Estimating the time required for water to travel through headwater catchments from where it recharges to where it discharges into streams (the transit time) is important for understanding catchment behaviour. This study uses tritium (3H) activities of stream water to estimate the mean transit times of water in the upper Latrobe and Yarra catchments, southeast Australia, at different flow conditions. The 3H activities of the stream water were between 1.26 and 1.99 TU, which are lower than those of local rainfall (2.6 to 3.0 TU). 3H activities in individual subcatchments are almost invariably lowest at low streamflows. Mean transit times calculated from the 3H activities using a range of lumped parameter models are between 7 and 62 years and are longest during low streamflows. Uncertainties in the estimated mean transit times result from uncertainties in the geometry of the flow systems, uncertainties in the 3H input, and macroscopic mixing. In addition, simulation of 3H activities in FEFLOW indicates that heterogeneous hydraulic conductivities increase the range of mean transit times corresponding to a specific 3H activity. The absolute uncertainties in the mean transit times may be up to ±30 years. However, differences between mean transit times at different streamflows in the same catchment or between different subcatchments in the same area are more reliably estimated. Despite the uncertainties, the conclusions that the mean transit times are years to decades and decrease with increasing streamflow are robust. The seasonal variation in major ion geochemistry and 3H activities indicate that the higher general streamflows in winter are sustained by water displaced from shallower younger stores (e.g., soils or regolith). Poor correlations between 3H activities and catchment area, drainage density, mean slope, distance to stream, and landuse, imply that mean transit times are controlled by a variety of factors including the hydraulic properties of the soils and aquifers that are difficult to characterise spatially. The long mean transit times imply that there are long-lived stores of water in these catchments that may sustain streamflow over drought periods. Additionally, there may be considerable delay in contaminants reaching the stream.