Endogenous antithrombin associated with microvascular endothelium. Quantitative analysis in perfused rat hearts

A recirculating Langendorff heart preparation is used to characterize the endogenous antithrombin associated reversibly with murine vascular endothelium. Rat hearts are perfused clear of blood and then recirculated with a physiological salt solution. Addition of heparin educes antithrombin activity continuously into the perfusate during 6 min of recirculation. This process contrasts with a more rapid equilibration of the system as assessed by displacement of [125I]thrombin with hirudin or with a heparin-antithrombin mixture. Perfusion of washed hearts with [125I]factor Xa, which evidences no significant binding to the coronary endothelium, identifies a minor fraction of the endogenous antithrombin that reacts immediately with factor Xa, i.e., at a rate indicative of heparin enhancement. This rapid-reacting antithrombin is not reproducibly detected with [125I]thrombin, which binds preferentially to thrombomodulin in this system. The amount of antithrombin reacting rapidly with factor Xa is too low to detect as a burst of antithrombin activity eluted into the perfusate when the hearts are perfused with heparin. It is concluded that the murine myocardial microvasculature harbors at least two pools of antithrombin, the minor of which is in an activated configuration characteristic of association with heparin. The major pool is in a more slowly accessible compartment or configuration.