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Biomedical subjects

J G Forte

Publications and source records attributed to J G Forte.

At least 109 records · Page 6Linked to original sources
PubMed1978

A study of H+ transport in gastric microsomal vesicles using fluorescent probes.

Fluorescent amines, 9-aminoacridine, acridine orange and quinacrine, were used as probes for a pH gradient (deltapH) across gastric microsomal vesicles. Analysis of probe uptake data indicates that 9-aminoacridine distributes across the membrane as a weak base in accordance with the deltapH. On the other hand, acridine orange and quinacrine show characteristics of binding to membrane sites in addition to the accumulation in response to deltapH. A discussion of the advantages and limitations of the probes is presented. Application of these probes to pig gastric microsomal vesicles indicates that that K+-stimulated ATPase is responsible for the transport of H+ into the vesicles and thus develops a deltapH across the membrane. The deltapH generated by the K+-ATPase has a definite requirement for internal K+. The proton gradient can be discharged slowly after ATP depletion or rapidly either by detergent disruption of the vesicles or by increasing their leakiness using both H+ and K+ ionophores. On the other hand, the sole use of the K+ ionophore, valinomycin, stimulates the ATP-induced formation of deltapH by increasing the availability of K+ to internal sites. This stimulation by valinomycin requires the presence of permeable anions like Cl-. Analysis of the Cl- requirement indicates that in the presence of valinomycin the net effect is the accumulation of HCl inside the gastric vesicles. With an external pH of 7.0, the ATP-generated deltapH was calculated to be from 4 to 4.5 pH units. The results are consistent with the hypothesis that the K+-stimulated ATPase drives a K+/H+ exchange across the gastric vesicles. Since other lines of evidence suggest that these gastric microsomes are derived from the tubulovesicular system of the oxyntic cell, the participation of the ATP-driven transport processes in gastric HCl secretion is of interest.

Acridines↗
PubMed1978

Na+ transport by mammalian stomach.

Gastric mucosas from newborn pigs (0--20 days) and rabbits (0--20 days) were used for in vitro investigation of active Na+ transport during resting (no HCl secretion) conditions. As measured with 22Na+, these tissues actively absorb Na+ from the mucosal-to serosal (m-t-s) bathing solution during both open-circuit and short-circuit current (Is) conditions. In the nonsecreting state, net Na+ transport accounts for 40--60% of Isc. The remaining current is provided by net s-to-m flux of Cl-. Amiloride (2-5 X 10(-5) M) in the mucosal solution abolishes this active Na+ transport by inhibiting m-to-s fluxes of Na+ (JNams). In vivo-in vitro experiments showed that active Na+ transport is a normal function of the resting mammalian stomach. Decreasing pH of the mucosal solution below pH 5 reversibly causes decreased current-generating capability of the tissue. Pretreatment of the tissue with amiloride abolishes this pH effect. The implication is that the low pH affects the Na+-entry step into cells. "Titration curves" of current vs. pH had an apparent pK approximately 4.0. Ouabain and K+-free solutions both cause decreases in active Na+ and Cl- current. Calculations indicate that a shunt may account for only a small (less than 30%) percentage of total transepithelial conductance.

Amiloride↗
PubMed1977

Gastric adenosine triphosphatases: a review of their possible role in HCl secretion.

In the search for gastric ATPases that might be related to the mechanism of HCl secretion, an interesting and rather unique K+-stimulated ATPase has been discovered. This enzyme is isolated from oxyntic cells and has been associated with the apical plasma membrane and/or tubulovesicular system. Membrane vesicles containing the K+-stimulated ATPase transport H+ into the vesicular lumen under the appropriate conditions of ATP, Mg2+, and KCl. This process can be measured by pH electrode, binding of certain metachromatic dyes to "energized" sites, or accumulation ratios of substances with appropriate pK values. Vesicular interior can be acidified to pH 3.5 or below. At the present time, it is difficult to distinguish between an electrogenic H+ pump and an electroneutral H+/K+ exchange mechanism. A hypothetical scheme for the gastric H+ secretory mechanism is proposed which fits much of the data from studies on the K+-ATPase, vesicular transport, and intact gastric mucosa.

Acridines↗
PubMed1976

Studies on the phosphorylated intermediates of a K+-stimulated ATPase from rabbit gastric mucosa.

A density gradient-purified microsomal membrane preparation from rabbit fundic gastric mucosa was used for a detailed study of the K+-stimulated ATPase and associated intermediate reactions. Membranes incubated with gamma-[32P]ATP show the rapid incorporation of 32P into phosphoprotein. Phosphoprotein levels were markedly reduced (1) when ATP hydrolysis went to completion or (2) upon addition of unlabeled ATP, thus suggesting the participation of a rapid turnover phosphorylated intermediate in the gastric microsomal ATPase. Addition of K+, Rb+ or Tl+ greatly reduced the level of the intermediate while stimulating ATPase activity; the observed affinities of these cations were similar for the effects on both ATPase and intermediate levels, with Tl+ greater than K+ greater than Rb+. Neither ATPase nor intermediate were stimulated by Na+, and ouabain was without effect on the reactions, thus differentiating this system from the (Na+ + K+)-ATPase. Addition of various inhibitors showed differential effects on the partial reactions of the gastric ATPase system. N-ethylmaleimide and Zn2+ showed characteristics of completely abolishing the K+-stimulated component of ATPase as well as the effects of K+ in reducing the level of intermediate, thus suggesting that these agents exert their inhibitory effect on a phosphoprotein phosphatase partial reaction. F- abolished the K+-stimulated ATPase, but its more complex effects on the intermediate suggested an additional reaction step within the domain of the phosphorylated intermediate. Results are consistent with a model system for the gastric microsomal ATPase involving a Mg2+-dependent protein kinase, a phosphorylated intermediate(s), and a K+-stimulated phosphoprotein phosphatase.

Adenosine Diphosphate↗
PubMed1975

Histamine-stimulated hydrogen ion secretion by in vitro piglet gastric mucosa.

1. A new preparation of gastric mucosa isolated from new-born piglets is described. The piglet gastric mucosa was easily separated from the serosal muscle layers by a "blistering" technique which appeared to cause minimal trauma to the tissue and which allowed extended study in vitro in a suitable chamber. Normal resting p.d. was approximately minus 30 mV (mucosal side negative with respect to serosal side), resistance about 100 omega. cm-2 and H+ secretion was absent or occurred at very low rates (0-1mu-equiv/cm-2. hr). 2. Maximally stimulating doses of histamine (1-6 times 10-5 M) caused H+ secretion to increase (up to 15 muequiv/cm-2. hr), p.d. to increase and resistance to decrease. A close correlation was observed between the increase in H+ secretion and decrease in transmucosal resistance. The threshold dose of histamine appeared to be 10-8 M; concentrations 10-4 M and higher reduced H+ secretion somewhat. 3. Pentagastrin ( 10-9-10-7 M) and acetylcholine (10-7-10-5 M) did not significantly stimulate the piglet gastric mucosa. Pentagastrin concentrations above 4 times 10-6 M reversibly inhibited H+ secretion of histamine-stimulated mucosa. High concentrations of acetylcholine (above 4 times 10-4 M) did not affect histamine-stimulated H+ secretion, but a significant reduction in p.d. was observed. 4. This investigation demonstrates the utility of the piglet gastric mucosa for in vitro studies of the mechanism H+ secretion and the action of secretagogues. From a consideration of such factors as the thinness of tissue and ease of preparation it is suggested that neonatal animals may represent a good source of in vitro mammalian gastric tissue.

Acetylcholine↗
PubMed1975

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.

Anaerobiosis↗
PubMed1975

Effects of bile salts on amphibian gastric mucosa.

The effect of five different bile salts on the mucosal or serosal surface of isolated bullfrog gastric mucosa was investigated. Serosal exposure (30 min) to 15-40 mM taurocholate (TCA) or taurodeoxycholate (TDCA), 10-40 mM deoxycholate (DCA), or 2.5-7.5 mM cholate (CA) typically produced a marked increase in resistance (R) and a decrease in potential difference (PD), short-circuit current (I-sc), and H+ secretion. Flux of 36Cl from serosal to mucosal solution decreased progressively. The flux of [14C]sucrose across the mucosa was slightly decreased. Microscopy showed condensation of the parietal cells. Mucosal exposure (30 min) at concentrations above 2.5 mM DCA or CA and 15 mM TCA and TDCA caused a marked decrease in R, PD, I-sc, and H+ secretion, and the flux of [14C]sucrose across the mucosa was increased. Extreme destruction of the surface mucous cells was the primary histologic finding. The effect of dehydrocholate was identical to that of TCA. Mucosal bile salts cause an increased permeability primarily by injury of the surface epithelium whereas serosal bile salts affected primarily the oxyntic cells.

Animals↗
PubMed1975

Acid-base balance in amphibian gastric mucosa.

It has been established that H+ secretion can be maintained in frog stomach in the absence of exogenous CO2 by using a nutrient bathing fluid containing 25 mM H2PO4 (pH approximately equal to 4.5) or by lowering the pH of a nonbuffered nutrient solution to about 3.0-3.6. Exogenous CO2 in the presence of these nutrient solutions uniformly caused a marked decrease in H+ secretion, PD, adn short-circuit current (Isc) and an increase in transmucosal resistance (R). Elevation of nutrient [k+] to 83 mM reduced R significantly but transiently without change in H+ when nutrient pH less than 5.0, whereas R returned to base line and H+ increased when nutrient pH greater than 5.0. Acidification of the nutrient medium in the presence of exogenous CO2 results in inhibition of the secretory pump, probably by decreasing intracellular pH, and also interferes with conductance at the nutrient membrane. Removal of exogenous CO2 from standard bicarbonate nutrient solution reduced by 50% the H+, PD, and Isc without change in R; K+-free nutrient solutions reverse these changes in Isc and PD but not in H+. The dropping PD and rising R induced by K+-free nutrient solutions in 5% CO2 - 95% O2 are returned toward normal by 100% O2. Our findings support an important role for exogenous CO2 in maintaining normal acid-base balance in frog mucosa by acting as an acidifying agent.

Acid-Base Equilibrium↗
PubMed1975

Ultrastructural and physiological changes in piglet oxyntic cells during histamine stimulation and metabolic inhibition.

Neonatal pig gastric mucosa was studied in order to correlate electrophysiological and secretory parameters with ultrastructural changes in membrane components of oxyntic cells. The non-stimulated tissue had a transmucosal resistance of about 130omega - cm2 while the oxyntic cells were characterized by numerous cytoplasmic tubulovesicles and short microvilli extending into patent glandular and canalicular lumina. Upon histamine-stimulation, the average rate of H+ secretion was 8.1 mueq - cm2 - hr-1 and the resistance decreased to 77omega - cm2. The changes were coupled with an immense elaboration of the oxyntic cell apical and canalicular surfaces with a concomitant decrease of tubulovesicles. Thus, the observed decrease in resistance was correlated to large increases in secretory membrane area. Anoxia inhibited H+ secretion while resistance increased to 211omega - cm2. Anoxic oxyntic cells were characterized by swollen mitochondria and occlusion of the lateral intercellular space and basal infoldings. Little change in the configuration of the secretory surfaces was noted, thereby suggesting that restriction of lateral and basal membranes might be responsible for the observed resistance increase. An electrical analogue of gastric mucosa is proposed on the basis of these morphological observations.

Animals↗
PubMed1975

Unique enzymes of purified microsomes from pig fundic mucosa. K+-stimulated adenosine triphosphatase and K+-stimulated pNPPase.

Microsomal fractions from homogenates of pig gastric fundic mucosa showed high levels of K+-stimulated adenosine triphosphatase (ATPase) and K+-stimulated phosphatase. Similar preparations from antral mucosa showed virtually no such activity. Because of mitochondrial contamination the fundic microsomes were further separated by sucrose density gradient centrifugation. A low density band of membranes (peak 1.12 to 1.13 g per ml) possessed all of the K+-stimulated enzyme activities. Morphological features and the abundant glycoproteins of the low density microsomes suggested they might be derived from the tubulovesicles of oxyntic cells. Mitochondrial and ribosomal markers were associated with membranes with much higher densities (greater than 1.22). The K+-stimulated ATPase has a pH optimum of 7.5 and required Mg++, but neither Na+ nor ouabain had any appreciable effect on the activity. Stimulation of basal ATPase by K+ ranged from 1.5 to 3.0-fold with an apparent Ka for activation between 0.2 to 0.4 mM K+. Addition of various K+ ionophoretic substances (e.g., gramicidin) produced further stimulation of K+-ATPase up to 6 times the basal rate. The mean activities for seven separate preparations of purified low density pig fundic microsomes were as follows (micromoles of ATP hydrolyzed per mg protein per hr +/- SEM); basal ATPase, 15.8 +/- 2.8; plus 10 mM K+, 29.3 +/- 4.5; plus 10 mM K+ and 10(-5) M gramicidin, 45.2 +/- 5.2. Neither the basal ATPase nor the K+-stimulated rates were altered by HCO3- or Cl-. The occurrence of these active and unique enzyme activities in the oxyntic region of gastric mucosa suggest some relation with secretory activity. Possible functional roles are discussed.

4-Nitrophenylphosphatase↗