A tight match appeared to be heading for stalemate when Brighton boss introduced on-loan Reading forward Rougier in place of Paul Brooker after 62 minutes.
Rougier made a decisive impact on 65 minutes The Trinidad & Tobago international latched on to a short pass from Graham Barrett to toe-poke the ball past Millwall goalkeeper Tony Warner from close range.
Rougier's shortage of goals for Reading made his contribution all the more remarkable. The 31-year-old netted only once in 31 outings for the Royals last season and has scored twice in 12 appearances for them so far this time.
Brighton just about deserved victory, which followed up crucial wins at home to Wolves and at Bradford.
Bobby Zamora hit the base of the post in the first half with a low drive from 15 yards.
Millwall, missing nine players through injury or illness, looked dangerous on the break and at set-pieces but they rarely threatened the veteran Dave Beasant on his return in goal for Brighton.
Beasant, 44 next month, had an anxious moment in stoppage time when a shot from the Lions' Belgian substitute Christophe Kinet drifted just wide.
Millwall's squad is so stretched that 15-year-old Moses Ashikodi was introduced in the closing stages in place of Neil Harris, who made his comeback from a long-term ankle problem.
Ivar Ingimarsson was impressive at the heart of the Brighton defence on his home debut and Steve Coppell's improving side have kept consecutive clean sheets since the Icelandic international signed for them on loan for a month from Wolves.