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PubMed1982

Effect of nafenopin on the uptake of bilirubin and sulfobromophthalein by isolated perfused rat liver.

Hepatic uptake of bilirubin and sulfobromophthalein has kinetic characteristics suggesting facilitated diffusion. Because these compounds demonstrate mutual competition for uptake, a shared uptake mechanism has been presumed. Previous studies in isolated perfused regenerating liver revealed depressed uptake of bilirubin, sulfobromophthalein, and asialoorosomucoid, a desialylated glycoprotein which enters hepatocytes by receptor-mediated endocytosis. This study was designed to determine whether or not depressed transport seen in liver regeneration occurs in other states of hepatocellular proliferation. Rats were pretreated with nafenopin (200 mg/kg . day x 2), a drug that causes rapid hepatocellular proliferation similar to that seen in regeneration. Twenty-four hours after nafenopin treatment, liver weight increased by 40%. Influx, efflux, and sequestration rate constants in isolated perfused liver were quantitated by a computer fit to the model of Goresky. Results 1 day after nafenopin treatment revealed no change in transport parameters for bilirubin and asialoorosomucoid, but 55% and 49% reductions in influx of sulfobromophthalein and conjugated bilirubin, respectively. These studies suggest that hepatocellular proliferation alone is not responsible for the transport alterations seen during liver regeneration. Nafenopin effectively unmasks differences in uptake of bilirubin and other more water soluble organic anions such as sulfobromophthalein and conjugated bilirubin, suggesting that their uptake mechanisms are partially independent.

Animals↗
PubMed1996

Influence of trifluoperazine and verapamil on the isolated perfused rat kidney.

1. Isolated rat filtering kidneys were perfused with Ringer bicarbonate solutions containing either trifluoperazine (TFP, 50 microM) or verapamil (VER, 100 microM) to prevent tissue dysfunctions observed during perfusion. 2. Water, sodium and chloride kidney contents diminished under both treatments as compared with control preparations, and potassium content increased. 3. When albumin concentration was increased (10 g%) in the perfusion medium (nonfiltering kidney preparation) these effects of TFP or VER were not observed. 4. Lipid peroxidation and LDH release diminished significantly under 50 microM TFP but only slightly under 100 microM VER.

Animals↗
PubMed1970

On the mode of action of ACTH on the isolated perfused adrenal gland.

1. Isolated cat adrenal glands were perfused with Locke solution, and the corticosteroid outputs in response to adrenocorticotrophin (ACTH) were studied.2. Steroid outputs varied with the ACTH concentration, as well as with the duration of exposure to a given ACTH concentration.3. Omission of calcium from the perfusion medium markedly depressed ACTH-evoked steroid release. The steroid output was directly related to the extracellular calcium concentration up to 0.5 mM.4. During a constant exposure to ACTH, steroid output was maintained for at least 2-3 hr, provided that calcium was present in the perfusion medium.5. Strontium, but not barium or magnesium, replaced calcium in maintaining the secretory response to ACTH.6. Magnesium depressed ACTH-evoked secretion in the presence of calcium, and this depression of secretion was antagonized by increasing the calcium concentration.7. Prolonged perfusion with sodium-free or potassium-free solutions did not markedly inhibit steroid output in response to ACTH. Excess potassium (56 mM) did not produce a consistent or marked increase in spontaneous steroid output and did not affect the response to ACTH.8. The steroid content of adrenal glands perfused with Locke solution and exposed to ACTH was about 10% of the amount which was secreted. By contrast, adrenal glands perfused with calcium-free media and exposed to ACTH contained much higher amounts of steroid, despite the negligible amount secreted.9. These data suggest that calcium plays a critical role in the mechanism of corticosteroid secretion from the adrenal cortex.

Adrenal Cortex Hormones↗
PubMed1975

Comparison of alpha- and beta-cell secretory responses in islets isolated with collagenase and in the isolated perfused pancreas of rats.

The inhibitory actions of somatostatin (100 ng./ml.) on insulin release, stimulated by high glucose (20 mM), and on glucagon release, stimulated by arginine (15 mM), were studied with two in vitro systems: the isolated perifused rat islets prepared by the collagenase procedure and the isolated perfused rat pancreas. Suppression of arginine-induced glucagon release by glucose (20 mM) and glyceraldehyde (5 mM) was also assessed in both systems. With the perfused pancreas, somatostatin caused 32 per cent inhibition of glucose-mediated insulin release and inhibited arginine-induced glucagon release by 72 per cent. In the same system, glucose and glyceraldehyde were similarly potent inhibitors of arginine-induced glucagon secretion. In contrast to the isolated perfused pancreas, there was no significant somatostation suppression of glucose-induced insulin release or arginine-induced glucagon release whether the inhibitor was present prior to or was added during stimulation by glucose or arginine. Furthermore, glucose was only minimally active and glyceraldehyde ineffective in inhibiting glucagon secretion due to arginine in the perifusion system. The most plausible explanation for the difference in the endocrine response of islet cells in the two types of widely used in vitro systems is that the alpha and beta cells have lost inhibitory receptors in the plasma membrane as a result of the collagenase isolation technic.

Animals↗
PubMed1986

Impaired parathyroid hormone-stimulated adenosine 3',5'-monophosphate release by isolated perfused bones obtained from vitamin D-deficient rats.

The present studies were designed to explore the mechanism underlying skeletal refractoriness to PTH in a vitamin D-deficient animal by assessment of PTH-stimulated cAMP release from isolated perfused bone. In vitamin D-deficient (-D) rats both basal and PTH-stimulated cAMP release were markedly diminished, compared with that in vitamin D-replete (+D) rats. Isolated perfused bones from -D rats that had undergone parathyroidectomy 2 days before death still showed reduced cAMP release in response to PTH, compared with +D bones. To investigate which factors in terms of Ca, endogenous PTH, or vitamin D might primarily be responsible for the impaired PTH-stimulated cAMP release from -D bones, some -D rats were switched to a diet identical to the vitamin D-deficient diet but with high Ca content (4%) for 2 or 5 weeks before death. This schedule maintained normocalcemia despite vitamin D deficiency. PTH-stimulated cAMP release in these rats was increased to a level intermediate between that in -D rats and +D rats, indicating partial restoration of the impaired response to PTH in -D rats. These data indicate that skeletal refractoriness to PTH in vitamin D-deficient animals might, in part, be due to the impaired activation of adenylate cyclase, which cannot be explained entirely by hypocalcemia or associated secondary hyperparathyroidism. Vitamin D deficiency per se, therefore, may play a key role in the impaired cAMP response to PTH.

Animals↗
PubMed1987

Hyperthermic pelvic isolation-perfusion in the treatment of refractory pelvic cancer.

Hyperthermic isolation-perfusion (I-P) was used to treat 27 patients with refractory pelvic cancer. All patients except one achieved pelvic isolation as manifested by selective pelvic heating and by pharmacologic monitoring. Patient response was good, with rapid pain relief in 75% and tumor control as detected by physical examination, computed tomographic scan, and decline in carcinoembryonic antigen levels. Pelvic drug exposure averaged 7.8 times that of systemic drug exposure. Of the 20 patients with recurrent rectal adenocarcinoma, one complete response (duration, eight months), seven partial responses (average duration, greater than or equal to 10 months), four patients with stable disease (average duration, greater than or equal to 12 months), and five with disease progression were observed. Three patients could not be evaluated due to late deaths as a consequence of their disease. There were two postoperative deaths in the remaining seven patients, one due to drug toxicity and one due to probable cardiac arrhythmia. Pelvic I-P has evolved with the avoidance of laparotomy and increased drug dose. We conclude that hyperthermic I-P for pelvic cancer is a safe, effective procedure and an excellent therapeutic option for patients with persistent pelvic cancer.

Adenocarcinoma↗
PubMed1995

Lack of luminal or basolateral uptake and transepithelial transport of mercury in isolated perfused proximal tubules exposed to mercury-metallothionein.

The lumen-to-bath and bath-to-lumen transport, cellular uptake, and toxicity of inorganic mercury bound to metallothionein (203Hg-MT) were studied in isolated perfused S1, S2, and S3 segments of the renal proximal tubule of rabbits. Evidence of very mild toxicity was displayed in some of the segments perfused through the lumen with 18.4 microM inorganic mercury in the form of Hg-MT. The toxic response was restricted primarily to mild swelling of the epithelial cells localized at the end of the tubular segments where the perfusion pipette was inserted into the lumen. The cells in the proximal portions of perfused S2 segments appeared to be most severely affected in that a few blebs would on occasion come off the epithelial cells. Mild cellular swelling was also observed in some S2 and S3 segments that were exposed to 18.4 microM inorganic mercury in the form of Hg-MT in the bath. The swelling was more generalized, involving all the epithelial cells along the perfused segment. Very little, or no, measurable lumen-to-bath or bath-to-lumen transport of Hg as Hg-MT could be detected in any of the 3 perfused segments of the proximal tubule during 40-45 min of perfusion. The complex of Hg-MT appeared to behave in a manner similar to that of the volume marker [3H]-L-glucose. The lack of tubular transport of Hg as Hg-MT was confirmed by little or no measurable uptake and accumulation of inorganic mercury in the tubular epithelial cells. Thus, our findings indicate that the Hg-MT complex is not taken up avidly in isolated perfused S1, S2, or S3 segments of the proximal tubule.

Animals↗
PubMed1997

Isoflurane inhibits endothelium-mediated nitric oxide relaxing pathways in the isolated perfused rabbit lung.

PURPOSE: The role of volatile anaesthetics on nitric oxide (NO)-dependent relaxation is unclear in the pulmonary circulation. We examined the effects of isoflurane on NO-dependent relaxation in isolated perfused rabbit lungs. METHODS: Eighteen rabbit lungs were perfused in a constant-flow recirculation manner. In study 1 (n = 12), acetylcholine (ACh, 4 x 10(-10)-10(-8) M) or nitroglycerine (NTG, 6 x 10(-10)-10(-8) M) was cumulatively injected into the pulmonary artery in the absence or presence of isoflurane (1, 2 MAC). In study 2 (n = 6), ACh was injected as in study 1 in the presence or absence of N omega-nitro-L-arginine methyl ester (L-NAME, 100 microM), an NO synthesis blocker. In all experiments, indomethacin was administered to prevent formation of vasoactive prostanoid metabolites, and the pulmonary vessels were preconstricted with prostaglandin F2 alpha (PGF2 alpha) infused before ACh or NTG injection. The ACh- or NTG-induced relaxation was expressed as % decrease in PGF2 alpha preconstriction. RESULTS: Isoflurane at 2 MAC attenuated the dose-dependent relaxation to ACh at doses of 4 x 10(-9) M and 4 x 10(-8) M from 27.8 +/- 4.3% and 38.8 +/- 5.3% to 17.0 +/- 3.5% and 25.5 +/- 4.9%, respectively (P < 0.05). Isoflurane did not change the dose-dependent relaxation to NTG and L-NAME abolished the ACh-induced relaxation. CONCLUSION: Isoflurane inhibited NO-dependent relaxation in the pulmonary circulation at a site distal to the endothelial cell receptor-mediated responses but proximal to guanylate cyclase activation of vascular smooth muscle. Acetylcholine-induced relaxation in isolated perfused rabbit lungs was regulated primarily by NO.

Acetylcholine↗
PubMed1983

Influence of taurocholate on hepatic clearance and biliary excretion of asialo intestinal alkaline phosphatase in the rat in vivo and in isolated perfused rat liver.

We have used canine intestinal alkaline phosphatase, an asialoglycoprotein, as a tool to study the effect of taurocholate on hepatic asialoglycoprotein transport. After bolus injections of 20 U of alkaline phosphatase into anesthetized rats in vivo and isolated perfused rat livers, a biexponential disappearance pattern was observed; 6.2% of the dose and 0.8% of the dose was excreted in bile, respectively. Taurocholate infusion in vivo and in isolated perfused livers resulted in a virtually complete abolition of the plasma disappearance of alkaline phosphatase in the second slow phase of the curve, whereas the primary disappearance rate was not affected. Wash out experiments showed release of exogenous alkaline phosphatase from the liver, which was strongly increased by taurocholate. Liver perfusion experiments with two subsequent doses of alkaline phosphatase indicated that the taurocholate effect was completely reversible. It is concluded that taurocholate enhances exocytosis of interiorized alkaline phosphatase from liver into plasma. The taurocholate effect on hepatic exocytosis of the asialoglycoprotein may also occur when bile salt concentrations in the systemic or portal circulation, or both, are elevated due to certain liver diseases and postprandial bile salt absorption.

Alkaline Phosphatase↗
PubMed1993

Effect of toremifene on antipyrine elimination in the isolated perfused rat liver.

Toremifene is a triphenylethylene antioestrogen with significant antitumor activity. It is structurally very similar to tamoxifen. Both drugs undergo extensive hepatic metabolism, and tamoxifen is known to inhibit hepatic mixed-function oxidases (MFO). Using the isolated perfused rat-liver model, we investigated the effect of toremifene on the elimination of antipyrine, a standard marker of MFO activity. Perfusate consisted of 20% red cells in a modified Krebs-Henseleit buffer, and 80 ml was recirculated at 14 ml/min for 3 h. High but clinically relevant steady-state toremifene levels of 3 and 10 micrograms/ml were achieved using bolus plus constant infusion into the reservoir. Elimination of 2.5 mg antipyrine was not inhibited by steady-state toremifene, but methanol (maximal perfusate concentration, 1.29%), the vehicle used for toremifene administration, caused a statistically significant increase in the antipyrine elimination half-life (mean, 1.4 +/- 0.2 h for controls vs 2.2 +/- 0.3 h for methanol; P < 0.05, n = 4). Whereas the methanol had no apparent effect on liver viability as assessed by bile flow and perfusate back-pressure, toremifene at a steady-state concentration of 10 micrograms/ml caused a statistically significant decrease in bile flow (value at 180 min, 0.22 +/- 0.05 ml/h as compared with 0.52 +/- 0.06 ml/h in the methanol control; P < 0.05) and a statistically significant increase in perfusate back-pressure (value at 180 min, 17.5 +/- 1.8 cm vs 11.0 +/- 2.6 cm in the methanol control; P < 0.05). Therefore, toremifene used at high doses can impair liver function in the isolated perfused rat liver, but it does not have any effect on antipyrine elimination.

Animals↗
PubMed1993

A maintained afterload model of ischemia in erythrocyte-perfused isolated working hearts.

The aim of this study was to develop an erythrocyte-perfused isolated working heart model of global ischemia that would allow controllable reductions in coronary perfusion pressure but with maintained afterload to facilitate the temporal association of myocardial ischemia with ventricular contractile or metabolic dysfunction. Isolated working rat hearts were perfused at 37 degrees C with a washed, resuspended porcine erythrocyte perfusate (40% hematocrit) at a preload of 10 mmHg and afterload of 75 mmHg, paced at 300 beats per min. A bifurcation above the aorta permitted imposition of two one-way valves, one opened by ventricular ejection in systole and closed by aortic afterload in diastole, the other closed by aortic pressure from ventricular ejection but opened to allow coronary perfusion in diastole according to an adjustable pressure head (75 mmHg initially). Ischemia was imposed by lowering the coronary perfusion pressure to 35 mmHg while maintaining diastolic afterload at 75 mmHg. The presence of erythrocytes provides oxygen delivery in the normal physiological range. This model permits the severity of ischemia to be graded by any specified level. The maintained afterload model of ischemia is a simple enhancement of the isolated working heart permitting a separation of workload and coronary perfusion pressure variables. This provides greater control over and direct monitoring of experimental ischemic events in progress.

Animals↗
PubMed1999

Differential uptake of FDG and DG during post-ischaemic reperfusion in the isolated, perfused rat heart.

Fluorine-18 2-fluoro-2-deoxyglucose (FDG) and 2-deoxyglucose (DG) are widely used as tracers of glucose uptake in the myocardium. Although there is agreement that the two analogues behave similarly to glucose under control conditions, there is growing evidence that some interventions (e.g. insulin stimulation or ischaemia/reperfusion) cause differential changes in their behaviour. The addition of a two-surface coil nuclear magnetic resonance (NMR) probe and a dual-perfusion cannula to our recently developed PET and NMR dual-acquisition (PANDA) system allows us to collect PET (FDG) images and phosphorus-31 NMR (2-deoxyglucose-6-phosphate) spectra simultaneously from each independently perfused coronary bed of the heart. We have used this technique to study the effect of regional ischaemia/reperfusion on FDG and DG uptake in the isolated, perfused rat heart. During control perfusion, FDG uptake was almost identical in both coronary beds. When one coronary bed was made ischaemic, FDG uptake ceased on that side but continued on the control side. Reperfusion failed to restore FDG uptake. In contrast, NMR spectra showed that, during reperfusion, the uptake and phosphorylation of DG did not differ between the two coronary beds. The results thus demonstrate that regional myocardial ischaemia/reperfusion has different effects on the uptake of FDG and DG in the isolated, perfused rat heart.

Animals↗
PubMed1983

Experimental hyperthermic isolation-perfusion using cis-diamminedichloroplatinum(II).

The intent of this study was to determine whether or not local control and/or cure of rabbits bearing the VX-2 carcinoma could be achieved with regional hyperthermia and chemotherapy. A model of isolation-perfusion used cis-diamminedichloroplatinum(II) (DDP). Five different experimental groups were studied, each group receiving progressively smaller tumor inocula and shorter treatment intervals. Although local control and cure improved as the tumor inocula became smaller and treatment interval became shorter, there was no benefit demonstrated when compared to sham-operated animals. DDP appeared to be well tolerated in animals perfused as normothermic temperatures. However, animals perfused at hyperthermic temperature with DDP experienced necrosis of normal as well as neoplastic tissue, resulting in early demise of the animal. This final observation suggests that the amount of DDP utilized in clinical isolation-perfusion should be approached with caution.

Animals↗
PubMedDate not supplied

The effect of heart rate on the termination of electrically induced ventricular fibrillation in the isolated perfused rat heart.

Ventricular fibrillation (VF) which is normally sustained in large animals and humans, is transient in small animals. The purpose of the present study was to evaluate the possible effect of changing cardiac rate on spontaneous ventricular defibrillation. In isolated perfused rat heart, VF was electrically induced during normal spontaneous rhythm of the heart at normal rate and at various ventricular pacing rates. It was found that: 1) Electrically induced VF in isolated perfused, non-ischemic rat heart spontaneously terminated in 88% of the hearts; 2) Ventricular pacing rhythm of spontaneous rate plus 10% caused VF to be sustained in 26% of the hearts (which defibrillated spontaneously during normal rates); 3) Ventricular pacing at 200% of the basic rate led to sustained VF in about half the VF episodes (14 out of 33, p less than 0.005). In the remainder, which defibrillated spontaneously, a sustained VF could be achieved by further increase in ventricular pacing rate; 4) Slow pacing rate, as a result of the surgical production of atrioventricular (A-V) block, enhanced the probability of spontaneous defibrillation (21 of 21 episodes after slow pacing vs 24 of 34 episodes following pacing at previous normal sinus rhythm, p less than 0.05). Selective modulation of conduction velocity, refractory period or both, achieved by changes in ventricular pacing rate was assumed to play an important role in determining whether electrically-induced VF would be transient or sustained.

Animals↗
PubMed1993

New experimental technique to study blood cardioplegia in the isolated, perfused rat heart.

Blood cardioplegia has been extensively studied clinically and in the large animal experimental model. We describe here a modification of the original Langendorff technique to study continuous warm blood cardioplegia in the isolated, perfused rat heart. The excised heart is mounted on the perfusion apparatus and perfused with Krebs-Henseleit buffer. Prearrest hemodynamics are recorded. The shed blood in the mediastinal cavity (8 to 12 mL) is collected, filtered, and reconstituted into cardioplegic solution (hematocrit, 0.20; K+, 15 mmol/L). Hearts are arrested and maintained at 37 degrees C by continuous recirculation of blood cardioplegia for 1 hour. The blood cardioplegia system consists of a Silastic tubing oxygenator, peristaltic pump, and two filters (40 microns pore size). The heart is reperfused with Krebs-Henseleit solution, and postarrest hemodynamics are recorded. Percentage recovery of peak left ventricular pressure, heart rate, and coronary flow were 98.5 +/- 3.1, 102 +/- 4.2, and 98.5 +/- 4.5 (mean +/- standard error of the mean; n = 6), respectively. Myocardial oxygen consumption during arrest was 57 microL.min-1.g-1 dry wt, which is 10% of the myocardial oxygen consumption of a beating heart in in-vivo and ex-vivo models. These results suggest the feasibility of studying blood cardioplegia in the isolated, perfused rat heart model under controlled conditions. Continuous warm blood cardioplegic arrest provided excellent myocardial protection for 1 hour in this model.

Animals↗
PubMed1978

A compartmental model for hepatic transport of taurocholic acid in isolated perfused rat liver.

In order to characterize the transport of bile acids through the liver and to study the influence of drugs on these processes, a kinetic model for hepatobiliary transport of taurocholic acid (TC) using the isolated perfused liver was developed. After the system was brought to a steady state by infusing TC at a constant rate, a tracer dose of 14C-TC was injected into the medium. The medium disappearance of 14C-TC followed a first-order kinetic with a single rate constant. The plot of the biliary secretion rate of radioactivity versus time revealed a curve composed of at least three exponential components. From the described results and the present knowledge of hepatobiliary transport of bile acids we proposed a three compartment model, composed of a perfusion medium compartment and two liver compartments. Parameters calculated from the model constants agreed well with model-independent estimations. The influence of bromosulfophthalein (BSP) on the kinetic parameters was studied to compare the result with the known effect of BSP on hepatic transport of taurocholic acid. BSP decreased the constant describing the fractional transfer of taurocholic acid from medium into the liver, which is in agreement with the inhibition of hepatic uptake of taurocholic acid by BSP. Thus a three compartment model may adequately define the hepatobiliary transport of taurocholic acid in the isolated perfused rat liver.

Animals↗
PubMed1986

Effect of 24-hour preservation with oxygen free radical scavengers on isolated-perfused canine heart-lungs.

The effectiveness of 24-hour hypothermic machine perfusion with TP-V (a hyperosmolar colloid solution containing dextrose, sucrose and ATP-MgCl2) alone, or in combination with oxygen free radical scavengers, was evaluated in isolated-perfused canine heart-lungs. Heart-lungs were perfused at 4 degrees C in either TP-V (n = 6), TP-V/Allopurinol (500 mg/L) (n = 6), or TP-V/Allopurinol (500 mg/L) & Catalase (5000 U/L) (n = 5). Lung inflation was maintained with 100% nitrogen. Following preservation, the heart-lungs were perfused with an albumin-mannitol perfusate for 3 hours at 37 degrees C, for functional, hemodynamic, and laboratory determinations. Cold preservation with TP-V/Allopurinol, and TP-V/Allopurinol & Catalase resulted in physiologically normal LDH levels during the 3-hour normothermic isolated perfusion test period. Significantly lower enzyme activity for CPK was evident at 0 (p less than .005) and 3 hours (p less than .05) of perfusion, while no significant differences in lactate production were seen among the groups. In addition, pH, PCO2, PO2, and left ventricular, aortic, and coronary artery pressures all remained within normal physiologic range, with no significant differences seen among the three groups. 99m Technetium scans demonstrated adequate patency among the heart-lungs, with better flow seen in those perfused with TP-V/Allopurinol & Catalase. Histological specimens confirmed a decrease in myocardial and pulmonary damage when Allopurinol and/or Catalase was used. It appears that oxygen free radical scavengers provide some protection from canine heart-lungs which have been hypothermically preserved for 24 hours.

Allopurinol↗
PubMed1991

Reverse cholesterol transport in the rat. Studies using the isolated perfused spleen in conjunction with the perfused liver.

1. A new method combining the use of an isolated perfused extrahepatic tissue with a perfused liver was developed as a model system for the study of reverse cholesterol transport. Rat spleens, initially labelled in vivo with [3H]cholesterol, were perfused for 3 h with whole blood. The spleen was then replaced with an isolated rat liver, whose uptake of cholesterol from the spleen-derived blood and excretion of cholesterol into bile constituents were determined. 2. During spleen perfusion, a net release of cholesterol mass and radioactivity to lipoproteins was observed. 3. During liver perfusion, there was also a rapid exchange or transport of unesterified cholesterol between high-density lipoprotein (HDL) and the liver, in particular with HDL2 (d = 1.085-1.125). 4. The liver showed an increased uptake of cholesteryl ester from serum that had previously been used in spleen perfusion. 5. Approximately half of the [3H]cholesterol released by the spleen was recovered in erythrocytes. During subsequent liver perfusion there was a substantial uptake of radioactivity from the erythrocytes, although less than that recorded from serum lipoproteins. 6. In all experiments there was significant excretion of [3H]cholesterol into bile; most (85%) was in bile acids. Thus the complete process of reverse cholesterol transport is observed in this dual-perfusion system.

Animals↗