This proposal constitutes a further expansion of ongoing ultracytochemical studies of endothelial cells (ECs) of micro-blood vessels (MBVs) of the blood-brain barrier (BBB) in normal and pathological brain. The goal of the present proposal is to characterize some physico-chemical properties of the luminal and abluminal plasma membrane (PM) of the EC using various markers detectable with the electron microscope. Anionic sites will be localized by cationic probes (cationic ferritin, alcian blue) and characterized by digestion with specific enzymes (neuraminidase, chondroitinase, pronase etc.); glycoconjugates will be detected by ferritin or peroxidase labelled lectins specifically recognizing such residues like Betra-D-galactosyl, Alpha-L-fucosyl, Beta-N-glucosaminyl and sialyl, etc.; plasmalemma-bound enzymes (alkaline phosphatase, adenylate cyclase) will be detected by ultracytochemical methods. These experiments will be performed on normal animals (mice and rats), on animals with damaged BBB (mechanical trauma, cold lesion injury, injection of cationic substance) and on scrapie-infected mice. The main attention will be focused on the participation of the PM in enhanced pinocytotic activity associated with luminal and abluminal (reverse) transport across the affected vessels. An infusion edema model will be used for studying the reverse and bidirectional transport across MBVs in rat brain. This model has special importance in understanding the role played by the microvasculature in the resolution phase of brain edema. For comparison, some MBVs, where a BBB does not exist (area postrema, choroid plexus, skeletal muscles) will be examined. It is expected, that the examination of the above mentioned features of the plasmalemma of ECs in various experimental and pathological conditions can help understand the role of cellular constituents of the endothelium in maintaining the function of the BBB. The use of morphological methods can also help to examine cellular elements located in close proximity to abluminal side of ECs (smooth muscle cells, pericytes, astrocytic perivascular andfeet) and their eventual influence on transport activity of the endothelium.
