1. ¹Institute for Surgical Research, Ludwig-Maximilians-University, Munich, Germany
2. ²Department of Neurosurgery, Ludwig-Maximilians-University, Munich, Germany
3. ³Department of Neurosurgery, Albert-Ludwigs-University, Freiburg i.B., Germany

Correspondence: Dr E Uhl, Department of Neurosurgery, Klinikum Grohadern, Ludwig-Maximilians-University, München D-81377, Germany. E-mail: eberhard.uhl@med.uni-muenchen.de

Received 12 December 2005; Revised 21 March 2006; Accepted 10 April 2006; Published online 24 May 2006.

Abstract

Activation of platelets induces interactions with platelets, endothelial cells, and leukocytes. In vivo observation of these interactions in the cerebral microcirculation is rare. The purpose of the present study was to develop a model enabling the in vivo observation of platelet kinetics in the cerebral microcirculation. Intravital fluorescence microscopy was performed in the Mongolian gerbil. Platelets of a donor were labeled ex vivo with carboxyfluorescein diacetat-succinimidylester (CFDA-SE), providing long-term fluorescence. Platelet function was tested ex vivo by flow cytometric analysis and in vivo by analyzing platelet-endothelium interactions. Labeled platelets stimulated with adenosine diphosphate ADP (200 mol/L) or thrombin (1000 U/L) showed aggregation in flow cytrometric analysis, whereas unstimulated platelets were not aggregated. Irradiation of the brain surface after intravenous injection of the photosensitizing dye Photosan first induced rolling and firm adherence of platelets on arteriolar and venular endothelium, followed by the formation of a thrombus obstructing the vessel. Quantitative analysis (n 100 m^-1 min^-1) before and after 6 mins of irradiation showed 2.63.2 versus 29.028.9 rolling, and 0.00.0 versus 1.72.3 firm adherent platelets in arterioles, and 3.93.3 versus 36.620.9 rolling and 0.00.0 versus 13.68.9 firm adherent platelets in venules. Thus, we conclude that ex vivo labeling of platelets with CFDA-SE does not activate platelets. Platelet aggregation and adhesion was achieved by platelet-specific stimulation such as ADP, thrombin or irradiation. In vivo assessment of physiologic and pathophysiologic mechanisms of platelets in the cerebral microcirculation can be achieved in this model.