A woman in a wedding dress, the Bride (Uma Thurman), lies wounded in a church, having been attacked by the Deadly Viper Assassination Squad. She tells their leader, Bill (David Carradine), that she is carrying his baby. He shoots her.
Marauding bandits approach a rural mountain village, but their chief decides to spare it until after the harvest because they had raided it before. The plan is overheard by a farmer who tells the rest of village. Lamenting their fate, three farmers ask Gisaku, the village elder and miller, for advice. He declares they should hire samurai to defend the village. Since they have no money to offer, Gisaku tells them to find hungry samurai.
Edo, 1630. Tsugumo Hanshirō arrives at the estate of the Ii clan and says that he wishes to commit seppuku within the courtyard of the palace. To deter him Saitō Kageyu (Rentarō Mikuni), the Daimyo's senior counselor, tells Hanshirō the story of another rōnin, Chijiiwa Motome – formerly of the same clan as Hanshirō.
In the Edo period of Japan, Isaburo Sasahara (Toshiro Mifune) is a vassal of the daimyo of the Aisu clan. Sasahara is the most skilled swordsman in the land, whose only rival in ability is his good friend Tatewaki Asano (Tatsuya Nakadai). Isaburo is in a loveless marriage with a shrew of a woman. One day one of the daimyo's advisors orders Isaburo's elder son Yogoro (Go Kato) to marry the daimyo's ex-concubine, Ichi (Yoko Tsukasa), even though she is the mother to one of the daimyo's sons. With much trepidation, the family agrees. In time, Ichi and Yogoro find love and happiness in the marriage and a daughter Tomi is born.
It is 1860, and the era of the Tokugawa shogunate is coming to a close. A rōnin, or masterless samurai, wanders into a small town divided by a gang war between two gangsters, Seibei and Ushitora. Ushitora used to be Seibei's right-hand man, until Seibei decided that his son Yoichiro would succeed him. Tazaemon, the silk merchant and mayor, backs Seibei, while Tokuemon the sake brewer is allied with Ushitora. Gonji, the owner of an inn, advises the stranger to leave while he can, but after sizing up the situation, the rōnin tells Gonji that the town would be better off with both sides dead, and that he intends to stay.
Hidetora Ichimonji, a powerful warlord, experiences a dream reminding him that he's showing his age and decides to divide his kingdom among his three sons: Taro, Jiro, and Saburo. Taro, the eldest, will receive the prestigious First Castle and become leader of the Ichimonji clan, while Jiro and Saburo will be given the Second and Third Castles. Hidetora will retain the title of Great Lord and Jiro and Saburo are to support Taro.
The film begins with two bedraggled peasants, Tahei and Matashichi (Minoru Chiaki and Kamatari Fujiwara). Through conversation, they reveal that they had intended to fight alongside the Yamana clan, but turned up too late, were taken for soldiers of the defeated Akizuki clan, and forced to bury the dead. After quarreling and splitting up, the two are both captured again and forced to dig for gold in the Akizuki castle with other prisoners.
Generals Miki and Washizu are Samurai commanders under a local lord, Lord Tsuzuki, who reigns in the castle of the Spider's Web Forest. After defeating the lord's enemies in battle, they return to Tsuzuki's castle. On their way through the thick forest surrounding the castle, they meet a spirit, who foretells their future. The spirit tells them that today Washizu will be named Lord of the Northern Garrison and Miki will now be commander of the first fortress. She then foretells that Washizu will eventually become Lord of Spiderweb Castle, and finally she tells Miki that his son will also become lord of the castle.
At the start of the film, the main character, Iguchi Seibei, becomes a widower when his wife succumbs to tuberculosis. His wife receives a grand funeral, more than what a low-ranking samurai such as Seibei could afford. Seibei works in the grain warehouse, accounting for stores inventory for the samurai clan. His samurai colleagues mock him behind his back with the nickname Tasogare (Twilight). When evening approaches, Seibei rushes home to look after his senile elderly mother and two young daughters, Kayano and Ito, instead of bonding with his supervisor and other samurai colleagues over customary nights of dinner, geisha entertainment, and sake drinking. Even though he is a samurai, Seibei neglects his appearance, failing to bathe and being shabbily dressed. The well-being of his young daughters and medicine for his mother take priority over new clothes or the monthly bath fee and his daughters say they are both happy, even without a mother.
A woman in a wedding dress, the Bride, lies wounded in a church in El Paso, having been attacked by the Deadly Viper Assassination Squad. She tells their leader, Bill, that she is carrying his baby. He shoots her.
The pregnant Bride and her groom rehearse their wedding. Bill, the Bride's former lover, father of her child, and leader of the Deadly Viper Assassination Squad, arrives unexpectedly. On Bill's orders, the Deadly Vipers kill everyone at the wedding, but the Bride survives and swears revenge.
Nine young samurai, including his nephew, believe that the lord chamberlain, Mutsuta, is corrupt after he tore up their petition against organised crime and said maybe he himself was behind the crimes. One of them tells the superintendent of this and he agrees to intervene. As the nine meet secretly at a shrine and discuss their problem, a ronin (Mifune) emerges from another room where he has been resting. The ronin has overheard their plans, and suggests that the superintendent is in fact the real corrupt official. While at first the samurai are insulted by his claims, they soon find themselves surrounded by the superintendent's men, proving that he was correct. The ronin hits a few of the attackers and bluffs their leader into leaving, taking only a small amount of money in reward to buy food because he is starving; however, after realizing that Mutsuta and his family must now be in danger, he decides to help the samurai bring down the corrupt officials.
Lord Shingen Takeda meets with his brother Nobukado, and a thief whom the latter met by chance and spared from crucifixion. Nobukado believes the thief's uncanny resemblance to Shingen would prove useful. The brothers then agree that he would prove useful as a double, and they decide to use the thief as a kagemusha.
The story follows the life of Ryunosuke Tsukue (Tatsuya Nakadai), an amoral samurai and a master swordsman with an unorthodox style. Ryunosuke is first seen when he kills an elderly Buddhist pilgrim who he finds praying for death. He appears to have no feeling. Later, he kills an opponent in self-defense in a fencing competition that was intended to be non-lethal, but became a duel after he coerced his opponent's wife to have sex with him in exchange for throwing the match and allowing her husband to win. His opponent finds out about the affair prior to the match, and is shown giving his wife a notice of divorce. His rage at Ryunosuke during the match causes him to take an illegal lunging attack after the judge proclaims a draw, and Ryunosuke, the better swordsman, parries and kills him with one stroke of his bokken. Ryunosuke flees town after killing the man, and cuts down many of the dead opponent's clansmen who attack him as he is leaving. His opponent's ex-wife asks to go along with him. To make a living, Ryunosuke joins the Shinsengumi, a sort of semi-official police force made up of rōnin that supports the Tokugawa shogunate through murder and assassinations.
The disgraced former executioner, or Kogi Kaishakunin to the shogun, Ogami Ittō is now living rough on the land with his three-year-old son Daigoro, travelling the countryside as a hired killer. Pushing his son in a baby cart, he stops at a bathhouse looking for a room and a bath, and is eagerly welcomed in by a young woman. However, the manager of the house sees Ittō as a dirty vagabond and scolds the young woman for letting him in the house. Overhearing this, Ittō goes to the baby cart and retrieves a bundle and hands it to the manager for safe keeping – it is 500 gold pieces, earned from a recent job. The manager's tone quickly changes, but when he tries to wash Daigoro's feet, the boy kicks water at the old man and tromps across the floor, leaving wet footprints.