Food allergens are protected from degradation during CD23-mediated transepithelial transport

BACKGROUND: CD23 (FcepsilonRII) is expressed by intestinal epithelial cells (IEC) following allergic stimulation and increases the uptake of IgE/allergen complexes. The aim of this study was to further analyze the role of CD23 in the intraepithelial processing of food allergens during transepithelial transport. METHODS: Balb-C mice were sensitized intraperitoneally with horseradish peroxidase (HRP) or beta-lactoglobulin (beta-LG) in the presence of pertussis toxin. In control and sensitized mice, 3H-HRP, intact HRP, or 14C-beta-LG fluxes were measured across jejunal segments mounted in Ussing chambers, in the presence or absence of mucosal anti-CD23 antibodies. HPLC analysis of serosal buffer was performed to detect HRP- or beta-LG-derived radiolabelled metabolites generated during transepithelial transport. RESULTS: In HRP-sensitized mice, 3H-HRP fluxes and intact HRP fluxes (3,836 +/- 476 and 290 +/- 86 ng/h x cm2, respectively) were significantly increased compared to control mice (1,677 +/- 297 ng/h x cm2, p < 0.01, and 106 +/- 23 ng/h x cm2, p < 0.02, respectively). HPLC analysis indicated the presence of intact HRP in the serosal compartment already 10 min after addition of HRP to the mucosal compartment, a result not observed in the control mice. In the presence of anti-CD23 antibodies, intact HRP fluxes were significantly decreased (131 +/- 27 ng/h x cm2) compared to control values in sensitized mice (290 +/- 86 ng/h x cm2, p < 0.02), suggesting that CD23 is involved is this 'protected' transport pathway. A similar protection during intestinal transport was observed for beta-LG in beta-LG sensitized mice. CONCLUSIONS: These results confirm that CD23 is involved in the rapid transepithelial transport of intact allergens in sensitized animals, and indicate that CD23 opens a 'protected' transport pathway in IECs.     

Effects of trifluoperazine, ouabain, and ethacrynic acid on intestinal calcium transport

Bidirectional steady-state calcium fluxes were measured in vitro under short-circuited conditions in segments of rat duodenum and descending colon. The calcium-activated ATPase (Ca-ATPase) inhibitor trifluoperazine (TFP, 0.1 mM) reduced net calcium absorption in both tissues by decreasing the absorptive flux from mucosa to serosa (Jm leads to s) without consistently altering the secretory flux from serosa to mucosa. 1,25-Dihydroxyvitamin D3 administration (50 ng/day for 4 days) increased net calcium absorption by increasing Jm leads to s, and TFP reduced Jm leads to s to the same extent across tissues from vehicle- or 1,25-dihydroxyvitamin D3-treated animals. Na-K-ATPase inhibitors ouabain and ethacrynic acid both reduced short-circuit current without affecting calcium fluxes. These data suggest that Ca-ATPase, located in the basolateral membrane of intestinal epithelial cells, plays a role in the transepithelial transport of calcium. More general effects of TFP on intestinal epithelium may also contribute to the reduction in calcium fluxes. Duodenal and descending colon calcium transport appears independent of transcellular sodium transport mediated by Na-K-ATPase.     

The intestinal zinc transport in aged rats

To determine if there are any age-related alterations in the intestinal zinc absorption that may contribute to zinc deficiency, we studied the zinc transport across the jejunal segments of 3-, 12- and 24-month-old Fisher 344 male rats using the Ussing chamber technique. We also evaluated the effect of 5 microM arachidonic acid in tissue bathing media. The zinc transport from mucosa to serosa in 24-month-old rats (55.0 +/- 3.5 nmol/h/cm2) was significantly greater than the transport in 12-month-old (30.1 +/- 6.3 nmol/h/cm2) and 3-month-old rats (41.0 +/- 5.2 nmol/h/cm2). There was no age-dependent differences in the zinc transport from the serosa to the mucosa. The addition of 5 microM arachidonic acid to the serosal side only resulted in a significant decrease in zinc transport rate from serosa to mucosa. The magnitude of change in the 24-month-old rats (36.0 +/- 3.9 vs. 26.5 +/- 3.2 nmol/h/cm2) was similar to the change seen in 12-month-old rats (34.5 +/- 4.6 vs. 20.1 +/- 3.5 nmol/h/cm2) and 3-month-old rats (34.5 +/- 4.6 vs. 26.1 +/- 3.9 nmol/h/cm2). The results indicate that intestinal zinc transport is increased in the aged rats, and that arachidonic acid or one of its metabolites may be important regulators of net zinc absorption through their effect on intestinal zinc secretion rates.     

Bicarbonate ions in active sodium transport across toad bladder.

The conductance, ga, and electromotive force, E, of active ion transport across toad bladders mounted as sacs were estimated from electrical measurements made before and after addition of sufficient ouabain (1.89 X 10(-3) M) to eliminate spontaneous potential. The ratio of net sodium transport (estimated from bidirectional fluxes) to external current in bladders voltage clamped to 0 mV was significantly less than unity in a normal medium containing HCO3- and Cl- ions, and also when Cl- was replaced with SO42-. However, when acetazolamide was added or when HCO3- was replaced by phosphate, short-circuit current and net sodium transport became equal. Spontaneous potential, E, and ga were all reduced about 20% by these maneuvers. The response of bidirectional sodium fluxes to voltage clamping at 0 mV or 120 mV in a bicarbonate-free medium was otherwise similar to that observed in a normal medium: net flux varied linearly with potential and calculated fluxes in the active transport path indicated a value significantly greater than unity for the empirical constant Q in the equation for change in the flux ratio, f, with change in potential, psi, viz., delta ln f = Q(ZF/RT)delta psi, similar to the high value for this constant that we have found in a bicarbonate-containing medium. We conclude that bicarbonate ions facilitate active sodium transport and also may be actively transported from serosa to mucosa in Dominican toads. However, coupling between bicarbonate and sodium fluxes does not account for the high value for Q for sodium in the active transport path.     

Effects of sugar and amino acid transport on transepithelial fluxes of sodium and chloride of short circuited rat jejunum

1. Using the Ussing Zerahn (1951) technique the relation between short circuit current (I(sc)) and unidirectional ion fluxes across the short circuited rat jejunum has been examined.2. These parameters of jejunal function have been measured in absence of sugars and amino acids and at 10 mM glucose, 28 mM glucose, 28 mM galactose and 20 mM proline. Sulphate has been used as substituent for chloride on both sides of the gut wall and potassium has been used on the serosal side to substitute for sodium. With regard to the effects of proline on I(sc) and ion fluxes, investigations were made to see whether sulphate and potassium substitutions significantly affected proline transport.3. The purpose of the study was to examine the I(sc) seen in sugars and amino acid free conditions and the Delta I(sc) induced by these substances. Further, it was our purpose to examine the validity of extending to amino acids and sugars in general the suggestion that jejunal transport of these substances stimulates an electroneutral secretion of sodium chloride.4. In the absence of sugars and amino acids at least two-thirds of an I(sc) of 50-70 muA/cm(2) are accounted for by a process of chloride secretion located in the deep parts of the epithelium. The remaining one third or less of the I(sc) can be accounted for in terms of net flux of sodium. The results further indicate that a state of hypoxia in the deep part of the epithelium is the cause of the very low I(sc), p.d. on electrical resistance usually found for sacs of everted rat jejunum.5. The increase in I(sc) induced by proline and glucose is best accounted for by the simultaneously induced increase in mucosa to serosa flux of sodium. There is no compelling evidence that proline should increase I(sc) by a stimulation of chloride secretion.6. Proline, glucose (and galactose) stimulate unidirectional fluxes of sodium and chloride. The effects on serosa to mucosa fluxes equal or exceed those on mucosa to serosa fluxes. The effect of proline on mucosa to serosa flux of thiourea exceeds by far that on serosa to mucosa flux. This and the effects of potassium and sulphate substitution lead to the conclusions in (7) and (8).7. Transport of sugars and amino acids induces or stimulates in rat jejunum in vitro an electroneutral secretion of sodium chloride.8. This effect suffices to explain why, in spite of marked increases in I(sc), increments in net flux of sodium are not induced by transport of proline and glucose. It also explains why the effects of sugar and amino acid transport on water transport in vitro are much less than in vivo.     

Quantitative determination of macromolecular transport rate across intestinal Peyer's patches

We used horseradish peroxidase (HRP) (mol wt, 40,000) to compare in vitro, in Ussing chambers, the rates of protein transport across segments of piglet jejunum with and without Peyer's patches. The mean HRP transport rate across intestinal segments with a patch, 25.2 +/- 4.2 SE ng . min-1 . cm-2 (22 animals), was increased threefold (P less than 0.0005) compared with control (no patch) tissue, 7.9 +/- 1.0 ng . min-1 . cm-2 (n = 29). Neither rate showed saturation with increasing concentrations of HRP; both were inhibited 75-95% by a temperature drop from 37 to 15 degrees C. Transport across patch-containing tissue was inhibited 48 +/- 6% (n = 5, P less than 0.0025) by 1 mM NaF, but NaF had no consistent effect on the transport across tissue without Peyer's patches. We conclude that HRP transport is increased across Peyer's patches. This transport is dependent on metabolism and does not involve specific receptors. These findings support the concept that the Peyer's patch serves an antigen-sampling function in the gut.     

Transport and electrical phenomena in resting and secreting piglet gastric mucosa.

1. Gastric mucosae were isolated from piglets (0-5 days old) and mounted in a chamber where electrical properties and secretory function could be measured. Unlike many previously reported mammalian in vitro preparations, pig gastric mucosae were stable and physiologically responsive for many hours after isolation. 2. With similar Ringer solutions bathing both surfaces, the isolated piglet gastric mucosa maintained a p.d. with the mucosal surface 30-35 mV negative with respect to the serosal surface. Limitation of access of Na+ from the mucosal bathing solution to the tissue (e.g. replacement of Na+ on mucosal side with choline or treatment with 10- minus 5 M amiloride) produced a decrease in p.d. and increase in mucosal resistance consistent with an hypothesis of Na+ transport from mucosa to serosa. 3. Isotopic flux measurements (36Cl and 24Na) and net H+ secretory rate were performed during open and short-circuit conditions, while the tissue was at rest and after stimulation of HCl secretion by 6 times 10- minus 5 M histamine. Up to 90% of the respective short-circuit current for resting or secreting mucosae was accounted for as the algebraic sum of Cl minus, H+ or Na+ fluxes. 4. The net transport of Na+ which occurred from mucosa to serosa during rest (ca. 4-7 muequiv/cm2.hr) was somewhat reduced during HCl secretion (ca. 2-7 muequiv/cm2.hr). This active transport of Na+ was more resistant to anaerobiosis than was H+ or Cl minus transport. 5. An active transport component of Cl minus from serosa to mucosa was clearly demonstrable in the non-secreting preparations (ca. 3-9 muequiv/cm2.hr). Active Cl minus transport was stimulated three- to fourfold after H+ secretion was stimulated by histamine. Anaerobiosis promptly reduced Cl minus and H+ transport. An exchange diffusion component was demonstrated for Cl minus which appeared to be prominent during H+ secretory activity and was considerably diminished in resting mucosae. 6. Large changes in mucosal resistance were associated with conditions of rest, histamine stimulation and anaerobic conditions; mean values were 113, 74 and 197 omega.cm2, respectively. Electrical conductance of the isolated gastric mucosa was due primarily to partial ionic conductance of Cl minus (60-65%) and Na+ (10-15%). The partial conductance of H+ was extremely low. The observed increase in tissue conductance associated with H+ secretory activity and the changes in the long-time constant p.d. transient to a current pulse are discussed in terms of the relative contribution of the serosal and mucosal plasma membrane surfaces.     

Segmental differences of short-chain fatty acid transport across guinea-pig large intestine.

Unidirectional fluxes of short-chain fatty acids (SCFAs) were measured under short-circuit current conditions across guinea-pig caecum, proximal and distal colon. Fluxes increased linearly with concentration. In the caecum with equal concentrations on both sides of the mucosa the serosal-to-mucosal (sm) fluxes were nearly twice the mucosal-to-serosal (ms) fluxes for all SCFAs; in the distal colon ms fluxes were always higher than sm fluxes. Thus, in the caecum the net effect was secretion while in the distal colon net absorption occurred. In caecum, ms fluxes decreased with chain length while sm fluxes were similar for the three fatty acids, acetic, propionic and butyric acid. In the distal colon both unidirectional fluxes increased with chain length. Fluxes across the mucosa of the proximal colon were intermediate to those in caecum and distal colon. A paracellular transport of short-chain fatty acids is not present. The results indicate that other processes are involved in transcellular transport of SCFA besides non-ionic diffusion.     

Acid secretion in fetal rat stomach in vitro

In vivo fetal rat stomach produces HCl 48 h before birth. This study examines the mechanisms of H+ secretion from days 19 to 21 before birth. Isolated fetal stomachs were mounted as flat sheets in Ussing chambers for measurement of the transepithelial H+ fluxes (JH+) and short-circuit current (Isc), as indexes of the active ionic fluxes, and for measurement of total ionic conductance (G) and unidirectional mannitol fluxes from serosa to mucosa (JMans leads to m), as indexes of passive permeability. The results indicate that JH+ was absent at day 19 but reached 0.75 +/- 0.1 and 0.75 +/- 0.09 mueq . h-1 . cm-2 at days 20 and 21, respectively. Concomitantly, Isc increased significantly (56%) between days 19 and 20 in the direction of anion secretion or cation absorption. Parallel reductions in G (45%) and in JMans leads to m (66%) were observed between days 19 and 20. In conclusion, the simultaneous appearance of active H+ secretion and decreased passive transepithelial permeability strongly suggests that both processes are involved in the mechanism of acidification of the fetal rat stomach before birth.     

Active sodium uptake by the skin of foetal sheep and pigs.

1. Simultaneous measurements of unidirectional sodium fluxes across foetal skin incubated in vitro with identical solutions ([Na] = 150 mM) bathing either side showed a flux ratio (influx/efflux) of 1-40+/-0-08 in twenty-seven sheep skins, which was significantly different from unity (P less than 0-001). The gestational ages ranged from 47 to 98 days (term = 147 days). Similar experiments on eight foetal pig skins at 58 days gestation (term = 114-118 days) gave a mean flux ratio of 1-10 +/- 0-03 (P less than 0-02). 2. Unidirectional sodium fluxes measured with dilute Ringer solution on the outside (mucosal) surface ([Na]0 = 100mM) gave influx to efflux ratios of 0-86 +/- 0-09 in seventeen sheep (P less than 0-05) and 1-07 +/- 0-26 in five foetal pigs; the value predicted for passive movement was 0-67. 3. Incubation with inhibitors, ouabain (10-4 M) or dinitrophenol (DNP) (10-4 M) gave a flux ratio for sodium which was not significantly different from unity in the absence of a gradient, or from 0-67 when the concentration gradient was applied. 4. Sequential measurement of unidirectional diffusional fluxes of tritiated water across foetal skin gave flux ratios of 0-98 +/- 0-02 in six sheep skins and 1-06 +/- 0-11 for four pig skins in control conditions. When the outside solution was diluted to give an osmotic gradient of 100 m-osmole. kg-1 across the skin a flux ratio of 0-95 +/- 0-07 was obtained for seven sheep and was not measured in pig skin. Hormones and inhibitors had no effect on the diffusional flux ratio for water in the presence or absence of an osmotic gradient. 5. Lysine vasopressin (ADH) (200 mu./ml.) increased influx and efflux of water in the presence and, to a lesser extent in the absence of an osmotic gradient in sheep skin. In pig skin prolactin (1 u./ml.) increased both influx and efflux, but ADH had no effect on diffusional water fluxes. 6. ADH increased sodium influx in sheep skin slightly but vasotocin (5-5 mu./ml.) was more potent, particularly in the presence of an opposing diffusion gradient. Vasotocin (55 mu./ml.) reduced sodium influx in pig skin ADH had no effect on influx or efflux and prolactin reduced sodium influx and efflux. Ouabain and DNP generally reduced permeability to both sodium and water in sheep skin but had no effect in pig skin.     

Chloride transport in glands of frog skin

The effects of beta-adrenergic stimulation on the bidirectional fluxes of Na+ and Cl- across the frog skin glands were determined. Isoproterenol elicited net serosal-to-mucosal fluxes of both Na+ (JNanet) and Cl- (JClnet) equal to 0.19 +/- 0.05 (SE) and 0.57 +/- 0.05 mueq X cm-2 X h-1, respectively. The residual current (JClnet - JNanet) of 0.38 +/- 0.05 mueq X cm-2 X h-1 closely approximates the isoproterenol-induced short-circuit current of 0.30 +/- 0.04 mueq X cm-2 X h-1. Furosemide added to the serosal side prior to isoproterenol inhibited the isoproterenol-induced net fluxes of both Na+ and Cl-. The addition of dibutyryl cAMP and 3-isobutyl-1-methylxanthine to the serosal side mimicked the action of isoproterenol by stimulating glandular short-circuit current. We conclude that an active Cl(-)-transport mechanism resides in the frog skin glands and is 1) stimulated by a beta-adrenergic agonist (its action is mimicked by cAMP) and 2) inhibited by the loop diuretic furosemide.     

The effects of Clostridium difficile crude toxins and purified toxin A on stripped rabbit ileal mucosa in Ussing chambers

Clostridium difficile crude toxins and purified toxin A had similar effects on stripped rabbit ileal mucosa in Ussing chambers. Both toxin preparations caused secretion of sodium and chloride ions by increasing serosa to mucosa (s----m) fluxes. Transmural potential difference and resistance decreased after toxin treatment. Onset of changes in electrical measurements and ion fluxes coincided with onset of histological changes. The response to theophylline was greatly reduced in toxin-treated tissue compared with control tissue.     

Protein synthesis in liver and small intestine in protein deprivation and diabetes

Protein synthesis (as a percent of the protein pool synthesized per day) has been measured in liver and small intestine of young male rats from the incorporation of 100 mumol [1-14C]leucine/100 g body wt into protein over 10 min. Dietary protein deprivation for 8 days depressed protein synthesis in liver (30%), jejunal mucosa (20%), and jejunal serosa (25%). In serosa, reduced levels of RNA relative to protein could account for altered synthesis; in liver and mucosa, the amount of protein synthesized per unit RNA was reduced. In liver of streptozotocin-diabetic rats protein synthesis was depressed 45%, whereas it was maintained in jejunal mucosa and serosa. Depressed synthesis in liver was accompanied by both a loss of RNA relative to protein and a reduction in the protein synthesized per RNA.     

Response of Amphiuma small intestine to theophylline: effect on bicarbonate transport.

The electrolyte transport properties of isolated proximal segments of Amphiuma small intestine and their response to theophylline were observed under various conditions. In the absence of theophylline the intestine generates a transepithelial potential (psi ms) serosa negative to mucosa when Cl- and HCO3- are present in the bath. Replacement of Cl- or HCO3- reduced the magnitude and usually reversed the sign of psi ms. Acetazolamide (10(-4) M) nearly abolished the serosa negative psi ms. Theophylline (10 mM) drove psi ms serosa positive, the magnitude depending on the bath Na+ and HCO3- concentrations. Simultaneously it increased the short-circuit current (Isc) and tissue resistance (Rt). The increase in Isc was not due to increase net Na+ transport in Cl-free buffer and was attributed to a residual ion flux. Acetazolamide reduced the Isc, Rt, and the net residual flux observed in theophylline-treated intestine. The magnitude of the acetazolamine effect on Isc was proportional to the Na+ and HCO3- concentrations of the bath. The results suggest that in the absence of theophylline, HCO3-, and Cl- transport are related. Furthermore, acetazolamide inhibits the movement of an ion, possibly HCO3-, secreted in response to theophylline.     

Sodium chloride transport across the chicken coprodeum. Basic characteristics and dependence on sodium chloride intake

1. The transport characteristics of the chicken coprodeum have been examined in vitro using the isolated mucosa. The short-circuit current (I_sc), the transepithelial electrical potential difference (p.d.), the unidirectional transmural fluxes (J_ms, J_sm) of sodium and chloride measured in the short-circuited state, and the unidirectional influx of sodium and chloride across the brush border membrane measured under open-circuit conditions have been studied. The effect of the sodium chloride contents of the diet on these parameters have been investigated.

2. The isolated mucosa depends functionally on the presence of glucose in the incubation media. This dependence reflects the need of glucose as a fuel. There is no indication of coupling between transport of sugars and sodium across the brush border membrane. For preparations from chickens on a low sodium diet a very high and stable I_sc can quantitatively be accounted for by the net transport of sodium. Influx of sodium across the brush border membrane is not significantly different from the net flux of sodium. By feeding the chickens a high sodium diet the I_sc is reduced by more than 95%, the net transport of sodium is abolished, and the transepithelial electrical conductance is reduced by more than 50%.

3. Both unidirectional transepithelial fluxes of chloride, and the serosa to mucosa flux of sodium appear to proceed through a paracellular shunt.

4. Under the conditions of the low sodium diet the paracellular pathway appears to be anion selective. Whereas, under the conditions of the high sodium regimen the paracellular route appears to be cation selective. After adaptation to a high sodium diet the influx of sodium across the brush border membrane is only moderately reduced. Consequently the decisive event in the adaptation must be localized elsewhere.

     

Cell volume‐induced changes in K+ transport across the rat colon

The effect of cell swelling and cell shrinkage on K⁺ transport across the rat colonic epithelium was studied by measuring unidirectional fluxes, uptake and efflux of ⁸⁶Rb⁺, a marker for K⁺. Exposure to a hypotonic medium stimulated the secretory, serosa‐to‐mucosa flux of K⁺, whereas exposure to a hypertonic medium inhibited the absorptive, mucosa‐to‐serosa flux of K⁺ in the distal, but not in the proximal colon. Neither manoeuvre had any effect on the uptake of K⁺ across the apical or the basolateral membrane. Cell swelling induced a sustained increase in the apical and basolateral K⁺ efflux from both colonic segments, whereas cell shrinkage reduced the efflux. Ba²⁺ (10^–2 mol l^–1) inhibited the swelling‐induced stimulation of the apical, quinine (10^–3 mol l^–1) that of the basolateral K⁺ efflux in the distal colon. Incubation of the tissue in Ca²⁺‐free buffer or La³⁺, which blocks Ca²⁺‐influx into the epithelium, strongly reduced the basal K⁺ efflux across the basolateral membrane. The same was observed with brefeldin A, a blocker of the transport of newly synthesized proteins out of the endoplasmatic reticulum. Swelling‐induced K⁺ efflux, however, was not reduced. In the presence of colchicine, an inhibitor of the polymerization of microtubules, swelling evoked only a transient increase in mucosal efflux, which, especially in the proximal colon, fell after 6 min to the level of the isotonic control period. These results demonstrate that the cell volume is involved in the regulation of transepithelial K⁺ transport across the rat colonic epithelium and suggest a role of the cytoskeleton in the control of a part of the volume‐sensitive K⁺ channels.     

The electrical properties and active ion transport across the urinary bladder of the urodele, Amphiuma means.

1. The electrical properties and the active transport processes of the isolated urinary bladder of the urodele, Amphiuma means, were studied by mounting this tissue as a flat sheet between two halves of a lucite chamber. The mean transepithelial potential difference was 70-2 +/- 2-3 mV (serosa positive), the mean short-circuit current was 10-9 +/- 0-5 micrionA/mg of dry weight and the mean transepithelial d.c. resistance was 6540 +/- 374 omega mg of dry weight. 2. The short-circuit current (Isc) accounted for 92% of the net 22Na+ flux from the mucosa to the serosa. The difference resulted from a transport of 36Cl- in the same direction as sodium. 3. The active sodium transport exhibited typical saturation kinetics, having a Km of 15-4 m-equiv/l. and approaching zero order at 60-70 m-equiv/l. The transepithelial potential difference increased linearly with the log of the mucosal sodium concentration at a rate of 50-3 mV per tenfold concentration change. 4. In the absence of the major anions (HCO3- and Cl-) from the bathing solutions, the electrical properties and the sodium influx decreased to less than 40% of their control values. The presence of only one of these two anions in the serosal bathing solution was sufficient to maintain these parameters. 5. Amiloride (10(-5)M) and ouabain (10(-6)M) inhibited the sodium transport 97% and 85% respectively. Amphotericin B (10(-6)M) stimulated the sodium transport 47%. Furosemide (10(-3)M) inhibited the chloride transport 43%. The sodium transport was insensitive to the action of two enurohypophyseal peptides tested, lysine-vasotocin and pitocin.     

Sodium and chloride transport by the intestine of the european flounderPlatichthys flesus adapted to fresh or sea water

1. Everted intestinal sacs prepared from Platichthys flesus adapted to sea water (SW) have higher rates of salt and water transport than did sacs prepared from fresh water adapted (FW) fish. 2. Intestines mounted in Ussing chambers gave stable open-circuit voltages and short-circuit currents after pre-incubation for 20--30 min of --1.91 +/- 0.14 (14) mV and --45.0 +/- 5.1 (14) muA/cm2, SW fish, and --1.24 +/- 0.14 (11) mV and --18.2 +/- 3.6 (11) muA/cm2, FW fish. 3. Isotope flux measurements indicated a net Na transport of 21.5 +/- 4.1 (11) neq/cm2-min in SW fish intestines, but no significant Na transport in FW fish (7.6 +/- 7.6 (9) new/cm2-min). Both preparations showed an active Cl transport, 26.6 +/- 6.1 (12) neq/cm2-min for SW and 18.5 +/- 9.7 (17) neq/cm2-min for FW fish. 4. No significant difference in the uptake of Na or Cl across the mucosa was observed between FW and SW fish. The uptake of both ions showed some saturation at high concentrations. 5. Interactions between Na and Cl uptake were small, although Cl uptake was significantly higher in Na-free solutions. 6. It is concluded that electrogenic Cl transport may be the dominant mechanism for water and salt transport in flounder intestine, and that adaptation to a saline environment involves regulation of pumping rather than Na or Cl entry across the mucosal membrane.     

Retrograde labeling of central nervous pathways with tritiated or Evans blue-labeled bovine serum albumin

In an attempt to develop a technique for the retrograde tracing of neuronal pathways which does not depend on enzyme activity of horseradish peroxidase (HRP), we have examined the retrograde transport of bovine serum albumin (BSA) by neurons of the central nervous system. Following injections of either tritiated or Evans blue-labeled BSA into the hippocampal formation of the rat, labeled neurons were found in the ipsilateral entorhinal area in a pattern which was highly reminiscent of that obtained with HRP. The retrograde transport of BSA provides a method for the retrograde tracing of central nervous pathways which is completely independent of the HRP method, making it possible to use both substances for studies of collateral innervation, and can be used with any fixative, or no fixative at all.     

Ion transport across the excised bullfrog lung.

Fluxes of ions and water across the short-circuited, excised bullfrog lung were determined by radioisotope techniques. The unidirectional flows of Na+, K+, Ca++, TcO4 minus, HCO3 minus, gluconate, rho-aminohippurate, dinitrophenolate, SO4 equal to, and water were symmetrical. Both HCO3 minus fluxes were reduced by acetazolamide. In contrast, Cl minus, Br minus I minus, and SCN minus movement from serosa to mucosa exceeded the flux in the opposite direction. Net Cl minus transport followed the kinetics of a saturable process and was inhibited by dinitrophenol and hypoxia. These results indicate an active secretion of halide anions and SCN minus into the lumen. Attempts to demonstrate Br minus anatagonism of Cl minus transport were equivocal. Cl minus transport accounted for 50 percent minus of the early short-circuit current but after 90 min the two measurements were equal. Incubation of the lung in bicarbonate-free Ringer revealed unequal decreases in the H+ concentration of the bathing solutions. Net "base" addition to the serosal solution was reduced by prior removal of the blood from the pulmonary vasculature. Therefore, "base" release could not be localized to the epithelia. The Na+, K+, Ca++ and Cl minus composition of the lung tissue was unchanged over 3 h. Since tissue and, hence cell Cl minus is lower than the concentration in the bathing solution the Cl pump is probably located in the luminal border of the alveolar epithelial cell.