We calculated N budgets and conducted nutrient uptake experiments to evaluate the fate of N in the aquatic environment of the Ipswich River basin, northeastern Massachusetts. A mass balance indicates that the basin retains about 50% of gross N inputs, mostly in terrestrial components of the landscape, and the loss and retention of total nitrogen (TN) in the aquatic environment was about 9% of stream loading. Uptake lengths of PO4 and NH4 were measurable in headwater streams, but NO3 uptake was below detection (minimum detection limit = 0.05 ?M). Retention or loss of NO3 was observed in a main stem reach bordered by wetland habitat. Nitrate removal in urban headwater tributaries was because of water withdrawals and denitrification during hypoxic events and in ponded wetlands with long water residence times. A mass balance using an entire river network indicates that basin-wide losses due to aquatic denitrification are considerably lower than estimates from several recent studies and range from 4 to 16% of TDN in stream loading. Withdrawals for domestic use restrict the runoff of headwater catchments from reaching the main stem during low base flow periods, thereby contributing to the spatial and temporal regulation of N export from headwater tributaries.