1. ¹Center for Molecular Imaging Research, Department of Radiology, Massachusetts General Hospital, Harvard Medical School, Charlestown, MA, USA
2. ²Cardiology Division, Department of Internal Medicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
3. ³Department of Radiology and Experimental Imaging, University of Texas, MD Anderson Cancer Center, Houston TX, USA

Correspondence: Ching H Tung, Center for Molecular Imaging Research, Massachusetts General Hospital, 149 13th St., Rm. 5410, Charlestown, MA 02129, USA. E-mail: tung@helix.mgh.harvard.edu

Received 7 June 2004; Revised 30 August 2004; Accepted 14 September 2004.

Abstract

An intravital microscopy imaging method was developed to visualize active cerebral thrombus and blood–brain barrier (BBB) disruption using Near Infrared Fluorescent (NIRF) probes. A circular craniotomy was made in CD-1 mice. Thrombi were formed by applying 10%-FeCl₃ to the entire exposed superior sagittal sinus (SSS, 5 mm), or to the posterior 2.5 mm of the SSS for 5 mins. Control animals were pretreated with heparin (50 U/kg) before thrombus induction. Three hours after thrombus formation, a FXIIIa-targeted NIRF imaging probe (A15) was intravenously injected, and the SSS was imaged by intravital microscopy. This was followed by injection of indocyanine green (ICG) to assess BBB permeability. The A15 optical probe bound to thrombus, and the fluorescent signal emitted by the bound agent corresponded well with histologically confirmed thrombus. A15 initially remained intravascular, followed by excretion and subsequent decrease in all tissues except for thrombus, where it was retained. The subsequent ICG was also intravascular immediately after injection, but then began to leak into the cerebral parenchyma at 3 to 5 mins. The sites of leakage were adjacent to thrombosed areas. Heparin pretreatment prevented thrombus formation and reduced ICG leakage significantly. This demonstrates the feasibility of simultaneous in vivo monitoring of thrombus and BBB permeability in an animal model of cerebral venous thrombosis.