Improved in situ rat liver perfusion technique

Collins, Fred G.; Skibba, Joseph L.

doi:10.1016/0022-4804(80)90084-0

Cited by 22 publications

(6 citation statements)

References 16 publications

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“…Liver anoxia was minimized (5-1Os) by commencing perfusion immediately, at a constant flow rate of l5ml-min-1, with 150ml of Krebs-Ringer bicarbonate buffer, pH7.4 (Krebs & Henseleit, 1932); this buffer also contained 2mM-CaCl2, 5 mM-glucose, 1% (w/v) bovine serum albumin, a physiological amino acid mixture (see Barnwell et al, 1983a) and 40% (v/v) of packed human red blood cells. This solution was recycled, gassed continuously with 02/CO2 (19:1) and maintained at 37 + 0.5°C within a thermostatically controlled cabinet similar to that recommended by Collins & Skibba (1980).…”

Section: Methodsmentioning

confidence: 99%

Rapid kinetic analysis of the bile-salt-dependent secretion of phospholipid, cholesterol and a plasma-membrane enzyme into bile

Lowe¹,

Barnwell²,

Coleman³

1984

Biochemical Journal

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Isolated rat livers were perfused under 'one-pass' conditions and bile was collected at 1 min intervals. After 1 min pulse, taurocholate appeared in the collected bile within 2 min, peak output occurring 2 min later. In contrast, the increased output of phospholipids and cholesterol was slower, peak output occurring 6-11 min after the original pulse of taurocholate. These results suggest that mixed micelles cannot be formed inside the cell or during passage of bile salts through the membrane, since bile salt and lipids should then parallel each other. The bile salts must therefore be pumped into the lumen and the lipids added subsequently, due to the actions of the bile salts in the canalicular lumen. It is suggested that the biliary lipid is obtained from microdomains of biliary-type lipid in the canaliculus membrane, which are vesiculated and solubilized by the action of bile salts. It is also suggested that this biliary-type lipid is brought continuously to the membrane via vesicle traffic; this traffic is increased during increased bile-salt output, and is a process that can be inhibited by colchicine.

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Section: Methodsmentioning

confidence: 99%

Rapid kinetic analysis of the bile-salt-dependent secretion of phospholipid, cholesterol and a plasma-membrane enzyme into bile

Lowe¹,

Barnwell²,

Coleman³

1984

Biochemical Journal

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“…This solution was gassed continuously with 02/C02 (19: 1) and maintained at 37 + 0.50C (Brauer et al, 1954). All perfusions were carried out in a thermostatically controlled cabinet similar to that recommended by Collins & Skibba (1980) for perfusions in situ.…”

Section: Methodsmentioning

confidence: 99%

Biliary protein output by isolated perfused rat livers. Effects of bile salts

Barnwell¹,

Godfrey²,

Lowe³

et al. 1983

Biochemical Journal

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The output of proteins into bile was studied by using isolated perfused rat livers. Replacement of rat blood with defined perfusion media deprived the liver of rat serum proteins (albumin, immunoglobulin A) and resulted in a rapid decline in the amounts of these proteins in bile. When bovine serum albumin was incorporated into the perfusion medium it appeared in bile within 20 min and the amount in the bile was determined by the concentration of the protein in the perfusion medium. The use of a defined perfusion medium also deprived the livers of bile salts and the amounts of these, and of plasma-membrane enzymes [5'-nucleotidase (EC 3.1.3.5) and phosphodiesterase I], in bile declined rapidly. Introduction of micelle-forming bile salts (taurocholate or glycodeoxycholate) to the perfusion medium 80 min after liver isolation markedly increased the output of plasma-membrane enzymes but had no effect on the other proteins. The magnitude of this response was dependent on the bile salt used and its concentration in bile; there was little effect on plasma-membrane enzyme output until the critical micellar concentration of the bile salt had been exceeded in the bile. A bile salt analogue, taurodehydrocholate, which does not form micelles, did not produce the enhanced output of plasma-membrane enzymes. This work supports the view that the output of plasma-membrane enzymes in bile is a consequence of bile salt output and also provides evidence for mechanisms by which serum proteins enter the bile.

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“…The rat liver may be isolated and perfused in situ or ex situ (i.e., attached to or removed from the animal carcass). In the in situ preparation, it may be difficult to maintain the carcass at a constant temperature, although a method for maintaining physiologic temperatures has recently been described (3). In contrast, ex situ perfusion in a temperature-controlled cabinet (2) allows for easy temperature control.…”

Section: Experimental Techniquementioning

confidence: 99%

The isolated perfused rat liver: Conceptual and practical considerations

Gores¹,

Kost²,

LaRusso³

1986

Hepatology

262

137

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PERSPECTIVESAlthough it has been over 100 years since Claude Bernard first reported the use of the isolated perfused rat liver (IPRL) (l), this model is still a valuable and commonly used tool for exploring the physiology and pathophysiology of the liver. Indeed, the IPRL remains an important experimental model despite the availability of newer techniques (e.g., liver slices, isolated and cultured cells, and isolated organelles) for evaluating hepatic function. This popularity is due to the fact that, in contrast to in uiuo models, such as the bile fistula rat, the IPRL allows repeated sampling of the perfusate, permits easy exposure of the liver to different concentrations of test substances and is amenable to alterations in temperature that would not be tolerated in uiuo. Furthermore, experiments can be done independent of the influence of other organ systems, plasma constituents and neural-hormonal effects. In contrast to other in uitro models, such as isoiated and cultured hepatocytes and isolated organelles, hepatic architecture, cell polarity and bile flow are preserved in the IPRL.Given these considerations, it seems timely to update and, in a selective manner, to review the IPRL, especially as it relates to the study of hepatic function, the advantages and disadvantages of different technical aspects of the model and the problem of reliably assessing organ viability. This manuscript is intended not only to help the reader establish this model, but also to allow him to choose those conditions which best fit his particular needs. In addition, it will provide sufficient conceptual information to permit the reader to critically evaluate work with the IPRL. However, the manuscript is not meant to be a comprehensive technical review, as these are readily available (2). Rather, it is a highly selective update focusing on concepts, practical considerations and applications. EXPERIMENTAL TECHNIQUEAlthough this review concerns itself with the IPRL, other animals such as the monkey, hamster, guinea pig, cat, rabbit, dog, sheep, calf and pig have all been used as liver donors. However, the rat is most commonly used because it is of convenient size and is relatively inexpensive to purchase and maintain; also, a large data base exists from studies using rat livers for comparison of observations.The rat liver may be isolated and perfused in situ or ex situ (i.e., attached to or removed from the animal carcass). In the in situ preparation, it may be difficult to maintain the carcass at a constant temperature, although a method for maintaining physiologic temperatures has recently been described (3). In contrast, ex situ perfusion in a temperature-controlled cabinet (2) allows for easy temperature control. The basic perfusion apparatus has three parts in series: a reservoir, a pump and an oxygenator. Peristaltic or pulsating perfusion pumps are generally used. While the peristaltic pump works well, the pulsating pump minimizes hemolysis in red blood cell-containing perfusates.Two types of oxygenators have been described ...

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Improved in situ rat liver perfusion technique

Cited by 22 publications

References 16 publications

Rapid kinetic analysis of the bile-salt-dependent secretion of phospholipid, cholesterol and a plasma-membrane enzyme into bile

Rapid kinetic analysis of the bile-salt-dependent secretion of phospholipid, cholesterol and a plasma-membrane enzyme into bile

Biliary protein output by isolated perfused rat livers. Effects of bile salts

The isolated perfused rat liver: Conceptual and practical considerations

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