These studies examined the effects of repeated intranasal administration of Cyclobenzaprine hydrochloride in rats and dogs. In rats, doses up to 1.05 mg/animal/day over 28 days showed no mortality or toxicity. Body weight, feed intake, ophthalmological and neurobehavioral assessments, and clinical pathology evaluations remained unaffected. Microscopic examinations revealed minimal non-adverse hyperplasia at the administration site in the nasal cavity. No histopathological changes were observed in other organs or tissues, establishing the NOAEL at 1.05 mg/animal/day. In dogs, doses up to 10.5 mg/animal/day over 14 days were well-tolerated, with only mild local irritation observed as nasal itching, which resolved quickly post-dosing. Body weights, food consumption, and comprehensive neurobehavioral assessments, including ECG examinations and reflex tests, showed no adverse effects. Hematological, clinical chemistry, and urinalysis variations were minimal and non-dose-dependent. Microscopic evaluations of the nasal cavity and other anatomical structures showed mild non-adverse changes, with no significant histopathological findings in the olfactory epithelium, olfactory bulb, or brain. These findings indicate that Cyclobenzaprine Hydrochloride Nasal Spray is well-tolerated with a NOAEL of 1.05 mg/animal/day in rats and ≥10.5 mg/animal/day in dogs, suggesting potential for safe intranasal administration in clinical use.