The carer, played by David Walliams, has traded in troublesome Andy Pipkin (Matt Lucas) and is now looking after Professor Stephen Hawking, 73, instead, in an upcoming sketch for Comic Relief.
The comedy will briefly make a comeback this Friday in aid of the charity, which will see the famous skit relocate to Cambridge University.
Sharing the exciting news on Twitter, the Britain's Got Talent star, 43, wrote: "#LittleBritain is returning for @rednoseday Lou is now the carer to Professor Stephen Hawking @BBCOne."
Filmed over two days in January, comedienne Catherine Tate also features as a nun.
David - who recently split from wife Lara Stone - last reprised his role as bumbling Lou for the Pride of Britain Awards in 2009.
Lenny Henry, Davina McCall and Claudia Winkleman will be among the hosts of Comic Relief's Red Nose Day at 7pm on BBC1.
Physicist Stephen recently saw his life turned into a movie starring Eddie Redmayne - who went on to win an Academy Award for his role.
He rushed to congratulate the actor via Facebook after his win, writing: "Congratulations to Eddie Redmayne for winning an Oscar for playing me in The Theory of Everything Movie. Well done Eddie, I’m very proud of you."
Little Britain's Lou and Andy 1769448735j2bzU9KX
As he took to the stage to claim the prize, the 33-year-old star said: "I don't think I'm capable of articulating quite how I feel right now. I'm fully aware that I'm a lucky, lucky man."
He dedicated the award to "all of those people around the world battling" motor neurone disease - the illness that left the professor in a wheelchair.
Eddie faced tough competition in the best actor category as he went head-to-head with fellow Brit Benedict Cumberbatch for his role in The Imitation Game.Â