Identification of an I(2) binding protein from rabbit brain.

Imidazoline-2 binding proteins exist as a heterogeneous population. The aim of this study was to isolate and identify I(2) binding proteins from rabbit brain using an affinity column synthesized with a highly selective I(2) ligand, 2-(2-benzofuranyl)2-imidazoline (2BFI). The results revealed an approximately 45-kD protein to be brain creatine kinase (EC 2.7.3.2). [(3)H]-2BFI (5nM) was able to bind specifically to the purified enzyme. This study has identified brain creatine kinase as a novel I(2) binding protein.

Novel ligands for the investigation of imidazoline receptors and their binding proteins.

New ligands for imidazoline receptors are described so that these receptors can be more fully explored and understood. BU224, (2-(4,5-dihydroimidaz-2-yl)-quinoline, shows high affinity and is selective for the imidazoline-2 (I(2)) class of receptors. BU224 was tested in the rat Porsolt forced swim paradigm where it was found to decrease time spent immobile and increase the time spent swimming, consistent with an antidepressant profile. BU224 was tritiated and, in radioligand binding studies, was found to label a single population of saturable sites with high affinity. In vitro brain autoradiography with [(3)H]BU224 also showed a pattern of distribution similar to the known labeling of I(2) receptors. A new series of four 2BFI (2-(benzofuranyl)-2-imidazoline) derivatives were investigated as potential ligands for imaging brain I(2) receptors using positron emission tomography (PET). At least two, BU20012 and BU20013, retained high affinity and moderate selectivity and penetrated the brain when administered peripherally in the mouse. 2BFI has undergone the Mannich reaction to immobilized diaminodipropyl amine to fabricate an affinity column, which was used to isolate a protein from rabbit brain; this protein was sequenced and identified as the enzyme creatine kinase.