Ralph is a young, adventurous talking mouse who lives with his family in a knothole in Room 215 of the Mountain View Inn. He finds his mother's constant worrying restrictive and yearns to explore the hotel beyond his small home. His deep fascination with human vehicles drives him to take dangerous risks, often prioritizing his own thrills over his family's safety.

Keith is a young boy from Ohio visiting California on a family road trip. He is imaginative and yearns for real independence, mirroring his mouse friend's desire for adventure. He possesses a toy motorcycle that magically moves when he makes engine noises, and he generously shares both his toys and his food with the hotel's hidden residents.

Mr. and Mrs. Gridley are Keith's parents, currently enduring a long family road trip from Ohio. Mr. Gridley is exhausted from driving and willing to compromise on accommodations, while Mrs. Gridley is highly anxious about the presence of mice in the Mountain View Inn. They care deeply for their son but impose restrictions that prompt his secretive behavior with his toys.

Matt is the 60-year-old caretaker and bellhop at the Mountain View Inn. Having worked at the hotel for many years, he is a calm, observant presence who knows the building's hidden rhythms. He possesses a rare, quiet openness that allows him to understand and communicate with the hotel's resident mice.

Ralph's Mother is a highly anxious mouse who lives in the wall of the Mountain View Inn. Following the tragic death of her husband from aspirin poisoning, she worries constantly about the safety of her children. She fears the hotel staff, vacuum cleaners, and the owls outside, preferring that her family remain quietly hidden at all times.

Uncle Lester is a male relative of Ralph who lives in the hotel walls. He is clever enough to establish a secure crumb-gathering spot in the housekeeper's office but generally behaves like a loudmouth. Despite his boastful nature, he actively avoids taking physical risks himself.

Aunt Sissy is a female relative of Ralph who resides in the hotel's bridal suite. She carries a slight sense of superiority due to her location but lives the same fearful, crumb-gathering existence as the rest of the mice. She maintains a strong disdain for humans despite eagerly enjoying the food they leave behind.

The Little Terrier is a small pet dog staying in Room 211 with his human caretaker. He possesses sharp senses and is highly aggressive toward the hotel's mice. His loud barking serves as a constant threat to Ralph's nighttime excursions, forcing the mouse to carefully plan his routes.

The Maid is a young summer employee at the Mountain View Inn. She is easily distracted and somewhat vain, often pausing her work to check her hair and makeup in the mirror. Her inattentive cleaning habits inadvertently create massive hazards for the tiny residents of the hotel.