Transmural Electrical Potential Difference Research Articles

David Wingate.

David Wingate.

Effects of ethanol and extract of cigarette smoke on the rabbit buccal mucosa

The combination of smoking and drinking alcohol has a high association with diseases of squamous epithelium within the human oral cavity. Therefore, a study was done to assess the impact of these agents alone or in combination on the squamous epithelium using as model the buccal epithelium from rabbit oral cavity. Buccal epithelium was mounted in Ussing chambers to monitor electrical parameters during exposure to ethanol (5-40%) or to Ringer extract of cigarette smoke (EOCS) from one to six cigarettes dissolved in 10 ml Ringer either alone or with combination. Exposure to EOCS reduced in a dose dependent manner above 2 cigarettes/10 ml transmural electrical potential difference (PD), short-circuit current (I(sc)), increased transmural electrical resistance (R). Morphology showed from generalize tissue edema to patchy necrosis with the increasing concentrations. Ethanol alone raised PD, I(sc) and R at lower concentrations (5%) and lowered PD, I(sc) and R at higher concentrations (40%). The combination of 5% ethanol, EOCS-1cigarette/10 ml reduced PD, I(sc) by 58% and increased R by 29%. Unlike exposure to 5% EtOH and EOCS-1, 10% EtOH combined with EOCS-1 produces a harmful effect by dropping PD and I(sc). Both, simultaneous, and sequential, use of these agents enhanced their negative impact on these parameters. The enhancement of these effects are not due to solubulization of additional tobacco products by EtOH or by or by EtOH enhancing smoking noxious effect. Histopathologic damage needs higher concentrations of ethanol and EOCS combination and changes were more profound compared to the sum of the isolated effects of both agents.

Electrical parameters and ion species for active transport in human esophageal stratified squamous epithelium and Barrett's specialized columnar epithelium

The human esophagus is lined by stratified squamous epithelium (ESSE), and in some subjects with reflux disease the distal esophagus becomes lined by Barrett's specialized columnar epithelium (BSCE). ESSE and BSCE differ both histologically and functionally, the latter evident by differences in their in vivo transmural electrical potential difference (PD), ESSE averaging -15 mV and BSCE being greater than -25 mV. In this report we examine the basis for this difference in PD. This is done by mounting endoscopic biopsies of ESSE from 25 subjects without esophageal disease and BSCE from 19 with Barrett's esophagus in mini-Ussing chambers for electrical recordings basally and after bathing solution ion replacement. The results show that the PD of human ESSE reflects a low level of active ion transport (5.1 +/- 0.8 muA/cm(2)) combined with a high level of tissue (electrical) resistance (344 +/- 34 Omega.cm(2)) and that of BSCE reflects a high level of active transport (43.6 +/- 11.6 muA/cm(2)) combined with a low level of resistance (69 +/- 8 Omega.cm(2)). Furthermore, active transport in ESSE was principally due to sodium absorption whereas in BSCE it was equally divided between sodium absorption and anion (chloride/bicarbonate) secretion, the latter through an apical membrane, 4-acetamido4'-isothiocyano-2,2'-stilbenedisulfonic acid-sensitive anion channel. As an anion-secreting tissue with bicarbonate secretory capacity more than fivefold greater than ESSE, BSCE is better suited than ESSE for defense of the esophagus against reflux disease.

Physiological and morphological effects of alendronate on rabbit esophageal epithelium

Alendronate, an aminobisphosphonate, produces as a side effect a topical (pill induced) esophagitis. To gain insight into this phenomenon, we assessed the effects of luminal alendronate on both esophageal epithelial structure and function. Sections of rabbit esophageal epithelium were exposed to luminal alendronate at neutral or acidic pH while mounted in Ussing chambers to monitor transmural electrical potential difference (PD), short-circuit current (I(sc)), and resistance (R). Morphological changes were sought by light microscopy in hematoxylin and eosin-stained sections. Impedance analysis was used for localization of alendronate-induced effects on ion transport. Luminal, but not serosal, alendronate (pH 6.9-7.2), increased PD and I(sc) in a dose- and time-dependent manner, with little change in R and mild edema of surface cell layers. The changes in I(sc) (and PD) were reversible with drug washout and could be prevented either by inhibition of Na,K-ATPase activity with serosal ouabain or by inhibition of apical Na channels with luminal acidification to pH 2.0 with HCl. An effect on apical Na channel activity was also supported by impedance analysis. Luminal alendronate at acidic pH was more damaging than either alendronate at neutral pH or acidic pH alone. These data suggest that alendronate stimulates net ion (Na) transport in esophageal epithelium by increasing apical membrane sodium channel activity and that this occurs with limited morphological change and no alteration in barrier function. Also alendronate is far more damaging at acidic than at neutral pH, suggesting its association with esophagitis requires gastric acid for expression. This expression may occur either by potentiation between the damaging effects of (refluxed) gastric acid and drug or by acid-induced conversion of the drug to a more toxic form.

Transmural potential changes associated with the in vitro absorption of theanine in the guinea pig intestine.

Theanine, L-N-ethylglutamine, is one of the major components of amino acids in Japanese green tea. To characterize the mode for intestinal absorption of theanine, the ionic dependency and kinetic properties of the theanine- and glutamine-evoked transmural electrical potential difference changes (delta PD) were investigated in vitro by using everted sacs prepared from the guinea pig ileum. Both theanine and glutamine applied to the luminal side induced dose-dependent increases in delta PD (increase in serosal positive value). The theanine- and glutamine-evoked delta PD values conformed to the Michaelis-Menten relationship, with delta PDmax not being different, whereas the half-saturation concentration was lower for glutamine (3.1 +/- 0.2 mM) than for theanine (21.4 +/- 0.6 mM). The theanine-evoked delta PD value was much smaller when theanine was applied in the presence of glutamine than when applied alone. The theanine- and glutamine-evoked delta PD values were both inhibited by removing Na+ from the luminal solution. These results suggest that the intestinal absorption of theanine and glutamine is mediated by a common Na(+)-coupled co-transporter in the brush-border membrane, the affinity of which is lower for theanine than for glutamine.

Effect of glycochenodeoxycholic acid on unidirectional transepithelial fluxes of electrolytes in the perfused human ileum.

Perfusion studies of the terminal ileum were performed in healthy volunteers to define the influence of a dihydroxy bile acid, glycochenodeoxycholic acid (GCDC) 2.5 mmol/l, on the mechanisms of electrolyte transport. Net movements of water and electrolytes, bidirectional fluxes of sodium, potassium, and chloride, and the transmural electrical potential difference (PD) were measured simultaneously. The results supported the notion of an active mechanism for sodium and chloride transfer. GCDC evoked net secretion of water and electrolytes, and decreased the mucosa to serosa flux of chloride, There was a tendency that the latter also applied to sodium and potassium. During bile acid perfusion active secretion of chloride occurred. PD was 16 +/- 4 mV, lumen negative, and was not influenced by GCDC. In conclusion, we propose a model for the GCDC effect.

Secretory reflexes in ileum and jejunum: absence of remote effects

We have tested the hypothesis that luminal secretagogues initiate neural reflexes that alter ion transport in small intestinal segments proximal or distal to the site of the secretory stimulus. Effects of secretagogues that act by different mechanisms were studied in vitro by measuring short circuit current ( I sc) of ileum or jejunum mounted in a unique flux chamber while proximal mucosa in neural continuity with the tissue was perfused with secretagogues (Na deoxycholate. Escherichia coli STa. 5-hydroxytryptamine, theophylline) or was stimulated electrically (EFS). No proximal stimulus affected distal I sc. We also studied in vivo adjacent segments of ileum or jejunum in neural continuity but with unconnected lumens. In anesthetized rabbits, we measured transmural electrical potential difference and fluid movement (Phenol red marker). Stimulation of proximal segments of ileum or jejunum with STa, or of ileum with 5-HT or Na deoxycholate did not affect distal transport. Stimulation of distal segments of ileum and jejunum with STa or 5-HT, or of jejunum with Ha deoxycholate did not affect proximal transport. We conclude that the secretion caused by luminal secretagogues in the rabbit small intestine is limited to the area of stimulation.

The acute effects of smokeless tobacco on transport and barrier function of buccal mucosa.

The acute effects of smokeless tobacco (ST) on buccal mucosal transport and barrier function were studied by means of in vivo and in vitro techniques. In humans, in vivo exposure to 0.5 g ST transiently increased the transmural electrical potential difference (PD). However, despite continued exposure, PD returned to baseline within 20 min. The mechanisms for these changes were explored by use of dog buccal mucosa mounted in Ussing chambers. Luminal exposure to a Ringer-extract of ST (EOST) increased PD and short-circuit current (Isc) and decreased electrical resistance (R), with changes reversible upon removal of EOST from the bath. Further, radioisotopic fluxes showed that the increase in Isc in EOST-exposed tissues was accompanied by increased absorption of electrolytes (Na, Cl, and other ions), and the decrease in R was accompanied by increased permeability to mannitol. Light microscopy of tissues exposed to EOST showed no morphological changes after exposure to 0.5 g of ST, but after exposure to 1.5-2.5 g of ST, dilated intercellular spaces were identified. Contact of aqueous media with ST led to the release of electrolytes and other soluble compounds into solution. To determine the effect of electrolyte release on buccal function, we exposed mucosae luminally to a solution with ion composition and/or osmolality similar to EOST or to one with an EOST previously dialyzed against Ringer. Solutions with similar ion composition and/or osmolarity changed PD, Isc, and R in a manner similar to EOST, while dialyzed-EOST had no effect. In addition, luminal nicotine produced effects different from EOST, decreasing PD and Isc and increasing R.(ABSTRACT TRUNCATED AT 250 WORDS)

Regional difference in intestinal adaptation after total colectomy as judged by the changes of mucosal Na-K ATPase, cyclic AMP, and transmural potential difference.

Intestinal adaptation and its regional difference after total colectomy were investigated in dogs by measuring mucosal Na-K ATPase, cyclic AMP, and transmural electric potential difference (PD). Twenty-four weeks after the total proctocolectomy, Na-K ATPase activity and PD increased significantly in all intestinal sites, whereas cyclic AMP showed no significant changes. The regional difference in the remaining intestine was examined in the jejunum, ileum, and interposed jejunum (neorectum). Na-K ATPase activity showed no significant regional difference, but the largest increase was found to occur in the ileum. PD also increased markedly in the ileum and there was significant difference between the ileum and other intestinal sites. These facts suggest that the increased active ion transport mediated by mucosal Na-K ATPase and transmural PD in the ileum is closely related to the intestinal adaptation occurring after total colectomy and indicates a greater potential of the ileum for adaptive compensation than either jejunum or neorectum.

AM and PM measurement of gastric potential difference and prostacyclin production in humans: effects of ranitidine.

It has been previously documented that aspirin induced gastric injury of healthy male volunteers was greater in the morning than in the evening. We have also reported that fasting gastric acid secretion rates and gastric retention times for solids were lower in the morning hours of the circadian cycle. We hypothesized, therefore, that defensive factors in the human stomach may exhibit circadian rhythmicity with greater vulnerability to noxious agents during the morning hours. It has also been hypothesized that an antisecretory agent, with reported protective effects, such as Ranitidine, would affect gastric defense mechanisms differently at different times in the circadian cycle. Transmural electrical potential difference (PD) and prostacyclin (PGI2) production by mucosal biopsy specimens were chosen as putative indicators of gastric defensive status. Accordingly, morning (1000) and evening (2200) studies were performed on 10 fasting healthy male subjects with and without oral Ranitidine (150 mg) given 3 hr before oro-gastric intubation for the measurement of PD and the removal of mucosal biopsy samples.(ABSTRACT TRUNCATED AT 250 WORDS)

Aldosterone-induced, amiloride-inhibitable short-circuit current in the avian ileum.

Transmural electrical potential difference, short-circuit current (Isc), and resistance were measured in vitro in the intestines of chickens. In birds maintained on a low-Na diet, there was a rise in Isc in the ileum and the colon but not in the duodenum or jejunum. A substantial portion of this Isc could be inhibited by the Na channel-blocking drug, amiloride. The low-Na diet results in elevated plasma levels of aldosterone and the effects of the diet on intestinal transport could be mimicked by administration of this hormone. The time course of the effects of these two treatments on the ileum and colon were quite different, as a much longer exposure was needed to elicit maximal effects in the ileum than in the colon, possibly due to the presence of two types of adaptation mechanisms. The responses observed may reflect an adaptation of mechanisms for increasing Na transport from the intestine. The presence of such a mechanism in the vertebrate ileum has until recently been in doubt.

Transmural electrical potential difference as an early marker in colon cancer.

It has been suggested that relative depolarization of the cell membrane occurs during carcinogenesis. A study was undertaken to determine the effect on the colonic transmural potential difference in CF1 mice following the administration of a carcinogen, a cytotoxic agent, and bile acids. Depolarization of the distal colonic mucosa was seen in mice treated with the carcinogen but not with the cytotoxic agent, and bile acids caused further depolarization of the mucosa. These experiments support the hypothesis that epithelial depolarization is a putative mechanism in carcinogenesis, which could have important implications for the early detection of colon cancer.

Effect of castration on prostaglandin-mediated changes in membrane potential and prostaglandin synthesis in guinea-pig seminal vesicle tissue.

Arachidonic acid stimulated an increase in transmural electrical potential difference (p.d.) in guinea-pig seminal vesicle tissue in vitro. Pretreatment with indomethacin abolished this response. Indomethacin pretreatment did not prevent the p.d. from increasing in response to theophylline. Changes in p.d. in response to arachidonic acid were greatly attenuated, and the response to theophylline was abolished in seminal vesicle tissue taken from castrated guinea-pigs. Seminal vesicles, aorta and ileum taken from castrated guinea-pigs synthesized and released more prostaglandins than did those from control animals. It is concluded that the effects of arachidonic acid on p.d. are mediated by its metabolism to prostaglandins; the inability of seminal vesicles from castrated animals to respond to arachidonic acid is not a result of a decrease in prostaglandin production, but is more likely a result of other degenerative changes attendant upon castration; and androgens appear to have some regulatory function on prostaglandin synthesis in a variety of tissues.

Changes of sugar-evoked transmural potential differences in intestine of rats with streptozotocin-induced diabetes.

The changes of the transmural electrical potential difference (delta PD) evoked by infusion of glucose, maltose and sucrose and the disaccharidase activities in the everted intestine were studied in diabetic rats. After the induction of diabetes by streptozotocin, delta PDs evoked by sugars and the enzyme activities were observed in the jejunum and ileum. delta PDs evoked by glucose, maltose and sucrose markedly increased both in the jejunum and ileum of diabetic rats. The Kt values for these sugars in diabetic rats were the same as those of control rats. The Vmax values were significantly increased in the ileum of diabetic rats. Maltase and sucrase activities in the ileum increased in diabetic rats. Highly significant linear correlations were found between the delta PDs evoked by glucose and the delta PDs evoked by maltose or sucrose both in the jejunum and ileum of control and diabetic rats. However, delta PDs evoked by maltose and sucrose did not correlate with maltase and sucrase activities in the jejunum. In the ileum, delta PDs evoked by sucrose correlated with the sucrase activity which was very low. These results suggest that the increase of transport of glucose derived from disaccharides in the diabetes induced by streptozotocin is mainly due to the increased activity of the glucose transport system, but not due to the increase of disaccharidase activities.

Effects of neuronal stimulation on mucosal transport in guinea pig ileum.

The effects of electrical stimulation of enteric nerves on mucosal transport parameters were determined in short-circuited flat sheets of guinea pig ileum. Rectangular bipolar stimulus pulses with a duration of 0.5 ms were applied continuously across Ag-AgCl foil electrodes positioned in parallel with the plane of the tissue. Electrical field stimulation (EFS) elicited a biphasic increase in the short-circuit current (Isc) and transmural electrical potential difference. The response to EFS consisted of an initial rapid rise and decline in Isc followed by a slower rise in Isc to a plateau. This response persisted with some decrement. The maximum change in Isc occurred at stimulus strengths of 3.2-5.2 mA and frequencies of 8-10 Hz. The stimulus-evoked increase in Isc was reversibly abolished by 0.1-0.15 microM tetrodotoxin, 22 mM magnesium, and 0.16 mM cadmium. Prior to EFS, net sodium absorption was 2.3 +/- 0.6 mu eq X cm-2 X h-1 in nine animals, and chloride secretion averaged 1.7 +/- 0.8 mu eq X cm-2 X h-1. EFS had no effect on unidirectional or net sodium fluxes or tissue conductance, whereas it increased active chloride secretion from 1.7 +/- 0.8 to 3.9 +/- 0.6 mu eq X cm-2 X h-1. Tetrodotoxin had no effect on resting sodium or chloride fluxes. These results show that EFS evokes an increase in chloride secretion in guinea pig ileum. These results suggest that EFS alters mucosal transport function by activating enteric neurons.

Mucosal function of the perfused ileum in patients with and without diarrhoea and dumping after vagotomy and pyloroplasty.

Perfusion studies of the terminal ileum were performed in patients who had previously had vagotomy and pyloroplasty performed for peptic ulcer disease and who at follow-up study had either diarrhoea or dumping, or no symptoms (the control group). The aim was to elucidate possible pathophysiological factors contributing to the sequelae. Net movements of water, electrolytes, and sugars, unidirectional fluxes of sodium and chloride, and the transmural electrical potential difference (PD) were measured simultaneously with and without the influence of chenodeoxycholic acid (CDC), 1.0 mmol/l, in the intestinal lumen. In patients without symptoms or with dumping, water and sodium chloride were absorbed during the control perfusion. Addition of CDC reduced the absorption or evoked secretion. In patients with diarrhoea the control perfusion disclosed a large spontaneous ileal secretion that was not influenced by CDC. The secretion was mainly due to a reduced mucosa-to-serosa flux of sodium and chloride and, to a lesser extent, to an increased serosa-to-mucosa flux. The diarrhoea patients showed an increased xylose absorption rate, suggesting an increased interepithelial leakiness. No change in PD was demonstrated. The effect of CDC on the unidirectional fluxes was identical in all three groups of patients. In conclusion, our findings suggest that the basic biological mechanisms are identical (and normal) in all patients, but the ileal epithelium of patients with diarrhoea secretes spontaneously and seems to be prestimulated by some at present unidentified agent.

The transmural potential of the newt ureter: evidence for amiloride-sensitive active sodium transport.

Transmural electrical potential difference (PD) of the newt ureter was measured with glass microelectrodes and its nature was studied pharmacologically or by ion replacement experiments. The PD in the presence of standard Ringer solution on both sides averaged -76.0 +/- 1.3 mV, lumen negative. The magnitude of the PD depended on the Na+ concentration of the luminal fluid; the PD increased in a saturable way with increase in the Na+ concentration. Amiloride added to the luminal fluid at 10(-4) M greatly reduced the PD and increased the transmural resistance. The half maximum inhibition was seen at 2.6 x 10(-6) M and the maximum inhibition at 10(-4) M where the PD reduced to 3.8% of control. Amiloride added to the external medium at 10(-4) M had no effect on both the PD and the resistance. Ouabain added to the external medium at 10(-3) M caused a gradual decrease in the PD, which fell to 36% of its initial value after 60 min. The specific transmural resistance (Rt) was estimated by recording spatially decaying electrotonic potentials along the ureter and applying cable analysis. The short-circuit current (Isc) was calculated from the PD and the Rt. The Rt and the Isc averaged 31.7 +/- 7.7 k omega X cm2 and 3.9 +/- 1.5 microA X cm-2, respectively. The results indicate that the epithelial cells of the newt ureter have amiloride-sensitive Na+ channels at the luminal membrane and are transporting Na+ actively, like epithelia of the urinary bladder and the skin of amphibia.

Esophageal Potential Difference Measurements in Esophageal Disease

To determine if esophageal transmural electrical potential difference measurements are of use for evaluating esophageal disease, we recorded potential difference in 129 patients with one or more of the following: heartburn, dysphagia, and chest pain. All potential difference studies were performed at the time of esophageal manometry using a Ringer-perfused catheter technique which yields accurate and reproducible results in healthy subjects. In 103 of the 129 patients, esophageal potential difference measurements could be correlated with findings at manometry, endoscopy, and biopsy. The remaining 26 patients had primary esophageal motor disease and were not biopsied. The results of this investigation showed: (a) that 94% of patients with gross endoscopic lesions have an abnormal esophageal potential difference, (b) that an abnormal esophageal potential difference (found in only 1 of 24 patients with normal mucosa) is highly specific for the presence of esophageal mucosal disease, (c) that the type of potential difference abnormality may suggest the nature of the mucosal abnormality, for example high potential difference with Barrett's esophagus and low potential difference with esophagitis or invasive carcinoma, and (d) that while an abnormal esophageal potential difference is highly sensitive for detecting gross esophagitis (38 of 40 patients), it is less sensitive for diagnosing microscopic esophagitis (8 of 16 patients). Based on these findings we conclude that the measurement of esophageal potential difference at the time of manometry can provide additional valuable information about the state of the esophageal mucosa.

Human Ileal Ion Transport In Vitro: Changes With Electrical Field Stimulation and Tetrodotoxin

To determine whether or not nerves lying in the lamina propria and submucosa affect ion transport by the human intestine, we studied the effects of electrical field stimulation on ion transport across the isolated ileum stripped of muscularis propria. Electrical field stimulation increased transmural electrical potential difference and the short-circuit current, and by increasing the serosal to mucosal movement of chloride (JsmCl), reduced the absorption of Cl (JnetCl). The movement of sodium and the residual ion movement (JnetR) were not affected. The neurotoxin, tetrodotoxin prevented all of the changes caused by electrical field stimulation, but did not prevent the responses to aminophylline, implying that tetrodotoxin affected electrically excitable cells rather than epithelial cells. Aminophylline increased the potential difference and shortcircuit current, reduced tissue conductance (G), and eliminated chloride absorption. In tissues treated with tetrodotoxin, aminophylline also reduced JmsNa, JnetNa, and JnetR; while increasing JsmCl, it augmented the reduction in JmsCl and caused chloride secretion. Atropine reduced baseline short-circuit current and reduced the short-circuit current response to electrical field stimulation. It was concluded that acetylcholine and other chemical transmitters that affect ion transport are released in the submucosa or lamina propria of the human ileum. The identity of the noncholinergic transmitters and the sequence of events leading to their liberation remain to be determined.

Human intestinal potential difference: recording method and biophysical implications.

1. The transmural electrical potential difference (PD) of the intact human small intestine was recorded with close attention to electrical symmetry, shielding from electro-magnetic waves and correction for junction potentials. 2. The PD is -12 mV (mucosa-negative) in the fasting jejunum and ileum and does not change during perfusion with isotonic NaCl. 3. Absorption of Na and Cl appears to be non-electrogenic and the 'resting' PD is probably generated by active anion secretion of fasting intestinal contents. 4. Diffusion potentials during isotonic D-mannitol perfusion indicated higher cation selectivity in the ileum than in the jejunum. 5. The calculated contribution of a free-solution path to total paracellular permeability is 55% in the jejunum but only 15% in the ileum. 6. No 'streaming' potential was detected during osmotic water flow, suggesting that the cation-selectivity of the channels is temporarily inactivated during dilatation of the lateral intercellular space.