Measurement of Intraocular Pressure Using Rebound Tonometry in Anesthetized Free-ranging South American Sea Lions (Otaria byronia)

Ocular disease occurs in both free-ranging pinnipeds and those in captivity under professional care. Keratopathy, cataracts, trauma, neoplasia, amyloid deposition, and fungal disease are commonly reported concerns (Hanson et al. 2009; Colitz et al. 2010a, b). Measurement of intraocular pressure (IOP) is part of any complete ophthalmologic examination, yet sparse reference data exist from wild marine mammals. Rebound tonometry is a practical, affordable, and easier method of IOP data collection compared to applanation tonometry in veterinary medicine, because it does not require topical anesthesia, making it well suited for application in awake patients (Cervino 2006). Intraocular pressure measurements obtained via rebound tonometry in awake cetaceans and California sea lions (Zalophus californianus) under professional care were higher than those reported for domestic terrestrial mammals (Colitz 2012; Mejia-Fava 2014). However, the impact of artificial aquatic environments on IOPs has not been evaluated, and these reference data may be inherently not representative of true normal values. Although effective examination of adult pinnipeds in the wild requires anesthesia, our null hypothesis was that reference intervals for free-ranging sea lions would be similar to those established for animals under professional care.

Punta San Juan (PSJ), a 54 ha marine protected area on Peru's southern coast, protects key rookeries of South American sea lions (Otaria byronia [OB]), which are locally endangered due to competition with commercial fisheries for prey, bycatch concerns, El Niño climatic events, disease epidemics, and oil spills (Cárdenas-Alayza 2016). The importance of the PSJ rookeries for the conservation of these species has prompted ongoing population health assessments at this location. Utilizing animals that were already being anesthetized for medical evaluations, our objective was to obtain IOP values using rebound tonometry in free-ranging OBs anesthetized using a common combination of induction agents that can be directly referenced when evaluating other anesthetized pinnipeds, and to evaluate the impact of body weight, sex, and year of data collection on results.

All work was performed at the PSJ marine protected area, Ica, Peru (15°22′S, 75°12′W) between 2011 and 2018 authorized under Peruvian permits 023-2011-, 022-2012-, 09-2013-, 24-2014-, 008-2015-, and 019-2016-SERNANP-RNSIIPG. Fifty-six (37 male, 19 female) unique individual OBs were anesthetized with injectable medetomidine (57±16 μg/kg), midazolam (0.33±0.1 mg/kg), and butorphanol (0.34±0.1 mg/kg) (MMB, Zoopharm, Windsor, Colorado, USA) with a consistent ratio of 1:6:6 of MMB on a milligram basis, delivered intramuscularly via dart (Daninject, Austin Texas, USA). Seven animals received supplementation with inhaled isoflurane, but IOP measurement was performed prior to isoflurane administration. Anesthesia was antagonized with flumazenil (WestWard Pharmaceuticals, Eatontown, New Jersey, USA, 1 mg per 300 mg midazolam intramuscularly), atipamezole (Zoopharm, 1 mg per 0.33 mg of medetomidine intramuscularly), and naltrexone (Zoopharm, 1 mg per 1.2 mg of butorphanol intramuscularly). Animals were allowed to fully recover prior to release at the site of capture.

Animals were determined to be in normal health based on physical examination and hematology and plasma biochemistry results. Unaided visual examinations of the ocular adnexa and cornea were performed while the eyes were naturally centrally rotated. Examinations were performed in bright outdoor ambient light, precluding comprehensive examination of all ocular structures and preventing examination of the posterior aspect of the globe. Animals with any gross ocular abnormalities were not included in data analysis.

We measured IOP using rebound tonometry (TonoVet^®, Icare^® Finland Oy, Vantaa, Finland) using the instrument's dog setting while animals were in ventral recumbency on a flat, level surface. The tonometer was consistently held in a horizontal position, with the probe parallel to the ground and starting 4–8 mm perpendicular to the corneal surface. The palpebrae were gently stabilized using care to not apply any pressure to the globe. All IOP measurements were obtained by the same examiner. Six individual measurements were obtained from the central cornea bilaterally with the tonometer calculating an average value after excluding the highest and lowest values. Results with deviations of ≤1.0 mmHg were accepted.

Mean (SD) IOPs in males and females are reported in Table 1. Pearson's correlation coefficient revealed no association between body weight and IOP and thus was not included in the following model. Multivariate analysis with IOP for each as the dependent variable and sex, year, and sex×year interaction was performed using commercial software (RStudio, Boston, Massachusetts, USA). No model had support for either eye, thus univariate analysis was performed to test the differences between sex (independent samples t-test) and year (one-way analysis of variance) in SPSS (version 24, IBM statistics, Chicago, Illinois, USA). Males had a significantly higher IOP in the right eye compared to females (P=0.02), but there was no difference in IOP of the left eye between sexes (P=0.096; Table 1). The IOP in the left eye was significantly different between years (P=0.01) with the only significant post hoc finding of higher IOP in 2014 compared to 2018 (P=0.038; Fig. 1). There was no difference in IOP in the right eye between years (P=0.054; Fig. 1).

Using similar methods, Mejia-Fava (2014) defined IOP ranges for awake, adult male California sea lions managed under professional care as 32.8±5.31 mmHg, right eye, and 34.3±4.86 mmHg, left eye (Mejia-Fava 2014). Use of the TonoVet at the instrument's horse setting was proposed based on a closer correlation in corneal thickness between equids and pinnipeds, but no studies have compared the accuracy of the settings to a gold standard (Mejia-Fava 2014). Given the similarity of our results using the instrument's dog setting, further investigation of the impact of this setting is indicated. Overall, IOP measurements obtained via rebound tonometry in both species of sea lions are higher than that reported in terrestrial mammals including dogs and horses (Gelatt and MacKay 1998; Knollinger 2005; Mejia-Fava 2014).

The use of either manual or chemical restraint, both of which can influence IOP measurement, to measure IOP in wild pinnipeds cannot be avoided. Manual restraint falsely elevates IOP measurement in other mammals by compression of vasculature in the neck (Klein et al. 2011). Previous studies in humans and dogs show variable results regarding how different anesthesia protocols affect IOP measurements (Kornbleuth 1964; Ghaffari et al. 2010; Rauser et al. 2016). Alpha-2 agonists (e.g., medetomidine) have minimal to no effect on IOP values in dogs and cats (Malmasi and Ghaffari 2016; Rauser et al. 2016). Midazolam also has little effect on IOP values in humans, and a similar lack of effect was noted in dogs given midazolam in combination with ketamine (Carter et al. 1999; Ghaffari et al. 2010). Butorphanol in combination with an alpha-2 agonist causes a transient increase and subsequent decrease in IOP, making its potential effect in our study harder to interpret (Rauser et al. 2012). Further studies measuring IOP in awake, compliant pinnipeds under professional care before and during anesthesia are warranted to determine the effect of anesthesia.

The challenges of working with free-ranging wildlife in a field environment created several additional, unavoidable limitations for our study. Field conditions, lack of biomicroscopy, and ambient light may have allowed some ocular lesions to be missed. Miosis from the strong ambient light further precluded evaluation of the lens or posterior chamber of the globe. Mydriatics are not effective in pinnipeds, and paralytics were not practical because animals needed to immediately navigate in the environment upon release (Miller et al. 2010). Standardization of data collection times relative to anesthesia was not possible due to immobilization logistics and priority of primary research projects.

Ocular diseases in free-ranging and professionally managed pinnipeds can result in decreased IOP from uveitis (caused by infection, cataracts, and trauma) or increases in IOP (caused by glaucoma and neoplasia). Established, normal values for IOP in pinniped species are important for the assessment and treatment of these conditions. Future research measuring the IOP in pinnipeds with clinical ocular pathology is warranted. The rebound tonometer appears to be an effective method for measuring IOP in anesthetized, free-ranging adult OB.