Distribution of anionic sites in glomerular basement membranes: their possible role in filtration and attachment

Abstract

Lysozyme (pI=11) has been used to identify anionic sites in the glomerular capillary wall. A solution of 1-3% lysozyme was perfused into the left kidney at varying rates. After perfusion, the kidney was fixed in situ and processed for electron microscopy. Lysozyme was seen as an electron dense deposit which was not present when succinylated lysozyme (pI=4.5) or myoglobin (pI= 6.9) was perfused instead of native lysozyme. First the epithelial cell plasma membrane was outlined by a 300-400 Å electron dense layer. Second, there were discrete dense deposits in the subepithelial portions (lamina rara externa) of the basement membrane which, in normal section, extended from the epithelial cell membrane to the lamina densa and, in grazing section, formed a continuous reticular pattern. The discrete binding sites in the lamina rara externa were also found in glomeruli that had been prefixed and then perfused with lysozyme and in isolated glomeruli incubated with lysozyme. Third, the lamina densa itself showed a homogeneous increase in density. Fourth, similar discrete dense areas were seen in the subendothelial light layer of the basement membrane, between Bowman's capsule and the parietal epithelium, and between the endothelium of peritubular capillaries and its basement membrane. The experiments show that, in addition to their location on the epithelial surface, anionic sites are present throughout the basement membrane and are distributed in a discrete reticular pattern in the lamina rara externa.