HCO 3_ secretion was investigated in interlobular duct segments isolated from guinea-pig pancreas using a semi-quantitative fluorometric method. Secretagogue-induced decreases in intracellular pH, following blockade of basolateral HCO 3 _ uptake with a combination of amiloride and DIDS, were measured using the pH-sensitive fluoroprobe BCECF. Apparent secretory HCO 3 _ fluxes were calculated from the initial rate of intracellular acidification.2. In the presence of HCO 3 _ , stimulation with secretin (10 nM) or forskolin (5 µM) more than doubled the rate of intracellular acidification. This effect was abolished in the absence of HCO 3 _ . It was also abolished in the presence of HCO 3 _ when DIDS and NPPB were applied to the luminal membrane by microperfusion. We therefore conclude that the increase in acidification rate is a useful index of secretagogue-induced HCO 3 _ secretion across the luminal membrane.3. Secretin, cholecystokinin (CCK) and bombesin each stimulated HCO 3 _ secretion in a dosedependent fashion. They evoked comparable maximal responses at about 10 nM and the EC 50 values were 0.5 nM for secretin, 0.2 nM for CCK and 30 pM for bombesin. Acetylcholine (ACh) was also effective, with a maximum effect at 10 µM.4. The stimulatory effect of CCK was blocked completely by the CCK 1 receptor antagonist devazepide but not by the CCK 2 receptor antagonist L365,260. The CCK analogue JMV-180 (Boc-Tyr(SO 3 H)-Nle-Gly-Trp-Nle-Asp-phenylethyl ester), which is an agonist of the highaffinity CCK 1 receptor but an antagonist of the low-affinity receptor, also stimulated HCO 3 _ secretion but with a smaller maximal effect than CCK. JMV-180 partially inhibited the response to a high concentration of CCK but not to a lower concentration, suggesting that both high-and low-affinity states of the CCK 1 receptor evoke HCO 3 _ secretion.5. The stimulatory effect of bombesin was blocked completely by the gastrin-releasing peptide (GRP) receptor antagonist D-Phe 6 -bombesin(6-13)-methyl ester (BME) but not by the neuromedin B (NMB) receptor antagonist D-Nal-cyclo[Cys-Tyr-D-Trp-Orn-Val-Cys]-Nal-NH 2 (BIM-23127).6. Secretagogue-evoked fluid secretion was also examined using video microscopy to measure the rate of swelling of ducts whose ends had sealed during overnight culture. Secretin, CCK, bombesin and ACh all evoked fluid secretion with maximal rates of approximately 0.6 nl min _1 mm _2, and with concentration dependences similar to those obtained for HCO 3 _ secretion.7. We conclude that CCK, bombesin and ACh stimulate the secretion of a HCO 3 _ -rich fluid by direct actions on the interlobular ducts of the guinea-pig pancreas and that these responses are mediated by CCK 1 receptors, GRP receptors and muscarinic cholinoceptors, respectively.

Genome-scale metabolic models (GEMs) have been widely used for phenotypic prediction of microorganisms. However, the lack of other constraints in the stoichiometric model often leads to a large metabolic solution space inaccessible. Inspired by previous studies that take allocation of macromolecule resources into account, we developed a simplified Python-based workflow for constructing enzymatic constrained metabolic network model (ECMpy) and constructed an enzyme-constrained model for Escherichia coli (eciML1515) by directly adding a total enzyme amount constraint in the latest version of GEM for E. coli (iML1515), considering the protein subunit composition in the reaction, and automated calibration of enzyme kinetic parameters. Using eciML1515, we predicted the overflow metabolism of E. coli and revealed that redox balance was the key reason for the difference between E. coli and Saccharomyces cerevisiae in overflow metabolism. The growth rate predictions on 24 single-carbon sources were improved significantly when compared with other enzyme-constrained models of E. coli. Finally, we revealed the tradeoff between enzyme usage efficiency and biomass yield by exploring the metabolic behaviors under different substrate consumption rates. Enzyme-constrained models can improve simulation accuracy and thus can predict cellular phenotypes under various genetic perturbations more precisely, providing reliable guidance for metabolic engineering.

Abstract. The present study was conducted to investigate the effects of histamine on the lipid metabolic disorder in mice loaded with restraint stress. When Kun Ming (KM) mice were exposed to restraint stress for 20 h, the histamine level in both plasma and cerebral regions significantly increased (P<0.01). Moreover, when a lipid emulsion (10% Intralipid ® ) was injected intravenously into the mice, the elimination period of plasma triglyceride was prolonged in the restraint group. Plasma triglyceride was 523 ± 44 mg/dl at 35 min after the Intralipid ® administration in the restraint stress group, while it was 436 ± 41 mg/dl in the restrained mice given histamine at a dose of 50 mg/kg. The improved plasma triglyceride metabolism was well explained by the observations of the significantly up-regulated hepatic triglyceride lipase (HTGL) activity and mRNA expression in response to histamine. These results suggested that the effects of stress-induced histamine on lipid metabolic disorder in mice loaded with restraint stress arose from its anti-stress action and promotion of lipase activity.

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