Mulgrew will play Warehouse protector Jane in four episodes of the show, TV Guide reports.
Speaking about Mulgrew, who played Captain Kathryn Janeway in Star Trek Voyager, Warehouse 13's executive producer Jack Kenny said: "I was an original Star Trek fan and loved the movies.
"I remember loving when they cast a female captain of the Enterprise and have found her work incredibly deep, thoughtful and compelling to watch. I'm thrilled to have her play a character who will play such an important overall role in our series."
Ryan - who starred in Star Trek: Voyager - will play Amanda in Warehouse 13. Kenny teased that she "has a past with someone in the Warehouse".
Star Trek: Deep Space Nine actor Rene Auberjonois will reprise his role as Hugo Miller, while Lindsay Wagner will return as Vanessa Calder.
Caprica stars Sasha Roiz and Alessandra Torresani have also landed roles in the new season of Warehouse 13.
"Sasha is a mysterious part of our Big Bad for the year," Kenny hinted. "His character is Marcus Diamond and first shows up in episode three. Alessandra guest stars as a girl in trouble in our ninth episode who needs Myka and Claudia's help."
The third season of Warehouse 13, which will also star Aaron Ashmore, will begin on July 11 on Syfy.
> 'Warehouse 13' spinoff in the works