Alkali Secretion Research Articles

Peculiarities of the urinary system functional state in neonates and its role in maintenance of the body homeostasis (literary review)

The urinary system activity is of great importance for homeostasis maintenance, which is determined by its participation in elimination of final metabolites, foreign and toxic substances from the body; providing stable blood volume and extracellular fluid; constant concentration of osmotic active substances and ions; regulation of acid-alkali balance, arterial pressure, erythropoiesis, blood clotting, modulation of hormone action; participation in the processes of protein, lipid and carbohydrate metabolism. Clinicians should know peculiarities of formation of the main kidney functions in neonates depending on gestation term and day of life, which can enable to diagnose renal damage in time. Antenatal development of the urinary system occurs during embryonic and fetal periods. During the embryonic period all the nephron structures are formed, and during the fetal period the system is maturing. In case of disorders of normal embryogenesis of the urinary system during the first three months of gestation severe embryopathy is formed. Its severity depends on the damage of an appropriate gene. During later periods fetopathy develops under the effect of various factors promoting disorders of the processes of renal differentiation, and renal pathology is characterized by dys- and hypoplasia. A leading factor of the kidney growth during postnatal ontogenesis is enlargement of the amount of cells but not their sizes. After birth the systemic circulation is reorganized, systemic blood pressure and hydrostatic pressure in glomeruli increase; a relative area of glomerular capillaries, the size of afferent and efferent arterioles increase; and arteriole resistance decreases. Glomerular-tubular imbalance is typical for neonates. It consists of morphological and functional prevalence of the glomeruli over proximal tubules in the immature nephron. The structure of the glomerular membrane changes during the postnatal period: endothelial cells become flat, fenestrations appear, the common basal membrane is formed between the endothelium and podocytes, podocyte legs are formed in the cells of the visceral layer of the glomerular capsule resulting in 40% enlargement of the filtration surface. The processes of tubular maturation in neonates consist of the formation of a dense location and structure of specific transport proteins, electrochemical gradient of the cellular membrane, sensitivity to hormones, phospholipid content, the volume of intracellular fluid and cellular metabolic activity associated with anatomical and functional nephron heterogeneity. Anatomical immaturity of the urinary system in neonates determines peculiarities of its functional state, that is, insufficiently adequate glomerular filtration rate, decreased ability to reabsorb filtered amino acids and minor peptides, phosphates, bicarbonates, glucose and non-organic ions, decreased secretion of organic acids, alkali and medicines, as well as reduced concentration ability of the kidneys.Awareness of peculiarities of the renal function development in healthy neonates is essential for correct interpretation of the results of laboratory and instrumental examinations, understanding the mechanisms of development of pathologic deviations.

PH-sensitive expression of calcium-sensing receptor (CaSR) in type-B intercalated cells of the cortical collecting ducts (CCD) in mouse kidney.

The localization and role of the calcium-sensing receptor (CaSR) along the nephron including the collecting ducts is still open to debate. Using the quantitative, highly sensitive in situ hybridization technique and a double-staining immunohistochemistry technique, we investigated the axial distribution and expression of CaSR along the nephron in mice (C57B/6J) treated for 6 days with acid or alkali diets. Under control condition, CaSR was specifically localized in the cortical and medullary thick ascending limb of Henle's loop (CTAL and MTAL), macula densa (MD), distal convoluted tubule (DCT), and CCD (TALs, MD > DCT, CCD). Along the CCD, CaSR was co-localized with an anion exchanger type 4 (AE4), a marker of the basolateral membrane of type-B intercalated cell (IC-B) in mice. On the contrary, CaSR was not detected either in principal cells (PC) or in type-A intercalated cell (IC-A). CaSR expression levels in IC-B significantly (P < 0.005) decreased when mice were fed NH4Cl (acid) diets and increased when animals were given NaHCO3 (alkali) diets. As expected, cell heights of IC-A and IC-B significantly (P < 0.005) increased in the above experimental conditions. Surprisingly, single infusion (ip) of neomycin, an agonist of CaSR, significantly (P < 0.005) increased urinary Ca excretion without further increasing the hourly urine volume and significantly (P < 0.05) decreased urine pH. CaSR, cloned from rat kidney, was localized in the basolateral membrane of IC-B and was more expressed during alkali-loading. Its alkali-sensitive expression may promote urinary alkali secretion for body acid-base balance.

Effect of Ketanserin and Methiothepin Implicate a 5‐HT‐like Receptors in Serotonin‐stimulated Alkali and Acid Secretion in Isolated, Perfused Midgut Preparations of Larval Yellow‐fever Mosquito Aedes aegypti

Previous work from our laboratory has shown that serotonin (5‐HT) is a major control factor for alkalinization of the anterior midgut (AMG) and acidification of the posterior midgut (PMG) of the larval mosquito, Aedes aegypti. Heretofore it has been impossible to associate the effect of serotonin on acid‐base transport with particular 5‐HT receptor(s) in the larval mosquito gut. Ketanserin (10μM), a specific antagonist for 5‐HT2‐like receptors, was effective against 5‐HT stimulation of alkali secretion by the AMG, but not the acid secretion by the PMG, whereas Methiothepin (10μM) which is active against 5‐HT1, 5‐HT6 and 5‐HT7 but not 5‐HT2‐like receptors, was ineffective against alkali secretion by the AMG but effective against acid secretion in the PMG. Since 5‐HT2 receptors are expected to act through an IP3/DAG pathway to secrete cytoplasmic Ca2+. This result is in accord with our recent demonstration that a Ca2+ 2nd message is required for initiation of 5‐HT stimulated alkali and acid secretion.

Signaling by intracellular Ca2+ and H+ in larval mosquito (Aedes aegypti) midgut epithelium in response to serosal serotonin and lumen pH

The midgut of larval mosquitoes (Aedes aegypti) mediates a cycle of alkali secretion in the anterior segment (AMG) followed by partial reacidification in the posterior segment (PMG); both processes are serotonin-dependent. Here we report that intracellular Ca2+(Cai2+) as indicated by Fura-2 fluorescence, is elevated in both tissues in response to serotonin, but the time courses differ characteristically in the two gut segments, and Ca2+-free solution abolishes the serotonin response in AMG, but not in PMG, whereas Thapsigargin, an inhibitor of endoplasmic Ca2+ transport, abolished responsiveness to 5-HT in PMG. These results suggest the origins for the Ca2+ signal differ between the two tissues. Quantitative real-time RT-PCR revealed expression of 5 putative 5-HT receptor types in AMG, including 5-HT2-like receptors which would be expected to initiate a Ca2+ signal. None of these receptors were highly expressed in PMG. Cyclic AMP (cAMP) is a secretagogue for both tissues, but H89, an inhibitor of Protein Kinase A (PKA), is also a secretagogue, suggesting that the stimulatory effect of cAMP involves a non-PKA pathway. Cytochalasins B and D block the effect of 5-HT in AMG, suggesting a vesicle-fusion mechanism of activation of the basal V-ATPase in this tissue. Finally, in PMG, elevation of luminal pH increases (Cai2+) and decreases intracellular pH as measured by BCECF fluorescence. These responses suggest that the rate of acid secretion by PMG might be responsive to local demand for luminal reacidification as well as to serosal serotonin.

Acidification of damaged cells is a consequence of damage to the gastric epithelium, and may contribute to surface pH increases after damage

Focal damage of the gastric surface epithelium causes a transient rise in extracellular pH (pHo) over the damage site, prior to epithelial repair. To identify sources of the pHo rise we measured intracellular pH (pHi) of cells in and around the damaged area. The exposed gastric mucosa of anesthetized mice was loaded with 5 μM SNARF‐5F‐AM, and tissue imaged on a two‐photon microscope stage, during tissue superfusion with buffered pH 3 saline. A 2‐3 cell region of the gastric epithelium was photodamaged (710 nm, 350 mW for 5‐10 sec). Damage was monitored as loss of NAD(P)H autofluorescence and disrupted cell structure (confocal reflectance). pHi was measured near the damage (5‐10 μm), far from the damage (100‐150 μm), or in the actual damaged area. Before damage, initial pHi values were similar in all cell groups (~pH 7.1). After damage, pHi values were similar in near and far cells (7.17 ± 0.06 and 7.15 ± 0.04), but decreased to 6.45 ± 0.08 in damaged cells (n=4). Adding 20 μM iv EIPA or 100 μM luminal SITS did not significantly change initial pHi values in any cell type. After damage, EIPA selectively acidified damaged cells vs control (6.00 ± 0.09 vs. 6.45 ± 0.08, n=4), but SITS did not affect damaged cell acidification (6.59 ± 0.3, n=4). We conclude that damaged gastric epithelial cells may activate basolateral Na/H exchange as part of transepithelial alkali secretion that causes the luminal pHo increase seen after epithelial damage.

Revisiting the cellular mechanisms of strong luminal alkalinization in the anterior midgut of larval mosquitoes

Here we critically review two recent hypotheses about the mechanism of strong alkalinization by the anterior midgut of mosquito larvae and our tests of these hypotheses. We present experimental evidence against the major components of transport models proposed in these hypotheses. Measurements of the transapical and transbasal proton electrochemical gradients provide an indication of driving forces faced by and generated by the transport mechanisms of the tissue. These measurements confirmed that basal V-ATPase energizes alkalinization. Serotonin stimulates the V-ATPase, as indicated by the ensuing increase in proton-motive force across the basal membrane. Moreover, the neurohormone resulted in a surprisingly large increase in the intracellular pH. The results of inhibitor studies indicate that, contrary to previous proposals, carbonic anhydrase is apparently not involved in supplying acid-base equivalents to the respective transporters. Furthermore, any apical processes proposed to be involved in alkali secretion or acid absorption must be Cl(-) independent and insensitive to DIDS, amiloride, Zn(2+) and ouabain. These results argue against the involvement of putative apical Cl(-)/HCO (-)(3) exchangers, apical H(+) channels, apical cation/proton exchangers and the importance of the apical Na(+)/K(+) pump. The studies analyzed here thus provide both a limitation and direction for further studies of the mechanism of strong alkalinization in this system.

HCO3−secretion in the esophageal submucosal glands

The mammalian esophagus has the capacity to secrete a HCO3−and mucin-rich fluid in the esophageal lumen. These secretions originate from the submucosal glands (SMG) and can contribute to esophageal protection against refluxed gastric acid. The cellular mechanisms by which glandular cells achieve these secretions are largely unknown. To study this phenomenon, we used the pH-stat technique to measure luminal alkali secretion in an isolated, perfused pig esophagus preparation. Immunohistochemistry was used to localize receptors and transporters involved in HCO3−transport. The SMG-bearing esophagus was found to have significant basal alkali secretion, predominantly HCO3−, which averaged 0.21 ± 0.04 μeq·h−1·cm−2. This basal secretion was doubled when stimulated by carbachol but abolished by HCO3−or Cl−removal. Basal- and carbachol-stimulated secretions were also blocked by serosal application of atropine, pirenzipine, DIDS, methazolamide, and ethoxzolamide. The membrane-impermeable carbonic anhydrase inhibitor benzolamide, applied to the serosal bath, partially inhibited basal HCO3−secretion and blocked the stimulation by carbachol. Immunohistochemistry using antibodies to M1cholinergic receptor or carbonic anhydrase-II enzyme showed intense labeling of duct cells and serous demilunes but no labeling of mucous cells. Labeling with an antibody to Na+-(HCO3−)n(rat kidney NBC) was positive in ducts and serous cells, whereas labeling for Cl−/HCO3−exchanger (AE2) was positive in duct cells but less pronounced in serous cells. These data indicate that duct cells and serous demilunes of SMG play a role in HCO3−secretion, a process that involves M1cholinergic receptor stimulation. HCO3−transport in these cells is dependent on cytosolic and serosal membrane-bound carbonic anhydrase. HCO3−secretion is also dependent on serosal Cl−and is mediated by DIDS-sensitive transporters, possibly NBC and AE2.

Regulated alkali secretion acts in tandem with unstirred layers to regulate mouse gastric surface pH

Background & Aims : The physical basis for the protective pH gradient at the gastric surface is unconfirmed. This study examined the role of mucus, the unstirred layer, and acid/alkali secretion in controlling gastric surface pH in vivo. Methods : Stomachs of anesthetized mice were exteriorized, and exposed gastric mucosa was imaged by confocal microscopy. Results : Accessibility of molecules at the gastric surface was determined by monitoring the decrease in probe fluorescence over time after dyes were removed from perfusate. On dye removal, Cl-NERF (400 molecular weight) fluorescence decreased more slowly at the gastric surface in the presence of mucus (rate constant [k] = 0.08 ± 0.02 per second) than in the absence of mucus (k = 0.15 ± 0.02 per second) or 90 μm distant from the surface (k = 0.22 ± 0.03 per second). In contrast, 70-kilodalton Cl-NERF/dextran washed from the gastric surface more slowly in the presence (k = 0.05 ± 0.01 per second) or absence (k = 0.09 ± 0.01 per second) of mucus compared with 90 μm from the tissue surface (k = 0.36 ± 0.08 per second). Two-photon uncaging of fluorescein near nonsuperfused gastric surface showed that diffusion was not slowed at the gastric surface compared with diffusion in free solution. Surface pH was measured by Cl-NERF ratio imaging. Increasing the superfusion rate decreased the thickness of the surface pH gradient without significantly changing surface pH values, suggesting a pH set point of approximately 4 for control of surface pH. Increasing perfusate pH buffers decreased surface pH toward perfusate values. Conclusions : Proton concentration at the gastric surface is the result of regulating acid/alkali secretion to a set point in combination with an unstirred layer and not by trapping of proton or small-molecular-weight buffers in the unstirred layer.

Endogenous cyclo-oxygenase activity regulates mouse gastric surface pH.

In the stomach, production of prostaglandins by cyclo-oxygenase (COX) is believed to be important in mucosal defence. We tested the hypothesis that endogenous COX activity is required for protective gastric surface pH control. Intact stomachs of anaesthetized mice were perfused with a weakly buffered solution (150 mM NaCl + 4 mM Homopipes) at pH values from 2.5 to 7.0. Gastric effluents were collected to measure pH and estimate amounts of acid or alkali secretion in nanomoles secreted per minute. A switch from net acid to net alkali secretion was seen in response to acidifying luminal pH with an apparent 'set point' between pH 4 and 5. At luminal pH 3, the net alkali secretion (12.7 +/- 2.8 nmol OH(-) equivalents min(-1)) was abolished (2.2 +/- 1.7 nmol OH(-) min(-1)) by the non-specific COX inhibitor indomethacin (5 mg kg(-1) I.P.). Similar inhibition was observed using a COX-1 inhibitor (SC-560; 10 mg kg(-1) I.P.), but not a COX-2 inhibitor (NS-398; 10 mg kg(-1) I.P.). Subsequent treatment with 16,16-dimethyl prostaglandin E(2) (dm-PGE(2); 1 mg kg(-1) I.P.) rescued the alkali secretion (21.8 +/- 2.7 nmol OH(-) min(-1)). In either the absence or presence of the H(+),K(+)-ATPase inhibitor omeprazole (60 mg kg(-1) I.P.), indomethacin blocked similar amounts of net alkali secretion (10.5 +/- 2.7 and 16.4 +/- 3.4 nmol OH(-) min(-1), respectively). We also used in vivo confocal microscopy to examine pH near the mucosal surface. The gastric mucosal surface of anaesthetized mice was exposed and mucosal surface pH was imaged using the fluorescence intensity ratio of Cl-NERF as a pH indicator. Results showed a switch from a continuous net acid to net alkali secretion by the stomach in response to changing superfusate pH from 5 to 3. At luminal pH 3, the relatively alkaline surface pH (4.3 +/- 0.1) was acidified (3.6 +/- 0.2) by indomethacin, and subsequent dm-PGE(2) restored surface pH (4.2 +/- 0.2). We conclude that the pre-epithelial alkaline layer is regulated by endogenous COX activity.

Coordinated regulation of gastric chloride secretion with both acid and alkali secretion.

Gastric secretion of hydrochloric acid requires protons and chloride, yet the mechanisms and regulation of gastric chloride secretion remain unclear. We developed an in vivo technique to simultaneously measure acid/base and chloride secretion into the gastric lumen of anesthetized rats. The cannulated stomach lumen was perfused with weakly pH-buffered chloride-free solution containing a chloride-sensitive fluorophore [5 microM N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE)]. Gastric acid and chloride secretion was detected in gastric effluents by 1) flow-through pH electrode and 2) MQAE fluorescence. Gastric effluent was also collected at 1-min intervals for independent determination of chloride amount by chloridometer. In all conditions, both optical and chemical determinations of chloride report similar amounts of secreted chloride. During luminal perfusion with pH 5 solution, net acid and chloride secretion into the lumen was observed. Pentagastrin stimulated both secretions. In contrast, proton pump inhibition (omeprazole) caused alkalinization of the gastric effluent, but chloride secretion was not diminished. During luminal pH 3 perfusion, net alkali secretion was observed, and chloride secretion at luminal pH 3 was greater than pH 5. When tissue is pretreated with omeprazole at luminal pH 3, the addition of prostaglandin E2 synchronously stimulates both alkali and chloride secretion. Results suggest that both acid and alkali secretions are separately coupled with chloride secretion.

Luminal acid elicits a protective duodenal mucosal response.

Measuring duodenal epithelial intracellular pH (pHi), blood flow and mucus gel thickness (MGT), we studied duodenal defense mechanisms in vivo so as to more fully understand the protective mucosal response to luminal acid. Exposure of the mucosa to physiologic acid solutions promptly lowered pHi, followed by recovery after acid was removed, indicating that acid at physiologic concentrations readily diffuses into, but does not damage duodenal epithelial cells. Cellular acid then exits the cell via an amiloride-inhibitable process, presumably sodium-proton exchange (NHE1). MGT and blood flow increase promptly during acid perfusion; both decrease after acid challenge and are inhibited by vanilloid receptor antagonists or by sensory afferent denervation. Bicarbonate secretion does not increase during acid perfusion but increases following acid challenge. Inhibition of cellular alkali uptake by anion transport inhibitors lowers pHi, and increases mucosal injury, whereas inhibition of apical alkali secretion alkalinizes pHi and diminishes injury. These observations support the following hypothesis: luminal acid diffuses into the epithelial cells, lowering pHi. Acidic pHi increases the activity of a basolateral NHE, acidifying the submucosal space and increasing cellular alkali loading. The acidic submucosal space activates capsaicin receptors on afferent nerves, increasing MGT and blood flow. With continued acid exposure, a new steady state with thickened mucus gel, increased blood flow, and a higher cellular buffering power protects the mucosa from acid injury. After acid challenge, mucus secretion, blood flow and pHi return to normal, while bicarbonate secretion increases. Through these integrated mechanisms, the epithelial cells are protected from damage due to repeated pulses of concentrated gastric acid.

Choosing sides in the battle against gastric acid.

The duodenum is the first segment of the small intestine, just distal to the gastric pylorus. This positions the epithelial lining of the duodenum directly in the path of destructive gastric acid leaving the stomach. Because pancreatic bicarbonate secretions only enter the gut lumen farther down the gastrointestinal tract, the early duodenum must defend itself, unaided, from this acidic stress. The duodenal defenses can be conveniently grouped as pre-epithelial (e.g., neutralization of acid in the lumen), epithelial (e.g., tight junctions, membrane impermeability to acid, secretion of alkali and mucus), and subepithelial (e.g., afferent and efferent neural mechanisms, blood flow). Each of these contributions to mucosal defense is potentially important to human health because acid damage is a crucial aggravating factor in formation and persistence of duodenal ulcers (1). In terms of duodenal defense, strong interest has centered on the avid bicarbonate secretion mediated by duodenal epithelial cells. This is unequivocally the first line of defense, as the bicarbonate reaching the gut lumen will act to neutralize intraluminal acid (producing CO2 and water) before it ever reaches the cells. Elegant and rigorous experiments have shown that, as a consequence of this neutralization, the spaces directly adjacent to the epithelium have close to a neutral pH (2, 3). This juxtamucosal alkaline layer also contributes to mucosal defense by directly shielding duodenal epithelial cells from luminal acid. The article by Akiba et al. in this issue of the JCI (4) is interesting and important because it raises a new possibility: that bicarbonate secretory mechanisms may have an even greater role in acid defense through their effects on intracellular pH of duodenal epithelial cells. The central observation reported by Akiba et al. is that two anion transport inhibitors (DIDS and NPPB) both block bicarbonate secretion into the lumen but that these agents have opposite effects on cellular damage in response to luminal acid (4). Most strikingly, NPPB actually reduces the damage in response to luminal acid. These results are not readily compatible with conventional models of extracellular defense, since inhibition of bicarbonate secretion will compromise both luminal neutralization of acid and the juxtamucosal alkaline layer. Despite uncertainty about the molecular specificity of the drugs, these new in vivo results clearly demonstrate a dissociation between bicarbonate secretion and cytoprotection.

Intragastric pH regulates conversion from net acid to net alkaline secretion by the rat stomach.

Our previous report showed gastric mucosal surface pH was determined by alkali secretion at intragastric luminal pH 3 but by acid secretion at intragastric pH 5. Here, we question whether regulation of mucosal surface pH is due to the effect of luminal pH on net acid/base secretions of the whole stomach. Anesthetized rats with a gastric cannula were used, the stomach lumen was perfused with weakly buffered saline, and gastric secretion was detected in the gastric effluent with 1) a flow-through pH electrode and 2) a fluorescent pH-sensitive dye (Cl-NERF). During pH 5 luminal perfusion, both pH sensors reported the gastric effluent was acidic (pH 4.79). After perfusion was stopped transiently (stop-flow), net acid accumulation was observed in the effluent when perfusion was restarted (peak change to pH 4.1-4.3). During pH 3 luminal perfusion, both pH sensors reported gastric effluent was close to perfusate pH (3.0-3.1), but net alkali accumulation was detected at both pH sensors after stop-flow (peak pH 3.3). Buffering capacity of gastric effluents was used to calculate net acid/alkaline secretions. Omeprazole blocked acid secretion during pH 5 perfusion and amplified net alkali secretion during pH 3 perfusion. Pentagastrin elicited net acid secretion under both luminal pH conditions, an effect antagonized by somatostatin. We conclude that in the basal condition, the rat stomach was acid secretory at luminal pH 5 but alkaline secretory at luminal pH 3.

Changes in luminal pH provoke conversion of whole stomach from net acid to net alkali secretion

Changes in luminal pH provoke conversion of whole stomach from net acid to net alkali secretion

Mucus and alkali secretion in the rat duodenum: Effects of indomethacin, Nω-Nitro- l-arginine, and luminal acid

Background & Aims Duodenal mucus and bicarbonate secretion appear to play an essential role in the protection of the duodenum. The aim of this study was to examine duodenal bicarbonate and mucus secretion and the effects of cyclooxygenase inhibition, nitric oxide synthase inhibition, and luminal acid. Methods Duodenal mucus gel thickness was measured using microelectrodes during intravital microscopy in anesthetized rats. Bicarbonate secretion was measured using back-titration. Results A continuous layer of mucus with a mean thickness of 284 ± 11 μm (n = 35) and a mean alkaline secretion of 0.18 ± 0.01 μmol · cm −2 · min −1 were found in untreated animals. Indomethacin decreased both mucus and bicarbonate secretion by about 35%. NO synthase inhibition with N ω-nitro- l-arginine reduced mucus secretion by about 21% but increased bicarbonate secretion by 39%. Exposure of the mucosal surface to 10 mmol/L HCl increased mucus secretion by 44% and bicarbonate secretion by 22%. Conclusions The duodenal mucus layer is continuous. It can be easily removed, and new secretion can be followed. Duodenal mucus secretion is strongly stimulated by luminal acid and endogenous prostanoids and less markedly by NO, whereas bicarbonate secretion is stimulated by acid and endogenous prostanoids and inhibited by endogenous NO

Neural influences on human esophageal and salivary alkali secretion

Esophageal secretion HCO3- ions occurs in opossum and man and may contribute to mucosal defense. Using a perfusion technique, neuroregulatory influences on esophageal and salivary HCO3- secretion were investigated in 24 healthy human subjects. The sight and smell of food increased median salivary HCO3- output from 424 to 573 mumol/15 min (P = 0.014), without significantly altering esophageal HCO3- secretion (74-105 mumol/15 min, P = 0.24). Atropine reduced both salivary (610 to 68, 17, 10, and 3 mumol/15 min in successive periods; P < 0.028) and esophageal HCO3- output (108 to 78, 35, 18, and 7 mumol/10 cm/15 min; P < 0.028, respectively. Following atropinization, cholinergic stimulation failed to increase salivary secretion but did "unmask" a small rise in esophageal alkali output (7 to 27 mumol/10 cm/15 min, P = 0.036), implicating a noncholinergic mechanism. Cold-induced pain activated sympathetic reflexes and reduced esophageal HCO3- output (91 to 64 mumol/10 cm/15 min, P = 0.041) without influencing salivary secretion. These observations support a role for the autonomic nervous system in modulating human esophageal and salivary HCO3- secretion.

Effect of topical oesophageal acidification on human salivary and oesophageal alkali secretion.

Recent human studies suggest that oesophageal HCO3- secretion, in conjunction with salivary HCO3- secretion and secondary oesophageal peristalsis, is important for the protection of oesophageal mucosa from refluxed gastric contents. This study evaluated simultaneously the responsiveness of oesophageal and salivary HCO3- secretion to oesophageal acidification in eight healthy subjects. A 10 cm segment of oesophagus was perfused at a constant rate of 5 ml/min with a specially designed tube assembly. Saline was used initially, and then 10 mM and 100 mM HCl. The perfusates contained 3H-polyethylene glycol (PEG) as a concentration marker to determine volumes. Corrections were applied for a small degree of contamination by swallowed saliva and refluxed gastric alkali. Oesophageal perfusion with 10 mM HCl did not cause symptoms (nausea and heartburn), but tripled the oesophageal HCO3- output from a baseline of 51 mumol/10 cm/10 min (p = 0.021), while doubling the rate of salivary HCO3- secretion from a median basal value of 140 mumol/10 min (p = 0.021). Oesophageal perfusion with 100 mM HCl was associated with symptoms of nausea and heartburn in all subjects. The median oesophageal HCO3- output increased 32 fold to 1659 mumol/10 cm/10 min (interquartile range 569 to 3373; p = 0.036), and salivary HCO3- secretion approximately tripled from basal values (p = 0.036). In conclusion, oesophageal acidification stimulates both salivary and oesophageal HCO3- secretion, responses which may be protective to the oesophageal epithelium.

Gastric mucosal injury in amplified by inhibition of nitric oxide synthase activity and attenuated by nitric oxide

Nitric oxide (NO) affects gastric mucosal blood flow, acid and alkali secretion and, thus, contributes to the maintenance of mucosal integrity. We, therefore, tested the effect of several irritants on NO synthase (NOS) activity and its inhibition on the extent of mucosal injury. Fasted rats were treated i.g. with 1 ml 96% ethanol (E), with or without cotreatment i.v. with L-NAME (50 mg/kg) or 3-morpholino-sydnonime (2 mg/kg) (SIN), a stimulator of NO generation, administered 30 min prior to E. Other rats were treated with iodoacetamide (IA), an SH alkylator, added to the drinking water (0.1%), with or without L-NAME (0.1 mg/ml) or i.g., with aspirin (AS) (100 mg/kg) or indomethacin (I) (20 mg/kg). Rats were sacrificed 30 min after E, two days after IA or 3 h after AS or I, the stomach removed, washed, mucosal damage assessed (ram 2) and mucosal NOS activity determined (nmol/g/min). The damage induced by all irritants was accompanied by significant decrease in mucosal NOS activity. Co-treatment with L-NAME amplified the sevedty of the damage induced by E and by IA. Co-treatment with SIN induced a 73% decrease in the severity of the damage induced by E. Treatment No. Lesions NOS None 20 0 5.8 + 0.2 Ethanol 24 59 + 8* 2.9 + 0,2* E +L-NAME 10 151 + 26** 1.6 + 0.2** E+ S I N 10 16+5* 3.3+0.2 Iodoacetamide 10 27 + 2* 3.2 + 0.4* IA + L-NAME 10 128+51* 2.5+0.1 Aspirin 10 15 + 4* 3.2 _+ 0.4* Indomethacin 10 10 + 4* 3.1 _+ 0.1 *significantly different from no treatment, p<0.05; ** from E or IA, p<O.05 Conclusions: The important contribution of NO to the maintenance of gastric mucosal integrity is indicated by: 1) amplification of gastric injury induced by inhibition of NO synthesis and its attenuation by NO donation; 2) decreased gastric mucosal NOS activity induced by all irritants. BRUSH CYTOLOGY: A RELIABLE METHOD IN DETECTING HELICOBACTER PYLORI INFECTION. E. Radice. M.Musella, F. D'Armiento. Dept. of Sperimental and Clinical Medicine, Medical School of Catanzaro, Italy

Model of bicarbonate secretion by resting frog stomach fundus mucosa. II. Role of the oxyntopeptic cells.

In the present publication we report mainly electrophysiological studies on oxyntopeptic cells of frog gastric mucosa which aim at clarifying a possible involvement of these cells in the process of resting gastric alkali (HCO3-) secretion, described in the preceding publication. The experiments were performed on intact gastric fundus mucosa of Rana esculenta mounted in Ussing chambers. After removal of the muscle and connective tissue layer oxyntopeptic cells were punctured from the serosal surface with conventional or pH-sensitive microelectrodes to measure, besides transepithelial voltage and resistance, the basolateral cell membrane potential, the voltage divider ratio, and the cell pH in response to secretagogues and/or changes in serosal ion concentration. Carbachol (10(-4) mol/l), which transiently stimulated HCO3- secretion by 0.22 mumol.cm-2.h-1, transiently acidified the cells by 0.09 +/- SEM 0.03 pH units (n = 6) and transiently induced an apical cell membrane anion conductance. According to the model of gastric HCO3- secretion presented in the preceding publication, this anion conductance could be involved in gastric HCO3- secretion, mediating, besides Cl- efflux, also apical HCO3- efflux. In addition carbachol stimulated basolateral Na+(HCO3-)n-cotransport, which according to the results from the preceding publication mediates basolateral HCO3- uptake for secretion. By contrast, cAMP-mediated secretagogues, such as histamine or others, which stimulate HCl secretion and transiently alkalinize the oxyntopeptic cells, were found to down-regulate the basolateral Na+(HCO3-)n-cotransporter.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization and regulation of acid-base secretory currents from individual epithelial cells

The turtle urinary bladder is composed of different epithelial cell types that are suspected to separately produce electrogenic acid and alkali excretion. We measured the electrical currents produced by individual cells, scanning a two-dimensional vibrating probe over the luminal surface of the bladder. Acidification (outward current) was produced by the type of epithelial cell rich in carbonic anhydrase (CA cells). The measured currents of these cells quantitatively accounted for the total epithelial acidification current. When alkali secretion was induced by adenosine 3',5'-cyclic monophosphate and acidification was inhibited (by luminal pH 4), we measured inward currents localized to a small number of epithelial cells in four bladders but found no localization in the other seven treated bladders. When alkali secretion was localized and induced without inhibiting acidification, we found both cells producing inward current and cells producing outward current, which demonstrated that the two transport functions can occur simultaneously. We conclude that net acid-base secretion can be determined by regulating the transport rates of separate cells.