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PubMed1984

Excretion and metabolism of dopa and dopamine by isolated perfused rat kidney.

The renal metabolism and handling of dopa and dopamine were studied with the use of the isolated perfused rat kidney to elucidate the source of urinary dopamine. Dopa added to the perfusate was rapidly metabolized, and a significant amount of urinary dopamine excretion was observed, whereas dopa was not detected in the urine. When the perfusate dopa concentration was near the plasma dopa concentration in normal rats, urinary dopamine excretion was comparable with physiological values. Carbidopa, a dopa decarboxylase blocker, added to the perfusate markedly reduced the urinary excretion of dopamine. When the physiological concentration of dopamine was added to the medium, the perfusate dopamine rapidly decreased and urinary dopamine excretion was much less than that seen in normal rats. In studies with nonfiltering kidneys, a marked decrease of dopa and slight increase of dopamine in the perfusate were observed. It is suggested that the process of glomerular filtration and tubular reuptake of dopa is not essential for renal dopamine formation. These data indicate that urinary free dopamine is mainly derived from plasma dopa, which is converted by dopa decarboxylase in the kidney.

Animals↗
PubMed1986

Effect of Bt2cGMP on action of cholecystokinin in isolated perfused rat pancreas.

We have examined the effects of Bt2cGMP on cholecystokinin stimulation of pancreatic exocrine secretion in the isolated perfused rat pancreas. Bt2cGMP produced a concentration-dependent inhibition of the stimulatory effects of cholecystokinin octapeptide (CCK-8, 100 pM) on both pancreatic juice flow and enzyme secretion. Adding 1 mM Bt2cGMP rapidly and completely abolished CCK-8-stimulated pancreatic juice and enzyme secretion. Adding 500 microM Bt2cGMP for 10 min at the termination of 1 nM of CCK-8 infusion caused an immediate and persistent inhibition of the residual response. In contrast, treatment with C-terminal CCK antiserum I had no influence on the residual response. To account for the ability of Bt2cGMP to function as a competitive antagonist of the action of CCK and the ability of the nucleotide to inhibit the residual stimulation caused by CCK, we feel that Bt2cGMP hindered the binding of CCK not only by reducing the association rate constant for hormone binding but also accelerating the dissociation rate.

Amylases↗
PubMed1993

Protective effects of histidine during ischemia-reperfusion in isolated perfused rat hearts.

We investigated the efficacy of histidine in reducing ischemia-reperfusion (I/R)-induced myocardial injury in isolated perfused rat hearts. In I/R hearts, the contractile function and coronary flow were 59 +/- 10 and 78 +/- 6% of control. Perfusion with histidine resulted in significant increase in contractility (94 +/- 4%) and coronary flow (92 +/- 4%). The incidence of arrhythmias during reperfusion was 100% (10 out of 10) in the I/R hearts with an average duration of 12.22 +/- 1.55 (SE) min. The duration of arrhythmias was shortened to 8.24 +/- 1.46, 2.15 +/- 0.9, and 2.49 +/- 1.50 min with 10, 25, and 50 mM histidine, respectively. The duration of sinus rhythm increased from 6.26 +/- 1.56 min in I/R hearts to 10.66 +/- 1.55, 14.99 +/- 1.61, and 17.18 +/- 0.95, and 11.73 +/- 0.93 min after perfusion with 10, 25, and 50 mM histidine, and superoxide dismutase (SOD)-catalase-mannitol, respectively. Electron microscopy revealed significant ultrastructural damage of myocytes in I/R hearts, which included swelling of the mitochondria and disruption of both the sarcolemma and the myofibrils. Histidine reduced the ultrastructural damage in a dose-dependent fashion. In general, the protective effect of histidine was superior than SOD-catalase-mannitol. We conclude that histidine protects myocardium against I/R damage most likely by a singlet oxygen scavenging mechanism.

Animals↗
PubMed1996

Differential changes in SP-A and disaturated phospholipids in the isolated perfused rat lung and in vivo.

Alveolar disaturated phospholipids (DSPA) increase in vivo in rats with hyperpnea and in isolated perfused lungs (IPL) in response to either salbutamol or increasing tidal volume (VT). Because surfactant protein-A (SP-A) may play a role in surfactant homeostasis, we have examined the relationship between SP-A and DSP in the alveolus lamellar bodies (LB-A), and in a vesicular (LB-B) lung subfraction. Whereas 2 h swimming increased total DSPA (approximately 48%), it had no effect on alveolar SP-A (SP-AA). In the IPL, salbutamol increased total DSPA (approximately 30%) and SP-AA (approximately 41%); increasing VT (2.5-fold) only increased DSPA (approximately 22%). SP-A and DSP also varied differentially in the tubular myelin-rich and -poor subfractions. In both the IPL and in vivo, we found inverse relationships between DSPA and SP-AA/DSPA, indicating that although SP-AA and DSPA are related, they vary independently. Whereas total SP-AA/DSPA varied between 0.046 and 0.074, it remained constant in LB-A (approximately 0.015) and LB-B (approximately 0.010), suggesting that DSP and SP-A are secreted differentially and that only a small portion of SP-AA is derived from lamellar bodies.

Animals↗
PubMed1984

Effects of low [Ca2+] and La3+ on PAH transport by isolated perfused renal tubules.

Effects of low medium calcium concentration and of lanthanum on p-aminohippurate (PAH) transport by isolated perfused snake (Thamnophis spp.) distal-proximal renal tubules were studied. Nominal removal of calcium from bath produced rapid, irreversible disruption of cells and depression of net PAH secretion (JPAH). However, nominal removal of calcium from lumen with calcium in bath reversibly depressed JPAH and apparent PAH permeability of luminal membrane without altering net fluid absorption (Jv) or intracellular PAH concentration [( PAH]cell). In the presence of calcium (1.8 mM), lanthanum (2 mM) in perfusate alone, in bath alone, or in both perfusate and bath reversibly depressed JPAH, apparent PAH permeabilities of luminal and peritubular membranes, and initial rate of PAH transport into cells without altering [PAH]cell or Jv. Variations in calcium concentration had little effect on depression of JPAH in sodium-free bath. These data suggest that calcium entry into cells, but possibly not cytosolic calcium concentration alone, is important for PAH transport from bath into cells and from cells into lumen and for normal passive permeability of peritubular membrane to PAH and that increased cytosolic calcium concentration is not primarily responsible for depression of JPAH in sodium-free medium.

Aminohippuric Acids↗
PubMed1987

Effect of acute renal failure on insulin disposition in the isolated perfused rat kidney.

The renal disposition of insulin in acute renal failure has not been evaluated. We used the isolated perfused rat kidney to test the hypothesis that acute renal failure (ARF) decreases renal insulin clearance. We used warm ischemia for 45 min, uranyl nitrate 5 mg/kg, ureter ligation, and nonfiltering kidneys as methods of inducing ARF. Comparisons were made with normal control kidneys. The concentrations of insulin in perfusate and urine was determined by radioimmunoassay. Acute renal failure caused significant reductions in glomerular filtration rate, sodium and potassium reabsorption, and an increased urine pH. Warm ischemia and uranyl nitrate toxicity caused a 50% decrease in the renal clearance of insulin. Nonfiltering kidneys cleared insulin at a rate 90% decreased from controls. Ureteral ligation caused a 32% decrease in insulin clearance. Filtration was necessary for insulin to be cleared from perfusate. We conclude that ARF decreased renal insulin clearance through a decrease in insulin uptake from both the tubular lumen and peritubular surface.

Acute Kidney Injury↗
PubMed1991

Effect of inhibitors of oxidative phosphorylation on erythropoietin mRNA in isolated perfused rat kidneys.

We have used RNase protection to measure oxygen-dependent changes in erythropoietin (EPO) mRNA in isolated perfused kidneys and to compare the effect of hypoxia with the response to inhibitors of oxidative phosphorylation. In well-oxygenated kidneys perfused for 2 h at 12 ml/min, with hematocrit of 0.09 +/- 0.005 and PO2 of 443 +/- 67 mmHg, EPO mRNA levels were similar to the baseline levels measured in nonperfused contralateral kidneys from the same animals. When perfusions were performed under identical conditions but at a PO2 of 32 +/- 4 mmHg, EPO mRNA increased approximately 16-fold. In contrast, graded concentrations of cyanide (10, 100, and 300 microM and 1 mM), antimycin (0.01, 0.1, 0.5, and 1 microM), and oligomycin (0.01, 0.1, and 1 microM) did not alter EPO mRNA in well-oxygenated perfused kidneys. However, in kidneys perfused at low PO2 with a high concentration of each inhibitor, EPO mRNA levels were increased, demonstrating that the ability to respond to hypoxia was retained. Thus inhibitors of oxidative phosphorylation did not mimic the effects of hypoxia, indicating that oxygen-dependent expression of the EPO gene in the kidney is not effected through hypoxic compromise of oxidative phosphorylation.

Animals↗
PubMed1987

Increased clearance rate of prednisone in the isolated perfused liver of uremic rats.

The effect of uremia upon the hepatic clearance rate of prednisone was studied in isolated perfused livers of normal and uremic female rats. The perfusion rate was kept at a constant flow rate of 25 ml/min. In order to induce uremia, one group of rats was 5/6 nephrectomized 3 weeks prior to the perfusions. The concentrations of prednisone and other steroids were measured by a normal phase HPLC technique. In experiments with an initial prednisone concentration of 500-700 micrograms/l, the hepatic clearance rate of prednisone was significantly (p less than 0.001) increased by 40% in livers obtained from uremic rats (11.1 +/- 0.49 ml/min/liver) as compared to control rats (8.0 +/- 1.17 ml/min/liver). No significant correlation was found between the hepatic clearance rate of prednisone and increasing initial prednisone concentrations from 141 to 1,953 micrograms/l in the perfusate. It is concluded that an adaptive mechanism may exist in uremia which results in an increase of the hepatic clearance rate of prednisone.

Animals↗
PubMed1976

Localization of glucocorticoid uptake in normal and ischemic myocardial tissue of isolated perfused cat hearts.

We studied the uptake of labeled dexamethasone (3H-Dex) or methylprednisolone (3H-MP) in isolated perfused cat hearts during the first hour of acute myocardial ischemia. Considerable amounts of 3H-Dex and 3H-MP were taken up by the plasma membrane (F1) fraction in control, border zone, and ischemic myocardial tissue. Lesser amounts were incorporated into the remaining cell fractions. A gradient of glucocorticoid uptake was observed that decreased from control tissue to ischemic tissue in all subcellular fractions (i.e., F1 to F5). Accordingly, supernatant fraction (S) to particulate (P) ratios of labeled glucocorticoid uptake increased from control to ischemic tissue, indicating that myocardial cell damage resulted in a decrease in glucocorticoid-binding capacity in subcellular fractions obtained from ischemic tissue. The activity of 5'-nucleotidase (5'ND), a plasma membrane marker in myocardial cells, also decreased from normal to ischemic tissue. Furthermore, we found that uptake of 3H-MP and 3H-Dex was associated with the retention of 5'ND activity in F1 fractions of both border zone and ischemic tissue. Similar protection of plasma membrane integritg occurred in the supernatant fraction as determined by changes in S/P ratios of 5'ND activity. These data provide support for the concepts that (1) plasma membrane changes occur soon after acute myocardial ischemia, and (2) the mechanism by which glucocorticoids exert a protective effect in myocardial ischemia may be related to membrane stabilization.

Animals↗
PubMed1979

Relationship between adenine nucleotide metabolism and irreversible ischemic tissue damage in isolated perfused rat heart.

The relationship between energy metabolism and the extent of irreversible ischemic damage was examined in an isolated perfused working rat heart. The amount of cardiac work recovered after reperfusion of hearts exposed to severe global ischemia was dependent upon both the duration of ischemia and the type of substrate provided (either 5 mM glucose or 5 mM glucose + acetate). There appear to be two distinct phases in the ability to recover mechanical function in the reperfused ischemic heart. The second phase corresponds to the onset of severe irreversible tissue damage. Irreversible mitochondrial damage was not found to correspond with the onset of heart failure since the ATP/ADP ratio remained constant in the reperfused myocardium. Furthermore, there does not appear to be a direct correlation between the total ATP content and the extent of irreversible damage, either during ischemia or following reperfusion. However, the total adenine nucleotide content during ischemia showed dramatic changes which correspond temporally with the initiation of the second phase of damage. The observation that the adenine nucleotide pool becomes further depleted during reperfusion suggests that alterations in the salvage pathway for adenine nucleotide synthesis have occurred. Loss of adenine nucleotides appears to be an excellent marker for irreversible heart failure. Acetate provides some protection the the ischemic myocardium. The mechanism by which acetate mediates this protective effect is discussed.

Adenine Nucleotides↗
PubMed1977

Inhibition of sodium transport by prostaglandin E2 across the isolated, perfused rabbit collecting tubule.

This study was designed to examine whether prostaglandin E2 can directly affect sodium transport across isolated perfused rabbit renal collecting tubules. Changes in transepithelial potential and isotopic sodium fluxes in response to peritubular prostaglandin E2 were measured. In addition, changes in transepithelial potential of the outer medullary collecting tubule in response to prostaglandin E2 were also measured. With few exceptions, all rabbits received 5 mg/day desoxycorticosterone acetate for 4-11 days before experimentation. The results of the experiments show that: (a) prostaglandin E2 inhibits the negative transepithelial potential in the cortical collecting tubule as well as the outer medullary collecting tubule; (b) prostaglandin E2 inhibits net sodium transport out of the lumen by inhibiting efflux while backflux is unaffected; (c) prostaglandin E2 produces this inhibition within 15 min, and the effects are dose dependent and reversible. These results suggest that prostaglandin E2 may modulate sodium transport in vivo and may contribute to the final regulation of sodium excretion.

Animals↗
PubMed1980

Effect of corticosterone on insulin and glucagon secretion by the isolated perfused rat pancreas.

The in vitro effect of corticosterone in insulin and glucagon secretion has been examined in the isolated perfused rat pancreas preparation. COrticosterone at physiological concentrations was found to inhibit acutely and strongly the secretion of insulin induced by both glucose and arginine and to potentiate the output of glucagon in a glucose-free medium or when induced by arginine. Phentolamine, an alpha-adrenergic blocking agent, diminished the strong inhibitory effect of corticosterone on insulin secretion. The present results demonstrate that corticosterone has direct effects on pancreatic islet cells, and they suggest that the inhibition of insulin secretion is to some extent related to the alpha-adrenergic receptors.

Animals↗
PubMed1981

Adrenergic modulation of insulin and glucagon secretion from the isolated perfused rat pancreas.

In order to observe the effect of the adrenergic system on pancreatic glucagon secretion in the isolated perfused rat pancreas, phenylephrine, an alpha-adrenergic agonist, and isoproterenol, a beta-adrenergic agonist, were added to the perfused solution. 1.2 microM phenylephrine suppressed glucagon secretion at 2.8 mM glucose, and it also decreased insulin secretion at 11.1 mM glucose. 240 nM isoproterenol enhanced glucagon secretion not only at 2.8 mM glucose, but also at 11.1 mM glucose, as well as insulin secretion at 11.1 mM. In order to study the role of intra-islet noradrenalin, phentolamine, an alpha-adrenergic antagonist, and propranolol, a beta-adrenergic antagonist, were infused with the perfused solution. 10 and 100 microM phentolamine caused an increase in insulin secretion, and 25 microM propranolol decreased insulin secretion, while they did not cause any change in glucagon secretion. From these results, it can be concluded that alpha-stimulation suppresses not only insulin but also glucagon secretion, while beta-stimulation stimulates glucagon secretion, as well as insulin secretion. Intra-islet catecholamine may have some effect on the B cell, whereas it seems to have no influence on the A cell.

Animals↗
PubMed1977

Effect of Verapamil on pancreatic glucagon release from the isolated, perfused canine pancreas.

The effect of Verapamil, a potent calcium antagonist, upon pancreatic glucagon release was investigated in the isolated, perfused canine pancreas. Verapamil at concentrations ranging between 10(-5)-10(-4) mol/l caused a dose-related inhibition of glucagon release at a glucose concentration of 25 mg/dl. The inhibition was an immediate and reversible phenomenon. The inhibitory effect was reduced when the Ca2+ concentration was increased from 1.3 to 5.0 mmol/l. The results are compatible with earlier findings from our laboratory demonstrating that calcium plays a key role in the stimulus-secretion coupling of glucagon release.

Animals↗
PubMed1995

Comparison of the disposition of two novel combined thromboxane synthase inhibitors/thromboxane A2 receptor antagonists in the isolated perfused rat liver.

1. The pharmacokinetics of two combined thromboxane synthase inhibitor/thromboxane A2 receptor antagonists were investigated in an isolated perfused rat liver preparation. 2. The clearance of both UK 101613 (15.4 +/- 1.9 ml/min) and UK 102333 (14.9 +/- 1.6 ml/min) was limited by hepatic bloodflow. Both hepatic uptake and biliary excretion of UK 101613 appeared to be active processes. The apparent biliary clearance of UK 101613 (10.3 +/- 2.3 ml/min) was less than perfusate clearance, indicating accumulation within the hepatocytes. 3. The efficiency of the active hepatic uptake of UK 101613 and UK 102333 explains the rapid removal of these compounds from the systemic circulation. The processes of biliary and/or metabolic clearance of UK 101613 and UK 102333 appear to be subsequent to and independent of perfusate clearance. 4. Although the compounds are of similar structure and have a similar hepatic extraction (> 0.9), UK 101613 had an apparent biliary clearance approximately twice that of UK 102333. This is due to the formation of a metabolite of UK 102333 which reduces the amount of parent compound available for biliary excretion. 5. Inhibition of this latter metabolism by ketoconazole (10 microM) resulted in the two compounds having comparable apparent biliary clearances.

Animals↗
PubMedDate not supplied

Metabolism of [14C]monocrotaline by isolated perfused rat liver.

The metabolism of the pyrrolizidine alkaloid [14C]monocrotaline [( 14C]MCT) was examined using the in situ isolated perfused rat liver. Hepatic tissue was perfused in a recirculatory fashion for 90 min and the distribution of metabolites between the bile and perfusate was analyzed. Monocrotalic acid (MCA) was found to be the major acidic metabolite of [14C]MCT, with trace amounts of 1-formyl-7-hydroxy-6,7-dihydro-5H-pyrrolizine, 6,7-dihydro-7-hydroxy-1-hydroxymethyl-5H-pyrrolizine (DHP), and 1-hydroxymethyl-7-oxo-6,7-dihydro-5H-pyrrolizine (tentative assignment) being identified in the perfusates using GC/MS. MCT N-oxide was also identified but represented less than 4% of the perfusate 14C. The simple necine base retronecine was not present at detectable levels in the perfusion medium. A large portion of the 14C recovered from both the bile and perfusate was not extractable, under acidic or basic conditions, into organic solvents. Using fast atom bombardment MS/MS, a portion of this material was identified as a glutathione conjugate of DHP. In addition, this nonextractable material retained a portion of the radioactivity that was equivalent to the acidic fraction. Given these findings and the absence of retronecine, the major pathway for the metabolism of MCT could potentially involve the production of MCT pyrrole, which subsequently reacts with cellular nucleophiles producing MCA in addition to highly water-soluble conjugated pyrroles and possibly macromolecular adducts.

Animals↗
PubMed1989

Differentiation of various cardiovascular drugs by means of specific myocardial and vascular load tests. Experiments at the isolated perfused heart.

A new method is described for examining the contractile function of myocardium and coronary vasculature in the isolated perfused, electrically stimulated guinea-pig and rat heart which is suitable for investigation of cardiovascular drugs. The criterion used for functional efficiency of myocardium and coronary vasculature is--in addition to the usual evaluation of cardiac force and coronary flow in steady-state--the reaction of the heart to perfusion pressure changes: abrupt increase of the perfusion pressure (PP) leads to an increase in cardiac force (= heterometric autoregulation, HA) and simultaneously to triggering of the myogenic autoregulation (MA). Using this PP loading two different functional tests can be performed: 1. By successive PP increases in 10-20 mmHg steps it is possible to determine a) an equivalent of Frank-Starling cardiac function curves, and b) pressure-flow (p-F) curves of the coronary vasculature, e.g. before and after drug application. 2. By a single PP step of e.g. 40 mmHg HA and MA are triggered before and after drug application, calculated as percentages of the initial value and plotted as concentration-response curves. Both methods produce a physiologically based description of cardiac and coronary mechanics. In particular, the behaviour of the coronary vasculature in this procedure represents a more sensitive criterion than total coronary flow in steady-state. Since HA and MA depend partly (HA) or completely (MA) on the slow transmembrane Ca++ inward current the method is particularly suitable for a basic differentiation of Ca++ and Na+ antagonistic drugs.

Animals↗
PubMed1979

The fate of intratracheally instilled 3H-styrene oxide in the isolated perfused rat lungs.

The metabolism and covalent binding of intratracheally instilled 7-3H-styrene oxide in the isolated perfused lungs of control and cigarette smoke-exposed rats were investigated. After intratracheal instillation of 100 nmoles of 7-3H-styrene oxide the lungs were perfused for 16 min and the nonrecirculating perfusate was collected in two fractions: 0--8 min and 8--16 min. About 40 percent of the instilled dose was in the 0--8 min perfusion medium, mainly in the form of unmetabolized styrene oxide. However, the radioactivity in the 8--16 min perfusion medium and lung tissue itself was mainly in the form of water soluble metabolites, probably glutathione conjugates. The amount of styrene glycol was very small. Exposure of rat to cigarette smoke had no effect on the metabolism of styrene oxide or on the amount of covalent binding to the protein and nucleic acid fractions of the perfused lungs.

Animals↗