In vitro studies using biochemical, pharmacological and molecular approaches demonstrated that the peripheral-type benzodiazepine receptor (PBR) is a mitochondrial protein, involved in the regulation of cholesterol transport from the outer to the inner mitochondrial membrane, the rate-determining step in steroid biosynthesis. In vivo animal models and ontogeny studies validated the role of PBR in steroidogenesis. Targeted disruption of the PBR gene in Leydig cells resulted in the arrest of cholesterol transport into mitochondria and steroid formation. Molecular modeling of PBR suggested that it might function as a channel for cholesterol. Indeed, cholesterol uptake and transport by bacteria cells was induced upon PBR expression. Amino acid deletion and site-directed mutagenesis studies identified a cholesterol recognition/interaction amino acid consensus sequence in the cytoplasmic carboxy-terminus of the receptor. In vitro reconstitution experiments demonstrated that the 18 kDa PBR protein binds with high affinity both drug ligands and cholesterol, suggesting that this protein might serve numerous functions considering the critical role of cholesterol in membrane biogenesis and human pathology. In this context, PBR expression correlated with the quality of kidney preservation, indicating that it might serve as an index of kidney and mitochondrial viability during ischemia-reperfusion injury. PBR overexpression was also found to be a prognostic indicator of the aggressive phenotype in breast, colorectal and prostate cancers. Moreover, in Alzheimer's disease brain specimens, PBR levels were increased and paralleled the elevated neurosteroid synthesis observed in specific brain areas. The role for PBR in these pathological conditions remains to be elucidated. paralleled the elevated neurosteroid synthesis observed in specific brain areas. The role for PBR in these pathological conditions remains to be elucidated.