"Wake Up, Girl Zoo!" is a short-form anime series that blends comedy and slice-of-life elements, centering around anthropomorphic animals working at a struggling zoo. The narrative revolves around the daily lives and challenges faced by the zoo's staff, who are depicted as animal-human hybrids. The main characters include a penguin, a lion, a giraffe, and other zoo inhabitants, each with distinct personalities and roles within the facility. The penguin, often the central focus, is earnest but clumsy, striving to improve the zoo's reputation and attract visitors. The lion, confident and charismatic, takes on a leadership role, while the giraffe, gentle and reserved, provides emotional support to the group.

The plot unfolds through episodic vignettes, highlighting the characters' efforts to revitalize the zoo and overcome various obstacles, such as budget constraints, low attendance, and interpersonal conflicts. Humor arises from the characters' exaggerated traits and their interactions, often parodying workplace dynamics and idol culture. Despite the comedic tone, the series occasionally touches on themes of perseverance, teamwork, and the importance of preserving wildlife. The narrative maintains a lighthearted approach, with each episode presenting self-contained stories that contribute to the overarching goal of saving the zoo from closure. The series balances its comedic elements with moments of heartfelt sincerity, creating an engaging and accessible viewing experience.