Control of hepatic nitrogen metabolism and glutathione release by cell volume regulatory mechanisms
Authors: HäUSSINGER Dieter , LANG Florian , BAUERS Kathrin , GEROK Wolfgang 
Published: March 2005

Urea synthesis was studied in isolated perfused rat liver during cell volume regulatory ion fluxes following exposure of the liver to anisotonic perfusion media. Lowering of the osmolarity in influent perfusate from 305 mOsm/l to 225 mOsm/l (by decreasing influent [NaCl] by 40 mmol/l) led to an inhibition of urea synthesis from NH4Cl (0.5 mmol/l) by about 60% and a decrease of hepatic oxygen uptake by 0.43 ± 0.03 μmol g−1 min−1 [from 3.09 ± 0.13 μmol g−1 min−1 to 2.66 ± 0.12 μmol g−1 min−1 (n= 9)]. The effects on urea synthesis and oxygen uptake were observed throughout hypotonic exposure (225 mOsm/l). They persisted although volume regulatory K+ efflux from the liver was complete within 8 min and were fully reversible upon reexposure to normotonic perfusion media (305 mOsm/l). A 42% inhibition of urea synthesis from NH4Cl (0.5 mmol/l) during hypotonicity was also observed when the perfusion medium was supplemented with glucose (5 mmol/l). Urea synthesis was inhibited by only 10–20% in livers from fed rats, and was even stimulated in those from starved rats an amino acid mixture (twice the physiological concentration) plus NH4Cl (0.2 mmol/l) was infused.
The inhibition of urea synthesis from NH4Cl (0.5 mmol/l) during hypotonicity was accompanied by a threefoldincrease of citrulline tissue levels, a 50–70% decrease of the tissue contents of glutamate, aspartate, citrate and malate, whereas 2‐oxoglutarate, ATP and ornithine tissue levels, and the [3H]inulin extracellular space remained almost unaltered. Further, hypotonic exposure stimulated hepatic glutathione (GSH) release with a time course roughly paralleling volume regulatory K+ efflux. NH4Cl stimulated lactate release from the liver during hypotonic but not during normotonic perfusion. In the absence of NH4Cl, hypotonicity did not significantly affect the lactate/pyruvate ratio in effluent perfusate. With NH4Cl (0.5 mmol/l) present, the lactate/pyruvate ratio increased from 4.3 to 8.2 in hypotonicity, whereas simulatneously the 3‐hydroxybutyrate/acetoacetate ratio slightly. but significantly decreased.
Addition of lactate (2.1 mmol/l) and pyruvate (0.3mmol/l) to influent perfusate did not affect urea synthesis in normotonic perfusions, but completely prevented the inhibition of urea synthesis from NH4Cl (0.5 mmol/l) induced by hypotonicity. Restoration of urea production in hypotonic perfusions by addition of lactate and pyruvate was largely abolished in the presence of 2‐cyanocinnamate (0.5 mmol/l). Addition of 3‐hydroxybutyrate (0.5 mmol/l), but not acetoacetate (0.5 mmol/l) largely reversed the hypotonicity‐induced inhibition of urea synthesis from NH4Cl.
The data are consistent with a cell‐swelling‐induced inhibition of the transfer of malate and reducing equivalents into the mitochondrial compartment, thereby inhibiting urea synthesis due to a limitation of aspartate supply for argininosuccinate synthesis. They suggest that alterations of liver cell volume and/or volume regulatory responses exertmarked effects on hepatic ammonia, amino acid, glutathione and carbohydrate metabolism in line with the previously raised hypothesis that liver cell volume changes and/or volume regulatory responses are important modulators of hepatic metabolism.