- Randle McMurphy: A free-spirited, rebellious con man, sent to the hospital from a prison work farm. He is guilty of battery and gambling. He had also been charged with — but never convicted of, due to the girl in question not wishing to testify so as not to implicate herself and her willingness to participate — statutory rape. McMurphy is transferred from a prison work farm to the hospital, thinking it will be an easy way to serve out his sentence in comfort. In the end, McMurphy attacks Nurse Ratched, sacrificing his freedom and his health in exchange for freeing the previously shackled spirits of the cowed patients on the ward.
Chief Bromden: The novel's half-Native American narrator has been in the mental hospital since the end of World War II. Bromden is presumed by staff and patients alike to be deaf and mute, and through this guise he becomes privy to many of the ward's dirtiest secrets. As a young man, the Chief was a high school football star, a college student, and a war hero. After seeing his father, a Native American chieftain, humiliated at the hands of the U.S. government and his white wife, Chief Bromden descends into clinical depression and begins hallucinating. Soon he is diagnosed with schizophrenia. He believes society is controlled by a large, mechanized system which he calls "The Combine."
- Richard Gray, author of A History of American Literature, said Bromden "supplies" the novel's "vision". Gray explains Bromden's "eye" "sees the inner truth" and Bromden "is an outsider, an innocent eye in a way like Huck Finn, but what he sees is far stranger, far more surreal." Gray explained Bromden's vision "may not be literally true but it is symbolically so because, to quote Emily Dickinson again, 'Much madness is divinest sense.'" The first chapter ends with the Chief pleading to the reader, "...you think this is too horrible to have really happened, this is too awful to be the truth! But, please. It's still hard for me to have a clear mind thinking on it. But it's the truth even if it didn't happen."
- Nurse Ratched (also known as "Big Nurse"): The tyrannical head nurse of the mental institution, who exercises near-total control over those in her care, including her subordinates. She will not hesitate to restrict her patients' access to medication, amenities, and basic human necessities if it suits her manipulative whims. Her favorite informant is the timid Billy Bibbit, whom she coerces into divulging the unit's secrets by threatening to complain about him to his mother. McMurphy's fun-loving, rebellious presence in Ratched's institution is a constant annoyance, as neither threats nor punishment nor shock therapy will stop him or the patients under his sway. Eventually, after McMurphy nearly chokes her to death in a fit of rage, Nurse Ratched has him lobotomized. However, the damage has already been done, and Nurse Ratched's rule is broken after McMurphy's attack leaves her nearly unable to speak, which renders her unable to intimidate her patients, subordinates and superiors.
- The "Black Boys" Washington, Williams, and Warren: Three black men who work as aides in the ward. Williams is a dwarf, his growth supposedly “stunted after witnessing his mother being raped by white men.” The Chief says Nurse Ratched hired them for their sadistic nature.
- Geever: the swing shift aide.
- Dr. John Spivey: The ward doctor. Nurse Ratched drove off other doctors, but she kept Spivey because he always did as he was told. Harding suggests that the nurse could threaten to expose him as a drug addict if he stood up to her. McMurphy's rebellion inspires him to stand up to Nurse Ratched.
- Nurse Pilbow: The young night nurse whose face, neck, and chest are stained with a profound birthmark. A devout Catholic who fears sinning, she blames the patients for infecting her with their evil and takes it out on them.
- Mr. Turkle: An elderly African American aide who works the late shift in the ward. He agrees to let McMurphy host a party and sneak in prostitutes one night.
- The Japanese Nurse: The nurse in charge of the upstairs disturbed ward, for violent and unmanageable patients. She is kind and openly opposes Nurse Ratched's methods.
The acutes are patients who officials believe can still be cured. With few exceptions, they are there voluntarily, a fact that angers McMurphy when he first learns of it, then later causes him to feel further pity for the patients, thus further inspiring him to prove to them they can still be strong despite their seeming willingness to be weak.
- Billy Bibbit: A nervous, shy, and boyish patient with an extreme speech impediment, Billy cuts and burns himself, and has attempted suicide numerous times. Billy has a fear of women, especially those with authority such as his mother. To alleviate this, McMurphy sneaks a prostitute into the ward so Billy can lose his virginity. The next morning, Nurse Ratched threatens to tell his mother; fearing the loss of his mother's love, Billy has an emotional breakdown and commits suicide by cutting his own throat.
- Dale Harding: The unofficial leader of the patients before McMurphy arrives, he is an intelligent, good-looking man who's ashamed of his repressed homosexuality. Harding's beautiful yet malcontent wife is a source of shame for him.
- George Sorensen: A man with germaphobia, he spends his days washing his hands in the ward's drinking fountain. McMurphy manages to persuade him to lead a fishing expedition for the patients after discovering he had captained a PT boat during World War II. Afterward, the three black boys maliciously forcibly delouse him, cruelly knowing the mental anguish this will cause him.
- Charlie Cheswick: A loud-mouthed patient who always demands changes in the ward, but never has the courage to see anything through. He finds a friend in McMurphy, who's able to voice his opinions for him. At one point McMurphy decides to fall in line when he learns his stay in the ward is indefinite and his release is solely determined by the Big Nurse. As a result, Cheswick drowns himself in the ward's swimming pool when he decides he himself will never escape the relentless Big Nurse.
- Martini: A patient who suffers from severe hallucinations.
- Scanlon: A patient obsessed with explosives and destruction. He is the only other non-vegetative patient confined to the ward by force aside from McMurphy and Bromden; the rest can leave at any time.
- Jim Sefelt and Bruce Fredrickson: Two epileptic patients. Sefelt refuses to take his anti-seizure medication, as it makes his teeth fall out and as such makes him self-conscious over his appearance. Fredrickson takes Sefelt's medication as well as his own because he is terrified of the seizures, and loses teeth due to the resulting overdosage.
- Max Taber: An unruly patient who was released before McMurphy arrived. The Chief later describes how, after he questioned what was in his medication, Nurse Ratched had him "fixed."
The chronics are patients who will never be cured. Many of the chronics are elderly and/or in vegetative states.
- Ruckly: A hell-raising patient who challenged the rules until the Big Nurse authorized his lobotomy. After the lobotomy, he sits and stares at a picture of his wife, and occasionally screams profanities.
- Ellis: Ellis was put in a vegetative state by electroshock therapy. He stands against the wall in a disturbing messianic position with arms outstretched.
- Pete Bancini: Bancini suffered brain damage at birth but managed to hold down simple jobs, such as a switch operator on a lightly-used railroad branch line, until the switches were automated and he lost his job, after which he was institutionalized. The Chief remembers how once, and only once, he lashed out violently against the aides, telling the other patients that he was a living miscarriage, born dead.
- Rawler: A patient on the Disturbed ward, above the main ward, who says nothing but "loo, loo, loo!" all day and tries to run up the walls. One night, Rawler castrates himself while sitting on the toilet and bleeds to death before anyone realizes what he has done.
- Old Blastic: An old patient who is in a vegetative state. The first night McMurphy is in the ward, Bromden dreams Blastic is hung by his heel and sliced open, spilling his rusty visceral matter. The next morning, Bromden learns Blastic died during the night.
- The Lifeguard: An ex-professional football player, he still has the cleat marks on his forehead from the injury that scrambled his brains. He explains to McMurphy, unlike prison, patients are kept in the hospital as long as the staff desires. It is this conversation that causes McMurphy to fall in line for a time.
- Colonel Matterson: The oldest patient in the ward, he suffers from severe senile dementia and cannot move without a wheelchair. He is a veteran of World War I, and spends his days "explaining" objects through metaphor.
- Candy: The prostitute McMurphy brings on the fishing trip. Billy Bibbit has a crush on her and McMurphy arranges a night for Candy to have sex with him.
- Sandra: Another prostitute and friend of Candy and McMurphy. She and Sefelt sleep together on the night she and Candy are sneaked into the ward late one night. Sefelt has a seizure while they are fornicating.
- Vera Harding: Dale Harding's wife. Described as an attractive lady with very large breasts. She is a primary cause of concern for Dale, who often worries about her fidelity. She reveals to the patients that actually Dale himself has affairs - with other men.