Increased Ion Transport Research Articles

Characterization of enteric functional changes evoked by in vivo anti-CD3 T cell activation.

Specific in vivo T cell activation initiated by treatment with anti-CD3 antibodies leads to diarrhea and structural damage of the intestinal mucosa. In this study, the effect of T cell-induced mucosal damage on jejunal epithelial ion transport, muscle contractility, and neuronal ACh release was assessed in Ussing chambers, organ baths, and a specialized perfusion apparatus, respectively. Time-matched control mice received hamster serum containing irrelevant antibodies. Jejunal segments from anti-CD3-treated mice displayed a significantly elevated epithelial baseline short-circuit current (which indicates increased ion transport) and a concomitant reduction in responsiveness to prosecretory stimuli (nerve stimulation, carbachol, and forskolin). Longitudinal smooth muscle displayed altered spontaneous contractile activity, length-tension relationships, and carbachol-stimulated contraction in tissues excised from mice 20 and 40 h posttreatment. Anti-CD3 treatment did not affect stimulated ACh release from myenteric plexus neurons. We conclude that specific T cell activation via anti-CD3 antibody results in dramatic alterations in jejunal epithelial and smooth muscle function. Such T cell-induced changes in intestinal function may contribute to the symptomatology of T cell-mediated enteropathies, including graft-versus-host disease, celiac disease, and idiopathic inflammatory bowel disease.

Investigation of the effects of platelet-activating factor (PAF) on ion transport and prostaglandin synthesis in human colonic mucosa in vitro.

1 We have investigated the effects of platelet-activating factor (PAF), an endogenous mediator of inflammation, on ion transport and prostaglandin synthesis in the human isolated colon. 2 Application of PAF to the serosal surface of human colonic mucosa induced a marked, concentration-dependent increase in ion transport. Mucosal application was without effect. 3 The secretory response to PAF was significantly inhibited by prior application of a specific PAF receptor antagonist WEB 2170, indicating that the response is dependent on PAF receptor activation. 4 The response to PAF was attenuated by prior application of indomethacin or piroxicam, implicating products of the cyclo-oxygenase pathway in the response. 5 The response to PAF was attenuated by the loop diuretic bumetanide, indicating an involvement of chloride ion secretion in the response. 6 Addition of PAF to the serosal surface induced a significant increase in serosal prostaglandin E2 (PGE2), but not 6-oxo-PGF1alpha release. There was no effect on mucosal application of PAF. 7 In summary, we have shown that PAF is a potent secretagogue in isolated preparations of human colon and that the response is dependent on a specific PAF receptor, cyclo-oxygenase products and bumetanide-sensitive chloride ion transport.

Fish gill respiratory cells in culture: a new model for Cl(-)-secreting epithelia.

Primary cultures of sea bass gill cells grown on permeable membranes form a confluent, polarized, functional tight epithelium as characterized by electron microscopy and electrophysiological and ion transport studies. Cultured with normal fetal bovine serum (FBS) and mounted in an Ussing chamber, the epithelium presents a small short-circuit current (Isc: 1.4 +/- 0.3 microA/cm2), a transepithelial voltage (Vt) of 12.7 +/- 2.7 mV (serosal positive) and a high transepithelial resistance (Rt: 12302 +/- 2477 omega x cm2). A higher degree of differentiation and increased ion transport capacities are observed with cells cultured with sea bass serum: numerous, organized microridges characteristic of respiratory cells are present on the apical cell surface and there are increased Isc (11.9 +/- 2.5 microA/cm2) and Vt (25.9 +/- 1.7 mV) and reduced Rt (4271 +/- 568 omega x cm2) as compared with FBS-treated cells. Apical amiloride addition (up to 100 microM) had no effect on Isc. The Isc, correlated with an active Cl- secretion measured as the difference between 36Cl- unidirectional fluxes, was partly blocked by serosal ouabain, bumetanide, DIDS or apical DPC or NPPB and stimulated by serosal dB-cAMP. It is concluded that the chloride secretion is mediated by a Na+/K+/2Cl- cotransport and a Cl-/HCO3- exchanger both responsible for Cl- entry through the basolateral membrane and by apical cAMP-sensitive Cl- channels. This study gives evidence of a functional, highly differentiated epithelium in cultures composed of fish gill respiratorylike cells, which could provide a useful preparation for studies on ion transport and their regulation. Furthermore, the chloride secretion through these cultures of respiratorylike cells makes it necessary to reconsider the previously accepted sea water model in which the chloride cells are given the unique role of ion transport through fish gills.

Comparison of electrospray and inductively coupled plasma sources for elemental analysis with mass spectrometric detection

Both the qualitative and quantitative aspects of electrospray (ES) and ICP sources were investigated using the same mass spectrometer (originally a VG PlasmaQuad I instrument). While it is well established that ICP-MS is a powerful tool for elemental analysis, electrospray ES-MS has recently become popular as a promising elemental analysis technique that can complement the detection capabilities of ICP-MS. The in-house constructed ES source efficiently produced the bare singly charged metal ion and the optimum ES source conditions varied depending on the charge reduction required to attain the singly charged state. The day-to-day signal reproducibility was excellent (5% RSD), and the removal of the photon stop increased ion transport to the detector. A comparison of the figures of merit for Rb, Cs, Ba, V, Cr, Ni, Co, Cu, Zn and U showed that the detection limits obtained by ES-MS (ng ml–1) are only 2–3 orders of magnitude higher than those found using the ICP source. Additionally, Ca was determined in NIST SRM 1643c (Trace Elements in Water) by ES-MS.

Increased ion transport in cultured nasal polyp epithelial cells.

To measure transepithelial bioelectric properties of cultured human nasal polyp and turbinate epithelial cells to test the hypothesis that nasal polyps have increased rates of ion transport. Cohort study. Private referral center. Individuals undergoing surgery for symptomatic nasal obstruction due to polyps caused by cystic fibrosis, nonatopic rhinosinusitis, or allergic rhinosinusitis. Epithelial cells were removed from separated polyp and turbinate samples by protease disaggregation and cultured on permeable collagen matrix supports. Transepithelial potential difference and resistance were measured daily. At the time of maximal transepithelial potential difference, the cultures were mounted in modified Ussing chambers and exposed to a sodium-positive channel blocker (amiloride hydrochloride) and to selected chloride-negative channel agonists (isoproterenol bitartrate, adenosine triphosphate). Maximal transepithelial potential difference, resistance, and equivalent short-circuit current. Bioelectric responses to amiloride, isoproterenol, and adenosine triphosphate. Polyp cultures had higher transepithelial potential difference and equivalent short-circuit current than turbinate cultures. The mediator responses were greater for polyp than for turbinate cultures. Polyp epithelia have increased Na+ absorption and Cl- permeability relative to turbinate epithelia. These results are consistent with the hypothesis that increased epithelial fluid absorption contributes to the development of nasal polyps.

The effect of atrial natriuretic peptides on ion transport by muscle-stripped and full-thickness preparations of rat colon

The effects of three atrial natriuretic peptides (ANPs) have been tested for their ability to increase ion transport in muscle-stripped and intact preparations of rat colon. Anartide (fragment 4–28 of human ANP), human ANP and rat atriopeptin III had no effect on short-circuit current in muscle-stripped preparations. Such insensitivity could not be explained by desensitisation, enzymic destruction or non-specific absorption. Short-circuit current was increased by 8-Br-cGMP and NaNP. The pharmacological activity of anaritide was demonstrated by abolition of phenylephrine-induced tone in rat aortic smooth muscle. In contrast to muscle-stripped colon, intact preparations responded with a tetrodotoxin-sensitive increase in short-circuit current to anaritide even though such preparations were less sensitive to acetylcholine and substance P. These results imply that selective neural damage or the lack of certain nerve pathways, caused by complete muscle stripping, will prevent the pro-secretory effects of anaritide in rat colon.

The routes of penetration of ions and 5-fluorouracil across human skin and the mechanisms of action of terpene skin penetration enhancers

The activation energies for ion movement and 5-fluorouracil diffusion across human epidermis in vitro have been measured to be 4.08 ± 0.16 and 20.6 ± 1.56 (mean ± SE) kcal mol −1, respectively. These data imply that ions travel across human epiderm largely through aqueous shunt routes and that 5-fluorouracil, a polar non-electrolyte, travels mainly through the intercellular lipid bilayers in the stratum corneum. Treatment of human epidermis with terpene penetration enhancers has been shown to increase electrical conductivity. The increase in ion transport suggests that terpenes open new polar pathways across the stratum corneum. A correlation between increases in ion transport and previously reported increases in 5-fluorouracil penetration suggests that terpene enhancers may create micro-pores in the intercellular lipids through which both ions and polar drugs may pass.

Membrane properties of erythrocytes in subjects undergoing multiple blood donations with or without recombinant erythropoietin

To examine the characteristics of 'young' human red cells, we studied blood from seven healthy male volunteers who developed systemic reticulocytosis during a 3-week blood donation period. Each of these subjects donated a total of 6 units (450 ml/unit) of blood (2 units/week for 3 weeks) with subcutaneous recombinant erythropoietin (SC rEPO; 200 U/kg daily for 3 weeks). Two of these subjects were also studied with a similar protocol in the absence of rEPO (4.5 +/- 0.5 units donated). SC rEPO administration was associated with an increased K content of the erythrocyte and with the appearance of hypochromic cells, which were initially normocytic and then became normochromic and microcytic. Measurements of cation transport revealed that, with the exception of the Na-K-Cl cotransport, all the systems studied increased their activities following blood donations with or without SC rEPO. The increase was highest in the K-Cl cotransport (2- and 5-fold for control and rEPO parts of the study, respectively), while the Na-K pump increased slightly in control and 40% with rEPO. The Na-Li countertransport increased 40% and 100% in the control and rEPO parts of the study, respectively. Concomitant with increased ion transport activity, electron microscopic studies of plasma and red cells of subjects receiving SC rEPO showed the presence of circulating exosomes and cytoplasmic multivesicular bodies. The transferrin receptor was detected in the circulating exosomes, thereby providing evidence that, as do nonhuman red cells, maturing human reticulocytes shed exosome-associated transferrin receptors.

Ultrastructural study of the choroid plexus of spontaneously hypertensive rats.

We previously gave an account of an increased ion transport activity in choroid plexus from spontaneously hypertensive rats. We have since examined this organ in scanning and transmission electronic microscopy. In the choroid plexus from young spontaneously hypertensive rats, the epithelial cells showed the following: a partial loss of the brush border and infoldings of basolateral membranes, an increased number of Golgi apparatus, vesicles, and mitochondria, and an activated nucleus. In adult hypertensive rats, the mitochondria had increased in number and tended to fill the cytoplasma while the nuclei had returned to a resting level. These ultrastructural changes furthermore suggest an increased secretory activity in the choroid plexus in spontaneously hypertensive rats.

Arginine vasopressin and atrial natriuretic peptide do not alter ion transport by cultured fetal distal lung epithelium.

Previous studies have shown that i.v. arginine vasopressin (AVP) decreases but does not stop lung fluid secretion in term fetuses not in labor. Although it has been presumed that the response to AVP results from augmented sodium transport, there is controversy whether AVP actually does affect sodium transport in mammalian lung epithelium. To determine if AVP or aldosterone could alone or together augment sodium transport in the perinatal lung, we studied primary cultures of fetal rat distal lung epithelium in Ussing chambers. The short circuit current of these sodium-transporting cells was not affected by the application of either 30 or 300 mU/mL AVP whether or not they were previously exposed to aldosterone (10(-6) M). Aldosterone also did not affect the baseline bioelectric properties. Short circuit current increased in response to 8-bromo cAMP (10(-4) M) and 3-isobutyl-1-methylxanthine (10(-3) M) to levels 169 +/- 16 (SEM) and 172 +/- 7% of respective baseline values. AVP had no effect in cells pretreated with 3-isobutyl-1-methylxanthine. Monolayers also did not respond to atrial natriuretic peptide (10(-11) to 10(-8) M). Monolayers of Na-absorbing A6 renal epithelium did increase short circuit current with either aldosterone or AVP. AVP increased endogenous cAMP levels in A6 but not fetal rat distal lung epithelium cells, suggesting that fetal rat distal lung epithelium lacks V2 receptors. These studies demonstrate that AVP does not increase ion transport in cultured fetal distal lung epithelium although these cells possess the necessary second messenger system.

Thrombin-Induced Membrane Depolarization of Platelets and Its Inhibition by Cetiedil<xref ref-type="fn" rid="fn1"><sup>1</sup></xref>

When 50 microM cetiedil alone was added to a platelet suspension, increase in Na+ content, decrease in K+ content, and depolarization of platelet membrane were observed without change in the intracellular concentration of free Ca2+ ([Ca2+]1) or in the morphology of platelets. The cetiedil-induced depolarization was attenuated by the reduction of extracellular sodium concentration, while sodium transport inhibitors such as procaine and tetrodotoxin failed to modify the depolarization. On the other hand, thrombin caused such changes in platelets as increases in Na+ content, 22Na space and [Ca2+]1, decrease in K+ content, and membrane depolarization. All these changes caused by thrombin were inhibited by cetiedil. It is suggested that cetiedil brought the increased ion transport and subsequent partial depolarization, which might lead to modification of the reaction of platelet membrane induced by thrombin.

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

Electrical parameters: PD, resistance, Isc and amiloride-sensitive-Isc 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.

Response of isolated rat jejunum to angiotensin peptides.

Following intravenous infusion, angiotensin I (ANG I), angiotensin II (ANG II), and angiotensin III (ANG III) enhance Na and water absorption across the jejunum by increasing sympathetic nerve activity. Increased absorption following intravenous infusion of angiotensin peptides may be secondary to an increase in ion transport. To test this hypothesis, the effect of angiotensin peptides on ion and water absorption from the isolated jejunum was studied. At a dose of 100 pM, serosal addition of ANG II stimulated jejunal Na and water absorption. In contrast, mucosal additions of the peptide were ineffective at doses up to 1 microM. ANG II enhanced jejunal absorption in the presence of prazosin. Thus ANG II does not appear to stimulate absorption from the isolated intestine through mediation of sympathetic fibers. ANG III and the substituted analogue [Sar1,Leu8]ANG II stimulated jejunal absorption at a dose of 100 pM. At a dose of 1 nM ANG I also stimulated jejunal absorption. The effect of ANG I on absorption could be prevented by prior treatment of the animals with the converting enzyme inhibitor MK 422. Thus ANG I must first be converted to ANG II to stimulate jejunal absorption.

Structural adaptation in initial collecting tubule following reduction in renal mass

Tubular function studies have shown that nephrectomy leads to a sharp increase in potassium secretion by initial collecting tubules. The aim of this study was to examine the relationship between potassium transport and the cellular morphology of the distal nephron. Clearance and ultrastructural studies were conducted in rats 10 days after 75% nephrectomy. Fractional urinary potassium excretion increased from 23.4 +/- 3.8% in controls to 63.9 +/- 10.1% in nephrectomized animals. Stereological analysis of principal cells in the initial collecting tubule revealed that following nephrectomy the surface density of the basolateral membrane increased from 2.32 +/- 0.12 to 4.11 +/- 0.20 micron2/micron3. While the basolateral membrane surface density of intercalated cells in this segment was not affected by nephrectomy, the luminal membrane surface density increased from 0.29 +/- 0.03 to 0.54 +/- 0.08 micron2/micron3. Ultrastructural changes were not observed in cells of the distal convoluted tubule. These observations demonstrate that cell type specific changes in membrane surface density accompany the adaptive increase in ion transport that occurs with a reduction in renal mass. The enhanced potassium excretion associated with nephrectomy may be partly mediated by a specific increase in the basolateral membrane of principal cells. The increase in luminal membrane of intercalated cells may be related to the increase in acid transport per surviving nephron following nephrectomy.

Reactions of Corn Root Tissue to Calcium

Washing corn (Zea mays L.) root tissue in water causes loss of about one-third of the exchangeable Ca(2+) over the first 10 to 15 minutes. Upon transfer to K(+)-containing solutions, the tissue shows a short period of rapid K(+) influx which subsequently declines. Addition of 0.1 millimolar Ca(2+) decreases the initial rapid K(+) influx, but increases the sustained rate of K(+) and Cl(-) uptake. It was confirmed (Elzam and Hodges 1967 Plant Physiol 42: 1483-1488) that 0.1 millimolar Ca(2+) is more effective than higher concentrations for the initial inhibition, and that Mg(2+) will substitute.The inhibition arises from a mild shock affect of restoring Ca(2+). With 0.1 millimolar Ca(2+) net H(+) efflux is blocked for 10 to 15 minutes and the cells are depolarized by about 30 millivolts. However, 1 millimolar Ca(2+) rapidly produces increased K(+) influx and blocks net H(+) efflux for only a few minutes; blockage is preceded by a brief net H(+) influx which may restore and increase ion transport by reactivating the plasmalemma H(+)-ATPase.Stimulation of electrogenic H(+)-pumping with fusicoccin eliminates the shock responses and minimizes Ca(2+) effects on K(+) influx. Fusicoccin also strongly decreases Ca(2+) influx, but has no effect on Ca(2+) efflux. Ice temperatures and high pH decreased Ca(2+) efflux, but uncoupler and chlorpromazine did not.It is suggested that the inhibitory and promotive actions of Ca(2+) are manifested through decreases or increases in the protonmotive force.

Triethyl tin-induced myelin oedema: an intermediate swelling state detected by X-ray diffraction.

X-ray diffraction was used to probe the effects of triethyl tin (TET) on the periodicity and amount of membrane disorientation in the lamellar myelin from respiring optic and sciatic nerves in vitro as well as from nerves of rats treated in vivo through their drinking water. The diffraction patterns show that in vitro TET at concentrations of 4-100 microM affects C.N.S. but not P.N.S. myelin structure. A planar, concentric membrane array with a 200 A period is detected in the C.N.S.; this ordered, swollen myelin contrasts with the vacuolar and vesicular structure seen in thin-sections in TET-induced oedema. No effects of short-term in vivo treatment with TET are observed in either the C.N.S. or P.N.S. The finding that carbonic anhydrase (CA) inhibitors have no effect on the TET-induced structural changes indicates that the swelling we observe is not related to a CA-dependent process. In comparison, the TET effect is prevented by replacing the mobile ions with isotonic sucrose. We conclude that TET-induced swelling in C.N.S. myelin arises from an increase in ion transport followed by obligatory fluid movement. Further, the ordered, swollen structure we detect may be an intermediate state that exists transiently in vivo in TET intoxication and that precedes the gross swelling and vacuolization usually observed.

Ion transport in tonic and phasic vascular smooth muscle and changes during deoxycorticosterone hypertension.

The washout of radioisotopes was studied in aorta, femoral artery and portal mesenteric vein from controls and rats made hypertensive with injection of deoxycorticosterone (DOC) acetate. Experiments were also conducted on tonic (pulmonary artery) and phasic (portal mesenteric vein) smooth muscle from rabbits. The rate constants for the washout of 42K, 36Cl and 24Na from aorta were significantly elevated in the DOC hypertensives. Acute removal of extracellular K slowed the washout of 24Na by 50%, however, the rate constant for the DOC group remained higher than the controls. Increased turnover of 42K was also observed in the femoral artery and in a vessel (portal mesenteric vein) not exposed to the effects of high blood pressure in the rat. The rate constants for the washout of 42K, 36Cl and 24Na from phasic smooth muscle in rabbit were greater than the tonic type. Ouabain significantly slowed 24Na washout, however, the rate constant for the portal mesenteric vein remained higher than the pulmonary artery. DOC hypertension was associated with increased ion transport at several vascular sites. A similar change was observed for the phasic vascular smooth muscle from normal rabbits. A question is raised whether increased spontaneous activity of arteries reported to accompany DOC hypertension is the consequence of a membrane in transition from tonic to phasic behavior.