Stimulative effect of skipjack tuna soluble extract on pepsin-like protease in the stomach of rockfish (Sebastes schlegelii) using an in vitro perfusion method

The effect of intra-gastric infusion of skipjack tuna (Katsuwonus pelamis) muscle soluble extract and of carnosine, anserine and histidine, on pepsin-like protease activity was studied in the isolated, externally batch-cultured stomach of the rockfish (Sebastes schlegelii). Stomach was isolated from the fish, then intragastrically infused with the above solutions or with a balanced saline solution (control solution) as the stimulant at 0.1 mL/min for 6h. Intra-gastric efflux was collected for measurement of pepsin-like protease activity. Skipjack tuna soluble extract, but not carnosine, anserine or histidine alone caused significant enhancement of pepsin-like protease activity during the infusion. Pepsin-like protease activity from skipjack tuna soluble extract responded immediately after infusion and was kept for 150 min after infusion. Stomach motility at the end of infusion was also observed. These results suggest that isolated stomach can receive mucosal side stimulants in vitro. It is likely that some effective components for stomach digestion in fish exist in skipjack tuna soluble extract.     

Direct interactions of mastoparan and compound 48/80 with GTP-binding proteins

The effects of mastoparan and compound 48/80 on the activities of alpha beta gamma-trimeric GTP-binding proteins (G proteins) were studied with purified Go and Gi-1 which had been reconstituted into phospholipid vesicles. Pertussis toxin-catalyzed ADP-ribosylation of Go or Gi-1 was inhibited by mastoparan or compound 48/80, suggesting that the G proteins were dissociated into their constituent alpha- and beta gamma-subunits in the presence of these compounds. The steady-state rate of GTP hydrolysis catalyzed by Go or Gi-1 was stimulated by the two compounds. Both the stimulations were due to increases in the rate of the GDP-GTP exchange reaction occurring on the G proteins. However, the modes stimulation of the GTPase activity depended on the type of G protein used, and the stimulations caused by the two compounds were differently affected by pertussis toxin-catalyzed ADP-ribosylation of G proteins. Moreover, the mastoparan-induced stimulation of the GTPase activity was partially inhibited by compound 48/80. Thus, the two histamine secretagogues mastoparan and compound 48/80 appear to activate G proteins differently, though they interact with the signal-transducing proteins, at least partly, at a common binding site.     

Microflow system promotes acetaminophen crystal nucleation

It is known that interfaces have various impacts on crystallization from a solution. Here, we describe crystallization of acetaminophen using a microflow channel, in which two liquids meet and form a liquid–liquid interface due to laminar flow, resulting in uniform mixing of solvents on the molecular scale. In the anti‐solvent method, the microflow mixing promoted the crystallization more than bulk mixing. Furthermore, increased flow rate encouraged crystal formation, and a metastable form appeared under a certain flow condition. This means that interface management by the microchannel could be a beneficial tool for crystallization and polymorph control.     

Chronic exposure to beta-hydroxybutyrate inhibits glucose-induced insulin release from pancreatic islets by decreasing NADH contents

To investigate the effects of chronic exposure to ketone bodies on glucose-induced insulin secretion, we evaluated insulin release, intracellular Ca2+ and metabolism, and Ca2+ efficacy of the exocytotic system in rat pancreatic islets. Fifteen-hour exposure to 5 mM d-beta-hydroxybutyrate (HB) reduced high glucose-induced insulin secretion and augmented basal insulin secretion. Augmentation of basal release was derived from promoting the Ca2+-independent and ATP-independent component of insulin release, which was suppressed by the GDP analog. Chronic exposure to HB affected mostly the second phase of glucose-induced biphasic secretion. Dynamic experiments showed that insulin release and NAD(P)H fluorescence were lower, although the intracellular Ca2+ concentration ([Ca2+](i)) was not affected 10 min after exposure to high glucose. Additionally, [Ca2+](i) efficacy in exocytotic system at clamped concentrations of ATP was not affected. NADH content, ATP content, and ATP-to-ADP ratio in the HB-cultured islets in the presence of high glucose were lower, whereas glucose utilization and oxidation were not affected. Mitochondrial ATP production shows that the respiratory chain downstream of complex II is not affected by chronic exposure to HB, and that the decrease in ATP production is due to decreased NADH content in the mitochondrial matrix. Chronic exposure to HB suppresses glucose-induced insulin secretion by lowering the ATP level, at least partly by inhibiting ATP production by reducing the supply of NADH to the respiratory chain. Glucose-induced insulin release in the presence of aminooxyacetate was not reduced, which implies that chronic exposure to HB affects the malate/aspartate shuttle and thus reduces NADH supply to mitochondria.     

Mother–young distance in Japanese Black cattle at pasture

Distance between dam and offspring (1–121 days old) in a herd of Japanese Black cattle (Bos taurus) grazing a tropical grass (Paspalum notatum) pasture (1.5 ha) was investigated during 7-h grazing periods over grazing seasons from May (spring) to October (autumn). The mother–young distance was not constant throughout the grazing period, repeatedly increasing and decreasing. Although significant periodicity was always detected in the mother–young distance, there was no consistent dominant cycle, indicating the complexity of the within-day pattern of mother–young distance. The mean mother–young distance over the grazing period increased as a calf aged, reaching a plateau at an age of about 33 days. The mean distance of a calf from its mother was usually shorter than that from a non-mother cow, with the difference between the mean distances decreasing sharply until a calf became about 35 days old. The results and literature show that mutual independence of mother and young rapidly develops in the first 30–50 days after parturition. Electronic Publication     

Coupling of the guanine nucleotide regulatory protein to chemotactic peptide receptors in neutrophil membranes and its uncoupling by islet-activating protein, pertussis toxin. A possible role of the toxin substrate in Ca2+-mobilizing receptor-mediated signal transduction

A chemotactic peptide stimulated the high-affinity GTPase activity in membrane preparations from guinea pig neutrophils. The enzyme stimulation was inhibited by prior exposure of the membrane-donor cells to islet-activating protein (IAP), pertussis toxin, or by direct incubation of the membrane preparations with its A-protomer (the active peptide) in the presence of NAD. The affinity for the chemotactic peptide binding to its receptors was lowered by guanyl-5'-yl beta, gamma-imidodiphosphate (Gpp(NH)p) reflecting its coupling to the guanine nucleotide regulatory protein in neutrophils. The affinity in the absence of Gpp(NH)p was lower, but the affinity in its presence was not, in the A-protomer-treated membranes than in nontreated membranes. The inhibitory guanine nucleotide regulatory protein of adenylate cyclase (Ni) was purified from rat brain, and reconstituted into the membranes from IAP-treated cells. The reconstitution was very effective in increasing formyl-Met-Leu-Phe-dependent GTPase activity and increasing the chemotactic peptide binding to membranes due to affinity increase. The half-maximal concentration of IAP to inhibit GTPase activity was comparable to that of the toxin to inhibit the cellular arachidonate-releasing response which was well correlated with ADP-ribosylation of a membrane Mr = 41,000 protein (Okajima, F., and Ui, M. (1984) J. Biol. Chem. 259, 13863-13871). It is proposed that the IAP substrate, Ni, couples to the chemotactic peptide receptor and mediates arachidonate-releasing responses in neutrophils, as it mediates adenylate cyclase inhibition in many other cell types.     

Effect of hydrogen peroxide in redox status estimation using nitroxyl spin probe

A procedure for estimating in vivo redox status using EPR and a hydrogen peroxide (H₂O₂)-dependent spin probe method is described. The mechanism of decreasing spin clearance in the selenium-deficient (SeD) rat is discussed. The in vivo decay constant of the nitroxyl spin probe in the liver region of SeD rats appeared to be slightly lower that of the selenium-adequate control (SeC) group, and was significantly smaller than that of normal rats. Bile H₂O₂ levels in normal rats were significantly lower than those in SeD rats. The in vivo decay constant of the spin probe in SeD rats depended on the bile H₂O₂ level. Furthermore, H₂O₂ was detected in the bile in all SeD rats, whereas bile H₂O₂ could be detected in only half of the normal rats. It was found that the in vivo decay constant of the spin probe in normal rats also depended on whether bile H₂O₂ was detected or not. In vivo decay constants were smaller in rats subjected to the surgical operation than in the nonoperated groups. The EPR signal of the nitroxyl radical in the liver homogenate was increased by addition of H₂O₂, which was administered 30 min before the rat was killed. It appears that H₂O₂ can oxidize the hydroxylamine formed following reduction of the spin probe in the liver.