The documented history of the Valley of the Muses begins in the sixth century BC, but its great period came later. Starting in the third century BC, the people of nearby Thespiae organized the Mouseia festival — held every four years, originally every year — and opened it to competitors from across Greece. Poets and musicians came to perform in games modeled on the great Panhellenic competitions at Olympia and Delphi. Winners dedicated their prizes — bronze tripods — to the sanctuary, and over time the valley accumulated a remarkable collection of votive offerings alongside statues of the Muses and portraits of celebrated poets. The festival was a statement about Boeotia's claim to cultural prestige: this region, home to Hesiod and Pindar, was not merely a farming landscape but the very ground on which Greek poetry had first found its voice.

In the second and first centuries BC, the character of the Mouseia shifted. Games were added in honor of the Roman emperor, and the festival acquired the additional name of the Great Kaisareia — the Great Imperial Games — because the emperor was now honored alongside, and eventually above, the Muses. Imperial patronage brought resources but also changed the festival's meaning. The sanctuary continued to accumulate sculpture: many statues, paintings, and reliefs of Muses, poets, and musicians filled the open-air space. Then, as monotheism spread through the Roman world and pagan festivals lost their legal standing, the valley was abandoned. The last ceremonies are unrecorded. The statues were eventually removed or toppled, the theatre silted up, and a small Christian church was built on the foundations of the Muses' own temple.

Modern excavation began in 1882, when the Greek archaeologist Panagiotis Stamatakis dug a test trench in the little church of Ayia Triada and found, beneath it, the rectangular foundations of the ancient temple of the Muses. He also identified the remains of a theatre on the hillside above. Systematic work followed: the French Archaeological School excavated in 1888, 1889, and 1890 and uncovered the principal monuments. The theatre, built at the end of the third or beginning of the second century BC, had its seating cut into the natural slope of the mountain; only the front row was marble. A long Ionic stoa, 96.7 metres in length, stood west of the temple and once sheltered the votive offerings given to the Muses. The bases of the nine Muse statues — the work of the poet and sculptor Onestos — were found during excavation, five of them still bearing inscribed names and epigrams.

The sanctuary was not without its military dimension. On the hill called Pyrgaki at the northern end of the valley, the excavators found the remains of a square tower dated to the fourth century BC — the Tower of Askra — along with traces of an enclosing circuit wall. The tower was probably built by the Thespians a little before 371 BC, the year of the Battle of Leuktra, when the Theban general Epaminondas destroyed Spartan power in a single afternoon on a plain not far away. The tower's purpose was to watch the Thebans, who were neighbors and rivals of Thespiae. Poetry festivals and military architecture: the Valley of the Muses held both at once, as places of gathering tend to do. Today the site is an open-air archaeological park, permanently accessible, the theatre outline visible on the green slope and the stoa foundations stretching across the valley floor beneath Mount Helicon's summit.