Gluconeogenesis in hepatocytes isolated from the skipjack tuna (Katsuwonus pelamis)

Abstract

Skipjack tuna (Katsuwonus pelamis) hepatocytes were isolated and determined to be viable based on trypan blue exclusion, energy charge, and lactate dehydrogenase leakage. Trypan blue staining and red blood cell contamination were always less than 0.1%, and lactate dehydrogenase leakage remained less than 1% at 25 °C for 10 h. The energy charge for freshly isolated cells ranged from 0.65 to 0.74. Alanine (10 mM) gave higher rates of gluconeogenesis and oxidation than lactate: alanine rates were 3.17 ± 0.65 μmol glucose/(g packed cells∙h) and 9.1 ± 1.19 μmol CO2/(g∙h), whereas lactate rates were 0.47 ± 0.08 and 1.8 ± 0.27, respectively. Phosphoenolpyruvate carboxykinase was 30% cytosolic and 70% mitochondrial, a finding that is in agreement with the liver's ability to use both lactate and alanine as gluconeogenic substrates. If hepatic gluconeogenesis is the major route of lactate removal postexercise, it is estimated that recovery would require 3800 h! Therefore, tuna liver is not the major site of glucose resynthesis and lactate clearance following burst exercise.