Lymphedema distichiasis (LD) is a form of primary lymphedema associated with mutations in the forkhead transcription factor gene FOXC2. Homozygous loss of Foxc2 in the mouse is associated with agenesis of secondary lymphatic valves as well as other cardiovascular and skeletal defects that contribute to lethality between E12.5 and P1. Foxc2+/- mice are characterized by distichiasis, reduced lymphatic drainage and lymph backflow, similar to patients with LD. We previously demonstrated a novel type of lymphatic pump dysfunction, termed valve lock, which can be experimentally induced if healthy single lymphangions or chains of lymphangions are subjected to pressure conditions that mimic those associated with the development of edema in peripheral extremities. Valve lock is due in part to a normal property of intraluminal lymphatic valves in which valve closure becomes more difficult as the vessel becomes distended (i.e. the adverse pressure gradient required for closure rises 20 fold). As we also recently reported, tests of isolated valves from Foxc2+/- mice indicate that about 50% of the valves have elevated back-leak and appear to be abnormally stiff. Here we tested whether lymphatic vessels from Foxc2+/- mice exhibit pumping deficits as a result of these valve abnormalities. Segments of collecting lymphatic vessels, each containing two valves, were isolated from the popliteal region of anesthetized adult C57Bl/6 (WT) or Foxc2+/- mice (Nembutal, 60 mg/kg, i.p.), cannulated at each end with a micropipette. Intraluminal pressure (PL) was measured between the valves with a servo-null micropipette, while tracking vessel diameter and monitoring the leaflet position of the output valve using video methods. A servo controller held input pressure (Pin) at 1 cmH2O while output pressure (Pout) was slowly elevated ramp-wise. The pumping limit (Plimit) was defined as the output pressure at which the output valve first failed to open. About half of the WT vessels simply failed to eject at high Pout (Plimit ~12 cmH2O); in the other half, valve lock occurred before this point (Plock). On average Foxc2+/- vessels exhibited lower values for Plimit compared to WT vessels (8.0±0.8 vs 13.9±0.2 cmH2O, respectively) and lower values for Plock (10.5±0.7 vs 15.9±0.1 cmH2O, respectively) and a higher susceptibility to valve lock (68% vs 25%). These new methods for studying mouse lymphatic vessels under defined conditions reveal that the abnormal valves in Foxc2+/- vessels render the collecting lymphatics more susceptible to pump failure. The same susceptibility may be characteristic of lymphatic vessels in human patients with LD and contribute to the peripheral edema from which those patients suffer.