The petition is calling on Spain's Ministry of Culture and the Mayor of Madrid to stop a real live bull from being used in the opera Moses and Aaron in the capital's Teatro Real (Royal Theatre).
The bull appears on stage in a 15 minute scene in the opera, which will run from May 24th to June 17th.
Using the animal "exposes the bull to lights and loud noises, constant movement and unnecessary stress both during the 15 minute scene in which it features and during the rest of the opera the its rehearsals," says the petition, which was launched on Change.org by Jaime Alcázar.
The petition is calling for the opera to be cancelled or for a statue of a bull to be used in place of a real animal.
The opera faced similar criticism when it was staged in Paris; animal rights groups presented a petition to Minister of Culture Fleur Pellerin calling for the opera to be closed because of the "stress on the animal" and the "suspicion that it is drugged to be able to better control the scene, which concerns a 1,500kg animal".
Madrid's Royal Theatre has said that the bull receives "no type of medicine, drug or painkiller" during the opera and has "two specialized carers to look after its wellbeing, transport and to provide veterinary care."