Identification and distribution of insulin receptors on cultured bovine brain microvessel endothelial cells: Possible function in insulin processing in the blood–brain barrier

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Abstract
The binding of 125 I-insulin to primary cultures of bovine brain microvessel endothlial cells was examined. Insulin binding was both time and temperature dependent and inhibited by excess unlabeled insulin. Furthermore, the specific binding of insulin was polarized to the apical side of the cell monolayers. Upon binding, the labeled insulin was internalized, with approximately 70% resistant to acid wash over a 90-min period. The inhibition of insulin internalization observed with cell monolayers exposed to either phenylarsine oxide or unlabeled insulin suggests a receptor-mediated endocytic process. Furthermore, the ability of chloroquine to reduce the metabolism of insulin indicates a significant portion of the peptide iis processed through a lysosomal pathway. In contrast to the fluid-phase endocytosis marker, Lucifer yellow, as much as 65% of internalized insulin undergoes apical to basolateral trancytosis in brain microvessel endothelial cells. While most of the effluxed insulin was degraded, as assessed by trichloroacetic acid precipitation, the results of the present study suggest insulin receptors within the brain microvasculature may be involved in the processing and transport of bloodborne insulin.