The Australian, who has recently complained of suffering from vertigo, crumpled as he made his way down the steep slope from tee to green on the par three 9th and needed urgent medical attention.
Day, who had started on the 10th, was 3 under for the championship when he fell to his left and struck his head on the ground as he landed.
The three-time PGA Tour winner remained flat on his back for several minutes while on-course medical staff attended to him.
Playing partner Jordan Spieth sportingly kept onlookers at bay and appealed for spectators not to take photographs out of respect for the 27 year old.
Day recovered sufficiently to complete the hole, and received a rousing cheer from the crowd as he climbed gingerly into a greenside bunker where his tee shot had landed. He was unable to get up and down and dropped a shot to finish his round on level-par 70 for a two-round total of 138, leaving him tied for 11th place.
However, he was clearly unsteady on his feet as he left the course and was taken to hospital for tests.
Day had withdrawn before the start of the Byron Nelson Championship three weeks ago due to severe dizziness and revealed on Tuesday that he had undergone numerous tests to try to identify the cause.
“I had three sleep studies done. I had a lot of blood tests done. I had an MRI on my head and my neck and everything came back negative,” he said.
“So I have no idea what that was, other than I just may have been exhausted. I was training so hard, I was doing two-a-days every day coming into tournaments and then on top of it I was doing practice, playing competitive golf and then trying to balance that with family as well.
“It’s just a full-time kind of gig there and I think I just ran out of gas and I wasn’t feeling good, so I had the shakes and the tingling up my arms. And the loss of energy and strength was probably caused by that. I’ve got severe sleep deprivation, so I guess that’s part and parcel of having a kid.”
Meanwhile, Masters champion Spieth kept his bid to make history firmly on track, while Tiger Woods headed towards just his fifth missed cut in 68 majors as a professional.
Spieth is looking to become only the sixth man in history after Craig Wood, Ben Hogan, Arnold Palmer, Jack Nicklaus and Tiger Woods to win the Masters and US Open in the same year.
The world No 2 can also become the first player since Gene Sarazen in 1922 to win multiple majors aged 21 or younger at Chambers Bay and cruised into the lead with four birdies in his first eight holes.
Starting on the back nine, Spieth birdied the 10th, 14th, 15th and 17th – he also three-putted the driveable 12th for par after missing from two feet – to reach six under par, one shot ahead of overnight leaders Henrik Stenson and Dustin Johnson.
However, the Ryder Cup star then hit the lip of a bunker with his second shot on the 18th, found more sand with his third shot and eventually carded a double-bogey six on the long par four, which had played as a par-five on Thursday. Spieth was heard on television berating himself for the “dumbest hole I’ve ever played in my life” after his second shot, but bounced back immediately with a birdie on the first, which was playing as a 593-yard par five.
A dropped shot on the 7th, however, was recovered at the his final hole, picking up a birdie in the midst of playing partner Day’s troubles.
At five under par, Spieth shared the lead with Stenson, Johnson and fellow American Daniel Summerhays, who appeared to be putting together the round of the day with six birdies and one bogey in 15 holes, but dropped back to three under par after a double bogey on the 7th.
South African Branden Grace and American Ben Martin were a shot behind, with Jamie Lovemark and JB Holmes setting the early clubhouse target on two under.
At the other end of the leaderboard, Woods was at least faring slightly better than he had during an opening 80, his worst ever score in the US Open and his third round in the 80s in his last six events.
The former world No 1 pulled his second shot to the 10th – his opening hole – on to such a steep slope that he lost his balance attempting to take his stance, but recovered from the resulting bogey with a birdie on the 12th.
Dropped shots on the 14th and 18th were followed by a birdie on the first, but more bogeys on the second and third left Woods 13 over par, with just Lucas Glover and Ryder Cup captain Darren Clarke below him on the leaderboard.
Spieth three-putted from long range on the seventh but could be thankful for small mercies after seeing playing partner Justin Rose run up a triple-bogey seven, and, at three over par, the 2013 US Open winner was suddenly facing a battle to avoid missing the cut.
Scotland’s Jimmy Gunn had a second successive 72 to finish four over, a shot inside the projected cut mark.