Hepatic glucose-6-phosphatase flux and glucose phosphorylation, cycling, irreversible disposal, and net balance in vivo in rats. Measurement using the secreted glucuronate technique

Abstract

Measurement of hepatic glucose production (HGP) by standard isotope dilution reveals only the net release of glucose from the liver, not the flux across glucose-6-phosphatase ([G6Pase] or total hepatic glucose output), hepatic glucose cycling (HGC), irreversible glucose disposal into glycogen in the liver (hepatic Rd), or net hepatic glucose balance. We describe two independent isotopic techniques for measuring these parameters in vivo, both of which use secreted glucuronate (GlcUA), HGC can be quantified by measuring a correction factor for glucose label retained in hepatic glucose-6-phosphate (G6P), sampled as GlcUA. A complementary technique for measuring total hepatic glucose output is also described (reverse dilution), requiring administration of no labeled glucose but instead a labeled gluconeogenic precursor and unlabeled glucose. Hepatic Rd is calculated by multiplying the rate of appearance (Ra) of hepatic UDP-glucose ([UDP-glc] based on dilution of labeled galactose in GlcUA) times the direct entry of glucose into hepatic UDP-glc and the fraction of labeled UDP-glc retained in the liver. The sum of hepatic Rd plus HGC represents the total hepatic glucose phosphorylation rate. Rats received intravenous (IV) glucose infusions at a rate of 15 to 30 mg/kg/min after a 24-hour fast. Despite a suppression of net HGP more than 50%, total hepatic glucose output was not significantly decreased, because of increased HGC. Total hepatic glucose output calculated by reverse dilution yielded similar results during IV glucose infusions at 15 mg/kg/min, although values were higher than obtained by the correction-factor method at 30 mg/kg/min. The fraction of labeled UDP-glc released into blood glucose, representing a hepatic glycogen cycle, decreased from 35% (fasted) to nearly 0% (IV glucose 30 mg/kg/min). Hepatic Rd was 1.4, 4.6, and 7.5 mg/kg/min (fasted and IV glucose 15 and 30 mg/kg/min, respectively); total hepatic glucose phosphorylation increased substantially (from 4.2 to 8.5 to 12.7 mg/kg/min) and net hepatic glucose balance changed from negative to positive during IV glucose. In conclusion, hepatic G6Pase flux, glucose phosphorylation, HGC, disposal of glucose into glycogen, and net glucose balance can be measured noninvasively in vivo under various metabolic conditions by techniques involving the GlcUA probe.