Apparently. In games like Grand Theft Auto V, American characters tend to mock Trevor's Canadian accent, and Trevor's backstory originally entailed that he grew up in the rural American backwoods, but when they hired Steven Ogg to portray Trevor, they decided to work Steven's "thick Canadian accent" into the character by changing the backstory to make it that Trevor came from Canada instead.
As a Canadian-American with an American accent, yeah there is definitely a difference but it's hard to notice if you speak another language. I think part of it comes from sort of a midwestern American accent that spread throughout Canada, but where I live people talk the same both sides of the border (unless it's French lol)