Nectomys squamipes is a semi-aquatic rodent with a wide geographic range in Brazilian forests and savannas. This species is a host for several parasites associated with aquatic environments, especially Schistosoma mansoni, and thus its spatial pattern may influence some diseases’ patterns. Here we present important aspects of water rat spatial behaviour, including technical and ecological aspects. Water rats were studied by live trapping in rivers and in gallery forests between January 2005 and August 2008, with a total sampling effort of 9511 trap-nights. We also studied some animals through radio tracking and the spool-and-line technique. Water rat movement was distributed evenly between the terrestrial and aquatic environments. Nectomys squamipes occurrence in small tributaries was higher than in larger and lotic rivers. Both home range, the conventional two-dimensional approach, and home length, a one-dimensional approach, proved to be useful to study N. squamipes, depending on the shape of the water body used by the individual. Although Nectomys squamipes’ home length was positively correlated with body weight, sex was also important to explain home length variation; at similar body weights, males have larger home lengths than females. Males explored larger areas during the rainy season, likely in search of females, as this is the water rat reproductive season in Atlantic forest. In contrast, females explored larger areas during the dry season, likely looking for additional food supplies as a result of deficient water rat food supplies during the Atlantic forest dry season. Females overlapped home lengths with more than one male, and males with more than one female, suggesting a promiscuous mating system for this species.