Rise In Isc Research Articles

Effects of Urocortins on Intestinal Ion Secretion in the Mouse Colon

Background and AimsUrocortins (Ucn1, Ucn2, and Ucn3) belong to the corticotropin‐releasing factor (CRF)‐like peptide family and bind to two subtypes of CRF receptors (CRF1 and CRF2) to exert their biological effects. Ucn1 binds to both CRF1 and CRF2 receptors with equal high affinity, whereas Ucn2 and Ucn3 bind exclusively to CRF2. CRF and urocortins are expressed in the gastrointestinal tract. CRF has been implicated in stress‐stimulated intestinal ion secretion and diarrhea. However, the effects of urocortins on intestinal ion secretion have remain uninvestigated. The aim of the present study was to investigate the role of urocortins in the regulation of intestinal ion secretion.MethodsMucosa/submucosa preparations from mouse colon were mounted in Ussing flux chambers for measurement of short‐circuit current (Isc) as an indicator of mucosal ion secretion.ResultsApplication of CRF, Ucn1, Ucn2, and Ucn3 to the basolateral side of the preparation increased baseline Isc in a concentration‐dependent manner with an EC50 (in M) of 1.28×10−6, 6.04×10−7, 2.89×10−7, or 2.98×10−7, respectively, with the rank order of potency: Ucn2≈Ucn3>Ucn1>CRF. At 1 μM, Ucn3 caused a greater rise in Isc (ΔIsc: 47.01±8.94 μA/cm2), compared with Ucn2 (17.71±2.63 μA/cm2), Ucn1 (7.06±4.33 μA/cm2), or CRF (8.92±4.11 μA/cm2). The CRF1 receptor antagonist NBI 27914 significantly suppressed Ucn1‐evoked ΔIsc, but had no effect on Ucn2‐ or Ucn3‐evoked ΔIsc. The CRF2 receptor antagonist antisauvagine‐30 significantly suppressed Ucn1‐evoked ΔIsc, and completely blocked Ucn2‐ or Ucn3‐evoked ΔIsc. The neuronal blocker tetrodotoxin had no effect on Ucn1‐, Ucn2‐, or Ucn3‐evoked DIsc.ConclusionsThe results suggest that urocortins stimulate colonic ion secretion independent the submucosal plexus of the enteric nervous system. Both CRF1 and CRF2 receptors are involved in Ucn1‐stimulated ion secretion, while Ucn2‐ and Unc3‐stimulated ion secretion is mediated by the CRF2 receptors. Ucn2 and Ucn3 are more potent than CRF in stimulation of colonic ion secretion and may play a dominant role in stress‐induced hyper secretion and diarrhea.Support or Funding InformationNIH R15 DK097460‐01A1 (SL) and UWL graduate research service and educational leadership grant (AK).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Anion secretion evoked by ATP in rabbit cornea and its suppression in dry eye model

To explore the regulation of anion secretion across rabbit corneal epithelia by extracellular ATP and to investigate the mechanism of benzalkonium chloride (BAC) induced dry eye model involved in this secretion, short circuit current and patch clamp were used. Changes in cytosolic Ca2+ concentration were measured using Fluo3. The cornea responded to apical application of ATP in short circuit current (Isc). Ion substitution experiments showed the rise in Isc due to Cl‐ and HCO3‐ secretion. Pretreatment with Cl‐ channel blockers, such as DIDS, NPPB and DPC attenuated the response. Moreover, the Isc was inhibited by U73122 and BAPTA/AM, indicating the requirement of PLC and intracellular calcium (Ca2+i) in the anion secretion. In whole‐cell patch clamp study, ATP stimulated a Ca2+‐activate Cl‐ channel and this current showed an outward rectifying I‐V relationship. In BAC‐pretreated epithelial cell, the whole‐cell current was suppressed. Dynamic changes of Ca2+i demonstrated the elevations of Ca2+i was smaller in BAC‐treated epithelia compared to control cells when they were challenged by ATP and Thapsigagin. These findings suggested ATP could induce anion secretion, and this secretion was inhibited in BAC‐evoked rabbit dry eye model. Abnormality of intracellular Ca2+ and Ca2+ activated Cl‐ channel may take charge of inhibitory anion secretion in the rabbit corneal epithelial cells of dry eye model.

Histamine-induced chloride secretion is mediated via H 2 -receptors in the pig proximal colon

The aim of this study was to investigate the action of histamine on function of epithelia in the pig proximal colon. Isolated epithelia of the pig proximal colon were prepared by slide-stripping and mounted in Ussing chambers. Short-circuit current (Isc) was measured after serosal addition of histamine (20 micromol/l) with or without pretreatment with histamine receptor antagonists (H1: chlorpyramine, 10 micromol/l; H2: famotidine, 100 micromol/l; H3: thioperamide, 10 micromol/l), a cyclooxygenase inhibitor (indomethacin, 10 micromol/l), or a neuronal conduction blocker (tetrodotoxin, 1 micromol/l). Alternatively, histamine receptor agonists (H1: 2-pyridylethylamine; H2: dimaprit; H3: R-alpha-methylhistamine, each 100 micromol/l) were added to the serosal side. Flux studies using 14C-mannitol, 22Na+ and 36Cl- were performed in the presence of 100 micromol/l histamine on the serosal side. Serosal application of histamine induced a rapid rise in Isc with a maximum 3 min after addition, followed by a slow decrease. Only pretreatment with famotidine decreased the epithelial response to histamine. Pretreatments with chlorpyramine, thioperamide, indomethacin or tetrodotoxin did not change histamine-induced increases in Isc. Action of histamine could be simulated by dimaprit, but not by 2-pyridylethylamine or R-alpha-methylhistamine. Histamine induced an increase in serosal-to-mucosal chloride flux leading to a decrease of chloride net absorption. Fluxes of sodium and mannitol were not affected by histamine. In contrast to the importance of H1-receptors in other gut epithelia, histamine acts directly via H2-receptors in the porcine proximal colon. Changes in Isc after histamine addition are primarily due to chloride secretion. The paracellular permeability is not influenced by histamine.

COX-dependent and -independent pathways in bradykinin-induced anion secretion in rat epididymis.

Lysylbradykinin (LBK) added to the apical or basolateral side of cultured rat epididymal monolayers stimulated a rise in short-circuit current (Isc) due to anion secretion. The concentration-response relationships for the apical and basolateral applications have EC50 value of 0.001 microM. The responses to apical or basolateral application of LBK were blocked by WIN64338, a specific B2 receptor antagonist, but not by Des-Arg9,[Leu8]-BK, a specific B1 receptor antagonist, indicating that the LBK effects were mediated through B2 bradykinin receptors. Experiments to desensitize the B2 receptors by repeated stimulation have demonstrated that the responses to apical or basolateral LBK were due to discrete receptors on the apical or basolateral surface. In epithelia clamped in the Ussing chambers, addition of LBK to the apical or basolateral surface evoked release of PGE2 into the apical and basolateral bathing solutions over the first 10 min following hormone addition. LBK added to the basolateral side elicited a greater release than it was added to the apical side. Pretreatment of the epithelia with piroxicam (5 microM) abolished PGE2 release elicited by apical or basolateral LBK and abrogated the Isc induced by basolateral LBK. However, the rise in Isc induced by apical LBK was reduced by 31.3% only. The anion secretion response to apical LBK was not affected by MDL-12330A, an adenylate cyclase inhibitor, but greatly attenuated by thapsigargin, an inhibitor of intracellular Ca2+ release. However, the reverse effects were seen for basolateral LBK. It is concluded that distinct pathways are involved in the stimulation of anion secretion by apical or basolateral LBK. The response to basolateral LBK was COX-dependent, mediated by PGE2 and involves cAMP as second messenger. In contrast, the response to apical LBK is largely COX-independent, not mediated by PCE2 and involves Ca2+ as intracellular messenger.

Oxygen-evoked Na+ transport in rat fetal distal lung epithelial cells.

Monolayer cultures of rat fetal distal lung epithelial (FDLE) cells generated larger spontaneous short circuit currents (ISC) when maintained (48 h) at neonatal alveolar PO2 (100 mmHg) than at fetal PO2 (23 mmHg). When cells were shifted between these atmospheres in order to impose a rise in PO2 equivalent to that seen at birth, no rise in ISC was seen after 6 h but the response was fully established by 24 h. Studies of basolaterally permeabilised cells revealed a small rise in apical Na+ conductance (GNa) 6 h after PO2 was raised but no further change had occurred by 24 h. A substantial rise was, however, seen after 48 h. Reporter gene assays showed that no activation of the -ENaC (epithelial Na+ channel -subunit) promoter was discernible 24 h after PO2 was raised but increased transcriptional activity was seen at 48 h. Studies of apically permeabilised cells showed that a small rise in Na+ pump capacity was evident 6 h after PO2 was raised and, in common with the rise in ISC, this effect was fully established by 24 h. The rise in ISC thus develops 6-24 h after PO2 is raised and is due, primarily, to increased Na+ pump capacity. The increase in GNa thus coincides with activation of the -ENaC promoter but these effects occur after the rise in ISC is fully established and so cannot underlie this physiological response. The increased transcription may be an adaptation to increased Na+ transport and not its cause.

Activation of cystic fibrosis transmembrane conductance regulator in rat epididymal epithelium by genistein.

The effect of genistein on anion secretion via cystic fibrosis transmembrane conductance regulator (CFTR) in cultured rat cauda epididymal epithelia was studied by short-circuit current (Isc) technique. Genistein added apically stimulated a concentration-dependent rise in Isc due to Cl(-) and HCO(3)(-) secretion. The genistein-induced Isc was observed in basolaterally permeabilized monolayers, suggesting that the Isc response was mediated by the apical anion channel. The response could be blocked by the nonspecific Cl(-) channel blocker, diphenylamine-2-carboxylate (DPC), but not by the Ca(2+)-activated Cl(-) channel blocker, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). Genistein did not increase intracellular cAMP, but H-89, a protein kinase A inhibitor, completely abolished the Isc response to genistein. Moreover, pretreatment of the tissues with MDL-12330A, an adenylate cyclase inhibitor, markedly attenuated the Isc response to genistein, but the response was restored upon the addition of exogenous cAMP. Ca(2+), protein kinase C, tyrosine kinase, and protein phosphatase signalling pathways were not involved in the action of genistein. It is speculated that genistein stimulates anion secretion by direct interaction with CFTR. This requires a low level of phosphorylation of CFTR by basal protein kinase A activity. It is suggested that genistein may provide therapeutic benefit to male infertility associated with cystic fibrosis.

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.

Nucleotide-evoked calcium signals and anion secretion in equine cultured epithelia that express apical P2Y2 receptors and pyrimidine nucleotide receptors.

1. Experiments with a spontaneously transformed equine epithelial cell line showed that certain nucleotides increased intracellular free calcium ([Ca2+]i) in cells plated on glass coverslips. The rank order of potency was ATP UTP > 5-Br-UTP, whilst UDP and ADP were ineffective. The response thus appears to be mediated by P2Y2 receptors. 2. Nucleotides also increased short circuit current (Isc) in cells grown into epithelial monolayers and the rank order of potency was UDP> UTP > 5-Br-UTP > ATP > ADP. The increase in [Ca2+]i and the rise in ISC thus have different pharmacological properties. Cross-desensitization experiments indicated that, as well as P2Y2 receptors, the monolayer cultures express at least one additional receptor population that allowed nucleotides to increase ISC. 3. The UDP-evoked increase in ISC was essentially abolished in BAPTA-loaded epithelia suggesting that this response is dependent upon increased [Ca2+]i. Moreover, experiments in which ISC and [Ca2+]i were measured simultaneously showed that the UDP- and ADP-evoked increases in ISC were accompanied by increases in [Ca2+]i. 4. When grown under conditions which favour the development of a polarized phenotype, these epithelial cells thus appear to express [Ca2+]i-mobilizing receptors sensitive to UDP and ADP that are not present in non-polarized cells on coverslips.

THE EFFECTS OF STROMELYSIN OR MATRIX METALLOPROTEINASE-3 (MMP-3) ON INTESTINAL SECRETORY FUNCTION

33 In inflammatory bowel disease there is extensive damage to the extracellular matrix (ECM) and loss of the transepithelial sodium gradient and hence absorption. Stromelysin (MMP-3) is produced in intestinal inflammatory states and is important in tissue degradation. We have previously used human fetal intestine to explore inflammatory mechanisms and here we study the effect of MMP-3 on its secretory function. 3 cm loops of 15 wk gestation gut or T84 cell monolayers were incubated with MMP-3 50 nM or saline (C) on the apical surface then mounted in Using Chambers. MMP-3 after 1, 2 and 3 hrs caused a rise in short circuit current (Isc) of 105±33, 125±5.8 and 145±99 μA/cm2 (p<0.005). Resistance fell from 22±8 to 9±7 ohms/cm2 and PD rose from 1.4±0.8 to 2.6±0.8 mV at 3 hrs. In Cl- free conditions Isc fell by 83%. In contrast in T84 cells the rise in Isc was insignificant (47 vs 53μA/cm2). Thus MMP-3 induces Cl- dependent secretion in intact intestine but not in T84 monolayers suggesting that this is not due to a direct effect on enterocytes. We speculate that disruption of the ECM by MMP-3 also induces secretion and that MMP inhibitors may have a therapeutic role in diarrhoeal states.

Regulation of anion secretion by prostaglandin E2 in the mouse endometrial epithelium.

The present study was an investigation of the regulation of anion secretion across cultured mouse endometrial epithelium by prostaglandin E2 (PGE2) using the short-circuit current (ISC) technique. The cultured endometrial monolayers responded to both apical and basolateral application of PGE2 with a sustained rise in ISC in a concentration-dependent manner. However, the potencies of apical and basolateral addition of PGE2 were different, with apparent EC50 of 200 and 4 nM, respectively. Replacement of Cl- or HCO3- in the bathing solution significantly reduced the ISC responses to both apical and basolateral addition of PGE2; however, the apical response exhibited greater dependence on HCO3- . Pretreatment with diphenylamine 2,2'-dicarboxylic acid, a Cl- channel blocker, significantly reduced both PGE2-induced ISC responses, while pretreatment with amiloride, a Na+ channel blocker, did not exert any effect. Forskolin, an adenylate cyclase activator, and 3-isobutyl-dihydro-testosterone-1-methyl-xanthine, a cAMP phosphodiesterase inhibitor, mimicked the ISC response to PGE2 while MDL12330A, an adenylate cyclase inhibitor, completely abolished the PGE2-induced ISC. The results of the present study indicate that the anion secretion across the mouse endometrial epithelium may be regulated by PGE2 involving a cAMP-dependent mechanism predominantly. The differential responses to apical and basolateral challenge with PGE2 also suggest that PGE2 of different origins may play different roles in uterine function.

Rapid modulation of electrolyte transport in Caco-2 cell monolayers by enteropathogenic Escherichia coli (EPEC) infection

Background and aims—The pathophysiology of enteropathogenic Escherichia coli (EPEC) diarrhoea remains uncertain. EPEC adhere to enterocytes and transduce signals which produce a characteristic “attaching and effacing” (A/E) lesion in the...

Short-chain fatty acids inhibit cAMP-mediated chloride secretion in rat colon.

Butyrate stimulates salt absorption in mammalian colon. We examined whether butyrate also affects Cl- secretion. Mucosal segments of distal colon of male Sprague-Dawley rats and T84 cells were studied in Ussing chambers. In control colon, 1 mM dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP) increased short-circuit current (Isc) and serosal-to-mucosal Cl- flux (JsmCl) by 3.2 +/- 0.8 and 2.9 +/- 0.8 mueq.cm-2.h-1, respectively. Mucosal or serosal 25 mM butyrate prevented DBcAMP-induced increases in Isc and JsmCl. Four and eight millimolar butyrate caused half-maximal inhibition of the increases in JsmCl and Isc, respectively. Butyrate also inhibited basal JsmCl (by 2.0 +/- 0.4 mueq.cm-2.h-1) but not carbachol-mediated Cl- secretion. The relative inhibitory potency at 25 mM of other short-chain fatty acids (SCFA) paralleled their degree of cellular metabolism: butyrate > acetate = propionate > isobutyrate. At 25 mM, all SCFA reduced mucosal intracellular pH (pHi) transiently by 0.1 pH unit. In intact T84 cells, 50 mM butyrate inhibited the DBcAMP-induced rise in Isc by 55%. In T84 cells with nystatin-permeabilized basolateral membranes, butyrate inhibited the increase in Isc by 82%. We conclude that butyrate inhibits basal and cAMP-mediated Cl- secretion by a mechanism independent of pHi, possibly located at the apical membrane.

Serotonin induces Cl- secretion in human jejunal mucosa in vitro via a nonneural pathway at a 5-HT4 receptor

We examined the effect of 5-hydroxytryptamine (5-HT) on Na+ and Cl- flux (J) and short-circuit current (Isc) in human jejunal mucosa. Segments of jejunum, taken at the time of gastric bypass surgery, were stripped of the seromuscular layers (and attached neural ganglia) and mounted as flat mucosal sheets in Ussing chambers under short-circuit conditions. 5-HT (0.1-100 microM) produced a concentration-dependent rise in Isc (mean effective concn = 2.5 microM). Using 22Na and 36Cl, we measured flux across control tissues and in those exposed to 5-HT. 5-HT decreased both net JNa and JCl and increased Isc (-1.1 +/- 0.6, -1.7 +/- 0.6, and 0.6 +/- 0.1 mueq.cm-2.h-1, respectively). Thus the 5-HT-induced rise in Isc could be accounted for by reduced net JNa and JCl. 5-HT induced a significant (P < 0.05) Cl- secretion (serosal-to-mucosal flux) when glucose was included in the buffer bathing the mucosal surface. Neither tetrodotoxin, the adrenergic receptor antagonists prazosin and propranolol, nor the cholinergic receptor antagonists atropine and hexamethonium inhibited the change (delta) in Isc induced by 5-HT. 5-Methoxytryptamine (5-MeOT) and zacopride, known 5-HT4 receptor agonists, induced significant delta Isc. The 5-HT receptor antagonists N-acetyl-5-hydroxytryptophyl-5-hydroxytryptamine (5-HT1P), ketanserin (5-HT2), and ICS-205-930 (preferential for 5-HT3 at 0.1 microM had no effect on delta Isc.(ABSTRACT TRUNCATED AT 250 WORDS)

Anti-immunoglobulin E-stimulated ion transport in human large and small intestine

Background: Mast-cell regulation of intestinal ion transport, previously shown in animals and cultured cells, was examined in surgically resected human bowel in this study. Methods: Changes in short-circuit current (Isc) in response to rabbit anti-human immunoglobulin (lg) E or control serum, histamine1 and electrical stimulation were measured in muscle-stripped, noninflamed segments of intestine mounted in Ussing chambers. Chloride-free buffer, pyrilamine, piroxicam, sodium cromoglycate, and tetrodotoxin were examined for their effect on Isc responses to these stimuli. Results: Within 1–2 minutes of adding anti-IgE serum, a specific monophasic rise in Isc (peaking at 7–10 minutes) was observed in large and small intestine. This response was reduced ~80% in chloride-free buffer and inhibited by the histamine1-receptor antagonist, pyrilamine, and the cyclo-oxygenase inhibitor, piroxicam, implicating histamine and prostaglandins as mediators of the ion transport changes. The mast-cell stabilizer, sodium cromoglycate, reduced anti-IgE responses in the small, but not large, intestine. Approximately 50% inhibition of anti-IgE responses in colon by the neurotoxin, tetrodotoxin, indicated that nerves were involved. Conclusions: These results suggest that activation of mast cells releases mediators that stimulate intestinal ion transport through direct epithelial action and via nerves. This study provides important evidence that immunoregulation of intestinal ion transport does occur in humans.

A 5-HT3 Receptor Agonist Induces Neurally Mediated Chloride Transport in Rat Distal Colon

Having previously demonstrated that serotonin (5HT)-induced chloride secretion in rat distal colon is mediated at both neural and nonneural receptors, we isolated the neural component of this response by adding the selective 5-HT3 receptor agonist, 2-methyl-5-hydroxytryptamine (2Me5HT), to in vitro sheets of rat distal colon with intact neural plexuses. Rats were sacrificed, and the distal colon excised, opened, cut into sections and mounted, all layers intact, as flat sheets in Ussing chambers under short-circuited conditions. 2Me5HT induced a prompt, significant (P < 0.01), concentration-dependent rise in short-circuit current (Isc; EC50 6.2 μM); 50 μM 2Me5HT decreased both net sodium and chloride absorption (-0.1 ± 0.5 and -2.1 ± 0.8 μEq/cm2 · hr, respectively); the difference (2.0 ± 0.8 μEq/cm2 · hr) in these changes was not statistically different from the rise in Isc (1.5 ± 0.3 μEq/cm2 · hr). Since the only significant change inunidirectional flux was the rise in electrogenic CI- secretion (P < 0.01), the ΔIsc induced by 2Me5HT may be used as a measure of electrogenic chloride secretion induced by the agonist. The rise in Isc induced by 2Me5HT was abolished by both 0.2 μM tetrodotoxin and 0.1 μM ICS 205-930 (a 5-HT3 antagonist) but was not inhibited by 1.0 M atropine, 100 μM hexamethonium, 10 μM phentolamine, 10 μM propranolol, 10 μM 5-HTP-DP (a 5-HT1p antagonist), or 0.1 μM ketanserin (a 5-HT2 antagonist). These results indicate that 2-methyl-5-HT is a highly selective agonist for neurally based 5-HT3 receptors in this model. Since 2Me5HT-stimulated secretion is inhibited by neither adrenergic nor cholinergic antagonists, it is likely that the secretomotor neurotransmitter for this transport change is a neuropeptide.

Phorbol esters stimulate and inhibit Cl- secretion by different mechanisms in a colonic cell line.

Phorbol 12-myristate 13-acetate (PMA) was found to increase both the short-circuit current (Isc) and the efflux of 125I- or 36Cl- in the colonic epithelial cell line HT-29.cl19A. Neither the PMA-provoked rise in Isc nor the stimulation of 125I- efflux was affected by the cyclooxygenase inhibitor indomethacin. The PMA-induced increase in Cl- efflux was not accompanied by a rise in adenosine 3',5'-cyclic monophosphate (cAMP) levels. A prolonged incubation with PMA (3 h), however, inhibited the PMA- and the cAMP-stimulated Isc by greater than 90%, whereas the cAMP-provoked 125I- and 36Cl- efflux was not inhibited. The long-term PMA treatment was found to inhibit the basal and cAMP-provoked 86Rb+ efflux by 65 +/- 9 and 86 +/- 7%, respectively. A 3-h incubation with PMA also strongly inhibited the Ca2+ ionophore A23187-induced increase in 86Rb+ efflux, whereas the A23187-stimulated 125I- efflux was only marginally inhibited. These data suggest that phorbol esters, presumably by activation of protein kinase C, can provoke Cl- secretion in HT-29.cl19A colonocytes independently of a prostaglandin- or cAMP-mediated pathway. Prolonged exposure to PMA, however, causes an inhibition of net electrogenic Cl- secretion by downregulation of the activity of K+ transporters.

Immunoregulation of Endometrial and Jejunal Epithelia Sensitized by Infection

The hypothesis was tested that the uterus of the rat orally infected with the parasite Trichinella spiralis becomes hypersensitized and that subsequent antigenic challenge affects functions in the endometrial epithelium. Results of experiments comparing the immunological responsiveness of isolated rat uterus with that of the jejunum supports our hypothesis. Antigenic challenge of uterus mounted in Ussing-type chambers causes an elevation in transuterine short circuit current (Isc) of 6.4 +/- 0.8 microA/cm2. The transduction of the antigenic signal to elicit the electrophysiological response involves 5-hydroxytryptamine (5-HT) working through a nerve-independent pathway. The antigen-stimulated rise in Isc peaks approximately 3 min after challenge. The uterine response is blocked by diisothiocyanostilbene-2,2'-disulfonic acid, an inhibitor of bicarbonate-chloride exchange. The antigen-evoked change in jejunal Isc is biphasic, peaking at 1.5 and approximately 4.0 min after challenge, and is about 10-fold greater in magnitude than the Isc in the uterus. The transductive pathway in the jejunum involves 5-HT, histamine and prostaglandin acting partly through intrinsic nerves. The jejunal response to antigen is inhibited by diphenylamine-2-carboxylate, a chloride channel blocker. Changes in net ion transport which are primed by infection and evoked by antigen are apparently triggered by local anaphylaxis in both the uterus and jejunum.

EFFECT OF ORAL OR PARENTERAL SENSITIZATION TO COW'S MILK ON MUCOSAL PERMEABILITY IN GUINEA-PIGS

The systemic and local immune responses and intestinal barrier function were examined in orally or parenterally milk-sensitized guinea-pigs. Both types of sensitization led to positive passive cutaneous anaphylactic responses and high IgG titers against β-lactoglobulin (β-lg) especially in parenterally immunized animals. In Ussing chambers, sensitized jejunum had higher short-circuit current (Isc) than control jejunum, with and without β-lactoglobulin challenge. The further increase in Isc induced by serosal β-lg treatment was higher in parenterally (23.2 ± 3.4 μA/cm2) than orally (10.9 ± 2.9 μA/cm2) sensitized animals. Barrier function was tested as the intestinal transport and degradation of Horseradish peroxidase (HRP) in the presence and absence of β-lg. There was a five-fold increase in degraded HRP transport in sensitized (39.4 ± 6.6 pmoles/h.cm2) versus control (7.37 ± 2.51 pmoles/h.cm2) animals, with and without (β-lg challenge. Serosally applied β-lg enhanced transport of intact HRP in sensitized but not in control animals. These results indicate that sensitization of guinea pigs to cow's milk permanently increases endocytic and electrogenic activities. The challenge with β-lg induced a further transient rise in Isc and increased intact HRP transport.

Effects of platelet-activating factor on ion transport in isolated rat jejunum.

In isolated normal rat jejunum, platelet-activating factor (PAF) induced a dose-dependent increase in short-circuit current (Isc) that was reduced in chloride-free buffer and inhibited by the Cl- channel blocker, diphenylamine-2-carboxylate. An immediate rise in Isc (early phase) occurred that fell to a new elevated base line by 15 min (late phase). These responses to PAF occurred only when experiments were conducted at or before approximately 9 A.M. Early phase responses were blocked by the specific PAF antagonists, BN52021 and WEB2086, and were inhibited by the neurotoxin, tetrodotoxin. Early and late phases were also reduced by cyclooxygenase inhibitors and by doxantrazole, a mast cell stabilizing drug. However, histamine and serotonin antagonists were ineffective. We conclude that PAF causes changes in ion transport that include Cl- secretion and acts on the epithelium possibly via an intermediate cell and enteric nerves. In addition, known PAF receptors are involved in one component of the response that appears to follow a circadian rhythm.

Luminal antisecretory effects of a beta-casomorphin analogue on rabbit ileum treated with cholera toxin.

Because the physiologic significance of the presence of the opioid peptides beta-casomorphins (beta-CMs) in enzymatic digestion of milk proteins is still undetermined, the effect of the nonmetabolized beta-CM analogue beta-[DAla2,4,Tyr5]CM-5-NH2 on water and electrolyte movements was studied in vivo and in vitro in rabbit ileum untreated or treated with cholera toxin (CT). When this analogue was introduced in vivo at a concentration of 10(-3) M into the lumen of rabbit ileal loops, it significantly stimulated net water absorption in untreated loops and reduced net water secretion in CT-treated loops. In vitro addition of this analogue (10(-4) M) to the serosal compartment of untreated ileum in an Ussing chamber reduced Isc (delta Isc = 0.44 +/- 0.05 muEq.h-1/cm2) and stimulated net Na and Cl absorption to the same extent. In CT-treated ileum, both serosal (10(-4) M) and mucosal (5.10(-4) M) addition of the analogue did not further modify the rise in Isc caused by CT but also stimulated net Na and Cl absorption. On the mucosal side, the effect of the analogue was accompanied by its transfer from the luminal to the blood side of the tissue. The transferred analogue was intact as shown by HPLC (Jm----s = 2.4 +/- 0.8 nmol.h-1/cm2). These results demonstrated that the beta-CM analogue stimulates intestinal absorption of electrolytes in rabbit ileum both in the basal state and after its stimulation by CT.(ABSTRACT TRUNCATED AT 250 WORDS)