (Isc) in a concentration-dependent manner in the rat distal colon but was ineffective in the proximal colon. 2. The action of NPY was dependent upon the presence of Cl- and HCO3- anions and was blocked by prior treatment of the tissue with a Cl- channel blocker. The decrease in Isc was associated with an increase in mucosa-to-serosa fluxes of Na+, Rb+ (K+) and Cl-, whereas the serosa-to-mucosa flux of Cl- was decreased. 3. The size of the inhibitory NPY effect was linearly correlated with the height of the basal Isc, i.e. it inhibited 55% of basal secretory Isc. 4. The action of NPY was unaffected by indomethacin and tetrodotoxin, when given alone, but was abolished, when the basal Isc was decreased to values near zero by a combination of both inhibitors. This inhibition could be overcome by restoring basal Isc with prostaglandin E2, indicating that the effect of NPY is not mediated by nerves or prostaglandins, but that NPY is only effective, when anion secretion is stimulated by the spontaneous release of neurotransmitters and prostaglandins. 5. NPY inhibited the increase in Isc induced by veratridine and prostaglandin E2, but it had no effect on the Isc induced by direct stimulation of the adenylate cyclase with forskolin, or on Isc induced by stimulation of the Ca(2+)-pathway with carbachol.(ABSTRACT TRUNCATED AT 250 WORDS)     

Appearance of segmental discrepancy of anion transport in rat distal colon

The present study investigated the segmental discrepancy of the rat distal colonic anion transport induced by luminal forskolin and the possible underlying mechanisms using short-circuit current recording technique and comparative quantity real-time PCR analysis. Forskolin-induced I(SC) in the segment next to lymph node (DC(1)) and the segment 4 cm away from lymph node (DC(4)) were 4.09+/-0.66 muA/cm(2) and 18.84+/-3.18 muA/cm(2) (n=13), respectively, which were blocked by luminal Cl(-) channel blocker, glybenclamide (1 mM) (n=5, p<0.01), as well as removal of extracellular Cl(-) and HCO(3)(-) in both DC(1) and DC(4) (n=5, p<0.001). Furthermore luminal pretreatment with K(+) blockers, TEA (5 mM) and glybenclamide (100 muM) didn't affect forskolin and bumetanide-enhanced I(SC). Reducing serosal Cl(-) concentration increased forskolin-induced I(SC) by 90% in DC(1) but decreased forskolin-induced I(SC) in DC(4) by 50%. Furthermore, pretreatment with serosal bumetanide, an inhibitor of Na(+)-K(+)-2Cl(-) cotransporter, enhanced forskolin-induced I(SC) by 87% in DC(1), from 4.09+/-0.66 muA/cm(2) to 7.65+/-0.53 muA/cm(2) (n=6, p<0.01), but inhibited forskolin-induced I(SC) by 50% in DC(4), from 29.19+/-4.51 muA/cm(2) to 15.06+/-4.10 muA/cm(2) (n=6, p<0.05). Pretreatment with luminal amiloride (10 muM), an inhibitor of ENaC, and serosal 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) (200 muM), an inhibitor of NBC, significantly inhibited the forskolin-induced I(SC) in DC(1) (n=6, p<0.05), but not in DC(4). Real-time PCR analysis did not show the significant differences between the two segments in the expression amounts of CFTR and NKCC mRNAs, however the expression of NBC mRNA in DC(4) was significantly higher than that in DC(1). In conclusion, the rat distal colon manifests a segmental discrepancy in anion transport stimulated by luminal forskolin. HCO(3)(-) might be predominantly involved in the forskolin-induced anion secretion in DC(1), but Cl(-) might be the main component of the anion secretion in DC(4).     

Cyclic AMP-dependent regulation of K+ transport in the rat distal colon.

1. The effect of agonists of the cyclic AMP pathway and of 293B, a chromanole-derived K+ channel blocker, on K+ transport in the rat distal colon was studied by measuring unidirectional fluxes, uptake, and efflux of Rb+ in mucosa-submucosa preparations and by patch-clamp of crypt epithelia from isolated crypts. 2. 293B concentration-dependently inhibited basal and forskolin-stimulated short-circuit current. In isolated crypts 293B blocked a basal K+ conductance but had no effect on cyclic AMP-evoked depolarization induced by the opening of apical Cl- channels. When the effect of cyclic AMP on Cl- conductance was prevented by substituting Cl- with gluconate, an inhibition of total cellular K+ conductance by forskolin and a membrane-permeable cyclic AMP analogue was unmasked. 3. Unidirectional ion flux measurements revealed that 293B suppressed the increase in JRbsm induced by forskolin. This, together with the inhibition of cyclic AMP-induced anion secretion indicates that the drug blocks K+ channels, presumably both in the apical and the basolateral membrane. Forskolin caused not only inhibition of K+ absorption, but also stimulation of K+ secretion. The inhibition was diminished, but not blocked, in the presence of inhibitors of the apical H(+)-K(+)-ATPase, vanadate and ouabain. Forskolin stimulated serosal, bumetanide-sensitive Rb+ uptake, whereas mucosal, ouabain/vanadate-sensitive uptake remained unaffected. 4. Efflux experiments revealed that forskolin caused a redistribution of cellular K+ efflux reducing the ratio of basolateral versus apical Rb+ efflux. 5. These results suggest that intracellular cyclic AMP exerts its effects on K+ transport by several mechanisms: an increase in the driving force for K+ efflux due to the depolarization induced by opening of Cl- channels, a stimulation of the basolateral uptake of K+ via the Na(+)-K(+)-Cl(-)-cotransporter, and a decrease of the ratio of basolateral versus apical K+ conductance leading to an enhanced efflux of K+ into the lumen and a reduced K+ efflux to the serosal compartment.     

Regulation of ion transport by 5-hydroxytryptamine in rat colon

1. 5-Hydroxytryptamine (5-HT) modulates the motility and secretion of the gastrointestinal tract. To examine the direct effect of 5-HT on the secretions of colonic epithelial cells, a short-circuit current was used to measure electrolyte transport in the rat stripped distal colon. A neuronal Na+ channel blocker and a cyclo-oxygenase inhibitor were routinely added in experiments to abolish the effects of the enteric nervous system and endogenous prostaglandin, respectively. 2. Basolateral application of 5-HT (10 micromol/L) induced an increase in the short circuit current (ISC). Removal of extracellular Cl-, HCO3- or both resulted in a 59.6, 76.4 and 90% reduction of 5-HT-elicited responses, respectively. The Ca(2+)-dependent Cl- channel blocker 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) had no effect on the 5-HT-induced increase in ISC, but the selective cystic fibrosis transmembrane conductance regulator (CFTR) channel blocker glibenclamide (1 mmol/L) inhibited 5-HT-induced increases in ISC by approximately 92.9%. Removal of apical Na+ reduced the 5-HT-induced increase in ISC by 33.3%. 3. Basolateral pretreatment with 100 micromol/L bumetanide (an inhibitor of the Na(+)-K(+)-2Cl- cotransporter), 200 micromol/L DIDS (an inhibitor of the Na(+)-HCO3- transporter or the Cl-/HCO3- exchanger) or both decreased the DeltaISC induced by 5-HT by approximately 75.5, 59.0 and 86.3%, respectively. Removal of basolateral Na+ also reduced the current evoked by 5-HT. 4. The selective 5-HT4 antagonist GR113808 (1 micromol/L) totally abolished the 5-HT-induced increase in ISC, whereas 2-methyl-5-HT (100 micromol/L) induced a weak ISC response. 5. In conclusion, the present study has demonstrated that 5-HT can elicit Cl(-)- and HCO3- anion secretion and Na+ absorption by acting directly on colonic epithelial cells via 5-HT4 receptors.     

Berberine inhibition of electrogenic ion transport in rat colon.

1. The effects of the alkaloid berberine on basal and stimulated ion transport were investigated in voltage-clamped rat colonic epithelia. 2. Berberine (100-500 microM) reduced basal short circuit current (SCC) when applied basolaterally but not when applied apically. 3. SCC responses to mast cell activation by anti-rat IgE were significantly attenuated in the presence of berberine. 4. Berberine, applied to the basolateral bathing solution, also reduced SCC responses to the following agents which stimulate chloride secretion in rat colon: carbachol, forskolin, sodium nitroprusside, dibutyryl cyclic-AMP, heat-stable E. coli enterotoxin, 8-bromo-cyclic GMP and thapsigargin. Calcium mediated ion transport responses appear to be more sensitive to berberine inhibition than those which are cyclic GMP-mediated, which in turn are more sensitive than cyclic AMP-mediated responses. 5. Berberine added apically was without effect upon forskolin-stimulated ion transport. Cytochalasin D treatment of the lumenal surface of rat colon conferred apical-side sensitivity to berberine. 6. Berberine (at concentrations up to 500 microM) was without effect on generation of cyclic AMP by forskolin or on generation of cyclic GMP by sodium nitroprusside in isolated mucosal segments. Protein kinase A activity stimulated by dibutyryl cyclic AMP was unaffected by berberine (at concentrations up to 500 microM). 7. The precise mechanism of action of berberine remains to be elucidated. However, its site of action appears to be distal to second messenger production and may be at a level common to all stimuli of colonic chloride secretion.     

Experimental inflammation of the rat distal colon inhibits ion secretion in the proximal colon by affecting the enteric nervous system

Intestinal inflammation causes hyporesponsiveness of the inflamed tissue to secretagogues but little is known about the behaviour of the areas proximal to the site of inflammation. We studied the responses of the proximal segment of the colon to carbachol, histamine, isobutylmethylxanthine (IBMX) and vasoactive intestinal peptide (VIP) in rats with trinitrobenzenesulphonic acid (TNBS)-induced, chronic inflammation of the distal colon. Macroscopic and biochemical analysis ruled out the presence of inflammation in the proximal colon. When mounted in Ussing chambers under voltage-clamp conditions, basal transport and conductance were not affected. However, the maximum response in the concentration/response curves (short-circuit current) for carbachol and histamine was reduced in TNBS-treated rats, without changes in the EC₅₀. This effect corresponded to reduced chloride secretion, as demonstrated by ion substitution experiments. The responses to IBMX and VIP were virtually unaffected. The inhibitory effect was abolished by pretreatment with the neural blockers tetrodotoxin and lidocaine but not indomethacin, suggesting that the enteric nervous system is responsible for the inhibition. In conclusion, chronic distal inflammation of the distal colon results in inhibition of calcium-dependent secretion in the proximal colon via a reduction of the contribution of the enteric nervous system.     

The effects of capsaicin upon electrogenic ion transport in rat descending colon

Summary Preparations of rat descending colon mucosa have been used to record changes in short circuit current (SCC) under voltage clamp conditions. When added to the basolateral compartment capsaicin (8-methyl-N-vanillyl-6-nonenamide, 0.1-3 M) caused an initial transient increase in SCC, followed by a more prolonged reduction in SCC, that lasted for 20–30 min.Repeated applications of 3 M capsaicin caused desensitisation of the initial secretory response. The antisecretory effects (i. e. reduction in SCC from the original baseline) remained, although they were significantly reduced. In some preparations described as non-responders, 3 M capsaicin did not elicit a secretory response. No desensitisation of the remaining antisecretory responses was observed in these tissues; in fact these reductions in SCC were consistently larger than those from tissues which responded with a secretory response.Tetrodotoxin (100 nM), hexamethonium (10 M), and yohimbine (50 M) had no significant effect upon either secretory or antisecretory responses. Ruthenium red (10 M) abolished the secretory response to 3 M capsaicin, but had no effect upon the antisecretory responses. Pretreatment of the tissues with 1 M substance P(SP) resulted in significant desensitisation to the peptide and abolished the secretory response to 3 M capsaicin. The antisecretory responses remained, and were significantly larger compared with responses from control tissues. Send offprint requests to H. M. Cox at present address     

Captopril inhibits sodium and water transport in the toad skin, a model of the distal nephron

Captopril (SQ 14225), an orally active angiotensin I-converting enzyme inhibitor (CEI), increases natriuresis and diuresis in man and experimental animals in vivo, as well as in the isolated perfused rat kidney, raising the possibility of a direct renal action of the drug. We tested this hypothesis by studying its effects in the isolated toad skin, a model of the distal nephron devoid of vascular and nervous influences. When added to the dermal bath, captopril caused a reversible, concentration-related decrease in short-circuit current (SCC), a measure of active transepithelial Na transport. Keeping the toads in 0.1 M NaCl for 4 or more days increased sensitivity to the drug, which then inhibited SCC maximally (49 +/- 12% at 3.4 X 10(-3) M, P less than 0.01, n = 10), suggesting its effect might be modulated by endogenous mineralocorticoid activity. Captopril also inhibited the increase in SCC and in osmotic water permeability caused by neurohypophyseal peptides (NHP). The increases in SCC by non-peptidic agents (nystatin, a polyene antibiotic, or norepinephrine, an adrenergic agonist) were not altered, ruling out a generalized toxic effect, or any significant inhibition of the Na pump by captopril. The apparently specific effect of the drug on the permeability responses to NHP seems to be exerted proximally to the apical border, since the response of the latter to other agents was preserved. The present data suggest SH groups may be involved, since other CEI lacking such groups (teprotide and MK-422) do not produce such effects. These observations support the notion that a direct tubular effect may be involved in the increased diuresis and natriuresis observed after administration of captopril.     

Neuronally mediated and direct effects of prostaglandins on ion transport in rat colon descendens

Summary Two preparations of rat colon descendens were used in order to localize the action sites of iloprost and prostaglandin E₂ (PGE₂). One preparation, the mucosa-submucosa preparation contained the submucosal and mucosal plexus whereas for the mucosa preparation in addition the submucosa with the submucosal plexus was removed. Iloprost (10^–6 mol · 1^–1) caused an increase in short-circuit current (Isc), potential difference (Pd) and tissue conductance (Gt) of the mucosa-submucosa preparation reflecting net Cl^– secretion as confirmed by unidirectional ion flux measurements. The Cl^– secretion was due to an increase in J _infsm ^supCl and a decrease in J _infms ^supCl . These effects were completely abolished by addition of 5 × 10^–5 mol · l^–1 atropine. Iloprost had only small and inconsistent effects in the mucosa preparation. In contrast PGE₂ (10^–6 mol · l^–1) increased Isc, Pd and Gt due to Cl^– secretion in both preparations. The Cl^– secretion was caused by an increase in J _infsm ^supCl and a decrease in J _infms ^Cl Only the PGE₂ effect in the mucosa-submucosa preparation but not in the mucosa preparation was inhibited by about 50% by atropine. The results suggest that the prostacyclin derivative iloprost induces a Cl^– secretion only by an activation of submucosal neurons whereas PGE₂ acts both on the epithelium and the submucosal plexus. The neuronal effects of prostaglandins appear to be, at least in part, mediated by muscarinic receptors. Send offprint requests to M. Diener     

Strontium transport in the rat colon

Summary Concentration dependence of strontium (Sr) fluxes across the colon ascendens and descendens of the rat were measured in a modified Ussing-chamber. Mucosa (m) to serosa (s) and s to m Sr fluxes across both colonic segments were linearly related to the Sr concentration from 0.125 mmol/l to 10 mmol/l. In the colon ascendens m to s Sr fluxes were slightly higher than the fluxes in the opposite direction, resulting in net Sr absorption. In the colon descendens s to m fluxes were higher than the ms fluxes, resulting in net Sr secretion. Neither Sr nor calcium (Ca) showed a concentration dependent interaction with respect to their unidirectional fluxes in both parts of the colon. Only in the colon ascendens Sr at the highest concentration (10 mmol/l) inhibited m to s calcium transport.Experiments, in which the voltage dependence of the unidirectional Sr fluxes was measured confirmed the results obtained from the concentration dependence: (1) The unidirectional fluxes of Sr across the colon ascendens and descendens were totally voltage dependent, i.e. diffusive. (2) In the colon descendens the voltage dependence of the s to m flux was steeper than the flux from m to s. It is hypothesized that this prevalence is caused by an anomalous solvent drag effect. 1.25-Dihydroxyvitamin D₃ [1.25 (OH)₂D₃] stimulated m to s calcium flux in the colon descendens but had no effect on Sr flux.The results demonstrate that Sr and Ca in the rat colon are transported by different mechanisms. In contrast to the Ca transport the Sr flux is only diffusive and insensitive to 1.25 (OH)₂D₃.Supported by the Deutsche Forschungsgemeinschaft SFB 38 Membranforschung Send offprint requests to U. Karbach at the above address     

P2Y2 receptor-mediated inhibition of ion transport in distal lung epithelial cells.

1 Rat foetal distal lung epithelial cells were plated onto permeable supports where they became integrated into epithelial sheets that spontaneously generated short circuit current (ISC). 2 Apical ATP (100 microM) evoked a transient fall in ISC that was followed by a rise to a clear peak which, in turn, was succeeded by a slowly developing decline to a value below control. Apical UTP evoked an essentially identical response. 3 UDP and ADP were ineffective whilst ATP had no effect when added to the basolateral solution. These effects thus appear to be mediated by apical P2Y2 receptors. 4 The rising phase of the responses to ATP/UTP was selectively inhibited by anion transport inhibitors but persisted in the presence of amiloride, which abolished the inhibitory effects of both nucleotides. Thus, apical nucleotides appear to evoke a transient stimulation of anion secretion and sustained inhibition of Na+ absorption. 5 Basolateral isoprenaline (10 microM) elicited a rise in ISC but subsequent addition of apical ATP reversed this effect. Conversely, isoprenaline restored ISC to its basal level following stimulation with ATP. Apical P2Y2 receptors and basolateral beta-adrenoceptors thus allow their respective agonists to exert mutually opposing effects on ISC.