The present study was conducted to evaluate the safety and the long-lasting anesthetic properties of Neosaxitoxin (NeoSTX) when injected into the intercoccygeal space in standing horses. A group of eighteen (n = 18) healthy client-owned mature crossbreed horses were enrolled in this study. Caudal epidural injection was ascertained by two methods, the "hanging drop" technique and radiographs. Horses were allocated into three groups (n = 6) each group receiving different dosages of NeoSTX. To assess the onset of action and duration of anesthesia, six horses received 20 μg in 2 mL, six 20 μg in 4 mL, and six 40 μg in 4 mL. Perineal sensitivity, tail and anal sphincter tone were evaluated to determine the anesthetic effect of NeoSTX. The safety of the toxin was assessed by conducting physical and neurological exams, chemistry panel, and complete blood count at different time points. Epidural injection of Neosaxitoxin showed significant differences in the onset of action and duration of anesthesia, displaying a long-lasting, dose-dependent nociceptive blocker effect with minor adverse effects. All horses did develop transient urine incontinence and fecal accumulation, which did not cause discomfort or changes in clinical parameters. This study demonstrated that Neosaxitoxin can be safely and efficiently used as an alternative for caudal epidural anesthesia and analgesia in horses, but more research is needed to evaluate its use in surgical procedures and pain control.