COUPLED GLUCOSE-TRANSPORT AND METABOLISM IN CULTURED NEURONAL CELLS -DETERMINATION OF THE RATE-LIMITING STEP

In brain and nerves the phosphorylation of glucose, rather than its tr ansport, is generally considered the major rate-limiting step in metab olism. Since little is known regarding the kinetic coupling between th ese processes in neuronal tissues, we investigated the transport and p hosphorylation of [2-H-3]glucose in two neuronal cell models: a stable neuroblastoma cell line (NCB20), and a primary culture of isolated ra t dorsal root ganglia cells. When transport and phosphorylation were m easured in series, phosphorylation was the limiting step, because intr acellular glucose concentrations were the same as those outside of cel ls, and because the apparent K-m for glucose utilization was lower tha n expected for the transport step. However, the apparent K-m was still severalfold higher than the K-m of hexokinase I. When [2-H-3]glucose efflux and phosphorylation were measured from the same intracellular g lucose pool in a parallel assay, rates of glucose efflux were three- t o-fivefold greater than rates of phosphorylation. With the parallel as say, we observed that activation of glucose utilization by the sodium channel blocker veratridine caused a selective increase in glucose pho sphorylation and was without effect on glucose transport. In contrast to results with glucose, both cell types accumulated 2-deoxy-D-[C-14]g lucose to concentrations severalfold greater than extracellular concen trations. We conclude from these studies that glucose utilization in n euronal cells is phosphorylation-limited, and that the coupling betwee n transport and phosphorylation depends on the type of hexose used.