Intestinal SGLT1-mediated absorption and metabolism of benzyl beta-glucoside contained in Prunus mume: carrier-mediated transport increases intestinal availability

The intestinal absorption of benzyl beta-glucoside (BNZ beta glc) contained in the fruit of Prunus mume SIEB. et ZUCC. (Rosaceae), which is traditionally used as a medicinal food in Japan, was studied in rat intestines. BNZ beta glc was absorbed from the mucosal to serosal sides. Its metabolite, benzyl alcohol (BAL), was also detected on both the mucosal and serosal sides. In the presence of phloridzin (Na(+)/glucose cotransporter (SGLT1) inhibitor) or in the absence of Na+ (driving force), BNZ beta glc absorption was significantly decreased. Transport clearance of BNZ beta glc across the brush border membrane decreased as its concentration increased. These results indicate that BNZ beta glc is transported by SGLT1. Metabolic clearance of BNZ beta glc also decreased as its concentration increased. The amount ratio of BNZ beta glc to BAL on the serosal side increased with the increase of BNZ beta glc concentration. The intestinal availability of BNZ beta glc was lower in the absence of Na+ than in the presence of Na+, indicating that the SGLT1-mediated transport of BNZ beta glc increases intestinal availability by decreasing the intestinal extraction ratio. This neutraceutical study concluded that intestinal carrier-mediated transport across the brush border membrane improves the intestinal availability of nutritionally, pharmacologically or physiologically active compounds that undergo intestinal metabolism (first-pass effect).     

Intestinal SGLT1-mediated absorption and metabolism of benzyl β-glucoside contained in Prunus mume: carrier-mediated transport increases intestinal availability

The intestinal absorption of benzyl β-glucoside (BNZβglc) contained in the fruit of Prunus mume SIEB. et ZUCC. (Rosaceae), which is traditionally used as a medicinal food in Japan, was studied in rat intestines. BNZβglc was absorbed from the mucosal to serosal sides. Its metabolite, benzyl alcohol (BAL), was also detected on both the mucosal and serosal sides. In the presence of phloridzin (Na⁺/glucose cotransporter (SGLT1) inhibitor) or in the absence of Na⁺ (driving force), BNZβglc absorption was significantly decreased. Transport clearance of BNZβglc across the brush border membrane decreased as its concentration increased. These results indicate that BNZβglc is transported by SGLT1. Metabolic clearance of BNZβglc also decreased as its concentration increased. The amount ratio of BNZβglc to BAL on the serosal side increased with the increase of BNZβglc concentration. The intestinal availability of BNZβglc was lower in the absence of Na⁺ than in the presence of Na⁺, indicating that the SGLT1-mediated transport of BNZβglc increases intestinal availability by decreasing the intestinal extraction ratio. This neutraceutical study concluded that intestinal carrier-mediated transport across the brush border membrane improves the intestinal availability of nutritionally, pharmacologically or physiologically active compounds that undergo intestinal metabolism (first-pass effect).     

Antimicrobial properties of analgesic kyotorphin peptides unraveled through atomic force microscopy

Antimicrobial peptides (AMPs) are promising candidates as alternatives to conventional antibiotics for the treatment of resistant pathogens. In the last decades, new AMPs have been found from the cleavage of intact proteins with no antibacterial activity themselves. Bovine hemoglobin hydrolysis, for instance, results in AMPs and the minimal antimicrobial peptide sequence was defined as Tyr-Arg plus a positively charged amino acid residue. The Tyr-Arg dipeptide alone, known as kyotorphin (KTP), is an endogenous analgesic neuropeptide but has no antimicrobial activity itself. In previous studies new KTP derivatives combining C-terminal amidation and Ibuprofen (Ib) - KTP-NH(2), IbKTP, IbKTP-NH(2) - were designed in order to improve KTP brain targeting. Those modifications succeeded in enhancing peptide-cell membrane affinity towards fluid anionic lipids and higher analgesic activity after systemic injection resulted therefrom. Here, we investigated if this affinity for anionic lipid membranes also translates into antimicrobial activity because bacteria have anionic membranes. Atomic force microscopy revealed that KTP derivatives perturbed Staphylococcus aureus membrane structure by inducing membrane blebbing, disruption and lysis. In addition, these peptides bind to red blood cells but are non-hemolytic. From the KTP derivatives tested, amidated KTP proves to be the most active antibacterial agent. The combination of analgesia and antibacterial activities with absence of toxicity is highly appealing from the clinical point of view and broadens the therapeutic potential and application of kyotorphin peptides.     

Absorption of intact beta-casomorphins (beta-CM) in rabbit ileum in vitro

The functional significance of the presence of opioid peptides in enzymatic digestion of bovine milk beta-casein remains unclear. Opiates modify intestinal electrolyte transport by acting on receptors located on the serosal side of the intestine. The aim of the present study is to determine under which conditions beta-casomorphins could act from the luminal side of the intestine. The effect of natural morphiceptin (beta-CM4-NH2) and the non metabolized analogue beta-[DAla2,4, Try5]-CM5-NH2 were studied on isolated rabbit ileum mounted in Ussing chambers. Both peptides caused a naloxone-reversible reduction in short-circuit current (lsc) and stimulated Na and Cl absorption after addition to the serosal side of the tissue. After mucosal addition, only the analogue (10(-3) M) crossed the epithelium intact (Jm-s = 3.5 +/- 1.2 nmol.h-1.cm-2) and reduced lsc. Morphiceptin, under the same conditions, was degraded by the intestinal mucosa without opiate action on electrolyte transport. Pretreatment of the ileum by 10(-3)M diisopropylfluorophosphate that inhibited brush-border dipeptidylpeptidase IV, prevented mucosal degradation of morphiceptin. Under these conditions, the peptide (10(-3)M) crossed the epithelium intact (Jm-s = 1.8 +/- 0.16 nmol.h-1.cm-2) and stimulated electrolyte absorption by means of an opioid mechanism. These results show that both natural morphiceptin and the protected analogue have an opiate activity on intestinal electrolyte transport. Their action from the lumen depends on their transfer intact to the serosal side of the intestine where opiate receptors are located. The limiting step in this transfer is at the brush-border membrane where dipeptidylpeptidase IV in particular seems to play a major role.     

Transport and metabolism of N6- and C8-substituted analogs of adenosine 3',5'-cyclic monophosphate and adenosine 3'5'-cyclic phosphorothioate by the isolated perfused rat kidney

The clearance and metabolism of N6-substituted (N6-dimethyl-), C8-substituted (8-bromo-, 8-p-chlorophenylthio- (PCPT-)), and exocyclic oxygen substituted phosphorothioate diastereomers (cAMPS(Sp)) and cAMPS (Rp)) of adenosine 3':5'-monophosphate (cyclic AMP, cAMP) has been studied in an isolated perfused rat kidney. The N6- and C8-substituted analogs of cyclic AMP (10-100 microM) were not cleared as rapidly as exogenous cyclic AMP and were metabolized: N6- and C8-substituted analogs of adenosine accumulated in perfusate and urine. All analogs exhibited net transtubular secretion, i.e. their urinary excretion rate greater than glomerular filtration rate. Probenecid (0.9 mM) included in the perfusate abolished transtubular secretion and inhibited the metabolism of PCPT-cyclic AMP, suggesting that cyclic AMP analogs, like cyclic AMP itself, penetrate the renal cell at the peritubular membrane by an organic acid transport system. The phosphorothioate diastereomers of cyclic AMP: cAMPS(Sp) and cAMPS(Rp) were cleared as rapidly from the perfusate as cyclic AMP, were extensively secreted (urinary excretion/ glomerular filtration greater than or equal to 10) and exhibited no metabolism. The latter analog would seem most suitable as an intracellular agonist for cyclic AMP-mediated phenomena in the rat kidney.     

Basolateral Mg2+/Na+ exchange regulates apical nonselective cation channel in sheep rumen epithelium via cytosolic Mg2+

High potassium diets lead to an inverse regulation of sodium and magnesium absorption in ruminants, suggesting some form of cross talk. Previous Ussing chamber experiments have demonstrated a divalent sensitive Na(+) conductance in the apical membrane of ruminal epithelium. Using patch-clamped ruminal epithelial cells, we could observe a divalent sensitive, nonselective cation conductance (NSCC) with K(+) permeability > Cs(+) permeability > Na(+) permeability. Conductance increased and rectification decreased when either Mg(2+) or both Ca(2+) and Mg(2+) were removed from the internal or external solution or both. The conductance could be blocked by Ba(2+), but not by tetraethylammonium (TEA). Subsequently, we studied this conductance measured as short-circuit current (I(sc)) in Ussing chambers. Forskolin, IBMX, and theophylline are known to block both I(sc) and Na transport across ruminal epithelium in the presence of divalent cations. When the NSCC was stimulated by removing mucosal calcium, an initial decrease in I(sc) was followed by a subsequent increase. The cAMP-mediated increase in I(sc) was reduced by low serosal Na(+) and serosal addition of imipramine or serosal amiloride and depended on the availability of mucosal magnesium. Luminal amiloride had no effect. Flux studies showed that low serosal Na(+) reduced (28)Mg fluxes from mucosal to serosal. The data suggest that cAMP stimulates basolateral Na(+)/Mg(2+) exchange, reducing cytosolic Mg. This increases sodium uptake through a magnesium-sensitive NSCC in the apical membrane. Likewise, the reduction in magnesium uptake that follows ingestion of high potassium fodder may facilitate sodium absorption, as observed in studies of ruminal osmoregulation. Possibly, grass tetany (hypomagnesemia) is a side effect of this useful mechanism.     

D-galactose accumulation in rabbit ileum. Effects of theophylline on serosal permeability.

The effects of theophylline and dibutyryl cyclic AMP, on in vitro unidirectional galactose fluxes across the mucosal and serosal borders of rabbit ileum have been studied. 1. When Ringer [galactose] = 2mM, theophylline and dibutyryl cyclic AMP reduce both mucosal-serosal and serosal-mucosal galactose flux by approx. 50%. The K1 for theophylline inhibition of flux in both directions is 2 mM. 1 mM dibutyryl cyclic AMP elicits a maximal inhibitory response. Concurrent with the inhibition in transmural galactose fluxes, theophylline and dibutyryl cyclic AMP increase the tissue accumulation of [galactose] and the specific-activity ratio R of 3H : 14C-labelled galactose coming from the mucosal and serosal solutions respectively. It is deduced that theophylline and dibutyryl cyclic AMP are without effect on the mucosal unidirectional permeability to galactose but cause a symmetrical reduction in serosal entry and exit permeability. 2. Reduction in the asymmetry of the mucosal border to galactose by reducing Ringer [Na], raising Ringer [galctose] or adding ouabain reduces the theophylline-dependent increase in galactose accumulation. 3. Hypertonicity in the serosal solution increases the permeability of the serosal border to galactose and reduces tissue galactose accumulation. Serosal hypertonicity partially reverses the theophylline-depedent effects on galactose transport. Replacing Ringer chloride by sulphate abolishes the theophylline-dependent effects on galactose transport. 4. It is considered that the theophylline-dependent increase in galactose accumulation results from the reduction in serosal permeability. This is shown to be a quantitatively consistent inference. 5. Further support for the view that the asymmetric transport of galactose in rabbit ileum results from convective-diffusion is presented.     

Coupling of volume and Na+ transport in frog skin epithelium

Whole skins and isolated epithelia were bathed with isotonic media (congruent to 244 mOsm) containing sucrose or glucose. The serosal osmolality was intermittently reduced (congruent to 137 mOsm) by removing the nonelectrolyte. Transepithelial and intracellular electrophysiological parameters were monitored while serosal osmolality was changed. Serosal hypotonicity increased the short-circuit current (ISC) and the basolateral conductance, hyperpolarized the apical membrane (psi mc), and increased the intracellular Na+ concentration. The increases in apical conductance and apical Na+ permeability (measured from Goldman fits of the relationship between amiloride-sensitive current and psi mc) were not statistically significant. To verify that the osmotically induced changes in ISC were mediated primarily at the basolateral membrane, the basolateral membrane potential of the experimental area was clamped close to 0 mV by replacing the serosal Na+ with K+ in Cl--free media. The adjoining control area was exposed to serosal Na+. Serosal hypotonicity produced a sustained stimulation of ISC across the control, but not across the adjoining depolarized tissue area. The current results support the concept that hypotonic cell swelling increases Na+ transport across frog skin epithelium by increasing the basolateral K+ permeability, hyperpolarizing the apical membrane, and increasing the electrical driving force for apical Na+ entry.     

CORRELATION BETWEEN PINOCYTOSIS AND HYDROOSMOSIS INDUCED BY NEUROHYPOPHYSEAL HORMONES AND MEDIATED BY ADENOSINE 3'',5''-CYCLIC MONOPHOSPHATE

The isolated urinary bladder of the toad responds to neurohypophyseal hormone with a net increase of water transport from the mucosal to the serosal solution in the presence of an osmotic gradient. This response is mediated intracellularly by cyclic 3',5'-adenosine monophosphate (AMP). The present study demonstrates that hydroosmotically active substances such as oxytocin, dibutyryl cyclic 3',5'-AMP, and theophylline, but not hydroosmotically inactive substances, induce the uptake of horseradish peroxidase from the mucosal solution. Peroxidase taken up by the mucosal cells is demonstrable in small tubules and vesicles, and eventually accumulates in lysosomes. The uptake of peroxidase from the serosal solution into similar bodies in the mucosal cells is not hormone-dependent. It is also shown that peroxidase does not penetrate the tight junction from either the mucosal or serosal solution. These results extend previous findings which implicated the apical membrane of the mucosal epithelium as the site affected by neurohypophyseal hormones. A mechanism based on secretory phenomena is proposed as a framework for future investigations of apical membrane permeability changes and pinocytosis.     

Regulation of the sodium permeability of the luminal border of toad bladder by intracellular sodium and calcium: role of sodium-calcium exchange in the basolateral membrane

Sodium movement across the luminal membrane of the toad bladder is the rate-limiting step for active transepithelial transport. Recent studies suggest that changes in intracellular sodium regulate the Na permeability of the luminal border, either directly or indirectly via increases in cell calcium induced by the high intracellular sodium. To test these proposals, we measured Na movement across the luminal membrane (th Na influx) and found that it is reduced when intracellular Na is increased by ouabain or by removal of external potassium. Removal of serosal sodium also reduced the influx, suggesting that the Na gradient across the serosal border rather than the cell Na concentration is the critical factor. Because in tissues such as muscle and nerve a steep transmembrane sodium gradient is necessary to maintain low cytosolic calcium, it is possible that a reduction in the sodium gradient in the toad bladder reduces luminal permeability by increasing the cell calcium activity. We found that the inhibition of the influx by ouabain or low serosal Na was prevented, in part, by removal of serosal calcium. To test for the existence of a sodium- calcium exchanger, we studied calcium transport in isolated basolateral membrane vesicles and found that calcium uptake was proportional to the outward directed sodium gradient. Uptake was not the result of a sodium diffusion potential. Calcium efflux from preloaded vesicles was accelerated by an inward directed sodium gradient. Preliminary kinetic analysis showed that the sodium gradient changes the Vmax but not the Km of calcium transport. These results suggest that the effect of intracellular sodium on the luminal sodium permeability is due to changes in intracellular calcium.     

Intestinal iodide secretion and its dependence upon mucosal I- permeability

Iodide secretion across different regions of rat small intestine has been investigated in vitro using the standard Wilson-Wiseman technique. Net I- secretion was observed along the entire small intestine, being significantly higher in the central region. Anaerobic conditions, ouabain (2 mM) and Na+ free Ringer solution prevented net I- secretion, whilst both theophylline (1 mM) and carbachol (0,1 mM) enhanced the observed basal intestinal I- secretion. Furthermore, Ca2+-deprived bathing solutions significantly reduced intestinal I- secretion. Epithelial I- uptake from both mucosal and serosal sides was measured by using a Ussing-type chamber technique. The initial rate of I- uptake across the mucosal membrane was significantly higher in the central region than in the proximal part of rat small intestine. No significant differences were observed in the rate of I- uptake from the serosal side. These studies suggest that mucosal I- permeability might determine the direction of net I- intestinal transport and that cytosolic Ca2+ may be a physiological regulator of intestinal I- transport.     

Absorption of intact morphiceptin by diisopropylfluorophosphate-treated rabbit ileum

The amidated beta-casomorphin morphiceptin Tyr-Pro-Phe-Pro-NH2 is an opioid peptide isolated from bovine milk beta-casein digests whose physiological significance remains unclear. Opiates are known to modify intestinal electrolyte transport by acting on receptors located on the serosal side of the intestine. The aim of the present study was to determine under what conditions morphiceptin can act from the luminal side. When added to the serosal side of untreated rabbit ileum in an Ussing chamber in vitro, 10(-3) M morphiceptin acted through an opiate mechanism to reduce simultaneously short-circuit current (delta Isc = 0.33 +/- 0.07 muEq.hr-1.cm-2) and stimulate net Na and Cl absorption (delta JnetNa = 1.62 +/- 0.11 and delta JnetCl = 2.07 +/- 0.08 muEg.hr-1.cm-2). After mucosal addition under the same conditions, morphiceptin was degraded without any opiate action on electrolyte transport. Pretreatment of the ileum by 10(-3) M diisopropylfluorophosphate, which inhibited brush-border dipeptidylpeptidase IV, prevented mucosal degradation of morphiceptin. Under these conditions, morphiceptin was able, when added mucosally, to cross the epithelium intact (Jm----s = 1.8 +/- 0.16 nmole.hr-1.cm-2) and to stimulate electrolyte absorption by means of an opioid mechanism (delta Isc = 0.22 +/- 0.02 muEq.hr-1.cm-2). These results showed that the action of morphiceptin from the lumen depends on its transfer intact to the serosal side of the intestine where the opiate receptors are located. The limiting step in this transfer is at the brush-border membrane, where dipeptidylpeptidase IV in particular seems to play a major role.     

Intracellular chloride activity and membrane potential in stripped frog skin (Rana temporaria)

The regulation of cell chloride activity in frog skin was investigated using double barrelled Cl--microelectrodes to measure cell membrane potentials and chloride activity in the isolated frog epidermis. Experiments were done under short-circuit conditions, impaling cells from the serosal side. The basic electrophysiological parameters of the isolated skin were similar to those reported in the literature for whole preparations. Intracellular chloride activity was on average 21.9 mM and membrane potential was about 57 mV, implying that chloride was distributed away from its electrochemical equilibrium (i.e., concentrated inside the cells). Chloride activity decreased after removal of either Cl- or Na+ from the serosal bathing solution, with no change in membrane potential. The chloride permeability of the serosal membrane was calculated to be 2.6 X 10(-6) cm X s-1 which represents about 1/4 of the total conductance of the serosal membrane. We suggest that an electrically silent sodium-dependent uphill transport of chloride is present at the basolateral membrane of the frog skin, which accounts for the non-passive distribution of chloride.     

Localization and characterization of transport-related elements in the plasma membrane of turtle bladder epithelial cells.

A mixed membrane preparation obtained from turtle bladder epithelial cells contains (Na+ + K+)-ATPase, adenylate cyclase and protein kinase, which interact with ouabain, norepinephrine and cyclic AMP, respectively. When such a preparation is obtained from bladders which had been preexposed to serosal fluids containing the tritiated form of 4,4'-diisothiocyano-2,2'-disulfonic stilbene, the subsequently isolated membrane proteins are enriched in tritium as well as in the afore-mentioned enzymes, none of which is inhibited. Free-flow electrophoresis separates the mixed membrane preparation into two distinguishable groups: one, construed as apical membranes, is enriched in norepinephrine-sensitive adenylate cyclase and cyclic AMP-sensitive protein kinase; the other, construed as basal-lateral membranes, is enriched in ouabain-sensitive ATPase and 4,4'-diisothiocyano-2,2'-disulfonic stilbene-binding proteins. The physiological counterparts of these enzymatically defined membrane markers are the mucosal sidedness of the transport effects of norepinephrine and cyclic AMP derivatives and the serosal sidedness of the transport effects of ouabain and disulfonic stilbenes in the intact turtle bladder. The discreteness and ion selectivity of each membrane-bound, transport-related element are discussed in relation to the corresponding characteristics of each transport process in vivo; the possibility of regulation of anion transport by adenylate cyclase-protein kinase system is also discussed.     

Effects of melittin on a model renal epithelium, the toad urinary bladder

The bee venom melittin, 10(-6) M, on the mucosal (urinary) side of the toad urinary bladder (in vitro), markedly decreased transepithelial potential difference, short-circuit current (Isc, sodium-dependent) and resistance. However, these effects were not seen when the toxin was placed on the opposite (serosal) side of the membrane preparation. The electrical effects were accompanied by a large increase in the transepithelial permeability to 22Na. The response was not changed by meclofenamic acid (which blocks formation of prostaglandins) but it was inhibited by La3+. In the presence of amiloride, which usually inhibits active Na transport and Isc, melittin, on the mucosal side, increased the Isc. The action of melittin appears to involve an interaction with anionic sites, which mediate its effects. Such sites appear to be present on the apical plasma membranes of the toad bladder epithelial cells, but they are not as abundant or they are inaccessible on the basal plasma membrane.     

Effect of various methods of oxygenation in the isolated small intestine on the tryptophan and glycine accumulation

The impact of various oxygenation types of intestinal preparations (mucosal, serosal and bilateral) on the transepithelial transport of amino acids and peptide according to the character of amino acids, their stereoisometry, incubation time, intestinal gradient and other factors, was investigated in fowl. Against-the-gradient-transport of L-tryptophan and glycine was observed at various oxygenation types, and its intensity decreased in the following sequence: bilateral, mucosal, serosal oxygenation. At various oxygenation types, the accumulation of L-tryptophan occurs more efficiently than that of DL-tryptophan and D-tryptophan. Using an in vitro bilateral intestinal oxygenation model system, we have demonstrated the plasma membrane of enterocyte microvilli to be rapidly impaired after oxidative stress. Glycine, but not L-tryptophan, attenuates oxidative injury in brush border membrane and alterations in amino acid transport activity. Overall, our data indicates that in vitro serosal oxygenation of the duodenum markedly improves glycine absorption, possibly involving the basolateral transporters.     

The Role of Potassium in Active Transport of Sodium by the Toad Bladder

Studies were carried out on the isolated urinary bladder of the toad, Bufo marinus, in order to explain the dependence of active sodium transport on the presence of potassium, in the serosal medium. Attempts to obtain evidence for coupled sodium-potassium transport by the serosal pump were unsuccessful; no relation between sodium transport and uptake of K⁴² from the serosal medium was demonstrable. Rather, the predominant effect of serosal potassium appeared to be operative at the mucosal permeability barrier, influencing the permeability of this surface to sodium. The mucosal effects of serosal potassium were correlated with effects on cellular cation content. When sodium Ringer's solution was used as serosal medium, removal of potassium resulted in significant decrease in tissue potassium content, commensurate increase in tissue sodium content, and marked depression of mucosal permeability and sodium transport. When choline replaced sodium in the serosal medium, removal of potassium resulted in only slight alterations of tissue electrolyte content, and effects on mucosal permeability and sodium transport were minimal.     

Physiology of the tracheal epithelium

The mucosa that lines the airways is covered with a fluid film forming a hypophase between mucus and cell surface. To study the function of this epithelium aims at describing the mechanisms by which fluid is normally produced. Another goal to be pursued consists in looking for the origin of pathological situations, such as cystic fibrosis, in which the functioning of epithelial cell is altered. The elucidation of transport mechanisms present in the apical and in the basolateral membrane results in a conceptual model that illustrates the asymmetrical functioning of epithelial cells. Recent discoveries enlarge our understanding of membrane transport processes; in particular, a concerted, reciprocal regulation of the activity of both membranes was shown to be exerted via the intracellular composition. The tracheal epithelium absorbs Na+ and secretes Cl-. These two transports are active and electrogenic; their sum corresponds approximately to the short-circuit current measured in vitro. Na+ absorption is sensitive to amiloride from the luminal side and also to ouabain added to the serosal compartment. The process is a primary active transport, analogous to that found in amphibian epithelia or in mammalian colon. Cl- secretion is abolished by furosemide (or bumetanide), by ouabain or by Na+ suppression in the serosal incubation solution. The mechanism is a secondary active transport: Cl- influx across the basolateral membrane is coupled to Na+ (probably through Na+, K+, Cl- symport); energy is dissipated by the Na+-K+-ATPase localised in the basolateral membrane. Thus, Na+ is recirculated across that membrane by the pump activity, which maintains a favorable gradient for influx via the symport. Cl- efflux takes place by diffusion through the luminal membrane. This model applies to other epithelia in which Na+-coupled Cl- secretion was shown to take place. It is confirmed by isotopic fluxes measurements and by electrophysiologic properties of the apical and the basolateral membrane. Various agents are known to influence ion transports. In particular Cl- secretion is stimulated by substances that increase the intracellular concentration of cyclic AMP. At the membrane level, the number of active Cl- channels in the apical membrane is primarily controlled, then the basolateral membrane K+ permeability. Yet, species differences are worth to note: the trachea of the cow is barely sensitive to agents that exert a marked action on dog trachea. The tracheal epithelium is used as an experimental model for studying cystic fibrosis, a disease in which the apical membrane is almost devoid of functional Cl- channels, so that Cl- permeability is quite low.(ABSTRACT TRUNCATED AT 400 WORDS)     

Localization and characterization of transport-related elements in the plasma membrane of turtle bladder epithelial cells

A mixed membrane preparation obtained from turtle bladder epithelial cells contains (Na⁺ + K⁺)-ATPase, adenylate cyclase and protein kinase, which interact with ouabain, norepinephrine and cyclic AMP, respectively. When such a preparation is obtained from bladders which had been preexposed to serosal fluids containing the tritiated form of 4,4′-diisothiocyano-2,2′-disulfonic stilbene, the subsequently isolated membrane proteins are enriched in tritium as well as in the afore-mentioned enzymes, none of which is inhibited. Freeflow electrophoresis separates the mixed membrane preparation into two distinguishable groups: one, construed as apical membranes, is enriched in norepinephrine-sensitive adenylate cyclase and cyclic AMP-sensitive protein kinase; the other, construed as basal-lateral membranes, is enriched in ouabain-sensitive ATPase and 4,4′-diisothiocyano-2,2′-disulfonic stilbene-binding proteins.The physiological counterparts of these enzymatically defined membrane markers are the mucosal sidedness of the transport effects of norepinephrine and cyclic AMP derivatives and the serosal sidedness of the transport effects of ouabain and disulfonic stilbenes in the intact turtle bladder. The discreteness and ion selectivity of each membrane-bound, transport-related element are discussed in relation to the corresponding characteristics of each transport process in vivo; the possibility of regulation of anion transport by adenylate cyclase-protein kinase system is also discussed.     

Effect of peptide conformation on membrane permeability.

The effect of peptide conformational constraint on the peptide permeation across the model membranes was examined by determining the permeability of pairs of cyclic and acyclic peptides related to c[d-Pen2, d-Pen5] enkephalin (DPDPE). The peptides were cyclized by formation of an intramolecular disulfide bridge between the second and fifth residues composed of either d-penicillamine or cysteine. In each case the acyclic peptide was three to seven times more permeable than corresponding cyclic peptide. The possibility that the differences in permeability of cyclic and acyclic peptides is based on the greater conformational freedom of the acyclic peptides in the presence of membrane was examined in more detail by isothermal titration calorimetric studies of Trp6-DPDPE and its acyclic analog. The membrane binding of the acyclic peptide is a more exothermic process than binding of its cyclic Trp6-DPDPE. The transfer of acyclic peptide from water to membrane is an enthalpy driven process, whereas the transfer of the cyclic peptide is driven by entropy.