Studies of hormonal regulation of metabolism using isolated hepatocytes.

A number of enzymatic methods have been developed to prepare hepatocytes using collagenase and hyaluronidase. However, best cell preparations are obtained by using only low concentrations of collagenase and exposing the liver to the enzyme for a very short period of time. These isolated cells with intact cell membranes and large numbers of microvilli on the cell surface respond to hormones at physiological concentrations suggesting that these microvilli contain hormone receptors. In addition, high glycogen content is essential to maintain the in vivo metabolic characteristics of the hepatocytes suggesting that intracellular glycogen plays an important role in the hormonal regulation of metabolism in hepatocytes. Studies with glucagon and insulin on carbohydrate metabolism show that the molar ratios of these hormones control gluconeogenesis and glycogenolysis. Furthermore, in vitro addition of insulin stimulates glycogen synthesis and activates glycogen synthase. Insulin also stimulates protein synthesis in cells containing high glycogen and maintains more normal parallel strands of polyribosomes. Studies with isolated hepatocytes from diabetic, hypophysectomized and adrenalectomized animals show a reduced glucagon response to glycogenolysis. This lack of glucagon response was not due to reduction in glycogen levels. Other hormones such as somatostatin and parathyroid also give rise to alterations in carbohydrate metabolism in isolated hepatocytes.