Roderick St. James (Hugh Jackman) is a pampered pet rat who lives in Kensington. When his owners go on vacation, that's when Roddy comes out to play. But one day, a common sewer rat named Sid (Shane Richie) comes spewing out the sink. That's when he decides to stay. And all of this goes on as the Soccer World Cup is playing, with England in a match against Germany. In one scene where it shows Sid watching the game, it announces the teams, with him cheering on England and booing Germany and throwing a hot dog at the TV. Roddy tries to get rid of Sid by telling him that the toilet is actually a jacuzzi, but Sid is no idiot. He plays along and tricks Roddy into going into the 'jacuzzi'. When he reaches the end of the pipes, he enters a rat version of London, complete with homemade Big Ben clocktower and everything. That's where he meets Rita (Kate Winslet) a streetwise scavenging rat who works the tunnels in her boat The Jammy Dodger. Initially, Rita dislikes Roddy, but she takes him on as her enemy The Toad (Ian McKellen) sends his 2 minions Spike (Andy Serkis) and Whitey (Bill Nighy) after her because she has stolen back her father's prized ruby. The Toad hates all rodents to the point of obsession, blaming them from his fall from the nice life, in the movie, he was once Prince Charles' pet. He decides to have them frozen with liquid nitrogen. However this plan fails, as during their escape, Rita steals the master control belt to control the floodgates. The Toad's plan is to open the floodgates during the World Cup's halftime, drowning the rats and the city in sewage. Then he can repopulate the city with his own tadpole offspring.
- Hugh Jackman as Roderick "Roddy" St. James - The main protagonist. A lonely pampered pet mouse living in a Kensington apartment with a wealthy English family who is flushed down the toilet by Sid into the sewers.
- Kate Winslet as Rita Malone. The deuteragonist. A street smart scavenger rat and the oldest child of a large family. She is the captain of The Jammy Dodger.
- Ian McKellen as The Toad. The main antagonist. A big, pompous amphibianwanting the entire rat population to be killed off so he can make room for his hundreds of offspring.
- Jean Reno as Le Frog The secondary antagonist. The Toad's French cousin. He masters martial arts and is the leader of a team of hench-frogs.
- Andy Serkis as Spike, one of the Toad's top two hench-rats. Being the Toad's hench-rat is Spike's first job, despite him being only a teenager. He is the smarter of the two.
- Bill Nighy as Whitey, another of the Toad's top two hench-rats. Whitey is an albinorat, once working in a lab and he is Spike's partner. Unlike Spike, Whitey is gentle and less violent, but is also rather slow-witted and dumb.
- Shane Richie as Sid, an over-weight and lazy rat from the sewers. He is an acquaintance of Rita and her family and the one who flushed Roddy down the toilet into the sewers.