Mrs. Frisby and the Rats of NIMH has been reissued in paperback under the title The Secret of NIMH, partly due to the film's name and because potential readers were deterred by a book centered on rats. O'Brien mentioned that people frequently questioned his choice to write about rats. He also pointed out that an early review of the book admitted the title initially discouraged the critic, but once past that hurdle, the review became one of the most enthusiastic. Despite this, rats are just a few of the novel's numerous engaging and believable characters, which also include humans, mice, owls, crows, and cats.

Mrs. Frisby and the Rats of NIMH narrates two stories at the same time: one follows Mrs. Frisby's efforts to save her home and her son Timothy from a severe illness; the other covers the rats of NIMH, their escape to the Fitzgibbon farm, and their plan to relocate to Thorn Valley. These stories are intertwined as Mrs. Frisby and the rats collaborate, each for their own reasons but towards similar goals.

Although it is never explicitly stated in the novel, NIMH stands for the National Institute of Mental Health, a government lab where some of these rats and mice were held for three years before their escape.

Mrs. Frisby and the Rats of NIMH delves into themes of loyalty, independence, and bravery, and explores the ultimate application of knowledge and science. The novel realistically presents both sides of issues and investigates the strengths and weaknesses of its characters. While the scientists of NIMH and Jenner, who deserts his fellow rats, act as antagonists in the story, they are not depicted as purely evil; both their positive and negative traits are portrayed. Everything, from scientific advancements to life at the Fitzgibbon farm, is shown as either beneficial or detrimental based on how it is used. Consequently, the novel explores life's complexities, where straightforward solutions are absent. To endure, characters must be brave and self-reliant, yet they also need to learn to depend on others for support.