The 51-year-old "Jeremy" hitmaker sang "Light Years" for the Canadian rocker during his concert at Wrigley Field in Chicago, reported Entertainment Weekly.
"There's something going on north of the border tonight and it involves somebody who we consider a family member just because he's part of the group of touring musicians that we've met over the years," Vedder said.
Downie, 52, performed his last show with the Tragically Hip the same night for a sold-out crowd, including Canadian Prime Minister Justin Trudeau in Kingston, Canada.
"So they're up there, going through the emotions up there. I just want to send them our energy from our gathering up to their gathering up there, and wish the best to Mr Gord Downie. We love you," Vedder added.