Recently, we discovered a non-AT1, non-AT2 angiotensin binding site in rodent and human brain membranes which, based on its pharmacological/biochemical properties and tissue distribution, is different from angiotensin receptors and key proteases processing angiotensins. In this study, the novel angiotensin binding site was localized to a specific brain cell type using radioligand receptor binding assays. Our results indicate that the novel binding site is expressed in mouse primary cortical neuronal membranes but not in primary cortical astroglial and bEnd.3 brain capillary endothelial cell membranes. Whole cell binding assays in neurons showed that the binding site faces the outer side of the plasma membrane. Consistent with our previous observations, the novel binding site was unmasked by sulfhydryl reagent p-chloromercuribenzoate (PCMB). This effect had a bell-shaped curve and was reversed by reduced glutathione, indicating that function of the binding site might be regulated by redox state of the environment. Density of the novel binding site measured by saturation binding assays was significantly increased in neuronal membranes of cells challenged in four in vitro models of cell death (oxygen-glucose deprivation, sodium azide-induced hypoxia, NMDA and hydrogen peroxide neurotoxicity). Additionally, our in vivo data from developing mouse brains showed that the density of the novel angiotensin binding site changes similar to the pattern of neuronal death in maturating brain. This is the first time that evidence is provided on the association of the novel angiotensin binding site with neuronal death, and future studies directed towards understanding of the function(s) of this protein are warranted.
- Received June 12, 2010.
- Revision received September 21, 2010.
- Accepted September 21, 2010.
- The American Society for Pharmacology and Experimental Therapeutics