Tachykinins Stimulate Release of Peptide Hormones (Glucagon-Like Peptide-1) and Paracrine (Somatostatin) and Neurotransmitter (Vasoactive Intestinal Polypeptide) from Porcine Ileum Through NK-1 Receptors

The effects of infusion of the two tachykinins,substance P (SP) and neurokinin A (NKA), and ofcapsaicin on the release of glucagon-like peptide-1(GLP-1), somatostatin, and vasoactive intestinalpolypeptide (VIP) were studied in isolated, vascularlyperfused ileal segments. SP (10-8 M) stimulated GLP-1,somatostatin, and VIP release to 141.8 ± 6.6% (N= 18), 230.3 ± 38.7% (N = 21), and 359.7 ±60.5% (N = 22) of basal output, respectively. NKA(10^-8 M) only stimulated VIP release (to181.2 ± 16.7% of basal release, N = 22). Theeffects of SP and NKA were blocked by the NK-1 receptorantagonist CP96345 (10^-6 M). Infusion of atropine(10^-6 M) had no effect on the SP-inducedGLP-1 release, but partly inhibited the effect of SP onsomatostatin and VIP release, and the effect of NKA onVIP release. Capsaicin infusions (10^-5 M) significantlystimulated both GLP-1, somatostatin, and VIP release to111.1 ± 4.5% (N = 9), 138.0 ± 15.8% (N =9) and 208.3 ± 63.8% (N = 8) of basal release,respectively. Simultaneous addition of receptor antagonists to all threetachykinin receptors (CP96345, SR48968, and SR142801,all at 10^-6 M) significantly inhibited theeffect of capsaicin on VIP release, whereas the releaseof GLP-1 and somatostatin was unaffected. Weconclude that tachykinins potently stimulate the releaseof GLP-1, somatostatin, and VIP in the porcine ileum viaNK-1 receptors. The effect on somatostatin and VIP is partly mediated via cholinergic neurons.Sensory neurons releasing tachykinins could be involvedin the regulation of VIPergic neurons.     

Kinetic studies on the formation of nitrosamines I

Summary The kinetics of nitrosation of dimethylamine (DMA) in aqueous perchloric acid solution haves been studied using a differential spectrophotometric technique. The rate law is Initial rate=e[DMA]₀ [nitrite] ₀ ² [H⁺]/(f+[H⁺])² where [DMA]₀ and [nitrite]₀ represent initial stoichiometric concentrations. At 310.0 K and =2.0 M, e=(2.2±0.2)×10^–5 M^–1 s^–1 and f=(1.28±0.02) ×10^–3M. The associated activation energy is 56±3kJ mol^–1. A clear inhibition of the nitrosation rate by ionic strength has been observed in which only the kinetic parameter (f) has an effective change. It is concluded that under the experimental conditions of this work only the dinitrogen trioxid is the effective carrier for the nitrosation.     

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.     

Immunosuppressive effect of chenodeoxycholic acid on natural killer cell activity in patients with biliary atresia and hepatitis C virus-related liver cirrhosis

Patients with severe liver diseases, such as liver cirrhosis and biliary atresia, have low natural killer (NK) cell activity. The relations between NK activity and measures of liver function, including serum levels of total bilirubin, total bile acids, bile acid components, aspartate aminotransferase, and alanine aminotransferase, and platelet count were examined in patients with biliary atresia (6 boys and 6 girls; mean age, 4.8 ± 5.7 years) and patients with liver cirrhosis due to hepatitis C virus infection (10 men and 2 women; mean age, 54.3 ± 13.8 years). Univariate analysis showed that platelet count was positively correlated with NK activity in patients with biliary atresia (r = 0.611, P < 0.05). Serum levels of free chenodeoxycholic acid were negatively correlated with NK activity both in patients with biliary atresia (r = –0.647, P < 0.05) and in patients with hepatitis C virus-related liver cirrhosis (r = –0.876, P < 0.01). None of the other free bile acids or conjugated bile acids or other indicators of liver function were correlated with NK activity. Multiple stepwise regression analysis showed that only levels of free chenodeoxycholic acid were independently correlated with NK activity. All patients with biliary atresia underwent liver transplantation from living related donors. NK activity had increased significantly two months after transplantation (from 24.1 ± 20.2% to 49.2 ± 12.5%, P < 0.01). In contrast, levels of free chenodeoxycholic acid in transplant recipients had decreased significantly two months after transplantation (from 1.22 ± 1.16 to 0.26 ± 0.21 mol/l, P < 0.05). In conclusion, in patients with biliary atresia or liver cirrhosis, NK activity in peripheral blood decreases, mostly because of free chenodeoxycholic acid.     

Paradoxical effects of endogenous and exogenous insulin on amino acid transport activity in the isolated rat pancreas: somatostatin-14 inhibits insulin action

Summary Regulatory effects of insulin, somatostatin and cholecystokinin on amino acid transport in the isolated perfused rat pancreas have been studied using a rapid dual isotope dilution technique. Uni-directional L-serine transport (15 s) was quantified relative to an extracellular tracer D-mannitol over a wide range of substrate concentrations. In pancreata perfused with 2.5 mmol/l D-glucose, a weighted nonlinear regression analysis of overall transport indicated an apparent K_m=14.4±1.6 mmol/l and V_max=25.9±1.4 mol ·min^–1·g^–1 (n=6). Although L-serine transport was stimulated during perfusion with 100 U/ml bovine insulin, endogenous insulin (7–25 ng·min^–1·g^–1) released during continuous perfusion with either 8.8 mmol/l or 16.8 mmol/l D-glucose had no such effect. Exogenous somatostatin-14 (250 pg/ml) or cholecystokinin octapeptide (CCK-8, 3 × 10^–11mol/l) appeared to increase only the K_m for transport. Only CCK-8 evoked a notable protein output (2.9±0.3 mg·30min^–1·g^–1) and juice flow (68±10l·30min^–1·g ^–1, n=3) from the exocrine pancreas. When pancreata were perfused with bovine insulin (100 U/ml) and somatostatin-14 (250 pg/ml), the stimulatory action of exogenous insulin on L-serine transport was abolished. If endogenous insulin and somatostatin, released concurrently in response to 16.8 mmol/l D-glucose, were conveyed to the exocrine epithelium via an islet-acinar portalaxis, it is conceivable that somatostatin modulates the stimulatory action of insulin on basolateral amino acid transport in the exocrine pancreas.     

Effects of islet amyloid polypeptide on hepatic insulin resistance and glucose production in the isolated perfused rat liver

Summary The impact of (pancreatic) islet amyloid polypeptide on glucose metabolism and insulin sensitivity was examined in isolated rat livers perfused in a non-recirculating system. Continuous infusion of 10^–7mol/l islet amyloid polypeptide affected neither basal nor glucagon (10^–9 mol/l)-stimulated glucose output by livers from fed rats, but it did increase the hepatic cyclic AMP release within 44 min (7.91±12.07 vs control: 0.07±0.03 pmol·100 g body weight^–1). The effect of the peptide on the ability of insulin to inhibit glucagon-induced hepatic glycogenolysis was measured in three experimental groups (n = 6). As expected glucagon (7×10^–11 mol/l) increased integral hepatic glucose release within 84 min (763.4±161.7 vs –25.7±73.2 mol · 100 g body weight^–1 in the control group, p<0.001), while insulin (100 mU/l) decreased the glucagon-stimulated glucose production (395.2±180.0 mol·100 g body weight^–1, p<0.01). Simultaneous infusion of 10^–7 mol/l islet amyloid polypeptide however, was not able to reverse insulin-dependent inhibition of glucagon-stimulated hepatic glucose output (370.0±102.5 mol·100 g body weight^–1, NS) or to enhance lactate-induced gluconeogenesis of livers from 24 h fasted rats (n = 8). The glucose production stimulated by 10^–9 mol/l glucagon was slightly greater in islet amyloid polypeptide-pre-treated livers than in a control group without addition of islet amyloid polypeptide (5 min: 3.60±3.36 vs 1.67±1.28 mol·min^–1·100 g body weight^–1). These results suggest that islet amyloid polypeptide neither directly affects hepatic glycogenolysis nor causes insulin resistance to hormone-sensitive glucose production, but may increase the size of the hepatic glycogen pool by enhancing gluconeogenesis.     

Liver and muscle insulin sensitivity,glycogen concentration and glycogen synthase activity in a rat model of non-insulin-dependent diabetes

Summary Mild diabetes was induced in adult rats with streptozotocin (45 mg/kg body weight), and insulin sensitivity, glycogen deposition and glycogen synthase activity assessed in liver and muscle 5 weeks later. Diabetic rats had significantly elevated fasting blood glucose concentrations (5.6±0.1 versus 3.6±0.1 mmol/l, p<0.001), and blood glucose concentrations 2 h after a 1 g/kg glucose load (12.0±0.6 versus 3.7±0.2 mmol/l, p<0.001). After a 20-h fast hepatic glucose output was significantly elevated (58±3 versus 47±3 mol·min^–1·kg^–1, p<0.05), and failed to suppress at high insulin concentrations during a euglycaemic clamp (hepatic glucose output 21±4 versus 2±4 mol·min^–1·kg^–1, p<0.02). Liver glycogen was lower in the diabetic rats by the end of the clamp (16±3 versus 38±6 mol/g wet wt, p<0.05). At the end of the clamp total glucose turnover was lower in the diabetic rats (107±4 versus 161±17 mol·min^–1· kg^–1, p<0.01), as was skeletal muscle glycogen synthase activity (0.46±0.04 versus 0.67±0.05 U/g wet wt, p<0.01) and glycogen concentration (22±2 versus 33±3 mol/g wet wt, p<0.05). Blood lactate and pyruvate responses suggested that glycolytic pathways were similarly affected. Thus, insulin insensitivity develops in both liver and skeletal muscle after 5 weeks of mild streptozotocin-induced diabetes.     

Dissociation between contraction and relaxation: The possible role of phospholamban phosphorylation

Summary The relationship between myocardial relaxation and phosphorylation of phospholamban, an intrinsic protein of sarcoplasmic reticulum (SR), was studied in perfused rat hearts beating at constant rate and perfused at constant coronary flow. The positive inotropic effect (increase in developed tension, T, and maximal rate of rise of tension, + ) of 3×10^–9 and 3×10^–8M isoproterenol (ISO) occurred together, with a proportionately greater increase in maximal velocity of relaxation, – . Thus, the + /– ratio decreased 0.23±0.04 and 0.41±0.05 respectively. Time to half-relaxation (t_1/2) and the time constant of relaxation (Tau) were also significantly decreased by ISO. Phospholamban phosphorylation (in pmol³²Pi/mg SR protein) increased from 23±3.3 (control) to 42±2.3 (3×10^–9M ISO) and to 186±19.3 (3×10^–8M ISO). When the negative inotropic action of nifedipine was just offset by either Ca²⁺ (N–Ca²⁺) or ISO (N–I), relaxation was faster when ISO was present. Perfusion with N–I significantly decreased + /– 0.18±0.05, t_1/2 14±3 ms and Tau 1.4±0.2 ms. Phospholamban phosphorylation significantly increased from 23±3.3 to 40±4.9 pmol 32 Pi/mg SR protein. N–Ca²⁺ did not elicit any significant change in these parameters nor in phospholamban phosphorylation. Thus, phospholamban phosphorylation appears closely related to myocardial relaxation and may be one of the important mechanisms by which contractility and relaxation are dissociated in vivo.This work was supported by grants # 3-106600/85 from CONICET and # 2109-1239/85 from CIC.     

Hemodynamic effects of acute and chronic administration of vapreotide in rats with cirrhosis

The aim of this study was to assess the hemodynamic effects of acute and chronic administration of vapreotide, a somatostatin analog, in rats with intrahepatic portal hypertension induced by dimethylnitrosamine (DMNA) administration. Acute effects were evaluated at baseline and 30 min after placebo (N = 13) or vapreotide (8 g/kg/hr, N = 13) infusions in DMNA rats. Chronic hemodynamic effects were evaluated using subcutaneous implants for five weeks in anesthetized DMNA rats (placebo: N = 13, vapreotide: N = 13) and in sham rats (placebo: N = 10, vapreotide: N = 10). Hemodynamic measurements included splenorenal shunt blood flow (SRS BF) by the transit time ultrasound (TTU) method and cardiac output by the combined dilution–TTU method. Acute administration of vapreotide significantly decreased SRS BF (–17.3 ± 19 vs –1.1 ± 14%, P < 0.05) and portal pressure (–8 ± 9 vs 0 ± 8%, p < 0.05) compared to placebo without systemic effects. Chronic administration of vapreotide significantly reduced the increase in SRS BF (2.4 ± 1.5 vs 1.2 ± 1.0 ml/min, P < 0.05) and cardiac index (50 ± 15 vs 33 ± 10 ml/min/100 g, P < 0.0001) while portal pressure and blood flow, and mean arterial pressure were not significantly changed compared to placebo. In conclusion, the acute administration of vapreotide decreased collateral circulation blood flow while chronic administration attenuated its development. Vapreotide seems to have a vasoconstrictive effect on collateral circulation.     

Peripheral and hepatic insulin sensitivity in subjects with impaired glucose tolerance

Summary Recent evidence suggests that the post-prandial hyperglycaemia in impaired glucose tolerance is primarily due to impaired suppression of basal hepatic glucose output. This in turn appears to be secondary to decreased first phase insulin secretion, although decreased hepatic insulin sensitivity, which is a feature of non-insulin-dependent diabetes mellitus, might also play a role. Eight mildly overweight subjects with impaired glucose tolerance and eight closely matched control subjects with normal glucose tolerance underwent an intravenous glucose tolerance test to assess first phase insulin secretion. Insulin sensitivity was examined by a 150-min hyperinsulinaemic-euglycaemic clamp. Somatostatin was infused from 150 min to suppress endogenous insulin secretion, and glucagon and insulin were replaced by constant infusion. Glucose with added dideuterated glucose (labelled infusion technique) was infused to maintain euglycaemia. First phase insulin secretion ( 0–10 min insulin area ÷ 0–10 min glucose area) was significantly decreased in the subjects with impaired glucose tolerance (median [range]: 1.2 [0.2–19.4] vs 9.1 [2.6–14.5] mU·mmol^–1; p<0.01). During the clamp, circulating insulin (93±8 [mean±SEM] and 81±10 mU·l^–1) and glucagon (54±4 and 44±6 ng·l^–1) levels were comparable. Total glucose disposal was decreased in subjects with impaired glucose tolerance (2.78±0.27 vs 4.47±0.53 mg·kg^–1·min^–1; p<0.02), and was primarily due to decreased non-oxidative glucose disposal. However, hepatic glucose output rates were comparable during the clamp (0.38±0.10 and 0.30±0.18 mg·kg^–1·min^–1). Therefore, the main defects in subjects with impaired glucose tolerance are decreased first phase insulin secretion and peripheral non-oxidative glucose disposal, but hepatic glucose output shows normal responsiveness to insulin.Abbreviations FPIS First phase insulin secretion - PG plasma glucose - NIDDM non-insulin-dependent diabetes mellitus - IGT impaired glucose tolerance - HGO hepatic glucose output - IVGTT intravenous glucose tolerance test - OGTT oral glucose tolerance test     

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.     

Insulin regulation of glucose and lipid metabolism in massive obesity

Summary Eight obese patients and 12 normal individuals underwent a euglycaemic insulin clamp (20 and 40 mU · m^2–1 · min^–1) along with continuous infusion of 3-³H-glucose and 1-¹⁴C-palmitate and indirect calorimetry. Basal plasma glucose concentration (4.7±0.3 vs 4.4±0.2 mmol/l) was similar in the two groups, whereas hepatic glucose production was slightly higher in obese individuals (1.11±0.06 vs 0.84±0.05 mmol/min) in spite of higher plasma insulin levels (17±2 vs 6±1 mU/l; p<0.01). Insulin inhibition of hepatic glucose production was impaired in obese subjects. Glucose disposal by lean body mass was markedly reduced both at baseline (11.7±1.1 vs 15.6±0.6 mol · kg^–1 · min^–1; p<0.05) and during clamp (15.0±1.1 vs 34.4±2.8 and 26.7±3.9 vs 62.2±2.8 mol · kg^–1 · min^–1; p<0.01) Oxidative (12.2±1.1 vs 17.8±1 and 16.1±1.1 vs 51.1±1.7 mol · kg^–1 · min^–1; p<0.05–0.002) and non-oxidative glucose metabolism (3.9±1.1 vs 15.0±2.8 and 12.8±3.3 vs 38.3±2.2 mol · kg^–1 · min^–1; p<0.01–0.001) were impaired. Basal plasma concentrations of non-esterified fatty acids (635±75 vs 510±71 mol/l) and blood glycerol (129±17 vs 56±5 mol/l; p<0.01) were increased in obese patients. Following hyperinsulinaemia, plasma non-esterified fatty acids (244±79 vs 69±16 and 140±2 vs 36±10 mol/l; p<0.01) and blood glycerol levels (79±20 vs 34±6 and 73±22 vs 29±5 mol/l; p<0.01) remained higher in obese subjects. Baseline non-esterified fatty acid production rate per kg of fat body mass was significantly larger in normal weight subjects (37.7±6.7 vs 14.0±1.8 mol/l; p<0.01) and insulin inhibition was reduced in obese patients (–41±9 vs –74±3 and –53±11 vs –82±3%; p<0.05). Basal plasma non-esterified fatty acid utilization by lean body mass was similar in the two groups (9.8±0.9 vs 8.8±2.0 mol · kg^–1 · min^–1), whereas during clamp it remained higher in obese patients (6.0±1.2 vs 2.8±2.5 and 4.9±1.3 vs 1.5±0.6 mol · kg^–1 · min^–1; p<0.1–0.05). Lipid oxidation was higher in obese individuals in spite of hyperinsulinaemia (3.7±0.3 vs 2.4±0.4 and 2.3±0.4 vs 0.9±0.3 mol · kg^–1 · min^–1; p<0.05– 0.02). An inverse correlation was found between lipid oxidation and glucose oxidation (r=0.82 and 0.93; p<0.001) and glucose utilization (r=0.54 and 0.83; p<0.05–0.001) both in obese and control subjects. A correlation between lipid oxidation and non-oxidative glucose metabolism was present only in normal weight individuals (r=0.75; p<0.01). We conclude that in obesity all tissues (muscles, liver, and adipose tissue) are resistant to insulin action. Insulin resistance involves glucose as well as lipid metabolism.     

Ketone bodies increase glomerular filtration rate in normal man and in patients with Type 1 (insulin-dependent) diabetes mellitus

Summary The purpose of this study was to investigate whether the administration of acetoacetic and hydrochloric acids in a group of control and Type 1 (insulin-dependent) diabetic patients influenced renal haemodynamics. Renal plasma flow increased from 657±88 to 762±81 ml·min^–1. 1.73 m^–2 in diabetic patients (p<0.01) and from 590±71 to 691±135 in control subjects (p<0.01). Glomerular filtration rate increased from 135±9 to 180±8 ml·min^–1·1.73 m^–2 in diabetic patients (p< 0.001) and from 117±8 to 145±7 in control subjects (p<0.01). Similar effects on renal haemodynamics, even if less pronounced, were observed with low dose acetoacetic but not with hydrochloric acid infusion. Total protein, 2-microglobulin but not albumin excretion rates were increased by acetoacetic acid. We conclude that an acute increase in blood concentration of ketone bodies within the range found in diabetic patients with poor metabolic control (1) increases renal plasma flow and glomerular filtration rate both in control subjects and diabetic patients and (2) causes a tubular proteinuria.     

Vasoactive intestinal peptide

Dose—response characteristics of feline corpus circular muscle were studiedin vitro for three neuropeptides individually and with vasoactive intestinal peptide. Bombesin, substance P, and cholecystokinin-octapeptide each elicited concentration-dependent isometric contractions that were reduced by 10^–8 M or 10^–7 M vasoactive intestinal peptide (P<0.01). The concentration of each neuropeptide producing a half-maximal response was increased more than one logfold to 10^–6 M by vasoactive intestinal peptide. Tetrodotoxin blocked responses to bombesin (P<0.001) and reduced responses to substance P (P<0.05), but had no effect on responses to cholecystokinin-octapeptide (P>0.1). These results demonstrate inhibition of neuropeptide responses of gastric smooth muscle and support vasoactive intestinal peptide as an inhibitory regulator of gastric motor function.     

Effects of Free Radicals and Leukocytes on Increases in Blood–Brain Barrier Permeability During Colitis

This study examined the role of leukocytes and free radicals on increases in permeability of the blood–brain barrier (BBB) in rabbits with acute colitis. Trinitrobenzene sulfonic acid (TNBS) was used to induce colitis in male New Zealand White rabbits. The extraction ratio of fluorescein was used as an index of the permeability of the BBB. The extraction ratio for fluorescein was 1.0 × 10^–3 ± 2 × 10^–4 ml/g and 1.1 × 10^–3 ± 3 × 10^–4 ml/g (mean ± se) for saline (N = 7) and ascorbic acid-treated (N = 8) rabbits with sham colitis. Conversely, TNBS-induced acute colitis increased the extraction ratio over 70% in saline (N = 8) and ascorbic acid-treated (N = 8) animals. Vinblastine significantly reduced the number of circulating leukocytes, whereas the permeability of the BBB was augmented by 54% in rabbits with TNBS-induced acute colitis (N = 8). Vinblastine had no effect on the permeability of the BBB in rabbits with sham colitis (N = 8). These data suggest that free radicals are not responsible for BBB disruption, and leukocytes may protect the BBB during acute colitis.     

Vasoactive intestinal polypeptide is a potent regulator of bile secretion from rat cholangiocytes.

BACKGROUND & AIMS: Vasoactive intestinal polypeptide (VIP) is a neuropeptide with diverse biological functions including stimulatory effects on bile secretion. The effects of VIP on bile secretion and its site of action were examined. METHODS: Choleretic effects of VIP were examined using isolated perfused livers, hepatocyte couplets, isolated bile duct units, and cholangiocytes from rat liver. RESULTS: VIP (100 nmol/L) produced a small increase in bile flow and bile salt output in taurocholate-supplemented isolated perfused livers but had no significant effect on bile flow in the absence of bile salt supplements or on fluid secretion in isolated hepatocyte couplets. In addition, VIP significantly increased bile pH, bicarbonate concentration, and output in the isolated perfused livers from both normal and 2 week bile duct-ligated rats, although bile flow increased only in the bile duct-ligated model. VIP also produced a dose-dependent increase in fluid secretion in isolated bile duct units, which was inhibited significantly by VIP antagonist, a specific VIP receptor inhibitor. This VIP-stimulated secretory response in isolated bile duct units was more potent than those produced by bombesin or secretin. Neither somatostatin nor substance P inhibited the VIP response in isolated bile duct units. In contrast to secretin, VIP had no significant effect on adenosine 3', 5'-cyclic monophosphate (cAMP) levels in isolated cholangiocytes. CONCLUSIONS: VIP is a potent stimulant of fluid and bicarbonate secretion from cholangiocytes via cAMP-independent pathways, suggesting that this neuropeptide plays a major regulatory role in biliary transport and secretion.     

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.     

Postnatal development of hepatic bile formation in the rabbit

To investigate postnatal maturation of hepatic bile formation, bile output was measured in four groups of rabbits: suckling infants at ages 10–14, 18–22, and 26–30 days, and adults. Bile output was collected directly from the common duct during three 1-hr periods: a basal period followed by intravenous infusion of 1 and then 2 mol/min/kg of glycodeoxycholic acid. [¹⁴C]Erythritol and [³H]inulin clearances measured canalicular bile flow and biliary permeability. Under basal conditions and with the exogenous bile acid, bile flow and bile salt secretion were lowest in 10-to 14-day-old infants and showed a gradual increase with increasing age. Bile salt-dependent flow, the linear increase in flow relative to bile salt secretion, was higher in the 10-to 14-and 17-to 22-day-old compared to the adult and 25-to 30-day-old groups. The ratio of chloride to bile salt secreted was also higher in the two younger groups. Bile salt-independent flow at theoretical zero bile salt secretion was absent in the younger groups, but evident in the adult and 25-to 30-day-old rabbits. Canalicular flow estimated by erythritol clearance was linearly related to bile salt secretion. Inulin clearance relative to erythritol clearance was higher in the 10-to 14-day-old infants than the adults. Thus, bile flow and bile salt secretion are reduced in the young infant but rise to near adult levels at the time of weaning, 25–30 days in the rabbit. The increase in flow results from increased bile salt secretion and the appearance of bile salt-independent flow. Biliary permeability as measured by inulin clearance is increased in the young, allowing increased water and electrolyte movement per unit of bile salt to maintain bile flow despite a depressed bile salt secretion rate.This work was supported by grants from the Medical Research Council of Canada and the Canadian Liver Foundation. Dr. Zahavi was the recipient of a Fellowship from the Alberta Heritage Foundation for Medical Research.     

In situ binding of islet hormones in the isolated perfused rat pancreas: evidence for local high concentrations of islet hormones via the islet-acinar axis

Summary Insulin and somatostatin reportedly affect pancreatic acinar cell function via specific receptor binding. Theoretically peri-insular levels depend on the islet-acinar portal system, but the actual hormone levels have never been demonstrated. Rat pancreata were perfused anterogradely or retrogradely with ¹²⁵I-insulin, -somatostatin, or -glucagon (each, 10^–11 mol/l). Tracer binding was determined from differences between influx and efflux radioactivity. Saturable binding was observed for insulin and somatostatin, but not for glucagon. Binding in the absence of unlabelled peptides was significantly higher during retrograde perfusion than during anterograde perfusion for insulin (25.9±2.6 vs 16.0±2.1%, mean±SD; each, n=4; p<0.001) and somatostatin (18.4±2.0 vs 13.6±1.2%; each, n=3; p<0.05). Non-specific binding was similar in both directions. These findings are attributable to endogenous hormones acting as unlabelled ligands competing with the tracers during anterograde perfusion. This conclusion was supported by the demonstration that endogenous insulin stimulation by d-glucose, but not by l-glucose, caused a decrease in labelled insulin binding only during anterograde perfusion. Displacement curves obtained during retrograde perfusion showed that interstitial concentrations of insulin and somatostatin were 7.5×10^–9 and 1.1×10^–9 mol/l, respectively. Thus, the exocrine pancreas is indeed exposed to locally high concentrations of islet hormones.Abbreviations SS Somatostatin - TCA trichloroacetic acid     

Effects of glycogen stores and non-esterified fatty acid availability on insulin-stimulated glucose metabolism and tissue pyruvate dehydrogenase activity in the rat

Summary The effects of increased tissue glycogen stores on insulin sensitivity, and on the response of insulin-stimulated glucose utilisation to an acute elevation in plasma fatty acid levels (1.5mmol/l), were investigated in conscious rats using the hyperinsulinaemic euglycaemic clamp. Studies were performed in two groups of rats; (a) fasted 24 h; (b) fasted 4.5 h, but infused with glucose for 4 h (0.5 g/h) of this period before the clamp (fed, glucose infused rats). Clamp glucose requirement and 3-³H-glucose turnover were 20–25% lower in the fed, glucose-infused rats. In these rats, elevation of plasma fatty acid levels resulted in impaired suppression of hepatic glucose output (residual hepatic glucose output: 41±4 vs 8±6 mol·min^–1·kg^–1. p < 0.001) but did not further decrease 3-³H-glucose turnover. Elevated nonesterified fatty acid levels had no significant effect on glucose kinetics in 24 h fasted rats. In the fed glucose-infused rats, at low plasma fatty acid levels, there was no deposition of glycogen in muscle during the clamp and liver glycogen levels fell. With elevation of non-esterified fatty acid levels muscle glycogen deposition was stimulated in both groups, and there was no fall in liver glycogen during the clamps in the fed glucose-infused rats. Increased non-esterified fatty acid availability during the clamps decreased pyruvate dehydrogenase activity in liver, heart, adipose tissue and quadriceps muscle, in both groups of rats. The findings are consistent with an inhibition of glycolysis in liver, skeletal muscle and heart by increased fatty acid availability. Increased glycogen synthesis, however, compensates for decreased glycolytic flux so that glucose turnover is not decreased. When liver glycogen stores are high, an acute increase in non-esterified fatty acid availability impairs suppression of hepatic glucose output. A chronic increase in non-esteriefid fatty acid availability may lead to insulin resistance by increasing glycogen stores.