sorbinil and Neuroblastoma

Astroglia-rich rat primary cultures can be grown in a glucose-free medium containing 25 mM sorbitol. After 10 days under these conditions, the total number of cells and DNA content are reduced to 50-60% of those of control cultures, but remain constant thereafter. The specific activities of the sorbitol pathway enzymes, sorbitol dehydrogenase and aldose reductase, are increased 2.5-fold and unchanged, respectively, if the cells are grown in the presence of sorbitol instead of glucose. Treatment with the aldose reductase inhibitor sorbinil does not decrease the number of cells cultured in the glucose-free medium in the presence of sorbitol. Fructose is as good a substrate for the glial cells as sorbitol, whereas out of a number of other polyols tested only xylitol can support the primary cultures for more than 3 days. Neither neuron-rich rat brain primary cultures nor rat glioma cells can be cultured in the sorbitol-containing medium in the absence of glucose. With sorbitol substituting for glucose in the culture medium, effects of glucose deprivation on cellular functions like sugar transport and metabolism can be investigated in glial cultures for an extended period of time.

Topics: Aldehyde Reductase; Animals; Astrocytes; Cell Division; Cells, Cultured; Culture Media; DNA Replication; Energy Metabolism; Fructose; Glioma; Imidazoles; Imidazolidines; L-Iditol 2-Dehydrogenase; Neuroblastoma; Neurons; Rats; Rats, Inbred Strains; Sorbitol; Tumor Cells, Cultured; Xylitol

Neuroblastoma cells were used to analyze the effect of galactose supplementation on myo-inositol metabolism, polyol accumulation, and Na+-K+ pump activity. Culturing cells in 30 mM galactose for a minimum of 1 wk led to a large accumulation of intracellular galactitol and a greater than 50% decrease in myo-inositol content. The effect of galactose on the intracellular content of galactitol and myo-inositol was concentration dependent. Extracellular myo-inositol accumulation and incorporation into phospholipid decreased by 20-30% in cells grown in 30 mM galactose. The decrease in myo-inositol accumulation is apparently due to a noncompetitive inhibition of high-affinity myo-inositol uptake. Treatment of the galactose-containing media with 0.4 mM sorbinil partially prevented the galactose-mediated decreases in myo-inositol metabolism and content. The galactitol content of the sorbinil-treated cells was significantly reduced compared with the galactitol levels in cells cultured in 30 mM galactose; however, galactitol levels remained significantly elevated over control cells. Exposing neuroblastoma cells to 30 mM galactose causes a decrease in the levels of phosphatidylinositol that is partially restored by the addition of sorbinil. The activity of the Na+-K+ pump was decreased by 20% in cells cultured in 30 mM galactose and was partially protected by sorbinil treatment. The effects of long-term galactose supplementation on myo-inositol metabolism, polyol accumulation, and Na+-K+-ATPase transport activity in cultured neuroblastoma cells are similar to the effects of high concentrations of glucose. These results provide additional evidence that the accumulation of polyol by neuroblastoma cells is partially responsible for alterations in myo-inositol metabolism and decreases in Na+-K+-ATPase transport activity.

Topics: Aldehyde Reductase; Animals; Cell Line; Galactose; Glucose; Imidazoles; Imidazolidines; Inositol; Mice; Neuroblastoma; Phosphatidylinositols; Polymers; Potassium Channels; Sodium Channels; Sodium-Potassium-Exchanging ATPase; Tumor Cells, Cultured

Neuroblastoma cells were used to analyze the effect of elevated glucose levels on myo-inositol metabolism and Na+/K+-pump activity. The activity of the Na+/K+ pump in neuroblastoma cells is almost totally sensitive to ouabain inhibition. Culturing neuroblastoma cells in 30 mM glucose caused a significant decrease in Na+/K+-pump activity, myo-inositol metabolism, and myo-inositol content, compared to cells grown in the presence of 30 mM fructose. Glucose supplementation also caused a large intracellular accumulation of sorbitol. The aldose reductase inhibitor sorbinil prevented the abnormalities in myo-inositol metabolism and partially restored Na+/K+-pump activity in neuroblastoma cells cultured in the presence of elevated glucose levels. These results suggest that the accumulation of sorbitol by neuroblastoma cells exposed to elevated concentrations of extracellular glucose causes a decrease in myo-inositol metabolism and these abnormalities are associated with a reduction in Na+/K+-pump activity.

Topics: Aldehyde Reductase; Animals; Cell Line; Glucose; Imidazoles; Imidazolidines; Inositol; Mice; Neuroblastoma; Rubidium; Sodium-Potassium-Exchanging ATPase; Tumor Cells, Cultured

The uptake of myo-inositol was investigated in femoral nerve fascicular preparations taken from control and streptozotocin-diabetic rats and in a clonal murine neuro-blastoma cell line (N1E-115), as a model of the neuronal component of the nerve preparation. Uptake was investigated in medium containing glucose, 5.6-25 mmol/l and inositol, 4 x 10(-5) mol/l. In the presence of glucose (25 mmol/l) myo-inositol uptake was decreased in nerve taken from streptozotocin-diabetic animals when compared to control (26.4 +/- 2.2 pmol/100 micrograms protein/2 h vs 55.1 +/- 2.4 pmol/100 micrograms protein/2 h, p less than 0.005). Uptake in both preparations was higher in the presence of insulin added during the uptake experiment (73.4 +/- 5.7 pmol/100 micrograms protein/2 h and 64.4 +/- 3.9 pmol/100 micrograms/2 h, respectively). Prior treatment of the animals with insulin or with the aldose reductase inhibitor, sorbinil also resulted in an increase in myo-inositol uptake in streptozotocin diabetic nerve preparations. In control nerve preparations and in N1E-115 cells raising the glucose concentration from 5.6 through 25 mmol/l was associated with decreased myo-inositol uptake, with an inhibitory constant (Ki) of 21.4 mmol/l and 20.4 mmol/l for femoral nerve and N1E-115 cells respectively. An increase in myo-inositol uptake was found in N1E-115 cells, following pre-treatment of cells in culture with sorbinil or inclusion of insulin during the uptake experiment. Altered myoinositol metabolism may play a role in the functional and structural changes characterizing diabetic neuropathy. The effects of hyperglycaemia on myo-inositol uptake in experimental diabetes may be modified by insulin or by inhibition of sorbitol accumulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Aldehyde Reductase; Animals; Cell Line; Diabetes Mellitus, Experimental; Femoral Nerve; Glucose; Imidazoles; Imidazolidines; In Vitro Techniques; Inositol; Insulin; Kinetics; Male; Neuroblastoma; Phloretin; Rats; Rats, Inbred Strains; Reference Values

Neuroblastoma cells were used to determine the effect of sorbinil on myo-inositol metabolism in cells exposed to elevated levels of glucose in culture. Exposing cells to elevated levels of glucose led to an increase in levels of intracellular sorbitol. The increase in sorbitol levels was dependent on the extracellular glucose concentration. In contrast, the myo-inositol content of cells was decreased in the presence of increasing concentrations of extracellular glucose. Increasing the concentration of glucose in the culture medium caused a decrease in myo-inositol uptake and in the incorporation of extracellular myo-inositol into phospholipid. The effect of elevated glucose levels on myo-inositol metabolism and sorbitol accumulation was blocked by addition of 0.4 mM sorbinil. The ability of sorbinil to block the decrease in myo-inositol metabolism and sorbitol accumulation caused by 30 mM extracellular glucose was dependent on its concentration. Maximal effects were obtained with 0.4 mM sorbinil. However, there was some variation in the degree of effectiveness among batches of sorbinil. These results at the cellular level suggest that the intracellular accumulation of sorbitol is responsible for the alteration of myo-inositol metabolism observed in neuroblastoma cells exposed to elevated glucose concentrations.

Topics: Animals; Cell Line; Fructose; Glucose; Imidazoles; Imidazolidines; Inositol; Kinetics; Mice; Neuroblastoma; Sorbitol; Tumor Cells, Cultured