Objective: Our goal was to assess the potential of adeno-associated virus (AAV) mediated gene delivery into coronary microvessels in vivo in a large animal. Methods and Results: Ten mongrel dogs were chronically instrumented and allowed to recover for 10 days. Dogs were re-anaesthetized, and the aorta was constricted by a hydraulic occluder (AOC), whereby left ventricular pressure increased by 30% and left circumflex coronary artery blood flow by 50%. Recombinant adeno-associated virus (AAV, serotype 2, CMV enhancer/chicken beta-actin promoter) encoding for green flourescent protein (GFP), was injected as a bolus into the left atrium during AOC at total titers of 1010 or 1012 infectious units (iu). Dogs were followed for 2 (n=4) or 4 weeks (n=6). Hemodynamics or body weight did not change. In LV tissue slices, a fluorscein labelled antibody to GFP stained endothelial and smooth muscle cells, but was absent in myocytes. To quantify transduction, slices were then stained with antibodies against, -smooth muscle actin or von Willebrand Factor. Approximately 4 % of arterioles and 2 % of microvessels stained positive for anti-GFP independent from viral titer or duration. Using regression analyses, the per percent of vessels transfected was proportional to the increase in LV systolic pressure during occlusion. Conlcusion: AAV is a potential vector for gene transfer into the coronary microcirculation in large animals including perhaps humans.