Mechanisms of protective activity of 16,16-dimethyl PGE2 and acetazolamide on gastric and duodenal lesions in rats

16,16-Dimethyl PGE₂ (16,16-dmPGE₂), given orally at 10–30 g/kg, had no effects on gastric acid secretion, or carbonic anhydrase activity, but did increase HCO₃ ^– secretion in both the stomach and duodenum of rats. 16,16-dmPGE₂, at nonantisecretory doses, potently inhibited indomethacin- and water-immersion stress-induced gastric lesions and mepirizole-induced duodenal lesions in rats. Acetazolamide, given orally at 50 mg/kg, markedly inhibited carbonic anhydrase activity, but had no effects on gastric acid secretion and the basal and 16,16-dmPGE₂-stimulated HCO₃ ^– secretion. Acetazolamide, at a nonantisecretory dose, had no effects on indomethacin-induced gastric lesions and mepirizole-induced duodenal lesions, but significantly inhibited water-immersion stress-induced gastric lesions. Combined administration of 16,16-dmPGE₂ and acetazolamide did not influence the protective activity of 16,16-dmPGE₂ on these lesions. The mechanism of the cytoprotective activity of 16,16-dmPGE₂ may involve an increase in HCO₃ ^– secretion (nonmediated by carbonic anhydrase), while mechanisms involved in the effects of acetazolamide are apparently different.     

Gastric motility is an important factor in the pathogenesis of indomethacin-induced gastric mucosal lesions in rats

Effects of atropine, cimetidine, and 16,16-dimethyl prostaglandin E₂ (16,16-dmPGE₂) on indomethacin-induced gastric lesions were investigated in rats by correlating their effects on gastric acid and HCO^}-₃ secretion and motility. Subcutaneously administered indomethacin (25 mg/kg) produced gastric mucosal lesions within 4 hr. In parallel studies, an equivalent dose of indomethacin inhibited gastric HCO^}-₃ secretion, and stimulated gastric motor activity measured as intraluminal pressure recordings, whereas acid secretion was unaffected. The lesions induced by indomethacin were significantly prevented by three agents: cimetidine (100 mg/kg), which reduced acid secretion; atropine (1 mg/kg), which reduced acid secretion and gastric motility; and 16,16-dmPGE₂ (10 g/kg), which reduced acid secretion and motility and increased gastric HCO^– ₃ secretion. If acid (150 mM HCl) was infused into the stomach (1.2 ml/hr) during indomethacin treatment, only the latter two agents significantly prevented the formation of gastric lesions in response to indomethacin. Since only the effect on gastric motility was common to these two agents (atropine and 16,16-dmPGE₂), the increased gastric motility may be an important pathogenetic factor in indomethacin-induced gastric lesions. The presence of acid as well as a deficiency of endogenous PGs may be prerequisite for later extension of the lesions but cannot account for the induction of mucosal lesions in rats following administration of indomethacin.     

Determination of acid-neutralizing capacity in rat duodenum

A model involving measurement of duodenal pH and acid-neutralizing capacity has been devised in anesthetized rats. A duodenal loop was made between the pyloric ring and the area just proximal to the outlet of the common bile duct (2 cm) and was perfused at a flow rate of 1.3 ml/min with HCl solution (1×10^–4 M, pH 4.0) made isotonic with NaCl. The pH of duodenal perfusate was continuously measured using a pH glass electrode of the flow type, and the amount of acid neutralized in the loop was titrated to pH 4.0 using a pH-stat method and by adding 10 mM HCl. Under normal conditions, the duodenal pH was kept around 6.0 as the result of neutralization in the loop (9 eq/hr). Subcutaneous administration of 16,16-dmPGE₂ (10 g/kg) significantly elevated the pH and increased acid-neutralizing capacity to 168.3% of normal levels. In contrast, indomethacin (5 mg/kg) and aspirin (200 mg/kg) as cyclooxygenase inhibitors or quinacrine (100 mg/kg) as a phospholipase A₂ inhibitor significantly decreased both the pH and acid neutralizing capacity. After sacrifice with saturated KCl (intravenously), the pH decreased to 4.3±0.2 and the neutralizing capacity was reduced to 30% of normal values. Basal HCO₃ ^– secretion in the proximal duodenum (5 eq/hr), when titrated to pH 7.4, was significantly stimulated by 16,16-dmPGE₂ and exposure of the mucosa for 10 min to 10 mM HCl. Neither indomethacin, aspirin, nor quinacrine had any effect on basal HCO₃ ^– secretion, but all significantly inhibited HCl-stimulated HCO₃ ^– secretion. These results suggest that endogenous prostaglandins play an important role in maintaining acid neutralizing capacity in the duodenum. The present system could be useful for screening drugs which may influence acid neutralizing capacity in the duodenum and for investigating the mechanisms of duodenal HCO₃ ^– secretion underin vivo conditions.     

Effects of hemorrhagic shock on alkaline secretion and mucosal tolerance to acid in rat duodenum

The effects of hemorrhagic shock (HE) on duodenal HCO₃ ^– secretion and mucosal tolerance to acid were investigated in anesthetized rats and compared with those of indomethacin. HE was performed by bleeding from the carotid artery to reduce arterial blood pressure to about 50 mm Hg (3 ml bleeding per 200 g of body weight) with a significant decrease in arterial pH and [HCO₃ ^–], and indomethacin was given subcutaneously in a dose of 5 mg/kg. The proximal duodenum (1.7 cm) secreted HCO₃ ^– at the rate of 1.5–1.8 eq/15 min (3.5–4.2 eq/cm/hr), and responded to luminal acid (10 mM HCl for 10 min) by a significant rise in HCO₃ ^– output. Indomethacin had no effect on basal HCO₃ ^– output but significantly inhibited the acid-induced HCO₃ ^– secretion, while under HE conditions duodenal HCO₃ ^– output significantly declined and failed to increase in response to luminal acidification. Subcutaneously administered 16, 16-dmPGE₂ (30 g/kg) significantly increased HCO₃ ^– secretion in the presence of indomethacin but had less effect on the impaired HCO₃ ^– output caused by HE. In contrast, intravenous infusion of NaHCO₃ (3 mmol/kg/hr) ameliorated the acid-base imbalance caused by HE, and significantly restored the impaired HCO₃ ^– responses induced by HE but not by indomethacin. Both HE and indomethacin induced extensive damage in the mucosa when the duodenal loop was perfused with 50 mM HCl for 1.5 hr, and these lesions were significantly reduced by NaHCO₃ infusion and 16, 16-dmPGE₂, respectively. These results suggest that HE impaired duodenal HCO₃ ^– secretion and reduced the tolerance of the mucosa to acid. This effect may be mainly a result of a decrease of HCO₃ ^– availability, but it is not accounted for by a deficiency of endogenous prostaglandins.     

Effects of nitric oxide synthase inhibitor NG-nitro-l-arginine methyl ester on duodenal alkaline secretory and ulcerogenic responses induced by mepirizole in rats

The inhibition of nitric oxide (NO) production by NO synthase inhibitors stimulates HCO ₃ ^– secretion in the rat duodenal mucosa. Therefore, we examined the effects of N^G-nitro-l-arginine methyl ester (l-NAME, the NO synthase inhibitor) and nitroprusside (the exogenous NO donor) on the duodenal HCO ₃ ^– and ulcerogenic responses in anesthetized rats. Animals were administered mepirizole (200 mg/kg, subcutaneously) for induction of duodenal ulcers, and gastric acid and duodenal HCO ₃ ^– secretions were measured with or without pretreatment withl-NAME (5 mg/kg, intravenously) or nitroprusside (4 mg/kg, intravenously). Mepirizole increased acid secretion, decreased the acid-induced duodenal HCO ₃ ^– secretion, and induced hemorrhagic lesions in the proximal duodenum. The inhibition of NO production byl-NAME potentiated the acid secretory response, increased the duodenal HCO ₃ ^– secretion, and prevented the duodenal lesions, and these changes were all antagonized by simultaneous administration ofl-arginine (200 mg/kg, intravenously) but notd-arginine. On the other hand, nitroprusside slightly reduced the acid response but further decreased the HCO ₃ ^– output, resulting in aggravation of duodenal lesions induced by mepirizole. These data suggest that the inhibition of endogenous NO production by the NO synthase inhibitorl-NAME increases duodenal HCO ₃ ^– secretion and protects the duodenal mucosa against acid injury.     

Pathogene sis of digitoxin-induced duodenal ulcers in pregnant rats

Late-stage pregnant rats (day 17) had higher rates of gastric acid secretion (45–55 eq/15 min) as compared to nonpregnant and middle-stage pregnant (day 10) rats (20–25eq/15 min). In contrast, basal rates of duodenal alkaline secretion were significantly lower (2–3 eq/15 min) in pregnant rats (day 10 and 17) than those in nonpregnant rats (5 eq/15 min), although the duodenal mucosa responded to acid with a significant rise in HCO₃ ^} output in these three groups of rats. In pregnant rats (day 17), a single injection of digitoxin, a Na⁺K⁺-ATPase inhibitor (10 mg/kg, subcutaneously), had no effect on basal acid and alkaline secretions, but significantly blocked the acid-induced HCO₃ ^}-secretion for more than 18 hr from 6 hr after administration. This drug, when given once daily for four days (10 mg/kg, subcutaneously), produced well-defined ulcers in the proximal duodenum with few lesions in the stomach of female rats, and the severity and incidence were significantly higher in late-stage pregnant rats than in the other two groups of rats. Following repeated administration of digitoxin (10 mg/kg) to late-stage pregnant rats (days 17–20), acid secretion significantly declined after two days of treatment, while the acid-induced HCO₃ ^}-secretion was significantly attenuated after one day of treatment and remained inhibited during the whole period. These results suggest that an impairment of the mechanisms related to acid-induced HCO₃ ^}secretion may be associated with the induction of duodenal ulcers caused by digitoxin in female rats, and the high incidence of these ulcers in late-stage pregnant rats may be due to acid hypersecretion.     

Role of Endogenous Nitric Oxide and Prostaglandin in Duodenal Bicarbonate Response Induced by Mucosal Acidification in Rats

Duodenal HCO₃ ^– secretion increases in response to mucosal acidification by luminal acid. Although this process is known to be mediated by endogenous prostaglandins (PGs), the role of nitric oxide (NO) in this response has been little studied. We examined the effects of indomethacin and N_G-nitro-l-arginine methyl ester (l-NAME) on the acid-induced HCO₃ ^– secretion in the rat duodenum, together with those on PGE₂ generation as well as luminal release of NO metabolites (NO_x). A proximal duodenal loop was perfused with saline, and the HCO₃ ^– secretion was measured at pH 7.0 using a pH-stat method and by adding 10 mM HCl. Mucosal acidification was performed by exposing the loop to 10 or 100 mM HCl for 10 min. Acidification of the duodenal mucosa stimulated the HCO₃ ^– secretion, with concomitant increase of mucosal PGE₂ contents and luminal release of NO_x, the response being much greater in case of 100 mM HCl. Indomethacin significantly inhibited the acid-induced HCO₃ ^– secretion as well as the PGE₂ biosynthetic response, without influence on the NO_x release. Pretreatment of the animals with l-NAME attenuated both the increase of mucosal PGE₂ contents and luminal release of NO_x following the acidification, resulting in a marked inhibition of the acid-induced HCO³> ^– response, and these effects were significantly antagonized by coadministration of l-arginine. Duodenal HCO₃ ^– secretion was also increased by mucosal exposure to NOR-3 (a NO donor), with concomitant increase of PGE₂ generation, but these effects were mitigated in the presence of indomethacin. In addition, the duodenal damage caused by mucosal perfusion with 100 mM HCl for 4 hr was markedly aggravated by pretreatment with l-NAME as well as indomethacin. These results suggest that both endogenous NO and PGs are involved in the mechanism for the acid-induced duodenal HCO₃ ^– secretion, and that NO may increase the HCO₃ ^– secretion by stimulating PG generation.     

Body temperature-dependent action of baclofen in rat stomach

The effect of baclofen (PCPGABA) on acid secretion, motility, and mucosa was investigated in the anesthetized rat stomach under various body temperatures (BT: 28–38° C), and they were compared with those of 2-deoxy-d-glucose (2DG), an acid stimulant through cytoglycopenia. Under these conditions PCPGABA induces lesions dose-dependently (>1 mg/kg, subcutaneously) in both the stomach and duodenum, and this action was dependent on BT; lowering of BT enhanced the ulcerogenicity. PCPGABA (3 mg/kg) had no effect on acid secretion at higher BT (36–38° C) but produced a marked increase of acid output at lower BT (30–32° C). 2DG caused a stimulation of acid output and gastric lesions without BT dependency, but the duodenal ulcerogenicity enhanced at lower BT. Gastric motility was enhanced significantly by these two agents to similar degrees, at either high or low BT. Neither PCPGABA nor 2DG affected alkaline secretion in the duodenum, while lowering of BT by itself reduced alkaline secretory responses. The above changes caused by PCPGABA and 2DG were blocked by both atropine and vagotomy. These results suggest that (1) acid stimulatory and ulcerogenic action of PCPGABA may involve a temperature-dependent process but does not relate to a cytoglycopenia, and (2) the vagus nerve mediating acid secretion and motility may be different in the temperature dependency.     

Duodenal acidity “sensing” but not epithelial HCO3? supply is critically dependent on carbonic anhydrase II expression

Carbonic anhydrase (CA) is strongly expressed in the duodenum and has been implicated in a variety of physiological functions including enterocyte HCO₃⁻ supply for secretion and the “sensing” of luminal acid and CO₂. Here, we report the physiological role of the intracellular CAII isoform involvement in acid-, PGE_2, and forskolin-induced murine duodenal bicarbonate secretion (DBS) in vivo. CAII-deficient and WT littermates were studied in vivo during isoflurane anesthesia. An approximate 10-mm segment of the proximal duodenum with intact blood supply was perfused under different experimental conditions and DBS was titrated by pH immediately. Two-photon confocal microscopy using the pH-sensitive dye SNARF-1F was used to assess duodenocyte pH_i in vivo. After correction of systemic acidosis by infusion of isotonic Na₂CO₃, basal DBS was not significantly different in CAII-deficient mice and WT littermates. The duodenal bicarbonate secretory response to acid was almost abolished in CAII-deficient mice, but normal to forskolin- or 16,16-dimethyl PGE₂ stimulation. The complete inhibition of tissue CAs by luminal methazolamide and i.v. acetazolamide completely blocked the response to acid, but did not significantly alter the response to forskolin. While duodenocytes acidified upon luminal perfusion with acid, no significant pH_i change occurred in CAII-deficient duodenum in vivo. The results suggest that CA II is important for duodenocyte acidification by low luminal pH and for eliciting the acid-mediated HCO₃⁻ secretory response, but is not important in the generation of the secreted HCO₃⁻ ions.     

Effect of duodenal ulcerogens cysteamine,mepirizole, and MPTP on duodenal myoelectric activity in rats

Increased gastric acid secretion, enhanced acid delivery to the duodenum, and reduced alkaline secretion in the proximal duodenum are relatively well-established pathophysiologic abnormalities in duodenal ulcer. Impaired duodenal motility, however, may also contribute to duodenal ulceration by altering the distribution of acid and alkaline secretions along the upper digestive tract. We tested the hypothesis that the duodenal ulcerogens cysteamine, MPTP, and mepirizole modify duodenal motility in the rat and that motility changes might be a common and early alteration in experimental duodenal ulceration. All three duodenal ulcerogens rapidly produced extensive changes in duodenal myoelectric activity and reduced the frequency of myoelectric slow waves. Cysteamine induced marked hypermotility for at least 6 hr; MPTP rapidly decreased motility and fragmented the myoelectric migrating pattern. Mepirizole induced biphasic changes: an early hypermotility phase of about 30 min was followed by profound hypomotility. These results indicate that marked alterations of duodenal motility are common during experimental duodenal ulceration. In light of the differential effect of the ulcerogens on duodenal motility, it remains to be determined how these changes influence acid neutralization in the proximal duodenum. Nevertheless, our results suggest that all three duodenal ulcerogens, which are different in structure, alter duodenal motility.Dr. Mangla did this work while on sabbatical leave at Harvard Medical School.     

Stimulation by Capsaicin of Duodenal HCO3 − Secretion via Afferent Neurons and Vanilloid Receptors in Rats: Comparison with Acid-Induced HCO3 − Response

We compared the HCO₃ ^– secretory response to capsaicin and mucosal acidification in rat duodenums, especially the relation to vanilloid receptor type 1 (VR1). A proximal duodenal loop was perfused with saline, and the HCO₃ ^– secretion was measured at pH 7.0 using a pH-stat method and by adding 10 mM HCl. The secretion was stimulated by exposing the loop to capsaicin (0.03–0.3 mg/ml) or 10 mM HCl for 10 min. Indomethacin subcutaneously or ruthenium red intravenously, a nonspecific VR1 antagonist, was given 60 or 10 min, respectively, before exposure to capsaicin or acid, while l-NAME was given intravenously 3 hr before these treatments. Capsazepine, another VR1 antagonist, was coapplied to the loop for 10 min with capsaicin or acid. Luminal application of capsaicin increased the secretion of HCO₃ ^– in a dose-dependent manner; this effect was markedly attenuated by chemical ablation of capsaicin-sensitive afferent neurons (CSN) as well as pretreatment with ruthenium red or capsazepine, and significantly mitigated by indomethacin or l-NAME (in an l-arginine-sensitive manner). The HCO₃ ^– secretion was also stimulated by mucosal acidification, and this response was attenuated by both capsaicin pretreatment, indomethacin and l-NAME, but not ruthenium red or capsazepine. Mucosal application of capsaicin as well as acid increased the mucosal PGE₂ content, and these effects were both significantly attenuated by indomethacin and l-NAME. These results suggest that both capsaicin and acid cause the CSN-dependent increase in duodenal HCO₃ ^– secretion mediated by NO and PG, yet the mode of their action differs in terms of the ruthenium red or capsazepine sensitivity. Although luminal H⁺ plays a modulatory role in duodenal HCO₃ ^– secretion, it is unlikely that the action results from the interaction of H⁺ with the ruthenium red- or capsazepine-sensitive site of VR1.     

COX and NOS isoforms involved in acid-induced duodenal bicarbonate secretion in rats

Duodenal HCO₃ ^– secretion increases in response to luminal acid, mediated by endogenous nitiric oxide (NO) as well as prostaglandins (PGs). In this study, we examined the effects of various inhibitors of cyclooxygenase (COX) or NO synthase (NOS) on the acid-induced HCO₃ ^– secretion in rats and determined the enzyme isoforms responsible for this response. A proximal duodenal loop was perfused with saline under urethane anesthesia, and the HCO₃ ^– secretion was measured at pH 7.0 using a pH-stat method and by adding 10 mM HCl. Mucosal acidification was performed by exposing the loop to 10 mM HCl for 10 min. Indomethacin, SC-560 (a selective COX-1 inhibitor) and rofecoxib (a selective COX-2 inhibitor) were given intraduodenally 1 hr before exposure to 10 mM HCl, while N ^G-nitro-l-arginine methyl ester (l-NAME: a nonselective NOS inhibitor) and aminoguanidine (a relatively selective inhibitor of iNOS) were given subcutaneously 3 hr before the acidification. The mucosal acidification increased the HCO₃ ^– secretion, with a rise in mucosal PGE₂ content and luminal release of NO. The HCO₃ ^– secretory and PGE₂ biosynthetic responses were significantly inhibited by indomethacin and SC-560, while rofecoxib had no effect on these responses. On the other hand, l-NAME, but not aminoguanidine, attenuated NO release following the acidification, resulting in inhibition of the acid-induced HCO₃ ^– secretion in a l-arginine-sensitive manner. Neither COX-2 nor iNOS mRNAs were observed in the mucosa before and 1 hr after acidification, while the gene expression of COX-1 and nNOS was constitutively detected in the mucosa and appeared to be slightly up-regulated after the acid stimulation. These results suggest that COX-1 and cNOS play as the respective key enzyme responsible for producing PG and NO following the duodenal acidification, both of which are involved in the mechanism for the acid-induced HCO₃ ^– secretion in the duodenum.     

Role of prostanoids in experimental duodenal ulcer in rat

The purposes of this study were to determine whether inhibition of cyclooxygenase is a mechanism by which cysteamine and mepirizole produce duodenal ulcers, identify qualitative or quantitative differences in prostanoid production between gastric mucosa and duodenum, and determine whether differences in cyclooxygenase sensitivity to inhibition by aspirin exist between these two tissues. In fed female rats, gastric mucosal prostaglandin E₂ (PGE₂) and prostacyclin (PGI₂) generation was 235±25 and 832±40 ng/g/min, respectively, whereas full-thickness duodenal PGE₂ and PGI₂ generation was 665±46 and 662±49 ng/g/min, respectively. Over an intraperitoneal dose range of 0– 25 mg/kg, aspirin-induced cyclooxygenase inhibition was dose-dependent and similar for the two tissues. Duodenal ulceration (16.7 mm²) produced by cysteamine, 425 mg/kg, was associated with a 46% reduction in duodenal PGE₂ generation, while having no effect on PGI₂ generation; however, cysteamine, 213 mg/kg, produced no visible duodenal mucosa injury yet reduced duodenal PGE₂ generation 39% compared to control values. In fed male rats, gastric mucosal PGE₂ and PGI₂ generation was 179± 18 and 813± 61 ng/g/min, respectivley, whereas duodenal PGE₂ and PGI₂ generation was 321± 27 and 454± 38 ng/g/min, respectively. Duodenal ulceration (7.7 ± 2.3 mm²) produced by oral mepirizole was associated with a 63% reduction in duodenal PGE₂ generation compared to control values, while having no effect on PGI₂ generation. Subcutaneous aspirin, 100 mg/kg, which reduced duodenal PGE₂ generation to a greater degree than either ulcerogen, given in conjunction with pentagastrin, did not produce visible duodenal ulceration. It therefore seems unlikely that reduced PGE₂ generation within the duodenum is the primary mechanism of gross injury associated with these two ulcerogens.Supported by grant AM 17328 from NIADDK.     

HCl-stimulated duodenal HCO 3 − secretion in conscious rat

Using an isolated loop of the proximal duodenum of conscious rats, the role of vasoactive intestinal peptide (VIP) in the duodenal HCO ₃ ^– response to HCl was examined, especially interactions with participating cholinoceptor mechanisms and prostaglandins. A 5-min perfusion with 150 mmol/liter HCl increased luminal VIP during 3 hr, with a peak output during and immediately after the acid challenge. The HCl-stimulated output was unaffected by atropine and hexamethonium, but was augmented by indomethacin from 13.6 (9.5–17.8) to 39 (20–85) fmol/cm/min. The HCO ₃ ^– secretion in response to graded doses of intravenous VIP (0.00625–6 nmol/kg/30 min) was dose-dependent to maximally 33.5±10.5 µmol/cm/hr. The HCO ₃ ^– secretion during a single intravenous infusion of VIP (12 nmol/kg/hr), 13.9±4.2 µmol/cm/hr, was unchanged by atropine, reduced to 10.0±3.5 µmol/cm/hr by hexamethonium, and augmented to 18.9±4.7 µmol/cm/hr by indomethacin. Exogenous VIP did not change the basal luminal output of PGE₂; neither did exogenous PGE₂ nor indomethacin affect the basal luminal output of VIP. HCl-induced increases in luminal outputs of VIP, substance P, and neurokinin A (the two latter with unknown roles) were differentially affected by atropine, hexamethonium, and indomethacin, indicating that the acid challenge released the peptides through controlled mechanisms. In conclusion, in the duodenal HCO ₃ ^– response to luminal HCl, VIP may have a stimulatory role, which partially depends on nicotinic, but not on muscarinic cholinoceptor mechanisms, and which is negatively modulated by prostaglandins.The study was supported by the Swedish Society of Medicine, The Professor Nanna Svartz Foundation, The Medical Research Council of the Swedish Life Insurance Companies, The Swedish Medical Research Council (7464), The Swedish Cancer Fund (2313) and by Funds of the Karolinska Institute.     

Effect of somatostatin-14 on duodenal mucosal bicarbonate secretion in guinea pigs

The role of somatostatin-14 in duodenal mucosal HCO ₃ ^– secretion was investigated in anesthetized, indomethacin-treated guinea pigs. Net HCO ₃ ^– output from the isolated, perfused (24 mM NaHCO₃ + 130 mM NaCl) proximal duodenum was measured during intravenous infusion (alone or in combination) of somatostatin-14, carbachol, vasoactive intestinal peptide (VIP), and prostaglandin E₂ (PGE₂). In homogenates of duodenal enterocytes, the effect of these agents on adenylate cyclase activity was studied. Basal duodenal HCO ₃ ^– secretion (3.5±0.2µmol/cm/10 min) was reduced dose dependently by somatostatin-14 (10^–11 mol/kg, 10^–9 mol/kg, and 10^–7 mol/kg). Carbachol, VIP, and PGE₂ (all 10^–8 mol/kg) increased basal duodenal HCO ₃ ^– secretion two- to threefold. Somatostatin-14 (10^–7 mol/kg) abolished the stimulatory effect of carbachol and VIP, but not that of PGE₂. Basal adenylate cyclase activity in isolated duodenal enterocytes (9.4±1.0 pmol cAMP/mg protein/min) was unaltered by somatostatin (10^–6 mol/liter) or carbachol (10^–3 mol/liter). VIP (10^–8 mol/liter) and PGE₂ (10^–7 mol/liter) increased adenylate cyclase activity two- to threefold, and these effects were unchanged by somatostatin-14 (10^–6 mol/liter). In conclusion, somatostatin-14 inhibits basal and carbachol- and VIP-stimulated duodenal HCO ₃ ^– secretion, and its mechanism of action is not via inhibition of adenylate cyclase activity in duodenal enterocytes.This study was supported by grants from the German-Israel Foundation for Scientific Research and Development (grant I-78-054.2/88), and the Israeli Ministry of Health.     

Role of carbonic anhydrase in basal and stimulated bicarbonate secretion by the guinea pig duodenum

The role of carbonic anhydrase in the process of proximal duodenal mucosal bicarbonate secretion was investigated in the guinea pig. In a series of experimentsin vivo, the duodenum was perfused with 24 mmol/liter NaHCO₃ solution (+ NaCl for isotonicity) to ensure that active duodenal HCO ₃ ^– secretion against a concentration gradient was measured. Acetazolamide (80 mg/kg) was infused intravenously to examine the role of carbonic anhydrase on basal and agonist-stimulated HCO ₃ ^– secretion. Acetazolamide abolished basal HCO ₃ ^– secretion and significantly decreased HCO ₃ ^– secretion after stimulation with dibutyryl 5-cyclic adenosine monophosphate (dBcAMP, 10^–5 mol/kg), dibutyryl 5-cyclic guanosine monophosphate (dBcGMP, 10^–5 mol/kg), prostaglandin E₂ (PGE₂, 10^–6 mol/kg), PGF₂ (10^–6 mol/kg), tetradecanoyl-phorbol-acetate (TPA, 10^–7 mol/kg), glucagon (10^–7 mol/kg), vasoactive intestinal polypeptide (VIP, 10^–8 mol/kg), and carbachol (10^–8 mol/kg). Utilizing a fluorescence technique, we could detect the enzyme carbonic anhydrase in equal amounts in villous and crypt cells of the proximal duodenal epithelium; no activity was demonstrated in tissues pretreated with acetazolamide. In conclusion, carbonic anhydrase is required for both basal and stimulated duodenal HCO ₃ ^– secretion.This research was supported by the German-Israel Foundation for Scientific Research and Development, Grant Number I-78-054.2/88.     

Induction of duodenal ulcers in rats under water-immersion stress conditions. Influence of stress on gastric acid and duodenal alkaline secretion

We investigated the influence of stress on gastric acid and duodenal HCO3- secretion in rats, and examined whether duodenal ulcers develop in rats under stress conditions in the absence or presence of acid hypersecretion caused by histamine. Either restraint alone or restraint plus water-immersion stress induced lesions in the stomach but not in the duodenum. However, subcutaneous administration of histamine dihydrochloride (40 mg/kg every 2.5 h for a total of three times) to stressed rats produced macroscopically visible damage in the proximal duodenum as well as in the stomach within 8 h of exposure to stress, and the incidence of duodenal lesions was 100% in the water-immersion group (24.8 +/- 3.8 mm2, n = 8). Histamine alone had no effect on either region. These lesions in the duodenum caused by water immersion plus histamine were prevented by subcutaneously administered cimetidine (30, 100 mg/kg) or 16,16-dimethyl prostaglandin E2 (10, 30 micrograms/kg) in a dose-related manner, but not by atropine (1, 3 mg/kg). Restraint decreased acid secretion by 40%, and additional water immersion restored the decreased acid secretion to normal levels. Basal duodenal HCO3- secretion was decreased to about 70% of normal values (5-6 microEq/15 min) in the restraint group, and after additional water immersion further declined to the values of 1.5-2 microEq/15 min. An increase of HCO3- secretion caused by acid was significantly inhibited by water immersion but not by restraint. Histamine significantly increased acid secretion but did not affect duodenal HCO3- secretion. In the rats treated with both water immersion and histamine, acid secretion was significantly reduced by either cimetidine (100 mg/kg) or 16,16-dimethyl prostaglandin E2 (30 micrograms/kg), whereas duodenal HCO3- secretion was significantly increased by 16,16-dimethyl prostaglandin E2. Atropine had little effect on either acid or HCO3- secretion. These results suggest that exposure of rats to stress decreases duodenal HCO3- secretion and increases the susceptibility of the mucosa to acid emptied from the stomach, thereby inducing duodenal ulcers if acid hypersecretion is concomitantly present.     

Exogenous prostaglandin protects against acid-induced deep mucosal injury by stimulating alkaline secretion in rat duodenum

In the anesthetized rat, exogenous acid (0.1–0.3 N HCl) perfused through the duodenum produced a dose-related increase in the severity of duodenal villous injury. Increasing the duration of perfusion of the 0.1 N HCl also increased the severity of the injury. The increase in the severity of the lesion score was due to an increase in the percentage of villi with damage extending to the lower half of the villus. 16,16-Dimethyl prostaglandin E₂ (dm PGE₂, 5 g/kg) administered subcutaneously significantly increased duodenal mucosal alkaline secretion and significantly reduced the duodenal villous injury produced by 0.1 N HCl. The reduction in the severity of the lesion score was due to a decrease in the percentage of villi with the deeper type of damage. These data indicate: (1) perfusion of the rat duodenum with 0.1 N HCl at 0.1 ml/min for 30 min provides a valid model for assessing deep duodenal villous injury, (2) exogenous prostaglandin enhances the resistance of the duodenal mucosa against acid induced deep villous injury, and (3) the enhanced resistance may be mediated at least in part by stimulation of duodenal alkaline secretion. The results support the hypothesis that stimulated duodenal alkaline secretion may play a role in defense of the duodenal mucosa against acid-induced deep villous injury.     

Role of capsaicin-sensitive sensory nerves in acid-induced bicarbonate secretion in rat stomach

The effect of capsaicin-sensitive afferent nerves on the alkaline secretory response induced by mucosal acidification was investigated in theex vivo stomachs of anesthetized rats. The stomach was mounted on a Lucite chamber and perfused with saline (pH 4.5) in the absence of acid secretion (omeprazole pretreatment: 60 mg/kg, intraperitoneal), and luminal pH and transmucosal potential difference (PD) were monitored simultaneously. Under these conditions the gastric mucosa responded to intravenous infection of prostaglandin E₂ (PGE₂: 300 g/kg) and mucosal acidification (0.2 N HCl for 10 min) by a significant increase of pH with a slight decrease of PD; the HCO₃ ^– output was 9.2±0.7 mol and 8.4±0.8 mol, respectively. The increased pH and HCO₃ ^– responses were significantly inhibited by prior administration of indomethacin (5 mg/kg, subcutaneously) or chemical deafferentation following capsaicin injections (total dose: 100 mg/kg, subcutaneously), whereas those induced by PGE₂ remained unchanged after either treatment. On the other hand, the mucosal application of capsaicin (0.3–6 mg/ml) increased the luminal pH and HCO₃ ^– output in a concentration-related manner, and this action was also significantly attenuated by either indomethacin or chemical deafferentation of capsaicin-sensitive sensory neurons. These results suggest that capsaicin-sensitive sensory nerves may be involved in the mechanism of acid-induced HCO₃ ^– secretion in the stomach, in addition to endogenous PGs, and these two pathways may interact somewhere in the stimulatory process.     

Regional differences of H+, HCO3-, and CO2 diffusion through native porcine gastroduodenal mucus

Gastroduodenal mucus may play a critical role in defending the epithelium from luminal acid and in the creation of a microenvironment suitable for H. pylori. We measured transmucus permeation of H⁺, HCO₃ ^–, and CO₂ with an in vitro perfusion chamber through freshly harvested or partially purified porcine gastric mucin. pH and CO₂ concentrations were measured with selective ion electrodes; HCO₃ ^– and CO₂ concentrations were derived. Viscosity was measured by rotational microviscometry. Mucin viscosity was directly related to concentration. There was a large variation in viscosity among native mucus from antrum, corpus, and duodenum. The highest viscosity was found in the antral mucus; duodenal mucus had the lowest. Diffusion coefficients of duodenal mucus for H⁺ and HCO₃ ^– were significantly lower than those from corpus and antrum. CO₂ diffusion coefficients were invariant. In conclusion, despite large variations in viscosity, antral and corpus gastric mucus were similar in terms of ion diffusion. Surprisingly, the low viscosity duodenal mucus was a more potent barrier to ion diffusion than was gastric mucus. Consequently, duodenal mucus may play a more important role in inhibiting ion diffusion than its gastric counterpart.