USA are bidding to retain the title and qualify for this year's Gold Cup.
It will be the squad's first meet-up since last year's World Cup.
"We come out of the World Cup knowing we're capable of more, and we're excited to keep moving forward in this group," said interim manager Anthony Hudson, who replaced former coach Gregg Berhalter in January.
"We want to defend our Nations League title and also qualify for this summer's Gold Cup, and to do that we need to get results," he added in a statement.
Chelsea's Pulisic heads the list of Premier League-based players called up, along with Fulham's Tim Ream (35) and Antonee Robinson (25), and Leeds United's Brenden Aaronson (22) and Weston McKennie (24).
Two dual-nationality players join the squad via Liga MX, Mexican-American winger Alex Zendejas (25) and Alan Sonora (24), who holds Argentine nationality.
Centre back Miles Robinson (26) is set to return after recovering from a ruptured Achilles tendon suffered last May.
With two matches remaining in Group D, the second-placed USA travel to Grenada on March 24 and three days later host top-ranked El Salvador in Orlando.
