Kratus and Bia, two servants of Zeus, carry in Prometheus and hold him against a rocky mountain in the Caucasus. Hephaestus, whose job is to chain Prometheus to the rock, follows them. Kratus states that this is a punishment for giving fire to human beings, and Prometheus must learn to like Zeus's rule. Hephaestus expresses his pity for Prometheus, lamenting the fact that he must bind his friend to the rock. Kratus urges him on, insisting that pity for Zeus's enemies is both useless and dangerous. Hephaestus finishes his task and leaves with Kratus and Bia.

Prometheus calls on nature to witness the suffering of a god at the hands of other gods, specifically the new ruler Zeus. He mentions that he has the gift of prophecy and knows all that will happen. He must live with his suffering because no one can fight fate. Prometheus hears the sound of wings and discovers that it is the Chorus of Oceanids, daughters of Oceanus, on winged chariots. The Oceanids express their sympathy for Prometheus's suffering, explaining that the new ruler of Olympus follows only his own laws. Prometheus prophesies that one day Zeus will be in danger and will be forced to befriend him to avoid it.

Asked what crime he has committed to deserve this punishment, Prometheus recounts the war between Zeus and the Titans. He had tried to help the Titans, but they refused his guile and decided to use force. Prometheus then offered his guile to Zeus, helping him win. Now Zeus punishes him because like all tyrants he distrusts his friends. Zeus had planned to destroy humanity but Prometheus stood in his way. Questioned further, Prometheus recounts that he gave humanity blind hope and also the gift of fire. The Chorus responds to this last admission, saying that he has sinned, but Prometheus replies that he did so willingly and will not now renounce his action. He asks the Chorus to come down to earth so he can tell them the whole story.

Oceanus flies in on a winged beast and says that Prometheus should stop provoking Zeus while he himself goes to the god to have Prometheus freed. Prometheus responds that talking to Zeus would be useless and Oceanus should not place himself in danger by getting involved. Oceanus argues that words are needed for healing, but Prometheus counters that the medicine must be applied at the proper time. Convinced by Prometheus's categorical refusal of his help, Oceanus departs.

The Chorus sings that the entire old world mourns for Prometheus and his brothers who also suffer at the hands of Zeus, especially Atlas who must hold the world. Prometheus summarizes everything he has done for humanity. He taught human beings agriculture, language, mathematics, harnessing of animals, and sailing. He also taught medicine, divination, and mining. He insists that all human arts come from him. Prometheus almost reveals his prophecy to the Chorus, but stops himself, saying only that it will free him because Zeus is a slave to necessity like all others. The Oceanids affirm their piety to Zeus, chide Prometheus for helping mortals who cannot help him back, and remember his happy wedding to their sister.

Io enters wearing cow horns. Screaming that a gadfly is biting her and the ghost of Argos pursues her, she calls on Zeus and asks why he tortures her like this. In response to her questions, Prometheus reveals who he is and why he is being punished. Io asks him to tell her about her future wanderings, but first the Chorus asks to know of her past suffering. Io tells how Zeus became infatuated with her and forced her father to drive her out of his house. Io was turned into a cow and guarded by Argos, but he was killed and returned as a ghost along with a gadfly that drove her around the world. Prometheus tells her of her future wanderings through Europe, Asia, and Africa, where she must constantly avoid dangerous peoples and monsters.

Prometheus reveals that Zeus, who makes Io suffer, will one day choose a mate whose son will depose his father. Only Prometheus can help Zeus prevent this. He reveals also that one day Io's descendant will free him. He then concludes the story of Io's journey, telling how at the end of her suffering Zeus will cure and impregnate her with a gentle touch of his hand. Prometheus talks about Io's descendants, who will become kings of the city Argos. Io runs off again tortured by the gadfly, and the Oceanids chant about the dangers of marrying above one's own rank, expressing the hope that Zeus never takes an interest in them.

Angered by Io's suffering, Prometheus shouts out that Zeus's own son will topple him. Zeus's messenger Hermes enters and orders Prometheus to reveal the identity of this son's mother. Prometheus mocks Hermes and says that he will tell nothing. Hermes accuses Prometheus of being overly obstinate, disobedient, and mad. Prometheus deflects each accusation with direct insults or sarcasm. Hermes warns that if Prometheus does not yield, a storm will send him to Tartarus. Then he will emerge only to have an eagle eat his liver every day, and this will not end until a god agrees to die for him. The Chorus advises Prometheus to yield, but he shows no fear of his destiny. Hermes orders the Oceanids away, but they reply that to betray a friend is the worst crime of all and vow to stay with Prometheus. As the earth begins to shake and thunder gathers around him, Prometheus calls on the elements to witness his suffering.