Visualization of the secretory canaliculi of human parietal cells with a peroxidase-labelled peanut lectin

Summary Peanut lectin reactivity was examined in normal fundic glands from human gastric samples, both at light- and electron-microscopic levels, using a peroxidase conjugate. Positive reaction was observed in the glycocalyx of parietal cell secretory canaliculi as well as in the mucous globules of mucous cells and in the luminal cell coat of chief cells. The presence of terminal galactose in the canalicular glycocalyx may be connected with the peculiar function of hydrochloric acid secretion. Peroxidase-labelled peanut lectin is proposed as a marker for visualizing the secretory canaliculus of parietal cells.     

CORRELATION OF THE FINE STRUCTURE OF THE GASTRIC PARIETAL CELL (DOG) WITH FUNCTIONAL ACTIVITY OF THE STOMACH

The fine structure of the parietal (oxyntic) cell in the gastric glands (corpus of the stomach) of the dog was examined under conditions of active gastric acid secretion and compared with cellular structure in the non-acid-secretory (basal) state. Animals, in both acute and chronic experiments, were equipped with gastric fistulae so that gastric juice could be collected for analysis of total acidity, free acidity, volume, and pH prior to biopsy of the gastric mucosa. The specimens of mucosa were fixed in buffered OsO₄ and embedded in n-butyl methacrylate and the thin sections were stained with lead hydroxide before examination in the electron microscope. A majority of parietal cells showed an alteration of fine structure during stimulation of gastric acid secretion by a number of different techniques (electrical vagal stimulation, histamine administration, or insulin injection). The changes in fine structure affected mainly the smooth surfaced vesicular elements and the intracellular canaliculi in the cytoplasm of the cell. The mitochondria also appeared to be involved to some extent. During acid secretion a greater concentration of smooth surface profiles is found adjacent to the walls of the intracellular canaliculi; other parietal cells exhibited a marked decrease in number of smooth surfaced elements. Intracellular canaliculi, always present in non-acid-secreting oxyntic cells, develop more extensively in cells of acid-secreting gastric glands. The surface area of these canaliculi is greatly increased by the elaboration of a large number of closely approximated and elongated microvilli. Still other parietal cells apparently in a different stage of the secretory cycle exhibit non-patent canaliculi lacking prominence; such cells have very few smooth surfaced vesicular elements. These morphological findings correlated with the acid-secretory state of the stomach provide evidence that the parietal cell participates in the process of acid secretion.     

Lectin-binding pattern in normal human gastric mucosa

Summary Normal human gastric mucosal cells were examined by light and electron microscopy using lectins as a probe. The ABC method was used with biotinylated lectins for light microscopy and HRP-labeled lectins for electron microscopy. The human gastric mucosal cells revealed specific binding patterns for each lectin by light microscopy. Among the lectins tested, in particular, DBA gave a characteristic pattern. It specifically stained the supranuclear region of surface epithelial cells and the perinuclear region of parietal cells. By electron microscopy, the stacked cisternae and the vesicles of the Golgi apparatus of the surface epithelial cells were positive for the DBA staining. These results show that the DBA-positive supranuclear region observed by light microscopy corresponds to the Golgi apparatus. In the parietal cells, DBA, RCA and ConA bound to the intracellular secretory canaliculi which are invaginations of the cell membrane running around the nucleus in the cytoplasm. Therefore, the tubular perinuclear positive region observed by light microscopy corresponds to the membranes of the intracellular secretory canaliculi. In addition, the ConA reagent stained the endoplasmic reticulum, Golgi apparatus, nuclear envelope, and cell membrane of the parietal cell, which explains the diffuse cytoplasmic staining observed at the light microscopic level with this lectin. Lectins have proved to be very useful for the evaluation of in situ cytochemical aspects of the glycoconjugates characteristic to human gastric mucosal cells.     

The unique ultrastructure of secretory membranes in gastric parietal cells depends upon the presence of H+, K+ -ATPase

Ion transporters play a central role in gastric acid secretion. To determine whether some of these transporters are necessary for the normal ultrastructure of secretory membranes in gastric parietal cells, mice lacking transporters for H+, K+, Cl-, and Na+ were examined for alterations in volume density (Vd) of basolateral, apical, tubulovesicular and canalicular membranes, microvillar dimensions, membrane flexibility, and ultrastructure. In mice lacking Na+/H+ exchanger 1 (NHE1) or the Na+-K+-2Cl- cotransporter (NKCC1), the ultrastructure and Vd of secretory membranes and the secretory canalicular to tubulovesicular membrane ratio (SC/TV), a morphological correlate of secretory activity, were similar to those of wild-type mice. In mice lacking Na+/H+ exchanger 2 (NHE2) or gastric H+, K+ -ATPase alpha- or beta-subunits, the SC/TV ratio and Vd of secretory membranes were decreased, though canaliculi were often dilated. In H+, K+ -ATPase-deficient parietal cells, canalicular folds were decreased, normally abundant tubulovesicles were replaced with a few rigid round vesicles, and microvilli were sparse, stiff and short, in contrast to the long and flexible microvilli in wild-type cells. In addition, microvilli of the H+, K+ -ATPase-deficient parietal cells had centrally bundled F-actin filaments, unlike the microvilli of wild-type cells, in which actin filaments were peripherally positioned concentric to the plasmalemma. Data showed that the absence of H+, K+ -ATPase produced fundamental changes in parietal cell membrane ultrastructure, suggesting that the pump provides an essential link between the membranes and F-actin, critical to the gross architecture and suppleness of the secretory membranes.     

Lectin-binding pattern in normal human gastric mucosa. A light and electron microscopic study

Normal human gastric mucosal cells were examined by light and electron microscopy using lectins as a probe. The ABC method was used with biotinylated lectins for light microscopy and HRP-labeled lectins for electron microscopy. The human gastric mucosal cells revealed specific binding patterns for each lectin by light microscopy. Among the lectins tested, in particular, DBA gave a characteristic pattern. It specifically stained the supranuclear region of surface epithelial cells and the perinuclear region of parietal cells. By electron microscopy, the stacked cisternae and the vesicles of the Golgi apparatus of the surface epithelial cells were positive for the DBA staining. These results show that the DBA-positive supranuclear region observed by light microscopy corresponds to the Golgi apparatus. In the parietal cells, DBA, RCA and ConA bound to the intracellular secretory canaliculi which are invaginations of the cell membrane running around the nucleus in the cytoplasm. Therefore, the tubular perinuclear positive region observed by light microscopy corresponds to the membranes of the intracellular secretory canaliculi. In addition, the ConA reagent stained the endoplasmic reticulum, Golgi apparatus, nuclear envelope, and cell membrane of the parietal cell, which explains the diffuse cytoplasmic staining observed at the light microscopic level with this lectin. Lectins have proved to be very useful for the evaluation of in situ cytochemical aspects of the glycoconjugates characteristic to human gastric mucosal cells.     

Subcellular distribution of ionic components in gastric mucosa of the guinea pig

Summary The topographical distribution of cations, anions and polyanions in the guinea-pig stomach has been studied by ultrastructural cytochemical methods. After fixation with the pyroantimonate-osmium tetroxide solution, variable-sized precipitates were localized in the basolateral extracellular space bordering parietal cells or chief cells but not in that bordering mucus-secreting cells. The basal lamina of all gastric cells disclosed a continuous layer of heavy antimonate deposits. Parietal cells disclosed uniformly fine deposits also on the apical plasmalemma both at the main lumen and in the intracellular canaliculi, and revealed, as well, coarse precipitates in the mitochondria. Fixation with a silver acetate-osmium tetroxide solution yielded nitric acid-resistant, silver deposits confined to the luminal surface of the apical plasmalemma in the main lumen and intracellular canaliculi, the lateral intercellular space, the outer surface of the basal plasmalemma and the basal lamina of the parietal cell.Staining with dialyzed iron demonstrated a glycocalyx rich in acid mucosubstance on the basolateral plasmalemma but not on the apical plasmalemma of parietal cells. In contrast, acid glycoconjugate was visualized on the apical plasmalemma of isthmus cells, mucous neck cells and the transitional cell between isthmus and mucous neck cells but little or no acidic glycoconjugate was demonstrated on the basolateral plasmalemma of these cells. The entire plasmalemma of gastroendocrine cells, unlike other epithelial cells, stained uniformly for acidic glycoconjugate. The dialyzed iron and high iron diamine methods stained the outer compartment of mitochondria in parietal cells intensely and that in other gastric cells lightly. These reagents stained the basal lamina of all gastric cells as did ruthenium red. The several characteristic cytochemical properties of parietal cells presumably relate to the unique secretory activity of these cells and are consistent with the view of the intracellular canaliculi of the parietal cell as the main route for hydrogen and chloride ion secretion.     

Effect of different photoperiods on the ultrastructure of the specific secretory cells and α-subunit mRNA level in the chicken pars tuberalis

The effect of different photoperiods on the specific secretory cells of the pars tuberalis was examined in male chicks. Animals were placed in one of three different photoperiod regimens: (1) normal control (light:dark = 12 h:12 h), (2) continuous light (L:D = 24 h:0), and (3) extended darkness (L:D = 1 h:23 h). The levels of common alpha-subunit mRNA in the pars tuberalis were examined by Northern blot analysis and compared with those in the pars distalis. In chicks exposed to continuous light for 1 week, alpha-subunit mRNA level in the pars tuberalis was decreased, although the level in the pars distalis was increased. Exposure to continuous light for 30 days also induced a decrease in alpha-subunit mRNA level in the pars tuberalis. On the other hand, in chicks exposed to extended darkness for 1 week, the alpha-subunit mRNA level of the pars tuberalis was markedly increased. In situ hybridization with digoxigenin-labeled common alpha-subunit cRNA probe also showed that the hybridization signals for alpha-subunit mRNA in the pars tuberalis cells become weak under continuous light for 30 days but they are very intense under extended darkness. Thus, the synthesis of alpha-subunits in the chick pars tuberalis was inhibited by continuous light but stimulated by extended darkness. These results were confirmed by semiquantitative electron-microscopic analyses. After exposure to continuous light for 30 days, many pars tuberalis (PT)-specific cells were filled with enlarged secretory granules, showing the reduction of secretory activity. On the contrary, extended darkness for 30 days induced hypertrophy of the PT-specific cells; the areas of cytoplasm and nucleus were significantly increased. In addition, secretory granules became small in size and exocytotic features were more frequent. Mitochondria and lysosomes were also increased in number. Thus, the synthetic and secretory activities of the PT-specific cells were increased under extended darkness. The data indicate that the specific cells of the pars tuberalis are responsive to photoperiodic changes in the chick.     

Gastric K+-stimulated p-nitrophenylphosphatase cytochemistry

A cytochemical study of gastric K+-stimulated p-nitrophenylphosphatase (K-NPPase) activity, corresponding to a K+-stimulated phosphoprotein phosphatase of H-K-ATPase system, has been made by a new cytochemical method. Sections of fixed guinea pig gastric mucosa in a mixture of 2% paraformaldehyde and 0.25% glutaraldehyde, were incubated with the incubation medium (1.0 M glycine-0.1 M KOH buffer, pH 9.0, 2.5 ml; 1.1 M KCl, 0.5 ml; 10 mM lead citrate dissolved in 50 mM KOH, 4 ml; levamisole, 6.0 mg; dimethyl sulfoxide, 2.0 ml; 0.1 M p-nitrophenylphosphate (Mg-salt), 1.0 ml; ouabain, 73.0 mg) for 30 min at room temperature. Under a light microscope the specific gastric K-NPPase reaction was distributed only in the parietal cells of the fundic glands. The electron microscopic cytochemistry showed that the gastric K-NPPase activity was localized on the membrane lining the apical surfaces, secretory canaliculi and tubulovesicles. On the other hand, ouabain-sensitive K-NPPase activity (Na-K-ATPase) was demonstrated to localize only in the basolateral membrane of parietal cells with Mayahara's method. These findings support the interrelationships between the apical surface membrane, secretory canalicular membrane and tubulovesicles, and the functional differentiation of the membrane between the secretory membrane and basolateral membrane.     

Fine reconstruction of the pancreatic ductular system at the onset of pancreatitis

The three-dimensional structure of the pancreatic ductular system (from the intercalated duct to the intercellular secretory canaliculus) is controversial and unclear. The aim of this study is to reveal the three-dimensional structure of the pancreatic ductular sysytem at the onset of pancreatitis. One day following rat pancreatic duct ligation, dilated lumina from the pancreatic ductular system were reconstructed by light microscopic and scanning electron microscopic examination of pancreatic tissue serial sections. The existence of the intra-acinar duct, which is formed only by centroacinar cells and interconnects the adjacent central lumina in an acinus, was demonstrated. The intercellular secretory canaliculi, which are the terminal parts of the pancreatic ductular system, anastomose and end blindly in the intercellular space located between adjacent lateral surfaces of the acinar cells. The intercalated ducts, the intra-acinar ducts, the central lumina, and the intercellular secretory canaliculi are arranged together in a complex connecting and branching system. However, there were no anastomoses found among the central lumina or acini.     

An electron microscopic and morphometric study of the gastric parietal cells of rats following administration of gastrin, histamine and cyclic 3',5'-adenosine monophosphate]

The ultrastructure of the gastric parietal cells was studied in rats affected by a genetic inductor of the hydrochloric acid secretion and its mediators in this process - histamine and cyclic 3', 5'-adenozinemonophosphate playing the role of the enzyme activators. All these stimulators caused similar changes in the cell structure. The square surface of the intracellular secretory tubules was shown to become 2,5 times larger, and that of the canaliculi - vesiculoid elements which seemed to be the cell membrane depot became 2,5 times less. It may be supposed that the gastric mucosa parietal cells are not the target-cells for pentagastrine (gastrine).     

Characterization of gastric mucosal membranes. X. Immunological studies of gastric (H+ + K+)-ATPase

Gastric mucosal homogenates from hog were fractionated by differential and density gradient centrifugation and free-flow electrophoresis. The two major membrane fractions (FI and FII) thus obtained are distinct both enzymically and in terms of transport reactivity. This heterogenicity extends to their antigenic activity. Purified antibodies which were raised against the K+-ATPase-containing H+ transport fraction FI were of two types: inhibitory and non-inhibitory. Inhibitory antibodies reduced the K+-ATPase activity by approximately 80% and the K+-p-nitro-phenylphosphatase activity by approximately 40% in a concentration-dependent manner, while the small Mg++-dependent component of the enzyme activity was unaffected. Antibodies inhibiting the K+-ATPase also inhibited H+ transport. These antibodies did not cross-react with the other major membrane fraction isolated by free- flow electrophoresis, FII, and gave a single band on rocket immunoelectrophoresis. Antibodies against this FII fraction also did not react with the K+-ATPase and were heterogeneous, giving at least four bands with rocket immunoelectrophoresis and inhibiting both the 5'- nucleotidase and Mg++-ATPase of this fraction. Immunofluorescent staining of tissue sections showed that the FI was derived from the parietal cell of gastric tissue and was localized to the supranuclear area of the cell. Staining of isolated rat gastric cell suspensions by FI antibodies confirmed the selectivity of the antibody and showed a polar, plasma membrane localization. FII antibodies also largely stained the parietal cells in tissue sections. In the 16 hog tissues tested, FI antibodies cross-reacted only with gastric fundus, thyroid and weakly with thymus. Immunoelectronmicroscopy showed that FI antibodies reacted strongly with the secretory membrane at the apical cell surface of the parietal cells and at the secretory canaliculi, weakly with the apical surface of the zymogen cell, and not with the basal-lateral surface of the cells. Thus, the protontranslocating ATPase is localized in the parietal cells and in the region postulated to be the site of acid secretion.     

Glycoconjugate distribution in the human fundic mucosa revealed by lectin- and glycoprotein-gold cytochemistry

The glycoconjugates of the human fundic mucosa were characterized at the ultrastructural level by means of direct (Helix pomatia agglutinin-gold complex) and indirect lectin techniques (Concanavalin A and horseradish peroxidase-gold complex; wheat germ agglutinin and ovomucoid-gold complex). Surface mucous cells and mucous neck cells secreted O-glycoproteins with N-acetylgalactosamine and N-acetylglucosamine residues at the non reducing terminus of the saccharidic chain. The secretory granules of the mucous neck cells showed condensed areas slightly reactive to ConA. The results obtained in the chief cells suggest that these cells secrete N-glycoproteins rich in mannose and/or glucose residues. "Transitional cells", presenting both morphological characteristics and lectin binding pattern intermediate to the mucous neck and chief cells have been observed. The surface of the intracellular canaliculi of the parietal cell was labelled by HPA, WGA and ConA. In the neck region of the gastric glands, immature parietal cells containing abundant mucous granules reactive to HPA, WGA and ConA were observed. The present results further corroborate the existence of a common cell precursor for surface mucous, mucous neck and parietal cells. In a further step, mucous neck cells gradually differentiate into chief cells the transitional cells being an intermediate stage.     

Role of ADP-ribosylation factor 6 (ARF6) in gastric acid secretion

ADP-ribosylation factor (ARF) proteins are monomeric GTPases that are essential for membrane transport and exocytosis in a number of secretory cells. We investigated ARF6, the activation of which is insensitive to brefeldin A, to determine whether it regulates membrane traffic in gastric parietal cells. ARF6 translocated from cytosol to tubulovesicle in the presence of GTPgammaS, a potential inhibitor of acid secretion in permeabilized cells, whereas under the Mg2+-chelated condition where activity of ARF-GTPase activating protein is inhibited, ARF6 translocated to the apical secretory membrane. Immunohistochemical examination revealed that ARF6 mainly located in parietal cell within the gastric glands, and it translocated from the cytosol to the intracellular canaliculi when the glands were stimulated. These results indicated that the distribution of ARF6 between cytosol and the two different membranes was regulated by its GTPase activity. In cultured gastric glands infected with adenovirus expressing ARF6 Q67L, a mutant lacking GTP hydrolysis activity, gastric acid secretion was inhibited. These results suggest that ARF6 regulates gastric acid secretion in parietal cell and that the GTP hydrolysis cycle of ARF6 is essential for the activation pathway.     

Mice with a targeted disruption of the AE2 Cl-/HCO3- exchanger are achlorhydric

The AE2 Cl-/HCO3- exchanger is expressed in numerous cell types, including epithelial cells of the kidney, respiratory tract, and alimentary tract. In gastric epithelia, AE2 is particularly abundant in parietal cells, where it may be the predominant mechanism for HCO3- efflux and Cl- influx across the basolateral membrane that is needed for acid secretion. To investigate the hypothesis that AE2 is critical for parietal cell function and to assess its importance in other tissues, homozygous null mutant (AE2(-/-)) mice were prepared by targeted disruption of the AE2 (Slc4a2) gene. AE2(-/-) mice were emaciated, edentulous (toothless), and exhibited severe growth retardation, and most of them died around the time of weaning. AE2(-/-) mice exhibited achlorhydria, and histological studies revealed abnormalities of the gastric epithelium, including moderate dilation of the gastric gland lumens and a reduction in the number of parietal cells. There was little evidence, however, that parietal cell viability was impaired. Ultrastructural analysis of AE2(-/-) gastric mucosa revealed abnormal parietal cell structure, with severely impaired development of secretory canaliculi and few tubulovesicles but normal apical microvilli. These results demonstrate that AE2 is essential for gastric acid secretion and for normal development of secretory canalicular and tubulovesicular membranes in mouse parietal cells.     

Differential response of pineal melatonin levels to light at night in laboratory-raised and wild-captured 13-lined ground squirrels (Spermophilus Tridecemlineatus)

Pineal melatonin levels were compared in laboratory-raised or wild-captured 13-lined ground squirrels (Spermophilus tridecemlineatus) that were either exposed to 10 h of darkness at night or to light which had an irradiance of 400 μW/cm². In laboratory-born squirrels the period of darkness was associated with a gradual rise in pineal melatonin levels with peak values being reached at 0200 h, 6 h after darkness onset. Thereafter, melatonin levels decreased and were back to low daytime levels by 0800 h, 2 h after light onset. The exposure of laboratory-raised animals to an irradiance of 400 μW/cm² during the night totally prevented the nocturnal rise in pineal melatonin levels in these animals. In wild-captured ground squirrels the period of darkness at night was associated with a rapid rise in pineal melatonin such that by 2200 h, 2 h after lights out, peak melatonin values were already attained; additionally, melatonin levels remained high throughout the period of darkness but returned to daytime values by 0800 h. Exposure of wild-captured squirrels to a light irradiance of 400 μW/cm² during the normal dark period was completely incapable of suppressing pineal melatonin levels. The difference in the sensitivity of the pineal gland of laboratory-raised and wild-captured ground squirrels may relate to their previous lighting history.     

Induction of c-fos protein-like immunoreactivity in the rat and hamster pineal gland after the onset of darkness

Induction of c-fos protein (FOS) after the onset of darkness was studied immunocytochemically in the rat and hamster pineal gland. The animals were kept on a 12:12 h light-dark cycle. Before the dark period no FOS staining was seen in either rat or hamster pineal cells. Five hours after the onset of darkness 342 +/- 18 pinealocytes/0.2 mm2 (mean +/- SD) displayed FOS-like immunoreactivity in the hamster pineal gland; in the rat pineal gland only 5 +/- 2 pinealocytes/0.2 mm2 showed a faint staining. Two hours later the density of FOS positive cells was decreased to 60 +/- 11/0.2 mm2 in the hamster but increased to 519 +/- 103/0.2 mm2 in the rat pineal gland. Three hours before the beginning of the light period no FOS positive cells were detected in either animal. Both the rat and hamster pineal gland showed a transient and temporally defined expression of c-fos protein in the middle of the dark period. This may be related to a more active functional state of pinealocytes, which is reflected in a peak of melatonin synthesis during the darkness.     

Radioautographic tracing of 3H-proline in the endodermal cells of the parietal yolk sac as an indicator of the biogenesis of basement membrane components

The biogenesis of basement-membrane components was investigated in the endodermal cells of the rat parietal yolk sac in 12.5-day pregnant rats; 3H-proline was injected into conceptuses. After various time intervals, the parietal yolk sac, including endodermal cells and the associated Reichert's membrane, was removed and processed for electron-microscopic radioautography. Silver grains were counted over endodermal cell organelles and Reichert's membrane. At 2 and 5 min after 3H-proline injection, endodermal cells showed heavy labeling in rough endoplasmic reticulum (rER). Silver grain density over the rER decreased from 2 to 20 min and then remained at a plateau. Grain density was moderate over the Golgi apparatus initially but rose to a peak at 2 hr and decreased by 4 hr and later. Grain density was negligible over secretory granules at 2 and 5 min and increased moderately with time to reach a maximum at 8 hr. Thus, radioautographic peaks occurred sequentially in rER, Golgi apparatus, and secretory granules. By 4 hr and later, silver grains accumulated over Reichert's membrane. These results indicated that endodermal cells incorporated labeled proline into substances which were processed from the rER through the Golgi apparatus, transported from there to the cell surface by secretory granules, and released for export to Reichert's membrane. To clarify the nature of the exported substances, the amount of label present in proline and hydroxyproline residues after 3H-proline injection was measured in Reichert's membrane with or without the associated endodermal cells. Within the cells, 61.8% of the labeled proteins were classified as "sedentary" and 38.2% as "exportable." Of the label exported to Reichert's membrane, 66.3% consisted of type IV collagen and the rest of other basement-membrane components. The results obtained with this model suggest that basement-membrane proteins, including type IV collagen, are elaborated by the associated cells through the classical pathway: rER-Golgi apparatus-secretory granules.     

Fine structure of the mandibular gland of the Djungarian hamster (Phodopus sungarus)

The mandibular gland of the Djungarian hamster was examined by light microscopy, and transmission and scanning electron microscopies. Its acinar cells reacted with periodic acid-Schiff (PAS) and were weakly stained with alcian blue (AB). There were intercellular canaliculi between the acinar cells. These cells therefore appeared to be seromucous. The acinar epithelium was composed of light cells containing various spherical secretory granules. The granular cells of the mandibular gland possessed many acidophilic granules exhibiting a positive reaction to PAS stain. They were frequently observed at the junction of the acini and intercalated ducts in all mandibular glands examined. All of these cells were light and contained secretory granules of varying size and density. The intercalated ducts consisted exclusively of light cells possessing a few round granules of high density in the apical region. The striated ducts were comprised of two portions--a secretory portion and a typical striated portion without secretory granules. The secretory portion consisted of light, dark and specifically light epithelial cells containing acidophilic granules, which exhibited a strongly positive PAS reaction. The epithelium of typically striated portions was composed of light and dark cells containing fine vacuoles in the apical region. The mandibular gland of the Djungarian hamster revealed no histological differences between sexes.     

Drebrin E2 is differentially expressed and phosphorylated in parietal cells in the gastric mucosa

Developmentally regulated brain proteins (drebrins) are highly expressed in brain where they may regulate actin filament formation in dendritic spines. Recently, the drebrin E2 isoform was detected in certain epithelial cell types including the gastric parietal cell. In gastric parietal cells, activation of HCl secretion is correlated with actin filament formation and elongation within intracellular canaliculi, which are the sites of acid secretion. The aim of this study was to define the pattern of drebrin expression in gland units in the intact rabbit oxyntic gastric mucosa and to initiate approaches to define the functions of this protein in parietal cells. Drebrin E2 expression was limited entirely or almost entirely to parietal cells and depended upon the localization of parietal cells along the gland axis. Rabbit drebrin E2 was cloned and found to share 86% identity with human drebrin 1a and to possess a number of cross-species conserved protein-protein interaction and phosphorylation consensus sites. Two-dimensional Western blot and phosphoaffinity column analyses confirmed that drebrin is phosphorylated in parietal cells, and several candidate phosphorylation sites were identified by mass spectrometry. Overexpression of epitope-tagged drebrin E2 led to the formation of microspikes and F-actin-rich ring-like structures in cultured parietal cells and suppressed cAMP-dependent acid secretory responses. In Madin-Darby canine kidney cells, coexpression of epitope-tagged drebrin and the Rho family GTPase Cdc42, which induces filopodial extension, produced an additive increase in the length of microspike projections. Coexpression of dominant negative Cdc42 with drebrin E2 did not prevent drebrin-induced microspike formation. These findings suggest that 1) drebrin can induce the formation of F-actin-rich membrane projections by Cdc42-dependent and -independent mechanisms; and that 2) drebrin plays an active role in directing the secretagogue-dependent formation of F-actin-rich filaments on the parietal cell canalicular membrane. Finally, the differential distribution of drebrin in parietal cells along the gland axis suggests that drebrin E2 may be an important marker of parietal cell differentiation and functionality.