Myo-inositol and D-glucose transport in rat glomerular and cultured mesangial cells

Abstract

Myo-inositol (MI) is the precursor of membrane-bound phosphoinositides important for transmembrane signaling. This study examines whether freshly isolated whole glomeruli can be used to characterize intact cell transport kinetics for MI and D-glucose. Transport properties of cultured mesangial cells are compared. Glomeruli greater than 95% tubule free were isolated from Sprague-Dawley rat kidney cortex by means of selective sieving. Scanning electron microscopy revealed substantial damage of glomerular epithelial cells, whereas endothelial and mesangial cells remained intact. Specific [3H]MI uptake (7-180 min, 37 degrees C) was observed in presence of 5.5 mM D-glucose when L-[14C]glucose was used as a marker of nonspecific uptake. With ouabain (3 mM), or when Na was replaced with N-methyl-D-glucamine, choline, or Li, specific MI uptake was reduced by 95%. A single high-affinity, Na-dependent MI transport site on glomerular cells with Km of 16.5 +/- 1.4 (SE) microM and Vmax of 947 +/- 56 (SE) fmol.mg protein-1.min-1 was observed for 0.75-100 microM MI. D-glucose competitively inhibited MI transport. Specific D-[3H]-glucose transport was Na independent. Phlorizin inhibition of D-glucose and MI uptake was in keeping with Na-independent D-glucose transport. Km and Vmax for MI uptake in cultured mesangial cells were 42.7 +/- 7.1 microM and 1,474 +/- 192 fmol.mg protein-1.min-1, respectively. We conclude that freshly isolated glomeruli can be used to study cellular transport function that may be modified in disease states.