T-cells may play a role in the evolution of ischaemic damage and repair, but the ability to image these cells in the living brain after a stroke has been limited. We aim to extend the technique of real-time in situ brain imaging of T-cells, previously shown in models of immunological diseases, to models of experimental stroke. Male C57BL6 mice (6-8 weeks) (n= 3) received a total of 2-5 x 10(6) carboxyfluorescein diacetate succinimidyl ester (CFSE)-labelled lymphocytes from donor C57BL6 mice via i.v. injection by adoptive transfer. Twenty-four hours later, recipient mice underwent permanent left distal middle cerebral artery occlusion (MCAO) by electrocoagulation or by sham surgery under isoflurane anaesthesia. Female hCD2-green fluorescent protein (GFP) transgenic mice that exhibit GFP-labelled T-cells underwent MCAO. At 24 or 48 h post-MCAO, a sagittal brain slice (1500 microm thick) containing cortical branches of the occluded middle cerebral artery (MCA) was dissected and used for multiphoton laser scanning microscopy (MPLSM). Our results provide direct observations for the first time of dynamic T-cell behaviour in living brain tissue in real time and herein proved the feasibility of MPLSM for ex vivo live imaging of immune response after experimental stroke. It is hoped that these advances in the imaging of immune cells will provide information that can be harnessed to a therapeutic advantage.
- brain imaging
- middle cerebral artery occlusion
- multiphoton microscopy