Eliza Dushku (Buffy the Vampire Slayer, Dollhouse) will provide the voice of Selina Kyle, as she does in Batman: Year One.
An image of the character from the 15-minute short has been released by TV Guide.
Dushku said: "I found the attitude for this character deep down in the Eliza Dushku archive of bad girls. They're in there somehow, somewhere for some reason, and I tap into them when I need them. They're characters with an edge."
The main film will also feature the voices of Battlestar Galactica's Katee Sackhoff as detective Sarah Essen, Southland's Ben McKenzie as Batman and Bryan Cranston in the role of Lieutenant James Gordon.
The straight-to-DVD feature is based on the seminal Frank Miller title of the same name, which retold the origin story of the Dark Knight.
Batman: Year One will be released in the US on October 18.