Ординатура / Офтальмология / Английские материалы / Advances in Understanding Mechanisms and Treatment of Infantile Forms of Nystagmus_Leigh, Devereaux_2008

VALLABH E. DAS

ABSTRACT

Alternating fixation is frequently observed in humans with strabismus. We examined this phenomenon in a monkey model for strabismus. An exotropic strabismus was induced in two infant monkeys using an alternate monocular occlusion paradigm for the first four months of life. When the animals were about three years old, we measured binocular eye movements, using the scleral search coil technique, as they performed a visually guided saccade task during monocular or binocular viewing. During binocular viewing, monkeys tended to fixate targets in the right hemifield with their right eye and targets in the left hemifield with their left eye, consistent with patterns of visual suppression in exotropia. An alternating saccade was executed when the target jumped across the midline. There were no significant differences in the amplitude–peak velocity or amplitude– duration main-sequence relationships between alternating and nonalternating saccades (monocular or binocular viewing). Saccade latency tended to be greater during binocular viewing than during monocular viewing. Our study establishes a strabismus monkey model useful for studying neural circuits involved in generating alternating fixation and alternating saccade behavior. Further, alternating saccade behavior may be used as a probe to study mechanisms of visual suppression in strabismus and mechanisms of target selection in normal individuals.

Loss of sensory or motor fusion early in postnatal development leads to binocular misalignment (strabismus).1 Various studies conclude that infantile forms of strabismus occur in as much as 5% of all children.2 For the last few years, our laboratory has pursued studies of eye movements in a primate model for sensory strabismus.3-6 In addition to horizontal misalignment, these strabismic animals also displayed A or V patterns of horizontal deviations and dissociated vertical deviation (DVD), all commonly observed in humans with strabismus.1 Investigation of motoneuron activity in the oculomotor nucleus identified a neural drive for abnormal eye movements associated with the A/V patterns and DVD. Our strategy thus far has been to utilize concepts, tools, and techniques derived from studies of binocular coordination in normals and apply them to examine binocular mechanisms in the strabismic monkey model.

Following along the lines of this general strategy, we examined new eye movement data regarding another strabismus phenomenon—alternating fixation—that we had observed in our animal model and were interested in exploring further. Some humans with strabismus, usually those who do not suffer from significant amblyopia, have the ability to fixate a target of interest with either eye and are often capable of spontaneously alternating (or switching) the eye of fixation. A saccadic eye movement in which the eye of fixation is switched is called an alternating saccade. The ability to alternate the eye of fixation depends on visual suppression mechanisms7 In exotropic strabismus, it is usually the temporal retina that is suppressed.8-11 It follows that when the

strabismic individual switches the eye of fixation, visual suppression must also be switched from one eye to the other. Steinbach studied this phenomenon and showed that switching of visual suppression coincided with the saccadic eye movement.7 This raises some questions about how an alternating saccade is triggered. In perhaps the only study in the literature that examined characteristics of alternating saccades in humans with exotropia, van Leeuwen et al.12 showed that some metrics of alternating and nonalternating saccades were similar. However, they chose a predictable selfgenerated horizontal saccade task and analyzed only the amplitude–peak velocity relationship in their subjects. Also, the patients in that study had already undergone strabismus correction surgery, which may have altered patterns of saccade alternation.

There were two main goals for our study. First, we wanted to establish our animal model as effective in producing alternating fixation and alternating saccade behavior. A second goal was to compare metrics of alternating and nonalternating saccades over a large range of horizontal and vertical amplitudes and orbital positions. A potential advantage of studying alternating behavior in a monkey model is that the animal would not have undergone strabismus surgery. Establishing a monkey model opens the avenue for identification of neural substrates involved in generating alternating saccade behavior using conventional neurophysiological methods. Some of these results have appeared before in abstract form.13

METHODS

Subjects and Rearing Paradigms

Behavioral data were collected from two strabismic (S1 and S2) juvenile rhesus monkeys (Macaca mulatta), each weighing between 8 and 11 kg. Monkeys with strabismus were reared at the Yerkes National Primate Research Center using an alternate monocular occlusion (AMO) paradigm.3,14 In the AMO rearing procedure, soon after birth (within the first 24 hours), an occluding patch (either opaque goggles or dark contact lenses) is placed in front of one eye for a period of 24 hours and thereafter switched to the other eye for the next 24 hours. The patch is alternated daily for a period of 4 to 6 months. In this method, binocular vision is severely disrupted during the first few months of life, the critical period during which the monkeys would normally develop proper eye alignment, stereovision, and binocular sensitivity in the brain.15-17

Surgical Procedures and Eye Movement

Measurements

Following special rearing, the animals were allowed to grow normally until they were approximately three

years of age. Sterile surgical procedures carried out under aseptic conditions using isoflurane anesthesia (1.25% to 2.5%) were used to stereotaxically implant a head-stabilization post. In the same surgery, a scleral search coil was also implanted in one eye using the technique of Judge et al.18 Later, in a second surgery, a second scleral search coil was implanted in the other eye. All procedures were performed in strict compliance with National Institutes of Health guidelines, and the protocols were reviewed and approved by the Institutional Animal Care and Use Committee at Emory University.

Binocular eye position was measured using the magnetic search coil method (Primelec Industries, Regensdorf, Switzerland).19,20 Calibration of the eyecoil signal was achieved by giving the monkey a small amount of juice or some other reward when the animal looked within a small region (±2° window) surrounding a 0.25° target spot that was rear-projected on a tangent screen 60 cm from the animal. All stimuli were under computer control. Calibration of each eye was performed independently during monocular viewing.

Experimental Paradigms and Data Analysis

Eye movement data were collected as the strabismic animals performed a visually guided saccade task in which the target appeared at random horizontal or vertical locations within a ±15° grid. Data were collected during both monocular and binocular viewing in separate experimental sessions. Binocular eye-position and target-position feedback signals were digitized at 1 kHz with 16-bit precision (PCI-6025E NI-DAQ board and LabVIEW software; National Instruments, Austin, Texas). The analysis of the saccade data was carried out using custom software built in MATLAB (MathWorks, Natick, MA). Velocity and acceleration signals were generated by digital differentiation of the position signal using a central difference algorithm. Position, velocity, and acceleration signals were filtered using software FIR filters (80 points; 0 to 80 Hz passband), also designed in MATLAB. Saccade onset was automatically determined by the software as the first time point at which eye acceleration was greater than 3 standard deviations from control-eye acceleration (eye acceleration during a period of fixation prior to the saccade), and saccade offset was determined as the last time point at which eye deceleration was less than 3 standard deviations from the same mean eye acceleration. Though detection of saccade onset and offset was automated, the investigator visually examined the velocity and acceleration traces of every saccade and had the option of either accepting or changing the computer selection. Typically, less than 10% of the computer’s marks were changed by the investigator. For the binocular viewing data, the investigator also made the

determination of whether the saccade was of the alternating/nonalternating variety, and this information was recorded along with the saccade parameters.

Following data collection and identification of saccade onset and offset, the data were parsed into six bins, depending on viewing condition and saccade type:

1.saccades during monocular right-eye fixation (OD);

2.saccades during monocular left-eye fixation (OS);

3.binocular viewing nonalternating saccades with the right eye fixating (OURR);

4.binocular viewing nonalternating saccades with the left eye fixating (OULL);

5.binocular viewing alternating saccades where the eye of fixation was switched from right eye to left eye (OURL); and

6.binocular viewing alternating saccades where the eye of fixation is switched from left eye to right eye (OULR).

Saccade metric parameters for each eye were calculated including amplitude, latency, peak velocity, and duration. Amplitude–peak velocity and ampli- tude–duration main-sequence relationships were plotted, and data were fit according to well-established equations. An exponential curve was fit to the ampli- tude–peak velocity data, and a linear regression was developed for the amplitude–duration data. Fitting was performed in MATLAB using routines available in the “curvefit” toolbox.

For saccade latency data, we developed histograms of the inverse of saccade latency and fitted a Gaussian curve to this data. We used the inverse of latency for fitting the Gaussian, because it has been shown that this parameter better represents a Gaussian process than saccade latency directly.21-23 The mean and standard deviation of this Gaussian fit was compared across the six saccade conditions using ANOVA.

RESULTS

The two animals included in the study were both exotropic. During right-eye viewing of a straightahead target, S1 showed an exotropia of 10°, and S2 an exotropia of 11°. During left-eye viewing of the same target, S1 showed an exotropia of 15°, and S2 an exotropia of 14°. A-patterns were observed in both animals and were similar to those observed in other animals reared using the AMO paradigm.3 The main focus of the study was alternating fixation behavior. Figure 6.1 shows some raw saccade traces during binocular viewing that illustrate the property of alternating saccades in these animals.

Figure 6.1 Panels (A) and (B) are examples of alternating saccades in exotropic animal S1 during a binocular viewing task. The eye of fixation is switched from the left eye (gray line) to the right eye (black line) as a consequence of the saccade (A). Target position is shown by the black dotted line. The eye of fixation is switched from right eye to left eye (B). (C) and (D) show examples of nonalternating saccades with either the right eye (C) or the left eye (D). OULR, binocular viewing alternating saccades with fixation switch from left to right eye; OURL, binocular viewing alternating saccades with fixation switch from right to left eye; OURR, binocular viewing nonalternating saccades with the right eye fixating; OULL, binocular viewing nonalternating saccades with the left eye fixating.

Spatial Pattern of Saccade Alternation

Previous studies have shown that in humans with exotropia, the temporal retina of the nonfixating eye is suppressed. We found a corollary in the spatial pattern of alternation behavior within the saccade task that was used. Figure 6.2 plots a spatial view of saccade beginning and ending locations where we observed alternating or nonalternating behavior during the binocular viewing random-saccade task in animal S1. Figures 6.2A and 6.2B show nonalternating saccades with either the right eye or the left eye fixating (OURR and OULL). Figures 6.2C and 6.2D show alternating saccades, where the eye of fixation was switched from right to left (OURL) or from left to right (OULR). In both animals, we observed that targets in the right hemifield were fixated by the right eye and targets in the left hemifield were fixated by the left eye. Therefore, target jumps that began and ended either in the right or left hemifield tended to be nonalternating saccades. When the target jumped across the midline (i.e., from the right to the left as in Figure 6.2C, or from left to right as in Figure 6.2D), an alternating saccade was generated. Approximately 1000

binocular viewing saccades (OURR, OULL, OURL, and OULR) and 1000 monocular viewing saccades (OD and OS) were collected in each animal. Of the binocular viewing saccades, 37% in monkey S1 and 25% in monkey S2 were of the alternating variety.

Comparison of Saccade Main-Sequence

Relationships

The next step was to analyze the metrics of alternating saccades and compare them to nonalternating saccades

and monocular viewing saccades. Figure 6.3 shows the amplitude–peak velocity main sequences from animals S1 and S2 during right-eye viewing for the six different saccade types (also see color insert). The figure illustrates that there is considerable overlap in the data for the different saccade types. We fit each set of data using an “exponential rise to maximum” curve. Figures 6.3A and 6.3B plot the estimated curves, along with the 95% prediction intervals. There is significant overlap in the prediction intervals, which suggests that there was no difference between the saccade types.

We also examined the amplitude–duration relationship in the different types of saccades in two animals. Figures 6.3C and 6.3D show these relationships. A linear fit was applied to the amplitude–duration data, and the regression line, along with 95% prediction intervals, is plotted. Once again there is significant overlap in the data and the prediction intervals among the different saccade types, which suggests that the saccade data in the various categories all came from the same population.

In summary, we did not observe any consistent differences in the saccade main-sequence relationship that suggested that alternating saccades were different from nonalternating saccades. Similarly, binocular viewing saccades did not show any consistent differences from monocular viewing saccades.

Comparison of Saccade Latency

We continued our analysis of saccade metrics by comparing saccade latency across the different saccade types. In normal humans and animals, a histogram of saccade latency follows a skewed distribution, with a rapid rise followed by a long tail-off. Carpenter et al.,23 in their Linear Approach to Threshold with Ergodic Rate (LATER) model for decision making, showed that the reciprocal of the saccade latency is representative of a Gaussian process. Therefore, for each of the saccade types in this study, we developed histograms of the inverse of saccade latency and then fit a Gaussian to the data. Figures 6.3E and 6.3F plot a histogram of the inverse of saccade latency for the different saccade types, along with the Gaussian fits. In animal S2, we found that saccades during binocular viewing were of significantly longer latency than saccades during either monocular viewing condition (difference in means, 8 to 15 milliseconds; one-way ANOVA; p > 0.05). In animal S1, we found that saccades during binocular viewing were of significantly longer latency than saccades during monocular left-eye viewing (difference in means, 7 to 18 milliseconds; one-way ANOVA; p > 0.05) but not in monocular right-eye viewing. Latency of nonalternating saccades was not consistently different from that of alternating saccades.

DISCUSSION

In this study, we sought to analyze alternating saccade behavior from the point of view of (a) establishing the AMO as a suitable animal model to study alternating fixation behavior and (b) comparing metrics of alternating saccade behavior to nonalternating saccades and monocular-viewing saccades.

Animal Model for Alternating Fixation Behavior

We have shown that AMO animals with exotropia demonstrated alternating fixation and alternating saccade behavior similar to that previously reported in humans11,12 and in monkeys with surgical exotropia.24 Thus the AMO strabismus model is appropriate for examining visual and ocular motor mechanisms that drive alternating saccade behavior. Even though we have not directly measured the degree of visual suppression at different parts of retina in the viewing and nonviewing eye, our results appear to suggest that the spatial pattern of saccade alternation is consistent with the pattern of visual suppression that is normally seen in exotropes.8-10 Therefore, alternating fixation and alternating saccade behavior may be used as probes to study visual suppression mechanisms in strabismus24 and how they may relate to developing a command signal for generating a saccade that brings one or the other eye onto the target. Some studies have shown that visual suppression is not complete.1,25 This might lead to a situation where the brain encodes two internal target representations (one for each eye) and must make a choice to generate a saccade that would bring either the previously fixating eye (nonalternating saccade) or the previously nonfixating eye (alternating saccade) onto the target. Therefore, studying these issues and understanding brainstem or cortical neural circuitry that might be involved in generating alternating saccades might also provide clues as to how selection between multiple target choices is carried out normally.

Metrics of Alternating Saccades Are Normal

It was not surprising that the main-sequence relationships of alternating and nonalternating saccades (monocular or binocular viewing) are similar. Perhaps it was more surprising that a systematic evaluation of the different saccade metrics had not been attempted previously. The basic implication of our result is that alternating and nonalternating saccades are governed by the same brainstem pulse-generation circuit. Any potential differences between alternating and nonalternating saccades must be examined upstream from the brainstem pulse generator.

Saccade Latency Differences in Alternating

Saccade Behavior

Our analysis suggested a tendency in monocular viewing saccades (OD and OS) to be of shorter latency than binocular viewing saccades (OURR, OULL, OULR, and OURL). This result may in fact be an indication that visual suppression is not complete. Previous studies that have examined target selection in normal animals

have consistently shown that saccadic latency is greater when multiple targets are presented to the subject as compared to a single target.26 In our experiments, during binocular viewing, the monkey may be presented with two retinal-error representations (one from each eye, which is equivalent to presenting two targets to a cyclopean eye). Even though the retinal-error representation from the partially suppressed eye is much weaker, the brain must still decide which eye to fixate on the target and must generate an appropriately sized saccade. The superior colliculus is implicated in target selection.27 The superficial layers of the superior colliculus may therefore be an appropriate structure to examine if, in fact, there are representations of visual error corresponding to each eye.

There was, however, no clear tendency for latency differences between alternating and nonalternating saccades. In the LATER model for saccade latency developed by Carpenter et al.,23 latency is proportional to the time taken for a decision signal to rise to a threshold value and trigger a saccade. In this model, the linearly increasing decision signal varies from trial to trial in a Gaussian manner. Presumably, in our experiments, the rate of rise of the decision signal is governed by strengths of competing inputs from both eyes. Therefore, distribution of latency may be governed more by the relative probability of generating an alternating or nonalternating saccade at a particular orbital position than whether the saccade itself was of the alternating or nonalternating variety. Therefore, in our exotropic monkeys, binocular viewing saccades that ended either far to the left or far to the right would probably have similar latencies, irrespective of whether they were alternating or nonalternating. However, saccades that ended in the middle of the ocular motor range may have longer latencies, because the probability of generating an alternating or nonalternating saccade is relatively close. A systematic evaluation of saccade latency across the two-dimensional ocular motor range may help to test this hypothesis.

ACKNOWLEDGMENTS We thank Sunal Patel, Tushar Jha, and Michelle Swann for their help with data collection and analysis. This work was supported by NIH RO-1 EY015312 (VED) and Yerkes Base Grant RR00165.

References

1.von Noorden GK, Campos EC. Binocular Vision and Ocular Motility: Theory and Management of Strabismus. 6th ed. St. Louis, MO: Mosby; 2002.

2.Lorenz B. Genetics of isolated and syndromic strabismus: facts and perspectives. Strabismus, 2002;10:147–156.

3.Das VE, Fu LN, Mustari MJ, Tusa RJ. Incomitance in monkeys with strabismus. Strabismus. 2005;13(1):33–41.

4.Das VE, Mustari MJ. Correlation of cross-axis eye movements and motoneuron activity in nonhuman primates with A-pattern strabismus. Invest Ophthalmol Vis Sci. 2007;48(2):665–674.

5.Das VE, Ono S, Tusa RJ, Mustari MJ. Conjugate adaptation of saccadic gain in non-human primates with strabismus. J Neurophysiol. 2004;91(2): 1078–1084.

6.Fu LN, Tusa RJ, Mustari MJ, Das VE. Horizontal saccade disconjugacy in strabismic monkeys.

Invest Ophthalmol Vis Sci. 2007;48:3107–3114.

7.Steinbach MJ. Alternating exotropia: temporal course of the switch in suppression. Invest Ophthalmol Vis Sci. 1981;20(1):129–133.

8.Horton JC, Hocking DR, Adams DL. Metabolic

mapping of suppression scotomas in striate cortex of macaques with experimental strabismus. J Neurosci. 1999;19(16):7111–7129.

9.Joosse MV, Esme DL, Schimsheimer RJ, Verspeek SA, Vermeulen MH, van Minderhout EM. Visual evoked potentials during suppression in exotropic and esotropic strabismics: strabismic suppression objectified. Graefes Arch Clin Exp Ophthalmol. 2005;243(2):142–150.

10.Joosse MV, Simonsz HJ, van Minderhout EM, Mulder PG, de Jong PT. Quantitative visual fields under binocular viewing conditions in primary and consecutive divergent strabismus. Graefes Arch Clin Exp Ophthalmol. 1999;237(7):535–545.

11.Sireteanu R. Binocular vision in strabismic humans with alternating fixation. Vision Res. 1982; 22(8):889–896.

12.van Leeuwen AF, Collewijn H, de Faber JT, van der Steen J. Saccadic binocular coordination in alternating exotropia. Vision Res. 2001;41 (25–26):3425–3435.

13.Patel SS, Jha RT, Swann MH, Das VE. Saccade alternation in non-human primates with strabismus. SFN Abstr. 2006;736.10..

14.Tusa RJ, Mustari MJ, Das VE, Boothe RG. Animal models for visual deprivation-induced strabismus and nystagmus. Ann N Y Acad Sci. 2002;956: 346–360.

15.Harwerth RS, Smith EL 3rd, Crawford ML, von Noorden GK. Behavioral studies of the sensitive periods of development of visual functions in monkeys. Behav Brain Res, 1990;41(3): 179–198.

16.O’Dell C, Boothe RG. The development of stereoacuity in infant rhesus monkeys. Vision Res. 1997;37(19):2675–2684.

17.Boothe RG, Dobson V, Teller DY. Postnatal development of vision in human and nonhuman primates. Annu Rev Neurosci, 1985;8:495–545.

18.Judge SJ, Richmond BJ, Chu FC. Implantation of magnetic search coils for measurement of eye position: an improved method. Vision Res. 1980;20:535–538.

19.Fuchs AF, Robinson DA. A method for measuring horizontal and vertical eye movement chronically in the monkey. J Appl Physiol. 1966;21(3): 1068–1070.

20.Hess BJ, Van Opstal AJ, Straumann D, Hepp K. Calibration of three-dimensional eye position using search coil signals in the rhesus monkey. Vision Res. 1992;32(9):1647–1654.

21.Carpenter RH. Contrast, probability, and saccadic latency: evidence for independence of detection and decision. Curr Biol. 2004;14(17): 1576–1580.

22.Reddi BA, Asrress KN, Carpenter RH. Accuracy, information, and response time in a saccadic decision task. J Neurophysiol. 2003;90(5): 3538–3546.

23.Carpenter RH, Williams ML. Neural computation of log likelihood in control of saccadic eye movements. Nature. 1995;377(6544):59–62.

24.Economides JR, Adams DL, Jocson CM, Horton JC. Ocular motor behavior in macaques with surgical exotropia. J Neurophysiol. 2007;98(6): 3411–3422.

25.Wensveen JM, Harwerth RS, Smith EL 3rd. Clinical suppression in monkeys reared with abnormal binocular visual experience. Vision Res. 2001;41(12):1593–1608.

26.McPeek RM, Keller EL. Short-term priming, concurrent processing, and saccade curvature during a target selection task in the monkey. Vision Res. 2001;41(6):785–800.

27.McPeek RM, Keller EL. Deficits in saccade target selection after inactivation of superior colliculus. Nat Neurosci. 2004;7(7):757–763.

ABSTRACT

Studies in humans and animals have shown that the cerebellum plays a vital role in calibrating eye movements to maintain gaze stability. In this chapter, we focus on the function of the cerebellar nodulus and uvula in stabilizing the foveal image when the object of regard is moving (smooth pursuit) and during linear motion of the head (the translational vestibulo-ocular reflex [TVOR]). Eye movements of two rhesus monkeys were recorded using the magnetic field search-coil method, before and after surgical ablation of the nodulus and uvula. Key findings included the following: (a) there was an asymmetric deficit of vertical smooth pursuit—downward, but not upward, pursuit was impaired; (b) the vertical TVOR was reduced for both upward and downward translation; and (c) the sustained, but not the initial, response to abrupt horizontal translations was impaired. Our results support a critical role for the nodulus and uvula in the control of vertical ocular following reflexes, and in the horizontal TVOR, possibly acting as an integrator of head acceleration.

The goal of gaze control is to stabilize images on the retina, in particular the fovea, in order to maximize visual acuity. This is accomplished by the eye movement control systems. When the head and the object being viewed are still, the fixation system holds the eyes steady in the orbit. When the object is moving, the pursuit system must determine the speed and direction of target motion and generate the appropriate

eye velocity to keep the target’s image on the fovea. Finally, when the head is moving (rotating or translating), the vestibular system calculates head motion and generates an eye movement of the appropriate amplitude and in the opposite direction, in order to stabilize the retinal (foveal) image. Finally, the vergence system maintains alignment of the two eyes.

The cerebellum plays a vital function in the control and calibration of all types of eye movements and thus is critical to optimizing gaze. In fact, impaired vision due to abnormal eye movements is a prominent symptom and cause of disability in humans with cerebellar diseases.

A number of prior studies in humans and animals have investigated the effects of cerebellar lesions on eye movements and have helped to delineate the specific roles of particular cerebellar substructures. For example, lesions of the dorsal vermis1 or the associated portions of the fastigial nuclei2 cause inaccurate (dysmetric) saccades because these areas are important for the calibration of saccade amplitude. Lesions of the flocculus and ventral paraflocculus, critical for the calibration of the rotational vestibulo-ocular reflex (RVOR), impair RVOR plasticity.3-8 Control of pursuit appears to be distributed among several areas of the cerebellum, including the flocculus and paraflocculus,9 the vermis10 and fastigial nucleus,11 the uvula,12 and the lateral cerebellar hemispheres.13 The ability to hold the eyes steady at a given position in the orbit is a function of the fixation and gaze-holding systems, which are impaired by lesions of the flocculus and paraflocculus, resulting in downbeat and gaze-evoked nystagmus.14