CELLULAR MECHANISMS OF GLUCOSE-INDUCED MYOINOSITOL TRANSPORT UP-REGULATION IN RAT MESANGIAL CELLS

Uptake of myo-inositol (MI) is necessary to maintain normal cellular phosphoinositide signaling and function. MI transport is upregulated in the cells of diabetic rat glomeruli compared with normal rat glomeruli [C.I. Whiteside, J.C. Thompson, and J. Ohayon. Am. J. Physiol. 260 (Renal Fluid Electrolyte Physiol. 29): F138-F144, 1991]. To identify mechanisms associated with upregulation of MI transport, rat mesangial cells were cultured in high (25.6 mM) vs. normal (5.6 mM) glucose. Specific Na+-dependent [H-3]MI uptake (> 97%), using L-[C-14]glucose as the nonspecific marker, was linear for 120 min in high and normal glucose. In high glucose, compared with normal glucose, there was no change in Michaelis-Menten constant values [29.1 +/- 0.6 vs. 30.3 +/- 0.7 mu M (SE)], whereas maximum velocity (V,,) was increased (2,024 +/- 52 vs. 1,132 +/- 115 fmol.mg protein(-1).min(-1), P < 0.001). Mannitol (20.0 mM), used as an osmotic control, had no effect on the upregulation of MI transport. Maximum upregulation of MI transport measured by V-max (control taken as 100%) was observed after 8 h of exposure to high glucose (222 +/- 6% above control, P < 0.0001) or galactose (20.0 mM) (194 +/- 6%, P < 0.0001) and was sustained for up to 48 h. The protein synthesis inhibitors cycloheximide (20 mu g/ml) or actinomycin D (5 mu g/ml), the F-actin depolymerizing agent cytochalasin D (2 mu g/ml), and the aldose reductase inhibitor Tolrestat (0.3 mM) independently prevented glucose- or galactose-induced upregulation of MI transport. Tolrestat inhibited excess sorbitol accumulation in mesangial cells exposed to high glucose. In the cells of glomeruli isolated from normal rats treated with and without Tolrestat (25 mg.kg body wt(-1).day(-1)), MI transport was upregulated after exposure in vitro to high glucose (25.6 mM) or galactose (20.0 mM) only in the absence of Tolrestat. These data imply that, in rat mesangial cells, high glucose stimulates upregulation of MI transport, which requires new protein synthesis and intact actin microfilaments. The mechanism appears linked to activation of the polyol pathway.