For nearly a century the Palazzo Te, its inhabitants, and its appropriators cooperated in the performance and practice of gender at the Renaissance court. After Ferdinando I Gonzaga's death in 1626 the palace seems to have languished under the subsequent reign of Vincenzo II Gonzaga, who was admittedly beset by other problems. Despite the fact that he had accused his wife of witchcraft, Vincenzo II had been unable to disentangle himself from his barren marriage to Isabella Gonzaga of Novellara. Under pressure from debtors, Vincenzo II sold the famed Gonzaga art collection to the British crown in 1627. Childless, impoverished, and in poor health, Vincenzo II made preparations for the duchy to pass to the francophone Gonzaga-Nevers branch of the family, and arranged the marriage of Maria Gonzaga, the only surviving child of Francesco IV Gonzaga and Margherita of Savoy, to Carlo II Gonzaga-Nevers.

Vincenzo II died on Christmas day 1627. The succession of Carlo II to the duchy of Mantua was contested by the Holy Roman Emperor Ferdinand II, who did not appreciate a French presence in Italy, as well as by Cesare II Gonzaga, Duke of Guastalla, and Charles Emmanuel, Duke of Savoy, each of whom believed that he should inherit Mantua. The War of Mantuan Succession began in 1628. As a result of the war, disease spread through the city, decimating the population, and conveniently carrying off Camilla Faa's son, Giacinto, who also posed a threat to the succession. The Sack of Mantua by imperial troops in 1630 ravaged what was left of the artistic and architectural patrimony of the city. The Palazzo Te became a barracks for imperial troops, who left behind graffiti and carried off its furnishings and movable goods. Some twenty years later Duke Carlo II Gonzaga-Nevers enlarged the gardens, adding a casino, and commissioned furnishings and wall hangings to replace those looted by the invading imperial troops.

However, the damage caused by the Sack of Mantua left a lasting impression of defeat and ruin, and the palace was no longer used by the Gonzaga dukes to welcome foreign visitors to their city. Writing almost a century later, the British traveler John Breval noted that the palace still bore the scars of the Sack, which had left it in a ‘naked and deplorable state’.