QUANTIFICATION OF DIFFERENT TRANSFERRIN RECEPTOR POOLS IN PRIMARY CULTURES OF PORCINE BLOOD-BRAIN-BARRIER ENDOTHELIAL-CELLS

Distribution of iron in the brain varies with region, cell type, and a ge. Furthermore, some neurological diseases are accompanied by an abno rmal accumulation of iron in specific areas of the CNS. These findings implicate a mobile intracerebral iron pool; however, transport of iro n across the blood-brain barrier and its regulation are largely unknow n. In an extensive series of experiments in primary cultures of porcin e blood-brain barrier endothelial cells, we separately quantified surf ace-bound and total cellular transferrin receptor pools. Although 90% of all transferrin receptors were located inside the cell, only 10% of these intracellular receptors actively took part in the endocytic cyc le. This large ''inactive'' intracellular transferrin receptor pool co uld either function as a storage site for spare receptors or be activa ted by the cell to increase its capacity for iron transport. Data were corrected for nonspecific binding by a separate biochemical assessmen t using a 100-fold excess of unlabeled ligand. Data were also analyzed in a nonlinear curve-fit program. This resulted in a less elaborate a nd less biased estimate of nonspecific binding.