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PubMed1992

Papaverine reduces the sodium permeability of the apical membrane and the potassium permeability of the basolateral membrane in isolated frog skin.

The effect of papaverine, an inhibitor of the phosphodiesterase responsible for breakdown of cAMP, on the transepithelial sodium transport across the isolated frog skin was investigated. Serosal addition of papaverine caused initially an increase in the short-circuit current (SCC), a doubling of the cellular cAMP content and a depolarization of the intracellular potential under SCC conditions (Vscc). The initial increase in the SCC was followed by a pronounced decrease both in the SCC and in the natriferic action of antidiuretic hormone (ADH), but papaverine had no inhibitory effect on the ability of ADH to increase the cellular cAMP content. As SCC declines, no hyperpolarization was observed. The I/V relationship across the apical membrane during the inhibitory phase, revealed that papaverine reduces the sodium permeability of the apical membrane (PNaa) as well as intracellular sodium concentration. These observations and the previously noted effect of papaverine on Vscc indicates that papaverine must have an effect on the cellular Cl or K permeability. The basolateral Na,K,2Cl cotransporter was blocked with bumetanide, which should bring the cellular chloride in equilibrium. Bumetanide had no effect on basal SCC and Vscc. When papaverine was added to skins preincubated with bumetanide, the effect of papaverine on SCC and Vscc was unchanged. Therefore, the depolarization of Vscc, observed during the papaverine-induced inhibition of the SCC, must be due to a reduction in the cellular K permeability. In conclusion, it is suggested that papaverine reduces the sodium permeability of the apical membrane and the potassium permeability of the basolateral membrane of the frog skin epithelium.

Animals↗
PubMed2006

Osmotic water permeability of plasma and vacuolar membranes in protoplasts I: high osmotic water permeability in radish (Raphanus sativus) root cells as measured by a new method.

Intra- and transcellular water movements in plants are regulated by the water permeability of the plasma membrane (PM) and vacuolar membrane (VM) in plant cells. In the present study, we investigated the osmotic water permeability of both PM (P ( f1)) and VM (P ( f2)), as well as the bulk osmotic water permeability of a protoplast (P ( f(bulk))) isolated from radish (Raphanus sativus) roots. The values of P ( f(bulk)) and P ( f2) were determined from the swelling/shrinking rate of protoplasts and isolated vacuoles under hypo- or hypertonic conditions. In order to minimize the effect of unstirred layer, we monitored dropping or rising protoplasts (vacuoles) in sorbitol solutions as they swelled or shrunk. P ( f1) was calculated from P ( f(bulk)) and P ( f2) by using the 'three-compartment model', which describes the theoretical relationship between P ( f1), P ( f2) and P ( f(bulk)) (Kuwagata and Murai-Hatano in J Plant Res, 2007). The time-dependent changes in the volume of protoplasts and isolated vacuoles fitted well to the theoretical curves, and solute permeation of PM and VM was able to be neglected for measuring the osmotic water permeability. High osmotic water permeability of more than 500 mum s(-1), indicating high activity of aquaporins (water channels), was observed in both PM and VM in radish root cells. This method has the advantage that P ( f1) and P ( f2) can be measured accurately in individual higher plant cells.

Cell Membrane Permeability↗
PubMed1997

Phase behavior and permeability properties of phospholipid bilayers containing a short-chain phospholipid permeability enhancer.

The thermodynamic phase behavior and trans-bilayer permeability properties of multilamellar phospholipid vesicles containing a short-chain DC10PC phospholipid permeability enhancer have been studied by means of differential scanning calorimetry and fluorescence spectroscopy. The calorimetric scans of DC14PC lipid bilayer vesicles incorporated with high concentrations of DC10PC demonstrate a distinct influence on the lipid bilayer thermodynamics manifested as a pronounced freezing-point depression and a narrow phase coexistence region. Increasing amounts of DC10PC lead to a progressive lowering of the melting enthalpy, implying a mixing behavior of the DC10PC in the bilayer matrix similar to that of a substitutional impurity. The phase behavior of the DC10PC-DC14PC mixture is supported by fluorescence polarization measurements which, furthermore, in the low-temperature gel phase reveal a non-monotonic concentration-dependent influence on the structural bilayer properties; small concentrations of DC10PC induce a disordering of the acyl chains, whereas higher concentrations lead to an ordering. Irreversible fluorescence quench measurements demonstrate a substantial increase in the trans-bilayer permeability over broad temperature and composition ranges. At temperatures corresponding to the peak positions of the heat capacity, a maximum in the trans-bilayer permeability is observed. The influence of DC10PC on the lipid bilayer thermodynamics and the associated permeability properties is discussed in terms of microscopic effects on the lateral lipid organization and heterogeneity of the bilayer.

Calorimetry, Differential Scanning↗
PubMed2006

Conformational flexibility, internal hydrogen bonding, and passive membrane permeability: successful in silico prediction of the relative permeabilities of cyclic peptides.

We report an atomistic physical model for the passive membrane permeability of cyclic peptides. The computational modeling was performed in advance of the experiments and did not involve the use of "training data". The model explicitly treats the conformational flexibility of the peptides by extensive conformational sampling in low (membrane) and high (water) dielectric environments. The passive membrane permeabilities of 11 cyclic peptides were obtained experimentally using a parallel artificial membrane permeability assay (PAMPA) and showed a linear correlation with the computational results with R(2) = 0.96. In general, the results support the hypothesis, already well established in the literature, that the ability to form internal hydrogen bonds is critical for passive membrane permeability and can be the distinguishing factor among closely related compounds, such as those studied here. However, we have found that the number of internal hydrogen bonds that can form in the membrane and the solvent-exposed polar surface area correlate more poorly with PAMPA permeability than our model, which quantitatively estimates the solvation free energy losses upon moving from high-dielectric water to the low-dielectric interior of a membrane.

Amino Acid Sequence↗
PubMed1998

Effects of vascular permeability factor on the permeability of cultured endothelial cells from brain capillaries.

The effects of vascular permeability factor (VPF) on the blood-brain barrier (BBB) permeability were studied in cultured capillary endothelial cells from bovine brain. When VPF was added to the abluminal or brain side of the BBB, a rapid increase in the BBB permeability was observed. This increase in the BBB permeability is dependent on the concentration of the VPF used and is reversible on removal of the VPF. Contrarily, when VPF was added to the luminal or blood side of the BBB, no increase in the BBB permeability was observed. These results suggest that the receptors for VPF are located on the abluminal side of the BBB.

Animals↗
PubMed1993

Vascular endothelial growth factor/vascular permeability factor expression in the rat uterus: rapid stimulation by estrogen correlates with estrogen-induced increases in uterine capillary permeability and growth.

In the uterus, estrogen causes a rapid increase in microvascular permeability, followed later by growth of the endometrium, including the richly vascular stroma. Vascular endothelial growth factor/vascular permeability factor (VEGF/VPF or VEG/PF) is an angiogenic protein that is not only a specific mitogen for endothelial cells, but also a potent stimulator of microvascular permeability. Because of these properties, it seems likely that VEG/PF might mediate estrogen-induced increases in uterine vascular permeability and blood vessel growth. Therefore, we determined whether the gene for VEG/PF is expressed in the rat uterus and if mRNA abundance is regulated by steroid hormones, using reverse transcription-polymerase chain reaction. The VEG/PF gene is alternatively spliced and gives rise to three transcripts coding for proteins of 188, 164, and 120 amino acids, which, in turn, form the active dimeric factors. Transcripts for VEG/PF mRNAs were detected in the uterus of the rat by reverse transcription-polymerase chain reaction. The mRNAs for the VEG/PF164 and VEG/PF120 subunits were the dominant forms expressed. Treatment with both estradiol (E2) and estriol (E3) rapidly induced an increase in the level of the two smaller transcripts. The increase was detectable as early as 0.5-1 h and peaked at 2 h. Levels of the two smaller transcripts then declined, but remained above control levels for 24 h. The degree of stimulation of VEG/PF mRNA levels was 8-fold at 2 h. VEG/PF188 mRNA levels were higher by 6 h compared to control values. The increase in VEG/PF mRNA levels in response to E2 was not contingent upon de novo protein synthesis, as it was not blocked by cycloheximide. The increase occurred as rapidly as that of the mRNA for Zif268, an estrogen-induced transcription factor. Progesterone also stimulated the expression (at 6 h) of VEG/PF164 and VEG/PF120, but not that of VEG/PF188. We conclude that the VEG/PF gene is expressed in the rat uterus, and that mRNA levels are rapidly enhanced by estrogen. This response suggests that VEG/PF may be involved in the estrogen-induced increase in permeability and proliferation of uterine blood vessels. The identification of VEG/PF as a primary response gene also suggests that VEG/PF expression may be a prerequisite for the subsequent expression or action of other growth factors in the uterus.

Animals↗
PubMed2001

Small bowel permeability in diagnosis of celiac disease and monitoring of compliance of a gluten-free diet (gut permeability in celiac disease).

BACKGROUND & AIM: Celiac disease is an autoimmune disease with the damage of the intestinal barrier. The aim of study was to measure gut permeability in patients with untreated celiac disease and during treatment with a gluten free diet. METHODS: 30 patients with celiac disease before and during treatment with gluten-free diet were investigated, 30 patients without organic damage of the gastrointestinal tract served as control. Small bowel permeability was measured using lactulose/mannitol and lactulose/D-xylose ratios. The saccharides were examined in the 5 hours collected urine using capillary gas chromatography. RESULTS: Small bowel permeability (indices lactulose/mannitol and/or lactulose/xylose) increased significantly in patients with untreated celiac disease. 23 patients were followed up before and during treatment with a gluten-free diet 2-6 months after beginning of this treatment and small bowel permeability (measured as indices lactulose/mannitol and/or lactulose/xylose) significantly decreased. CONCLUSION: Small bowel permeability test is a non-invasive test suitable for the diagnosis of celiac disease and monitoring of compliance to a gluten-free diet in these patients.

Adult↗
PubMed1977

Measurements of intestinal permeability using low molecular weight polyethylene glycols (PEG 400). II. Application to normal and abnormal permeability states in man and animals.

A new approach to the measurement of intestinal permeability using low molecular weight polyethylene glycols (PEG 400) has been applied to the study of normal and abnormal permeability states in man and animals. Successful assessments of gastric, jejunal, ileal, and colonic permeability suggest that the technique has application to any area of the gastrointestinal tract. The method has shown that bile acids alter mucosal permeability in the stomach and colon and that celiac sprue is associated with decreased intestinal permeability. These examples illustrate the potential of the technique for studying the function of diseases intestine in man.

Adult↗
PubMed2007

Impact of cremophor-EL and polysorbate-80 on digoxin permeability across rat jejunum: delineation of thermodynamic and transporter related events using the reciprocal permeability approach.

The effect of Cremophor-EL (Cr-EL) and polysorbate-80 (PS-80) on the transepithelial permeability of digoxin (DIG) has been evaluated using the reciprocal permeability approach to delineate thermodynamic and transporter related events. Permeability data were corrected for solubilization using the micellar association constant (Ka) obtained from Papp data generated in the presence of the nonspecific ATPase inhibitor sodium orthovanadate. In the presence of mucosal Cr-EL, a concentration dependent decrease in serosal-mucosal (S-M) and increase in M-S transport was observed. Whilst serosal Cr-EL resulted in a reduction in S-M DIG transport, no impact on M-S transport was apparent. For PS-80, the presence of either serosal or mucosal surfactant led to a decrease in secretory (S-M) DIG transport, however no effect on absorptive transport was evident. The data confirm the potential P-gp inhibitory effects of Cr-EL, but suggest that in contrast to Cr-EL, PS-80 is not a potent inhibitor of P-gp and is incapable of increasing absorptive drug transport, at least in excised rat intestinal tissue and at the concentrations tested. The data are also consistent with the involvement of additional transporters (both apical and basolateral) in the intestinal permeability of DIG, although more definitive data is required to confirm this possibility.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1985

Passive cation permeability of turtle colon: evidence for a negative interaction between intracellular sodium and apical sodium permeability.

The role of intracellular sodium in the regulation of apical sodium permeability was investigated in an electrically "tight" epithelium, the turtle colon. In the presence of low mucosal sodium (3 mM) and serosal ouabain, an inhibitor of the basolateral sodium pump, the apical membrane retained a substantial amiloride-sensitive, sodium conductance and the basolateral membrane exhibited a barium-sensitive potassium conductance in parallel with a significant sodium (and lithium) conductance. In the presence of a high mucosal sodium concentration (114 mM), however, inhibition of active sodium absorption by ouabain led to a disappearance of the amiloride-sensitive, transepithelial conductance that was due, at least in part, to a virtual abolition of the apical sodium permeability. Two lines of evidence indicate that this permeability decrease was dependent upon an increase in intracellular sodium content. First, raising the mucosal sodium concentration from 3-114 mM in the presence of ouabain reversibly inhibited the amiloride-sensitive conductance. The time course of the decline in conductance paralleled the apparent intracellular accumulation of sodium in exchange for potassium, which was monitored as a transient deflection in the amiloride-sensitive, short-circuit current. Second, the inhibitory effect of mucosal sodium-addition was markedly attenuated by serosal barium, which prevented the accumulation of sodium by blocking the electrically coupled, basolateral potassium exit. These results support the notion of a "negative feedback" effect of intracellular sodium on the apical sodium permeability.

Amiloride↗
PubMed1989

The effect of ferricyanide with iodoacetate in calcium-free solution on passive cation permeability in human red blood cells: comparison with the Gardos-effect and with the influence of PCMBS on passive cation permeability.

Freshly prepared human red blood cells incubated with 5 mM ferricyanide, 0.2 mM iodoacetate and 2 mM adenosine in the presence of 5 mM EGTA demonstrate comparable increases in Na+ and K+ permeability (ferricyanide effect). This effect is unrelated to the Ca2+-activated K+ channel (Gardos effect) since influx of Ca2+ from outside the cell is excluded. Also this effect is different from the non-specific Na+ and K+ permeability change elicited by PCMBS. These differences become obvious by using various reagents. For example, A23187 and quinidine exert opposite effects in Gardos and ferricyanide experiments, where A23187 and atebrin react oppositely in the latter and in PCMBS experiments. The ferricyanide effect described here does not involve formation of nonspecific channels. The change in Na+ permeability separately from K+ permeability under certain circumstances suggests a more specific effect.

4-Chloromercuribenzenesulfonate↗
PubMed1978

A protease-like permeability factor in the guinea pig skin. 2. In vitro activation of the latent form permeability factor by weakly acidic phosphate buffer.

Conditions for the in vitro activation of the latent form of a protease-like permeability factor in the pseudoglobulin fraction from guinea pig skin were examined. (1) The factor was activated by dialysis against 67 mM phosphate buffer at pH 5.8--6.4, not at pH 7.0--8.0. (2) High salt concentration (200 mM or greater phosphate buffer or 67 mM phosphate buffer containing 200 mM or greater KCl or NaCl) prevented the activation at pH 6.2. (3) High osmotic pressure (sucrose at 1 M) did not affect activation at pH 6.2. (4) Reconversion of the activated permeability factor into an inactive form was not observed under high salt conditions, under which the latent permeability factor was stable in its own form. (5) The molecular size of the latent permeability factor was estimated as approx. 80 000 by Sephadex G-100 gel filtration at high salt concentration.

Animals↗
PubMed2004

The dynamic placenta: II. Hypothetical model of a fetus driven transplacental water balance mechanism producing low apparent permeability in a highly permeable placenta.

In vitro and isotopic studies in vivo have reported the paradox that the human placenta is highly permeable, water exchanging at 3.6 litres per hour at 35 weeks of gestation, but clinical measurements in vivo show net transfer is minimal, around 2 ml/day. Current theories are based on osmotic pressure balances, but changes in maternofetal hydrostatic pressure change much faster than osmotic factors could respond. An alternative explanation might be that net transfer is not in fact the result of passive mechanisms, but is actively controlled by the fetus itself. The fetus is well equipped to monitor changes in blood volume, such as via sensors in venous and atrial stretch receptors to control ANF and hence urine production. Transplacental water regulation requires modification of transvillus pressures. Placental sub-chorial arteries and veins (of extra-embryonic origin) have different sensitivities from fetal body tissues to some vasoactive substances, and stem villous veins have unusually well developed vascular smooth muscle. It is thus theoretically possible for the fetus to modify subchorial venous resistances, and hence villous capillary pressure with a suitable circulating placental venous constrictive agent. A computer modelling study was undertaken using a fictitious placental venous constricting agent "fictensin", considered to be released by the fetus in proportion to disturbance of vascular volume. The effective placental permeability fell in proportion to the tightness of this fetal control mechanism, suggesting that the apparent placental permeability measured in vivo is a measure of fetal control, not true permeability. However, the range of compensatory pressures that the fetus can produce by this means is limited and failure of such a mechanism could allow flooding or dehydration of the feto-placental unit. This may shed new light on disorders such as polyhydramnios and fetal hydrops.

Animals↗
PubMed2004

Corneal permeability in a redesigned corneal holder for the bovine cornea opacity and permeability assay.

The bovine cornea opacity and permeability assay (BCOP) is a proposed alternative to the Draize rabbit test for potential eye irritants. In the standard BCOP, bovine corneas are mounted in a holder on a flat surface between two identical chambers. The flat configuration of the standard holder does not conform to the normal curved shape of the bovine cornea and it comes into direct contact with the cornea tissue. Mounting corneas in this holder causes extensive damage to both epithelial and endothelial corneal cell layers. Our laboratory has designed a new holder that allows the cornea to maintain its natural curvature and does not damage the cornea. Previous tests, using both the new and standard holders, and comparing corneal opacity, hydration and endothelial morphology, have shown that the new holder is a significant improvement over the standard holder. The present study extends the comparisons of the new and standard holders to measurement of corneal fluorescein permeability. The permeability (ng/cm(2)/min) of intact corneas, corneas with no epithelium, and corneas treated with 1% NaOH, isopropanol, acetone, 30% trichloroacetic acid or 30% sodium dodecysulfate for either 1 or 10 min was determined by measuring fluorescence of samples taken from the endothelial chamber after 90 min epithelial exposure to 0.04% sodium fluorescein. In all trials, the redesigned holders yielded not only lower permeability measurements but also decreased measurement variability. The data provide further evidence that the new holder is an improvement over the standard holder and should be incorporated into a new protocol for the BCOP.

Animal Testing Alternatives↗
PubMed1998

Inhibition by tranilast of vascular endothelial growth factor (VEGF)/vascular permeability factor (VPF)-induced increase in vascular permeability in rats.

We studied the effects of tranilast, an anti-allergic and anti-proliferative drug in clinical use, on VEGF/VPF-induced vascular permeability in a rat air pouch model. A large increase in vascular permeability was induced by injection of 4 ml of a 100 ng/ml VEGF/VPF solution into the preformed air pouch. Over a 15-min period, tranilast inhibited the VEGF/VPF-induced vascular permeability in a dose-dependent manner. This result suggests that tranilast, which we recently found to inhibit VEGF/VPF-induced angiogenesis, could also improve VEGF/VPF-dependent increases in vascular permeability.

Animals↗
PubMed2002

Regarding the sources of data analyzed with quantitative structure-skin permeability relationship methods (commentary on 'Investigation of the mechanism of flux across human skin in vitro by quantitative structure-permeability relationships').

We investigated the sources of data used in recently published predictive models of skin permeability. It was found that skin permeability coefficients for 63 compounds are poorly documented. We hypothesized that these coefficients were calculated using the simple two variable, three parameter 'Potts and Guy' regression equation and hence were not derived from experimental measurements. We therefore examined the distribution of residuals of these reported coefficients compared with the Potts and Guy predictions. The residuals cannot be described by a normal distribution. A substantial (51%) number of residuals equaled 0.00. Further analysis demonstrated that 89% (56 out of 63) of the skin permeability coefficients can be explained as being calculated by the Potts and Guy equation using different documented octanol-water partition coefficients, and/or transcription errors. The results strongly suggest that these 63 skin permeability coefficients are calculated and not experimentally determined-a conclusion subsequently confirmed by one of the developers of the data set. Continued use of these data would lead to biased model selection, underestimation of experimental variability, and overestimation of model predictive ability.

Linear Models↗
PubMed2006

Parallel artificial membrane permeability assay: a new membrane for the fast prediction of passive human skin permeability.

This work was devoted to the search for new artificial membranes allowing a rapid evaluation of passive human skin permeation of compounds with a parallel artificial membrane permeability assay (PAMPA). Effective permeability coefficients (Pe) determined for a set of compounds using the PAMPA technique with isopropyl myristate (IPM) and silicone oil, alone or in mixture, were compared to the corresponding human skin permeability coefficient values (Kp). A good correlation between Pe and Kp was found for compounds tested through a membrane consisting of 70% silicone and 30% IPM. Moreover, positive correlation between the membrane retention of compounds and stratum corneum/water partition coefficients (PSC) was established. These results showed that this new artificial membrane, defined as PAMPA-skin, is able to mimic the main barrier properties of human stratum corneum and can be used for the fast prediction of passive human skin permeability coefficients.

1-Octanol↗
PubMed1989

Cl- permeability of human sweat duct cells monitored with fluorescence-digital imaging microscopy: evidence for reduced plasma membrane Cl- permeability in cystic fibrosis.

Salt reabsorption by the human sweat duct is markedly reduced in cystic fibrosis (CF). We used fluorescence-digital imaging microscopy in combination with a halide-specific fluorescent dye [6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ)] to determine if this defective salt reabsorption is referable to a reduced plasma membrane Cl- permeability of the epithelial cells that line the sweat duct. Sweat duct cells were cultured from explants of normal and CF reabsorptive duct and loaded with SPQ, the fluorescence of which is specifically quenched by halide ions (Br- greater than Cl-) and provides a relative index of intracellular halide concentration. Two lines of evidence indicate that normal sweat duct cells exhibit a substantial permeability to Cl- and Br-. First, the replacement of extracellular Cl- with an impermeant anion (i.e., gluconate) resulted in a rapid and reversible increase in the intracellular fluorescence, as expected if the cells rapidly lost Cl- to the extracellular media. Second, the replacement of extracellular Cl- with Br- resulted in a rapid and reversible quenching of the intracellular fluorescence, as expected if the cells accumulated Br- (a more effective quencher of SPQ fluorescence) in exchange for Cl-. The rate of fluorescence change that was induced by either maneuver was inhibited by the Cl- channel blocker, diphenylamine-2-carboxylate (10 microM). Moreover, CF cells exhibited markedly reduced rates of fluorescence change in response to either maneuver. Our results document the utility of this imaging strategy for assessing the Cl- permeabilities of individual epithelial cells that are affected in cystic fibrosis and indicate that the defective salt reabsorption by the CF sweat duct is referable, at least in part, to a reduced plasma membrane Cl- permeability of sweat duct cells.

Adolescent↗