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D Cremaschi

Publications and source records attributed to D Cremaschi.

At least 55 records · Page 3Linked to original sources
PubMed1989

Enkephalin regulation of L-valine transport in rabbit ileum.

We studied the influence on ionic basal transport (Na+ and Cl-) and L-valine transport of two enkephalins which are not metabolized and act in delta and mu receptors respectively. Transports have been indirectly determined measuring the transepithelial electric potential and the short circuit current. DADLE does not significantly influence ion and amino-acid transport, while DAGO alters both of them in the presence of the myenteric plexus (muscle layers present) or inhibits only L-valine transport in the absence of the plexus (muscle layers removed).

Animals↗
PubMed1988

Uncoupling of Na+H+ from Cl-HCO3- exchange under some steady state conditions in rabbit gallbladder.

The transapical Cl- influx and transepithelial Na+ transport were measured in rabbit gallbladder. Only 11.7% of the transported Na+ was found to be accompanied by HCO3-. 10(-4) M SITS eliminated the HCO3- dependent fraction of Cl- influx (50%) but did not significantly alter intracellular Na+ activity and Na+ transport. Exposure to HCO3-free salines or to 10(-4) M acetazolamide about halved Cl- influx and Na+ transport. 25 mM SCN- reduced Cl- influx to zero, decreased intracellular Na+ activity, but only halved Na+ transport which under these conditions was abolished only in the absence of HCO3-. Exposure to a Cl- -free saline produced effects similar to those caused by SCN-. These results suggest that when Cl-/HCO3- exchange is inhibited at the apical membrane, Na+/H+ exchange and transepithelial Na+ transfer are unmodified if HCO3- is available for transport. The permanent uncoupling of the exchangers and the elevated transepithelial transport of Na+ are not due to an increased activity of the parallel Na+-Cl- cotransport but to a redirection of HCO3- flux toward the basolateral side.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1987

Isolation of apical plasma membrane in rabbit gallbladder epithelium by Percoll density gradient centrifugation.

The apical membranes of rabbit gallbladder epithelial cells were isolated by treating the homogenate with Ca2+ or Mg2+ and centrifuging the suspension in Percoll gradient. In this way brush-border membranes were obtained with enrichment factors ranging between 10 and 20 and yields of 15-30%. A second method is described with which membranes were isolated, without any preliminary treatment, first by differential centrifugation, then with Percoll gradient; the final membrane enrichment was over 15, however the yield was very low (3%). Many possible enzymatic markers of the apical plasma membrane were investigated: L-gamma-glutamyltransferase, alkaline phosphatase, leucine aminopeptidase, sucrase. The first appears to be that of choice. Apical membrane fraction could be also evidenced by autofluorescence or by labeling with Lotus tetragonolobus lectin. Preliminary experiments showed that apical plasma membranes isolated in this way form vesicles.

Animals↗
PubMed1987

The nature of the neutral Na+-Cl(-)-coupled entry at the apical membrane of rabbit gallbladder epithelium: I. Na+/H+, Cl-/HCO3- double exchange and Na+-Cl- symport.

Cl- influx at the luminal border of the epithelium of rabbit gallbladder was measured by 45-sec exposures to 36Cl- and 3H-sucrose (as extracellular marker). Its paracellular component was evaluated by the use of 25 mM SCN- which immediately and completely inhibits Cl- entry into the cell. Cellular influx was equal to 16.7 mu eq cm-2 hr-1 and decreased to 8.5 mu eq cm-2 hr-1 upon removal of HCO3- from the bathing media and by bubbling 100% O2 for 45 min. When HCO3- was present, cellular influx was again about halved by the action of 10(-4) M acetazolamide, 10(-5) to 10(-4) M furosemide, 10(-5) to 10(-4) M 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonate (SITS), 10(-3) M amiloride. The effects of furosemide and SITS were tested at different concentrations of the inhibitor and with different exposure times: they were maximal at the concentrations reported above and nonadditive. In turn, the effects of amiloride and SITS were not additive. Acetazolamide reached its maximal action after an exposure of about 2 min. When exogenous HCO3- was absent, the residual cellular influx was insensitive to acetazolamide, furosemide and SITS. When exogenous HCO3- was present in the salines, Na+ removal from the mucosal side caused a slow decline of cellular Cl- influx; conversely, it immediately abolished cellular Cl- influx in the absence of HCO3-. In conclusion, about 50% of cellular influx is sensitive to HCO3-, inhibitable by SCN-, acetazolamide, furosemide, SITS and amiloride and furthermore slowly dependent on Na+. The residual cellular influx is insensitive to bicarbonate, inhibitable by SCN-, resistant to acetazolamide, furosemide, SITS and amiloride, and immediately dependent on Na+. Thus, about 50% of apical membrane NaCl influx appears to result from a Na+/H+ and Cl-/HCO3- exchange, whereas the residual influx seems to be due to Na+-Cl- cotransport on a single carrier. Whether both components are simultaneously present or the latter represents a cellular homeostatic counter-reaction to the inhibition of the former is not clear.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1987

The nature of the neutral Na+-Cl(-)-coupled entry at the apical membrane of rabbit gallbladder epithelium: II. Na+-Cl- symport is independent of K+.

In the epithelium of rabbit gallbladder, in the nominal absence of bicarbonate, intracellular Cl- activity is about 25 mM, about 4 times higher than intracellular Cl- activity at the electrochemical equilibrium. It is essentially not affected by 10(-4) M acetazolamide and 10(-4) M 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonate (SITS) even during prolonged exposures: it falls to the equilibrium value by removal of Na+ from the lumen without significant changes of the apical membrane potential difference. Both intracellular Cl- and Na+ activities are decreased by luminal treatment with 25 mM SCN-; the initial rates of change are not significantly different. In addition, the initial rates of change of intracellular Cl- activity are not significantly different upon Na+ or Cl- entry block by the appropriate reduction of the concentration of either ion in the luminal solution. Luminal K+ removal or 10(-5) M bumetanide do not affect intracellular Cl- and Na+ activities or Cl- influx through the apical membrane. It is concluded that in the absence of bicarbonate NaCl entry is entirely due to a Na+-Cl- symport on a single carrier which, at least under the conditions tested, does not cotransport K+.

Animals↗
PubMed1986

Intracellular potassium as a possible inducer of amino acid transport across hamster jejunal enterocytes.

Brush border membrane potentials (Vm), intracellular K+ activity (aiK) and alanine uptake were measured in different parts of villi in mid-jejunal tissue taken from hamsters fed different amounts of food at high and low environmental temperatures. Basal villus enterocytes (Y cells) were found to have lower values for Vm and aiK than upper villus enterocytes (O cells). Alanine uptake was confined to O cells. The position on the villus where values for Vm and aiK changed, and where alanine uptake could first be seen to take place, depended on the energy intake and environmental temperature at which hamsters were kept. Na+-dependent alanine uptake and Vm were both higher in O cells of hamsters fed 10 kcal day-1 at 30 degrees C (10 k/30 degrees C) compared with animals fed 30 kcal day-1 at an environmental temperature of 12 degrees C (30 k/12 degrees C hamsters). The rates at which enterocytes migrated along the crypt-villus axis, measured separately in thymidine-labelling experiments, were 6.9 and 16.1 micron h-1 for 10 k/30 degrees C and 30 k/12 degrees C hamsters respectively. Both Vm and aiK were estimated, from these measurements, to have increased significantly by the time enterocytes became 30 h old. Alanine uptake began 15 h later. Neither of these parameters were influenced by previous adaptation conditions. The close temporal and variable positional relationship found between changes in aiK and onset of transport suggests that early changes in electrolyte composition might initiate a second phase of development enabling the enterocyte to absorb nutrients. The possibility that other ions besides K+ might also be involved in this aspect of regulation is also discussed.

Alanine↗
PubMed1985

Construction of K+- and Na+-sensitive theta-microelectrodes with fine tips: an easy method with high yield.

A new method is described to prepare theta-microelectrodes with tips up to 0.15 micron diameter controlled under scanning electron microscope. K+- and Na+-sensitive resins were tested. Method features are the following: i) hard drying of the glass, ii) rehydration of one channel and weak wetting of the other with a three-methylchlorosilane solution before pulling, iii) simultaneous presence of water and silane in the two channels during pulling, iv) gradual silanization from the tip to the shank. Selective and conventional channels did not affect each other and no displacements of resins were observed. The change of potential difference of the selective channel was more than -50 mV/decade. Apical membrane potentials and cell Na+ and K+ activities of the epithelial cells of rabbit gall-bladder (cell diameter: 5-10 micron) were measured with these theta-microelectrodes and with single-barrel microelectrodes of similar tip size: results obtained were not significantly different.

Animals↗
PubMed1984

Positional dependence of enterocyte membrane potential in hamster and rabbit enterocytes.

A technique is described allowing microelectrode impalement of enterocytes located at known positions along intestinal villi from rabbits and hamsters. Using this technique a 5 mV hyperpolarization in membrane potential is shown to occur as enterocytes migrate over the basal third of intestinal villi. The villus structure of the hamster ileum is similar to the rabbit, but the enterocyte lifespan in these two tissues differs considerably (enterocyte migration rates of 17.6 and 6.3 microns hr-1 for hamster and rabbit respectively). A correlation was found between the position an enterocyte occupied on the crypt-villus axis and the developmental state of the membrane potential. No such correlation existed when making comparisons on a time basis. These results are discussed both in terms of what is now known concerning different aspects of enterocyte development and in relation to what type of control mechanism might be generally responsible for initiating differentiation in this tissue.

Animals↗
PubMed1984

Developmental changes in intra-enterocyte cation activities in hamster terminal ileum.

Intracellular Na+ and K+ activities have been determined under visual control in enterocytes located at known positions along hamster ileal villi in vitro. The intracellular K+ activity found in enterocytes migrating over the basal third of villi was significantly less than that found in older enterocytes (83 and 113 mM respectively). The intracellular Na+ activity remained unaffected by the state of enterocyte development (32 mM throughout). Extracellular K+ and Na+ activities measured close to the luminal surface of the intestine were always similar to those measured in the bulk of the bathing medium. There was, however, a small difference in K+ activity detected between the bulk solution and solution near the luminal surface of enterocytes (3 . 4 and 3 . 6-3 . 7 mM respectively). Na+ activity in the villus core was also similar to that of the bathing medium. The brush border membrane content of sucrase-isomaltase, determined cytochemically, was found to increase rapidly as enterocytes migrated over the basal third of the villus and to plateau at a point roughly equivalent to that where intracellular K+ activities became maximal. The numerous ways by which enterocytes might control their intracellular K+ concentrations during development are emphasized. Attention is nevertheless drawn to the close correspondence seen between different aspects of brush border membrane development and the ability of enterocytes to maintain high internal concentrations of K+.

Animals↗
PubMed1983

Different sodium chloride cotransport systems in the apical membrane of rabbit gallbladder epithelial cells.

The kinetics of Cl- influx from the lumen to the cell and the paracellular pathway was examined in isolated rabbit gallbladder by measuring 36Cl uptake (45 s) and by correcting it for the extracellular space with 3H-sucrose. The paracellular fraction of the influx was studied by incubating the tissue in Na+-free saline or in solutions containing 25 mM SCN-; the kinetics turned out to be hyperbolic. The cellular fraction of the influx comprised three components. The first was immediately Na+-dependent and insensitive both to exogenous and endogenous cell bicarbonate; its sigmoidal kinetics revealed the presence of a carrier with three Cl- binding sites cooperating positively with one another, with strong interaction factors. The second cellular component was immediately Na+-dependent and sensitive to endogenous cell bicarbonate; the kinetics was hyperbolic with a maximum at 20 mM Cl- concentration and a substrate inhibition from 20 to 130 mM; it was completely inhibited by 10(-4)M acetazolamide. The third cellular component was slowly Na+-dependent and slowly sensitive to exogenous bicarbonate; its kinetics was hyperbolic, without substrate inhibition in the tested Cl- concentration range. On this basis, the presence of three Na+-Cl- cotransports is suggested: i) on a single carrier without any exchange with H+ and HCO3-, ii) on a single carrier with an exchange with H+ and HCO-, and, and iii) on two separate carriers in exchange with H+-and HCO3-.

Acetazolamide↗
PubMed1983

Bicarbonate effects, electromotive forces and potassium effluxes in rabbit and guinea-pig gall-bladder.

The stimulating effect of external HCO3- on Na+ salt transport has been examined in rabbit and guinea-pig gall-bladder by electrophysiological methods, as a sequel to a previous study carried out by radiochemical techniques. At steady state, cell K+ activity was found to be significantly reduced in the presence of HCO3-, whereas cell Na+ activity significantly increased; in parallel the apical membrane p.d. was depolarized; K+ equilibrium potential was higher than membrane p.d. in every case. The apical p.d. dependence on K+ was unaffected by HCO3-, but in the guinea-pig it was affected by Cl-. Rapid increases in HCO3- concentration on the luminal side caused a depolarization of the apical p.d. of the guinea-pig within about 30 sec, an effect that did not occur if the tissue was pre-treated with 10(-4) M-acetazolamide; the epithelial resistance and apical/basolateral resistance ratio were unchanged in all cases. The primary action of HCO3- is confirmed to be on the apical membrane; an HCO3- conductance does not seem to be present at this level, either in the rabbit or guinea-pig, nor does HCO3- affect Na+ influx through the apical conductive pathway, so that all the stimulating effects of the anion are confirmed to be on the neutral transports of Na+ salts; in spite of this, the apical electromotive force is modified due to the changed cell K+ activity. The rapid depolarization caused by the anion in the guinea-pig is in agreement with an HCO3- electrogenic secretion and/or a basolateral conductance for the anion. Polyelectrolyte dissociation from protons increases in the absence of external HCO3-: the negative charges are mainly counterbalanced by bound Na+ in the rabbit and by free K+ in the guinea-pig. K+ leakage from the cell into the lumen is calculated to be minimal in the rabbit and all K+ lost could be reabsorbed through the paracellular pathways; K+ efflux to the subepithelial layer via conductive routes is insufficient to account for the over-all K+ efflux.

Animals↗
PubMed1982

Membrane potentials of differentiating enterocytes.

A positional analysis of enterocyte membrane potential has been carried out using in vitro preparations of rabbit distal ileum. Young enterocytes were found to possess a microvillar membrane potential significantly less than that seen in older enterocytes. The length of enterocyte microvilli was also found to be significantly less in younger enterocytes. It is suggested that developmental changes in membrane potential, occurring during the early stages of enterocyte differentiation, probably reflect a changed permeability to ions associated with the establishment of a fully developed microvillar membrane. Other explanations for the observed findings are also considered.

Age Factors↗
PubMed1982

Amiloride-sensitive sodium channels in rabbit and guinea-pig gall-bladder.

1. The effects of low amiloride concentrations (less than 2.5 x 10(-5) M) on the apical membrane (Vm) and transepithelial (Vms) potential differences, transepithelial resistance (Rep) and apical/basolateral membrane resistance ratio (Rm/Rs) have been determined in the gall-bladders of the rabbit and guinea-pig. 2. Vm was hyperpolarized, Vms decreased towards negative values, Rm/Rs increased, but Rep remained unchanged. 3. K+ selectivity of the apical membrane was also checked by increasing luminal K+ concentration from 5.9 to 29.4 mM; the corresponding change in the apical electromotive force was much higher in the rabbit than in the guinea-pig. 4. Pre-incubations with Cl--and HCO3--free salines did not modify K+ selectivity in the rabbit, but nearly doubled it in the guinea-pig. 5. Pre-incubations with control solutions containing 10(-5) M-amiloride in the lumen converted the apical membrane of rabbit gall-bladder into a perfectly K+-perm-selective membrane; the same results were obtained in the guinea-pig only on bathing the tissue with Cl-- and HCO3--free salines together with 10(-5) M-amiloride. 6. It is suggested that in the apical membrane of rabbit gall-bladder K+ and Na+ conductances exist, whereas in guinea-pig gall-bladder both K+, Na+ and anion (Cl- and/or HCO3-) conductances are present; in both species conductive Na+ pathways are inhibited by amiloride. .

Amiloride↗
PubMed1981

Electrophysiology of pig distal colon measured during early post-natal development.

1. Brush border membrane (Vm) and transepithelial (Vms) potential differences, transepithelial resistance (Rep) and the brush border membrane/basolateral membrane resistance ratio (Rm/Rs) have been measured in distal colons taken from new-born and 4-day-old pigs. 2. Vm remained independent of the age of the animal. Substituting choline or Tris for Na+ hyperpolarized Vm; substituting SO4(2-) for Cl- or increasing the concentration of K+ depolarized Vm in colons taken from new-born pigs. K+ depolarization was not seen using colons taken from 4-day-old animals. 3. The effect of Na+ or Cl- replacement on Vms and Rm/Rs was in agreement with the hypothesis that the brush border membrane of the distal colon possessed a conductance for both these ions. The brush border membrane of the new-born, but not the 4-day-old pig colon possessed an additional conductance for K+. 4. These electrical measurements were repeated using 4-day-old pig distal colons in the presence of amiloride, to enable values for the electromotive force across the brush border membrane (Em) and the basolateral membrane (Es) to be calculated. The Em of the 4-day-old pig colon was found to be about 23 mV negative with respect to the mucosal solution. Cl- conductance appears to predominate in colons taken from these very young animals. 5. Separate experiments showed net transport of Na+ and measured short-circuit current to be partly dependent upon the presence of Cl. Na+ transport measured in the absence of Cl- was approximately equal to that calculated from the measurement of short-circuit current. 6. It is suggested that the brush border membrane of the neonatal pig colon is conductive to both Na+ and Cl-, that the transport of these ions are partly coupled in some way and that this alone is sufficient to explain the discrepancies noted previously when measuring Na+ flux and short-circuit current in Cl- -containing medium. The possible physiological importance of this transient transport function is discussed.

Animals↗
PubMed1980

Nonelectrolyte fluxes across gastric mucosa in relation to gastric stimulation. Is gastric juice secreted by osmosis or exocytosis?

The effects of histamine and thiocyanate, added to the serosal bathing solution, on unidirectional fluxes of some nonelectrolytes (thiourea, methylated thiourea derivates, mannitol), and on H+, pepsinogen and mucous secretion were investigated in frog (Rana esculenta) fundic gastric mucosa. Histamine (10(-4) M) increases significantly the outfluxes (serosa to mucosa fluxes) of only thiourea and its derivates (but not mannitol) and the stimulation is the greater the more lipidsoluble the nonelectrolyte is. Influxes (mucosa to serosa fluxes) of the same molecules are not affected. In parallel histamine stimulates H+-secretion but does not modify pepsinogen and mucous secretion. SCN- (10(-2) M) inhibits the histamine effect on thiourea outfluxes and on H+-secretion, while pepsinogen and mucous secretion are not affected. Colchicine (10(-4) M) pretreatment inhibits the histamine effect on outfluxes and H+-secretion. It is concluded that: (1) histamine induces a secretion of nonelectrolytes towards the lumen; (2) such secretion is correlated with the hormone-induced secretion of HCl and fluid; (3) this process is mediated by an exocytotic mechanism.

Animals↗
PubMed1979

Stimulation by HCO3- of Na+ transport in rabbit gallbladder.

Bicarbonate presence in the bathing media doubles Na+ and fluid transepithelial transport and in parallel significantly increases Na+ and Cl- intracellular concentrations and contents, decreases K+ cell concentration without changing its amount, and causes a large cell swelling. Na+ and Cl- lumen-to-cell influxes are significantly enhanced, Na+ more so than Cl-. The stimulation does not raise any immediate change in luminal membrane potential and cannot be due to a HCO3(-)-ATPase in the brush border. The stimulation goes together with a large increase in a Na+-dependent H+ secretion into the lumen. All of these data suggests that HCO3- both activates Na+--Cl- cotransport and H+--Na+ countertransport at the luminal barrier. Thiocyanate inhibits Na+ and fluid transepithelial transport without affecting H+ secretion and HCO3(-)-dependent Na+ influx. It reduces Na+ and Cl- conentrations and contents, increases the same parameters for K+, causes a cell shrinking, and abolishes the lumen-to-cell Cl- influx. It enters the cell and is accumulated in the cytoplasm with a process which is Na+-dependent and HCO3(-)-activated. Thus SCN- is likely to compete for the Cl- site on the cotransport carrier and to be slowly transferred by the cotransport system itself.

Animals↗
PubMed1979

Colchicine effect on the permeability of the whole epithelium and of isolated cells of frog skin.

The effect of 2 X 10(-5) M colchicine on epithelial cells isolated from frog skins was investigated. Three hours of treatment with colchicine did not change either Na+ and K+ content of isolated cells or nonelectrolyte permeability. When ADH (50 mU/ml) was added, thiourea uptake values became greater than without the hormone; the same values were found in the cells previously treated with colchicine. Na+ transepithelial transport, measured by means of short-circuit current, was inhibited by the antimitotic agent both under control conditions and after ADH stimulation. These results support the view that colchicine does not directly affect ADH action on membrane permeability, but influences some mechanism that controls ADH action on transepithelial transport. Intercellular junctions appear to be the location of such a mechanism.

Animals↗