Rapid and sensitive measure of gluconeogenesis in isolated bovine hepatocytes

Conference · Tue Mar 04 23:00:00 EST 1986 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:6898431

Azain, M J; Kasser, T R; Atwell, C A; Baile, C A

Available methods for determining glucose synthesis from radiolabelled precursors using ion exchange column chromatography limit the number of samples that can be processed. To facilitate this process, a rapid method for determining glucose synthesis from 3-carbon precursors was developed using suspensions of anion and cation exchange resins. Hepatocytes were prepared from calf liver by collagenase perfusion of the caudate lobe. Isolated cells were incubated with /sup 14/C-labelled lactate or propionate in the presence or absence of glucagen and/or palmitate. Glucose synthesis was determined by vortexing an aliquot of cell suspension with a 50% slurry of anion exchange resin (acetate form), followed by cation exchange resin. After centrifugation /sup 14/C-glucose was recovered in the supernatant and measured by scintillation counting. Using this method, more than 95% of unused labelled precursor was bound to the ion exchange resin and essentially 100% of /sup 14/C-glucose tracer was recovered in the supernatant. In hepatocyte suspensions, radioactivity recovered in the supernatants was confirmed to be glucose by pre-incubating aliquots of media with glucose oxidase prior to addition of ion exchange resins. The present system allows determination of hepatic gluconeogenesis, is sensitive to substrate and hormonal manipulations and has the capacity for processing several hundred samples per day.

• Research Organization: Monsanto Co., St. Louis, MO
• OSTI ID: 6898431
• Report Number(s): CONF-8604222-
• Journal Information: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Vol. 45:4; ISSN FEPRA
• Country of Publication: United States
• Language: English

Related Subjects

550501* -- Metabolism-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ALDEHYDES
ANIMAL CELLS
ANIMALS
CARBOHYDRATES
CARBON 14 COMPOUNDS
CATTLE
CHROMATOGRAPHY
COUNTING TECHNIQUES
DOMESTIC ANIMALS
GLUCOSE
HEXOSES
ION EXCHANGE CHROMATOGRAPHY
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
LIVER CELLS
MAMMALS
METABOLISM
MONOSACCHARIDES
ORGANIC COMPOUNDS
PRECURSOR
RUMINANTS
SACCHARIDES
SCINTILLATION COUNTING
SEPARATION PROCESSES
SOMATIC CELLS
TRACER TECHNIQUES
VERTEBRATES