The sudden announcement on Tuesday comes with Villa just four points above the relegation zone with five games remaining.
The central England club says Ian Culverhouse, assistant to manager Paul Lambert, and Gary Karsa, the head of football operations, have been suspended pending an internal investigation. Further details were not provided.
Villa says goalkeeper Shay Given and development coach Gordon Cowans have been temporarily promoted to assist Lambert.
Given says "as players, it's important that each and every one of us take responsibility right now and I'm delighted to play my part in helping the manager."
Villa's next game is against Southampton on Saturday.