The observation of the neuro-ophthalmic responses is an established basic tool for monitoring the depth of anesthesia in horses. However, their usefulness during balanced anesthesia in the clinic is questionable. The aim of this study was to compare the usefulness of neuro-ophthalmic responses during 3 commonly used anesthetic protocols. An observational study was performed on 22 horses, 5 maintained only with isoflurane (ISO group), 12 maintained with isoflurane and a constant rate infusion of ketamine (ISO+KET group), and 5 maintained with isoflurane and a constant rate infusion of detomidine (ISO+DET group). The occurrence of all five responses was noted: spontaneous palpebral and provoked palpebral response, corneal response, nystagmus and eye bulb rotation. The most consistent with the literature and most useful for anesthetic depth assessment was the observation of spontaneous palpebral reflex in all of the groups (p = 0.788) and the most significant differences were visible for the observation of the provoked palpebral reflex (p = 0.015) for all groups. The reflexes in the group ISO+DET were less consistent with the literature than in other groups, making them least useful. The group ISO seemed to have the reflexes’ observations most consistent with the literature expectations. The corneal reflex was always present and nystagmus always absent which confirmed the observations from literature. The above results suggest that neuro-ophthalmic responses, especially eye bulb rotation and provoked palpebral reflexes, should not be the only indicators in assessing the depth of anesthesia during maintenance with ketamine or detomidine with concurrent isoflurane.