Pena's blast in the first inning staked the Braves to an early lead while Upton's clout in the next frame put Atlanta ahead to stay at 3-2.
Paul Maholm (7-4) held the Nationals to three runs (two earned) on five hits over six frames to gain the win and boost Atlanta's lead in the East to 6 1/2 games.
Anthony Varvaro and Luis Avilan each logged a scoreless relief frame and Craig Kimbrel racked up his 17th save, securing the victory.
Washington's Nate Karns (0-1) absorbed the loss in his second major-league start, surrendering four runs (three earned) on seven hits over 4 2/3 frames.
Ian Desmond homered for the Nats.