Kinetic evidence for compartmentalization of myo-inositol in hepatocytes.
myo-Inositol uptake and conversion to phosphatidylinositol (PI) was studied in isolated rat hepatocytes. Uptake of myo-[2-3H]-inositol into the trichloroacetic acid (TCA)-soluble fraction showed no evidence of saturation, while incorporation into lipid had an apparent Km of 0.28 mmol/L for external myo-inositol. With 50 mumol/L myo-[2-3H]-inositol, approximately half of the radiolabel was found in lipid at 30 minutes. Glucose and galactose were weak inhibitors, while phlorizin at 1 mmol/L reduced uptake by 50%. Metabolic inhibitors reduced incorporation of myo-[2-3H]-inositol into lipid, but had no effect on uptake. Hepatocytes maintained myo-inositol levels of 0.4 mmol/L for 60 minutes when incubated with 50 mumol/L myo-inositol, but levels increased when incubated with 1 mmol/L myo-inositol. Efflux of label was studied in hepatocytes prelabeled for 20 minutes with myo-[2-3H]-inositol. Loss of label was initially rapid, but had slowed by 20 minutes, with much of the label remaining in the cells. Phlorizin inhibited the loss of myo-[2-3H]-inositol, while increasing myo-inositol concentration in the medium enhanced efflux. The effects of these agents on the rate of efflux was found in lipid rather than in the TCA-soluble myo-inositol fraction. These findings suggest that myo-inositol is compartmentalized within hepatocytes, with a bulk metabolically inert pool and a smaller active pool that equilibrates with extracellular myo-inositol via an energy-independent carrier-mediated mechanism, and is preferentially available for efflux or for synthesis of phosphoinositides.
Sigal
S H
SH
Division of Biochemical Development and Molecular Diseases, Children's Hospital of Philadelphia, PA 19104.
Yandrasitz
J R
JR
Berry
G T
GT
D000818:Animals; D002478:Cells, Cultured; D005690:Galactose / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D007294:Inositol / Q000032:analysis / Q000378:metabolism* / Q000493:pharmacokinetics; D007700:Kinetics; D008099:Liver / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism*; D008297:Male; D010695:Phlorhizin / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D010716:Phosphatidylinositols; D007294:Inositol; D010695:Phlorhizin; D005947:Glucose; D005690:Galactose