the source of carbon dioxide

8/9/2019 The Source of Carbon Dioxide

1/8

The Source of Carbon Dioxide for

Gastric Acid ProductionHOWARD STEER*

Department of General Surgery, Southampton General Hospital, Southampton UniversityHospitals NHS Trust, University of Southampton School of Medicine, Southampton,

SO16 6YD, United Kingdom

ABSTRACTThe source of carbon dioxide for the chemical reaction leading to the

production of gastric acid is unknown. The decarboxylation of an aminoacid releases carbon dioxide. Pepsinogens provide a rich source of theamino acid arginine. Both the source of carbon dioxide, arginine, and theconsequence of arginine decarboxylation, agmatine, have been studied.The site of carbon dioxide production has been related to the survival ofthe parietal cell. An immunohistochemical study has been carried out onglycol methacrylate embedded gastric biopsies from the normal stomachof 38 adult patients. The sections have been stained using polyclonal anti-body to pepsinogen II, polyclonal antibody to agmatine, and polyclonalantibody to Helicobacter pylori. Pepsinogen II and agmatine are found inthe parietal cell canaliculi. This is consistent with the production of car-bon dioxide from arginine in the parietal cell canaliculi. Evidence is pre-sented for the decarboxylation of arginine derived from the activation seg-ment of pepsinogen as the source of carbon dioxide for the production ofgastric acid. The production of carbon dioxide by the decarboxylation ofarginine in the parietal cell canaliculus enables the extracellular hydra-tion of carbon dioxide at the known site of carbonic anhydrase activity.The extracellular production of acid in the canaliculus together with the

presence of agmatine helps to explain why the parietal cells are notdestroyed during the formation of gastric acid. Agmatine is found in themucus secreting cells of the stomach and its role in acid protection of thestomach is discussed. Anat Rec, 292:7986, 2009. 2008 Wiley-Liss, Inc.

Key words: gastric acid; parietal cell; pepsinogen; agmatine

Detailed information on the production and secretionof gastric acid is available but, despite this knowledge,there are numerous unanswered questions. What is the

source of the carbon dioxide for the chemical reactionleading to the production of gastric acid? Parietal cellsare responsible for the production of gastric acid butwhy are they not destroyed by an acid which canachieve a pH of 1.2? Contact with such an acid shoulddestroy any living tissue.

The hydration of carbon dioxide is the chemical reac-tion which produces gastric acid. The proton pump isintimately involved in the gastric acid production, andthe current theory of gastric acid production envisagesan exchange of K1 for H1. However, elegant in vivoexperiments have indicated that the K1 compartment

which is related to acid secretion is small (Hirschowitzand Sachs, 1966). These experiments on dogs with gas-tric fistulas have shown that the insulin inhibition of

*Correspondence to: Howard W Steer M.A.(Oxon),B.Sc.(hons), Ph.D., F.R.C.S, Department of General Surgery,Southampton General Hospital, Southampton University Hospi-tals NHS Trust, University of Southampton School of Medicine,Southampton, SO16 6YD, United Kingdom. Fax: 02380798911.E-mail: [email protected]

Received 16 April 2008; Accepted 16 July 2008

DOI 10.1002/ar.20762Published online 24 October 2008 in Wiley InterScience (www.interscience.wiley.com).

2008 WILEY-LISS, INC.

THE ANATOMICAL RECORD 292:7986 (2009)


2/8

acid secretion can be reversed within 5 min by the intra-venous injection of only 0.5 mEq/kg K1, with the meanconcentration of the secreted gastric acid rising from126 to 153 mEq/L. The rise in gastric H1 output doesnot correlate with the intravenous K1 administration.Thus, the amount of K1 in this specific K1 compartment

is insufficient to account for the H

1

secretion on an ionfor ion exchange basis.Gastric acid secretion is associated with the transfer of

a large volume of water through the gastric mucosa(Durbin and Moody, 1965). In vitro experiments using froggastric mucosa have shown a significant associationbetween water transport through the gastric mucosa andhistamine-stimulated gastric acid secretion (Gerbino et al.,2007). The increase in cAMP-dependent [K1] is associatedwith an osmotic gradient resulting in the passage of waterinto the gastric glands and this occurs before acid secre-tion (Gerbino et al., 2007). Could this cAMP-dependent[K1] compartment be the small K1 compartment demon-strated by Hirschowitz and Sachs (1966)?

The hydration of carbon dioxide which produces gas-tric acid is catalyzed by the enzyme carbonic anhydrase.

The precise localization of carbonic anhydrase in thegastric epithelial cells has been established (Cross, 1970;Winborn et al., 1974). Carbonic anhydrase has beenidentified with the electron microscope in significantquantity on the luminal surface of the cell membrane ofthe parietal cell canalicular microvilli (Cross, 1970). Theenzyme has not been located on the cytoplasmic surfaceof the cell membrane of the parietal cell canalicular mi-crovilli. The hydration of carbon dioxide resulting in theproduction of gastric acid requires a significant supply ofcarbon dioxide. Although the source of carbon dioxide isunknown, textbooks suggest interstitial fluid as thesource. However, the possibility that this carbon dioxidecomes from the gastric vascular system is untenable.The gastric vascular system does not have any specific

modifications which would enable the stomach to be sup-plied with carbon dioxide enriched blood.

In investigating these questions, it has been decidedto examine the source of carbon dioxide for the criticalchemical reaction of the hydration of carbon dioxide cat-alyzed by the enzyme carbonic anhydrase.

The theoretical background to the conclusion that thedecarboxylation of amino acids is one of the principlesources of carbon dioxide for gastric acid production hasbeen outlined (Steer, 2005). Biochemically, basic aminoacids are ideally suited to act as a source of carbon diox-ide in the stomach and the most basic amino acid is ar-ginine (isoelectric point 11.15). A significant source of ar-ginine is present in the stomach.

Pepsinogens are the major proteins produced by the

chief cells of the stomach. These pepsinogens are convertedinto pepsins with the release of activation segment. Activa-tion segment of human pepsinogen I is made up of fortyseven amino acids. Six of these amino acids are arginine(Kageyama and Takahashi, 1980). This is a significantamount of arginine in a small protein molecule. The decar-boxylation of the arginine derived from a single moleculeof activation segment would result in the release of sixmolecules of carbon dioxide.

The current work has investigated the source of car-bon dioxide (pepsinogen) and the distribution of agma-tine the product of arginine decarboxylation in relationto human gastric parietal cells to explore the role of

these molecules in gastric acid production. Evidence ispresented that arginine decarboxylation takes place inthe parietal cell canaliculi which is extracellular. Theextracellular location of carbon dioxide production andthe presence of the strong base, agmatine, helps toexplain why parietal cells are not destroyed during gas-

tric acid production.Preliminary reports of this work have been producedand presented (Steer, 2005, 2007).

MATERIALS AND METHODS

Biopsies have been taken at upper gastrointestinal en-doscopy from 38 adult patients. The patients have beenfasting for 4 hr before endoscopy and biopsy. The gastricmucosa is endoscopically normal in all patients. Twenty-six patients have a final clinical diagnosis of no abnor-mality found, and 12 patients have a final clinical diag-nosis of gallstones. The gastric biopsies have been takenfrom the fundus of the stomach and embedded in resinfor immunohistochemical study (Britten et al., 1993).

The biopsies are histologically normal. The patients arenot infected with Helicobacter pylori as indicated by anegative CLO test (Kimberly-Clark, Ballard MedicalProducts, Draper, Utah) on duplicate biopsies and theabsence of any immunohistochemical staining for Helico-bacter pylori.

The process of glycol methacrylate (GMA) resinembedding will now be outlined. The biopsies are placedin ice acetone containing 2 mM phenyl methyl sulphonylfluoride and 20 mM iodoacetamide. Overnight fixation at2208C is followed by replacing the fixative with acetoneat room temperature for 15 min and then methyl benzo-ate at room temperature for 15 min. The biopsies havebeen infiltrated with processing solution (5% methylbenzoate in glycol methacrylate solutionGMA solution

A) at 48C. There have been three changes of processingsolution with 2 hr in each change before embedding infreshly prepared embedding solution (GMA solution A/benzoyl peroxide). The capsules containing resin embed-ded biopsies have been polymerized at 48C for 48 hr andstored in airtight boxes at 2208C. Sections, 12 mmthick, have been cut from the resin embedded specimensand have been stained with polyclonal antibodies (Steer,2005). Colocalization of pepsinogen II and agmatinehave been performed using serial thin sections (seelater). This study has involved using a sheep polyclonalantibody to human pepsinogen II (Abcam, Cambridge,U.K.), a rabbit polyclonal antibody to agmatine (1-amino-4-guanidobutane) (Chemicon International, ChemiconEurope, Chandlers Ford, Hampshire, U.K.) and a rabbit

polyclonal antibody to Helicobacter pylori (DakoCytoma-tion, Ely, Cambridge, U.K.).GMA resin offers numerous advantages over frozen

and paraffin preparations. The use of a noncross linkingfixative (acetone), together with processing at low tem-perature, ensures good antigen preservation; the tissuemorphology is excellent; thin sections (12 mm) can becut allowing for several sections to be made through onecell enabling colocalization studies of pepsinogen andagmatine and a large number of sections to be cut fromone sample, which is especially important when study-ing small mucosal biopsies, and embedded tissue can bestored relatively long term.

80 STEER


3/8

GMA embedded tissue is unsuitable for immunofluore-sence as it autofluoresces in the FITC channel.

The following immunohistochemical staining proce-dure has been used for sections of the GMA embeddedtissues. The sections have been washed in a solution of0.1% sodium azide and 0.3% hydrogen peroxide in

reverse osmosis water for 30 min. The sections havebeen washed three times with tris buffered saline (TBS)at 5 min intervals before inhibiting the nonspecific bind-ing sites by incubating the sections with 20% fetal calfserum in Dulbeccos modified Eagles medium (DMEM)containing 1% bovine serum albumin for 30 min. Theslides have been drained and then incubated with theprimary antibody at the appropriate dilution in TBS for18 hr at 48C. Evaporation has been prevented by placinga coverslip over the section. During this process, care istaken to exclude any air bubbles. The sections havebeen rinsed three times with TBS at 5 min intervals.The slides have been drained and the appropriate biotin-ylated second stage antibody applied at the appropriatedilution for 2 hr at room temperature. The slides havebeen rinsed three times with TBS at 5 min intervals

before incubation with streptavidin-biotin horseradishperoxidase complexes at appropriate dilution in Tris/HClfor 2 hr at room temperature. The slides have beenrinsed three times with TBS at 5 min intervals beforeapplying amino ethyl carbazole (AEC) solution for 20 minat room temperature. The slides have been rinsed withTBS and washed in running tap water for 2 min. Thesections have been counterstained with Mayers haema-toxylin for 2 min and washed in running tap water for5 min to blue. Crystalmount has been applied and thesections baked for 10 min at 808C until the crystal-mount is dry. The slides have been allowed to cool toroom temperature before mounting with a coverslip indistrene dibutyl phthalate xylene (DPX).

Endogenous peroxidase is blocked with 0.1% sodium

azide containing 0.3% (v/v) hydrogen peroxide. Nonspe-cific protein interactions are blocked with a solution ofDMEM containing 1% bovine serum albumin and 20%fetal calf serum. These are applied to both test slidesand negative control slides. This blocked nonspecificstaining as the negative control slides incubated withbuffer in place of primary antibody, but with the detec-tion stages, are negative.

Initially, all antibodies used in this study are opti-mized by titration by limiting dilution to determine theoptimum working dilution for each antibody. Matchedimmunoglobulin controls (rabbit IgG and sheep IgG) areused at the same concentration as the primary antibod-ies to check for nonspecific binding. These are negative.For each staining negative control slides are incubated

with buffer instead of primary antibody.Ethical approval for the study was obtained. Permis-sion to obtain the endoscopic biopsies as well as to per-form the immunohistochemical analyses were obtainedfrom the patients. The patients were undergoing endo-scopic examinations as part of the investigation of theirpresenting symptoms.

RESULTS

Pepsinogen II is found in the chief cells and the parie-tal cells of the stomach (Fig. 1). The presence of pepsino-gen II in the chief cells takes the form of a generalized

cytoplasmic staining, cytoplasmic granules, and cyto-plasmic vacuoles. Pepsinogen II is present in the canali-culi of the parietal cells (Fig. 1) with the site of openingof a canaliculus into the gastric gland lumen noted inFig. 1. There is no cytoplasmic staining for pepsinogenII in the parietal cells. All the pepsinogen II found in

the parietal cells is outside the cell membrane in thecanalicular lumen. The parietal cell canaliculi can be ei-ther nondilated or dilated. The presence of pepsinogen IIin the parietal cell canaliculi is not universal.

Agmatine is found in the parietal cells, the chief cells,and the mucus secreting cells of the stomach (Figs. 2and 3). The agmatine in the parietal cells is localized tothe lumen of the canaliculi (Figs. 2 and 3). A parietalcell canaliculus is seen opening into the lumen of a gas-tric gland (Fig. 3). The agmatine present in the parietalcell canaliculi is not universal and is similar to the find-ings for indicator dye (Bradford and Davies, 1950).There is no agmatine staining in the cytoplasm of theparietal cells. This contrasts with the findings in thechief cells and the mucus secreting cells of the stomach.

Agmatine is present in the cytoplasm of the chief cells

and takes the form of numerous discrete granules usu-ally located in the supranuclear part of the cytoplasm(Fig. 2). There is also strong staining for agmatine posi-tive material on the luminal surface of the chief cells(Fig. 2).

Agmatine is found in the mucus secreting cells of thestomach (Fig. 3). There is significant cytoplasmic stain-ing for agmatine in the supranuclear part of the mucussecreting cells. This agmatine staining is present in thatarea of the cytoplasm where mucus is located.

The endothelial cells of mucosal blood vessels stain foragmatine (Fig. 2).

The control sections for pepsinogen II and agmatineare negative with no positive chromogen staining. Thereis no Helicobacter pylori positive material in the biopsies.

DISCUSSION

The presence of the protein pepsinogen II in parietalcell canaliculi and the fact that not all parietal cellcanaliculi contain pepsinogen II is consistent with thepatchy distribution of indicator dye found in the classi-cal studies leading to the establishment of the role of pa-rietal cells in acid secretion (Bradford and Davies, 1950).Previously, a coagulable material has been noted in pari-etal cell canaliculi (Stohr, 1882; Harvey and Bensley,1912; Revell, 1912; Zimmermann, 1925; Ma, 1927). Thiscoagulable material is consistent with the presence of aprotein such as pepsinogen in the canaliculi. There is nocytoplasmic parietal cell pepsinogen. There is no evi-

dence from this work to suggest that pepsinogens areproduced by the parietal cells, but pepsins can be formedfrom pepsinogens outside the parietal cell cytoplasm inthe parietal cell canaliculi. Although only pepsinogen IIhas been examined in this study, a similar process mayoccur with respect to other pepsinogen molecules.

Carbon dioxide can be produced from the decarboxyl-ation of arginine. The activation segment of pepsinogenin the parietal cell canaliculi can provide a source of ar-ginine (Kageyama and Takahashi, 1980).

The decarboxylation product of arginine is agmatine. Agmatine has been recognized because the work of Pro-fessor Albrecht Kossel (Kossel, 1910). Agmatine had

81PEPSINOGEN AND GASTRIC CARBON DIOXIDE


4/8

Fig. 1. The parietal cell area of the fundus of the stomach. Pepsinogen II in the parietal cell canaliculi

is shown by the black arrows. The site of opening of a parietal cell canaliculus into the gastric gland

lumen is shown (*). Numerous chief cells (c) are seen. Space bar is 20 mm. Polyclonal antibody to pepsin-

ogen II.

82 STEER


5/8

Fig. 2. The parietal cell area of the fundus of the stomach. Agmatine in the parietal cell canaliculi is

shown by the black arrows. Numerous chief cells (c) and mucosal blood vessels (bv) are seen. Space bar

is 20 mm. Polyclonal antibody to agmatine.



6/8

Fig. 3. The parietal cell area of the fundus of the stomach. The opening of the parietal cell canaliculus

into the gastric gland lumen is indicated by the black arrow. The canaliculus contains agmatine. Agmatine

is seen in the gastric mucus secreting epithelial cells (Am). Space bar is 20 mm. Polyclonal antibody to

agmatine.

84 STEER


7/8

originally been extracted from herring roe (Kossel,1910), and for many years agmatine had been consid-ered to be absent from mammalian tissues. This misap-prehension was corrected in 1994 with the identificationof agmatine in mammalian tissue (Li et al., 1994). Anexamination of various mammalian organs has revealed

that the greatest concentration of agmatine is found inthe stomach (Raasch et al., 1995).The process of decarboxylation of arginine in the stom-

ach has been confirmed in the present study by findingthe end product of this reaction, namely agmatine, inthe parietal cell canaliculi. The decarboxylation of argi-nine provides a source of carbon dioxide. The hydrationof this carbon dioxide results in the release of hydrogenions. The decarboxylation of another basic amino acid,lysine, may also provide carbon dioxide for this processbut the possible role of lysine has not been studied. Inhuman pepsinogen I, the activation segment has fifteenbasic amino acids out of a total of forty seven aminoacids. As previously stated, six of these basic aminoacids are arginine but of the remaining nine basic aminoacids eight are lysine (Kageyama and Takahashi, 1980)

which has an isoelectric point of 9.59. Thus, if both argi-nine and lysine are involved in this process of decarbox-ylation, one molecule of activation segment would pro-duce fourteen molecules of carbon dioxide.

The small K1 compartment related to acid secretion(Hirschowitz and Sachs, 1966, 1967) is inconsistent withthe ion for ion exchange envisaged in the theory thatH1 are produced in the parietal cell cytoplasm andtransported through the cell membrane. However, thissmall K1 compartment would be consistent with the K1

dependent elevation in cAMP occurring before the onsetof acid secretion. The increase in cAMP permits the gen-eration of an osmotic gradient enabling water to bepassed into the parietal cell canaliculi (Gerbino et al.,2007). This water could then be used to hydrate the

carbon dioxide derived from arginine with the produc-tion of acid.

The episodic intake of food into the stomach leads tofluctuations in the demand for gastric acid. One of thefactors which determine the level of activity of the chem-ical reaction which produces gastric acid is the intake offood. Variations in food intake would result in quantita-tive physicochemical changes at the site of this chemi-cal reaction. Carbonic anhydrase has been located at theluminal surface of the cell membrane of the canalicularmicrovilli of the parietal cells (Cross, 1970). Since 1893(Golgi, 1893), it has been noted that acid secretion isassociated with changes in the apical surface membraneand the canalicular microvilli of parietal cells. Thesechanges which result in an increase in the secretory

membrane have been described in the frog, mouse, rab-bit, dog, and human (Golgi, 1893; Kasbekar et al., 1968;Frexinos et al., 1971; Helander and Hirschowitz, 1972;Ito and Schofield, 1974; Carlisle et al., 1978). In a quan-titative ultrastructural study using dogs with gastric fis-tulas, histamine stimulation causes a change in the se-cretory surface (apical and canalicular) of the parietalcells which increases from 0.19 m2/cm3 of the parietalcell mass to 1.92 m2/cm3 of the parietal cell mass(Helander and Hirschowitz, 1972). This increase in thesecretory membrane results in a greater surface area forcarbonic anhydrase activity. The increase in carbonicanhydrase activity would have the potential for the

hydration of more carbon dioxide and the production ofa greater quantity of gastric acid in the parietal cellcanaliculi. The dynamic process of parietal cell canalicu-lar changes related to HCl activators and inhibitors areassociated with cytoskeletal and cytochemical rearrange-ments as has been demonstrated by in vitro studies

(Agnew et al., 1999; Berg et al., 2007).For almost two centuries ever since acid has beenidentified in the stomach (William Prout 17851850), ithas been a dilemma explaining why the acid does notdestroy the stomach? The finding that the greatest con-centration of agmatine in the body is in the stomach(Raasch et al., 1995), the fact that agmatine is such astrong base and the cellular localization of the agmatinein the gastric mucosa (present work) makes agmatine astrong candidate for that protective role. This mucosaldefence role of agmatine is best illustrated by the gastricmucus secreting cells. When considering the source ofthe agmatine in the mucus secreting cells of the stom-ach, it is interesting to note that radiolabeled agmatineis taken up from the gastric lumen by the gastric mu-cosa (Molderings et al., 2002). The agmatine present in

the mucus secreting gastric cells could have been takenup by these cells from the lumen and this agmatinederived from the decarboxylation of arginine from theactivation segment of pepsinogen.

Helicobacter pylori infection of the stomach is associ-ated with a decrease in the amount of mucus in the mu-cus secreting cells of the stomach (Steer, 2005). Thischange is associated with a decrease in the amount ofagmatine in these mucus secreting cells (Steer, 2005).Such a decrease in the amount of agmatine in the epi-thelium of the helicobacter pylori infected stomachwould make this epithelium more vulnerable to damageby gastric acid.

The concept that gastric acid production involvescAMP changes at the cell membrane, the passage of

water into the parietal cell canaliculus and microvilloussurface changes would necessitate a finite time intervalbetween stimulation and acid production. In thisrespect, it is interesting to note that there is evidencefor a latent period between gastric stimulation and theonset of acid secretion. In the frog gastric mucosa, hista-mine stimulation is followed by a 10 min time lag beforeacid secretion (Kasbekar, 1967), and acid secretionoccurs 3 min after the passage of water into the gastricglands (Gerbino et al., 2007). After the onset of stimula-tion, the increase in the dogs parietal cell secretory sur-face occurs in the initial 10 min after the start of thehistamine infusion (Helander and Hirschowitz, 1974).Such experimental observations support the conceptthat gastric stimulation is followed by a latent period

during which physicochemical changes occur.The extracellular production of gastric acid by thehydration of carbon dioxide which has been derived fromthe decarboxylation of arginine is shown in Fig. 4. Thisextracellular, rather than intracellular, hydration of car-bon dioxide and the formation of agmatine wouldexplain why the parietal cells are not destroyed by theproduction of the strong acid. The current observationsand their interpretations concur with the principleexpounded by Hoerr and Bensley (1936) that gastric

juice (including gastric acid) . . .. . .. . . is a result of theinteraction of the various products on one another modi-fied by the living membrane over which the secretion



8/8

flows. The reaction space described by Hoerr andBensley (1936) for gastric acid production is in the parie-tal cell canaliculi.

ACKNOWLEDGMENTS

Grateful acknowledgement is made for the helpreceived from Dr. Susan Wilson, Linda Jackson, HelenRigden and Jon Ward of the Histochemistry ResearchUnit, University of Southampton School of Medicine,

Anton Page of the Biomedical Imaging Unit, Universityof Southampton School of Medicine / Southampton Uni-versity Hospital NHS Trust and Adie Falcinelli of theLearning Media, Southampton University Hospital NHS

Trust.

LITERATURE CITED

Agnew BJ, Duman JG, Watson CL, Coling DE, Forte JG. 1999.Cytological transformations associated with parietal cell stimu-lation: critical steps in the activation cascade. J Cell Sci 112:26392646.

Berg A, Redeen S, Sjostrand SE, Ericson A-C. 2007. Effect of nitricoxide on Histamineinduced cytological transformation in pari-etal cells in isolated human gastric glands. Dig Dis Sci 52:126136.

Bradford NM, Davies RE. 1950. The site of hydrochloric acid pro-duction in the stomach as determined by indicators. Biochem J46:414420.

Britten KM, Howarth PH, Roche WR. 1993. Immunohistochemistry

on resin sections: a comparison of resin embedding techniques forsmall mucosal biopsies. Biotech Histochem 68:271280.Carlisle KS, Chew CS, Hersey SJ. 1978. Ultrastructural changes

and cyclic AMP in frog oxyntic cells. J Cell Biol 76:3142.

Cross SAM. 1970. Ultrastructural localization of carbonic anhydrasein rat stomach parietal cells. Histochemie 22:219225.

Durbin RP, Moody FG. 1965. Water movement through a transport-ing epithelial membrane: the gastric mucosa. Symp Soc Exp Biol19:299306.

Frexinos J, Carballido M, Louis A, Ribet A. 1971. Effects of penta-gastrin stimulation on human parietal cells. An electron micro-scopic study with quantitative evaluation of cytoplasmic struc-tures. Dig Dis 16:10651074.

Gerbino A, Fistetto G, Colella M, Hofer AM, Debellis L, Caroppo L,Curci S. 2007. Real time measurements of water flow in amphib-ian gastric glands. J Biol Chem 282:1347713486.

Golgi C. 1893. Sur la fine organization des glandes peptiques desmammiferes. Arch Ital de Biol 19:448453.

Harvey BCH, Bensley RR. 1912. Upon the formation of hydrochloricacid in the foveolae and on the surface of the gastric mucousmembrane and the non-acid character of the contents of glandcells and lumina. Biol Bull 23:225249.

Helander HF, Hirschowitz BI. 1972. Quantitative ultrastructuralstudies on gastric parietal cells. Gastroenterology 63:951961.

Helander HF, Hirschowitz BI. 1974. Quantitative ultrastructuralstudies on inhibited and on partly stimulated gastric parietalcells. Gastroenterology 67:447452.

Hirschowitz BI, Sachs G. 1966. Reversal of insulin inhibition of gas-tric secretion by intravenous injection of potassium. Am J Dig Dis11:217230.

Hirschowitz BI, Sachs G. 1967. Insulin inhibition of gastric secre-tion: reversal by rubidium. Am J Physiol 213:14011405.

Hoerr NL, Bensley RR. 1936. Cytological studies by the Altmann-

Gersh freezing-drying method. 11. The mechanism of secretion ofhydrochloric acid in the gastric mucosa. Anat Rec 65:417435.

Ito S, Schofield GS. 1974. Studies on the depletion and accumula-tion of microvilli and changes in the tubulovesicular compartmentof mouse parietal cells in relation to gastric acid secretion. J CellBiol 63:364382.

Kageyama T, Takahashi K. 1980. Isolation of an activation interme-diate and determination of the amino acid sequence of the activa-

tion segment of human pepsinogen A. J Biochem 88:571582.Kasbekar DK. 1967. Studies of resting isolated frog gastric mucosa.

Proc Soc Exp Biol Med 125:267271.Kasbekar DK, Forte GM, Forte JG. 1968. Phospholipid turnover

and ultrastructural changes in resting and secreting bullfrog gas-tric mucosa. Biochim Biophys Acta 163:113.

Kossel A. 1910. Uber das Agmatin. Zeitschrift fur PhysiologischeChemie 66:257261.

Li G, Regunathan S, Barrow CJ, Eshraghi J, Cooper R, Reis DJ.

1994. Agmatine: an endogeneous clonidine-displacing substancein the brain. Science 263:966969.Ma WC. 1927. A method for the demonstration of the intracellular

secretion canaliculi of the parietal cells of mammals. Anat Rec

35:337340.Molderings GJ, Heinen A, Menzel S, Gothert M. 2002. Exposure of

rat isolated stomach and rats in vivo to (14C) agmatine: accumula-tions in the stomach wall and distribution in various tissues.Fundam Clin Pharm 16:219225.

Raasch W, Regunathan S, Li G, Reis DJ. 1995. Agmatine, the bacte-rial amine, is widely distributed in mammalian tissues. Life Sci56:23192330.

Revell DG. Personal Communication (see Harvey and Bensley, 1912).Steer HW. 2005. Acid/base changes in the stomach. In: Howard W.

Steer, editor. The Stomach, Helicobacter pylori and acid secretion.U.K. (ISBN 0 9550009). Chapter 7:193217.

Steer HW. 2007. Why does gastric acid fail to destroy those cells

producing this acid? Surg Prac 11:A9, abstract 19.Stohr P. 1882. Zur kenntnis des feineren baues der menschlichenmagenschleimhaut. Archiv F mikrosk Anat 20:221245.

Winborn WB, Seelig LL, Jr, Girard CM. 1974. Variation in the pat-tern of carbonic anhydrase activity in the cells of the gastricglands. Histochemistry 39:289300.

Zimmermann KW. 1925. Beitrag zur kenntnis des baues und derfunction der fundusdrusen im menschlichen magen. Asher-SpiroErgb Physiol 24:281307.

Fig. 4. Diagrammatic representation of the decarboxylation of argi-

nine derived from pepsinogen and the extracellular production of gas-

tric acid in the parietal cell canaliculus. CA, carbonic anhydrase.

86 STEER

the source of carbon dioxide

Documents