Action of insulin on the subcellular metabolism of polyphosphoinositides in isolated rat hepatocytes

Journal Article · Tue Aug 05 04:00:00 EDT 1986 · J. Biol. Chem.; (United States)

OSTI ID:5021234

Sakai, M; Wells, W W

The effect of insulin on /sup 32/Pi incorporation into phospholipids in various subcellular sites of isolated rat hepatocytes was investigated. After labeling the phospholipids of hepatocytes from rats previously starved for 24 h with /sup 32/Pi (10 mu Ci/10(6) cells) for 90 min, either saline or insulin (32 nM) was added. Following incubations of 1, 5, and 30 min, chilled cells were rapidly washed, homogenized in the presence of inhibitors of phospholipid degradation, and fractionated into the major subcellular organelles. Phospholipids were extracted from plasma membranes, microsomes, lysosomes, mitochondria, and nuclei with acidic chloroform:methanol. The aqueous deacylation products were separated by anion exchange high performance liquid chromatography, and the /sup 32/ incorporated into all the major diacylglycerophospholipids was determined. In parallel experiments, the specific radioactivity of /sup 2/Pi and (gamma-/sup 32/P)ATP was determined. The results revealed that insulin had no effect on the turnover of the major phospholipids, including the polyphosphoinositides, of all subcellular compartments analyzed relative to the control. In addition, there were no significant differences in the amount and /sup 32/P labeling of cellular orthophosphate between saline- and insulin-treated cells. The specific radioactivity of (gamma-32P)ATP was increased by 20% after 30-min treatment with insulin, requiring appropriate correction of /sup 32/P-labeled phosphatidic acid, phosphatidylinositol 4-phosphate, and phosphatidylinositol 4,5-bisphosphate for estimation of mass changes at near steady-state labeling of cellular ATP.

• Research Organization: Michigan State Univ., East Lansing
• OSTI ID: 5021234
• Journal Information: J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 22; ISSN JBCHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BODY
CELL CONSTITUENTS
DAYS LIVING RADIOISOTOPES
DIGESTIVE SYSTEM
ESTERS
GLANDS
HORMONES
INSULIN
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
LIPIDS
LIVER
LIVER CELLS
LYSOSOMES
MAMMALS
METABOLISM
NUCLEI
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANOIDS
ORGANS
OXYGEN COMPOUNDS
PEPTIDE HORMONES
PHOSPHATES
PHOSPHOLIPIDS
PHOSPHORUS 32
PHOSPHORUS COMPOUNDS
PHOSPHORUS ISOTOPES
RADIOISOTOPES
RATS
REACTION KINETICS
RODENTS
SOMATIC CELLS
TRACER TECHNIQUES
UPTAKE
VERTEBRATES