The hotel pranks on the PLA main site are funny but the page isn't very clear as to how to get through to these rooms. Now, it varies from hotel to hotel and ITT I'll explain which ones to call and which ones not to call.
First of all, you want to avoid the big fancy hotels. Places like Marriott and Hilton are difficult because the receptionist will usually ask for the name of the guest that resides in the room you are trying to call to prove that you aren't some dickhead trying call random people. A good choice for hotel prank is cheap motor lodges like Best Western, Comfort Inn, etc. The front desk will usually transfer you to the room you want in a heartbeat, but the best choice is Motel 6 because you don't deal with a receptionist at all; just punch in the room number and off you go.
Now that you've gotten through to someone's room, what do you say? There's lots of ways to mess with people in hotels. Tell them you work in housekeeping and tell them that they need to start flushing the toilet when they are done. Tell them they are eating too much fiber. Call in at 4 in the morning until the guest wakes up. When he answers tell him he sounds tired and needs to get some sleep. Call room 666 and ask if Satan is there. If you call a room and get ahold of an elderly couple, tell them that they need to start using the Do Not Disturb sign. When they ask why, tell them that Terry, the bellhop that started working yesterday walked in on them today while they were having sex and now he is in a coma. Call someone before they go to bed and tell them that the hotel is currently having a very severe roach problem. Tell them that the inner walls are basically filled with big flying roaches about an inch long and they've been very troublesome lately. Tell them that you've had a few guests leave because of it, one of them finding roaches in their coffee, and another waking up in the night screaming because roaches had crawled all over his face and into his mouth. Tell them that the board of motor lodges requires that you inform all guests about the problem until it is resolved. If you are convincing enough, they might leave the hotel. Call someone and ask them if they have used any of the towels. Tell them that a mentally challenged man that used their room last was using the towels to wipe himself after using the toilet. If he has used the towels, tell him he should probably go take a few more showers and use his own towel this time.