Measurement of Position Acuity in Strabismus and Amblyopia: Speciﬁcity of the Vernier VEP Paradigm

Sean I. Chen,1,2 Anthony M. Norcia,1 Mark W. Pettet,1 and Arvind Chandna2

PURPOSE. An objective measure of positional acuity is desirable each of these studies vernier offsets were introduced and in the nonverbal clinical population. This study was conducted withdrawn from the stimulus, resulting in a combination of to investigate the specificity of the vernier VEP as a measure of vernier offset and motion-related responses. In each case, con- positional acuity, evaluating the potential confound of asym- trol conditions were used to verify which aspects of the re- metric motion responses that may be present in some groups sponse were specific to vernier offsets. Studies using the tran- of patients. These motion responses could masquerade as po- sient VEP23–25 found that vernier offset stimuli produced an sition-specific responses, since they occur at the same re- evoked response to very small offsets that did not produce a sponse frequency as the vernier-related response. response to motion-control conditions. 26–29 METHODS. Twelve observers with early-onset esotropia (EOE), The steady state vernier VEP involves the periodic 30 children with untreated amblyopia, and 15 control children introduction and withdrawal of vernier offsets in a high-con- underwent swept vernier VEP acuity testing accompanied by a trast square-wave grating target (Fig. 1A). The response con- swept motion control stimulus. The control condition was tains two components: one related to the magnitude of the used to detect the presence of artifactual responses not related misalignment (positional information) that occurs at the odd to position sensitivity. The patients with EOE were selected for harmonics of the stimulus frequency (first, third, fifth, and so high levels of motion asymmetry as documented with oscillat- forth) and the other related to the motion of the stimulus that ing gratings presented monocularly. As a measure of motion occurs at the even harmonics. These components can be confound (penetration), the proportion of first-harmonic re- readily differentiated by using spectral analysis.26 The control sponses recorded in the control condition was determined. condition utilizes the same amount of motion, but the moving RESULTS. The penetration rate in the vernier condition in each elements are placed symmetrically with respect to the static study group (EOE: 0.93%; amblyopes: 4.26%; normal subjects: reference elements (Fig. 1B). This results in a symmetric mis- 2.40%) and the entire group (2.85%) was acceptably low. The alignment–misalignment stimulus, with no change from col- level of penetration was not significantly influenced by the linear to noncollinear. The response of normal observers to the presence of amblyopia. control condition comprises only even harmonics. In these observers, the odd harmonics to the alignment–misalignment ONCLUSIONS The vernier VEP paradigm, when applied in the C . stimulus are thus specific to the relative position of the static manner described, can be interpreted as a measure of position and moving elements and they reflect the output of a position- sensitivity. The presence of motion asymmetry or untreated sensitive mechanism.26 amblyopia does not affect the validity of vernier measurements A difficulty in applying the steady state vernier VEP in made. (Invest Ophthalmol Vis Sci. 2005;46:4563–4570) DOI: clinical populations is the possible presence of motion re- 10.1167/iovs.05-0792 sponses that are not symmetric. A clear example of asymmetric motion VEP responses occurs in patients with early-onset es- t is well-known, on the one hand, that grating acuity system- otropia (EOE) in which the monocular response to rapidly Iatically underestimates optotype acuity losses in amblyo- 1–7 oscillating gratings is dominated by odd-harmonic components pia. On the other hand, losses on vernier acuity more closely 30–35 match the loss of optotype acuity than do losses on grating that reflect the asymmetry of motion processing. A similar 8,9 10–22 asymmetry is found in the monocular response of normal acuity. These results have prompted several attempts to 30,32,33,36 develop psychophysical vernier acuity tests for pre- or nonver- infants. Odd harmonic responses due to asymmetric bal subjects. motion processing or other sources not related to position Visual evoked potentials (VEPs) provide another means of cues could be confused with odd harmonics arising from the assessing visual function in nonverbal subjects. VEPs specific to processing of the vernier offsets. vernier offsets have been measured in several studies.23–29 In Asymmetry to horizontally induced motion has been exten- sively investigated using steady state VEP,30–38 transient VEP,39,40 oculomotor parameters (optokinetic nystagmus, pursuit eye movements),39–47 and psychophysical and/or percep- From the 1Smith-Kettlewell Eye Research Institute, San Francisco, tual studies.48–52 2 California; and the Vision Assessment Unit, Department of Ophthal- There have been fewer reports of vertical motion asymme- mology, Royal Liverpool Children’s Hospital, Liverpool, United King- try, however,43,45,49,52,53 and to our knowledge there is only dom. one report of vertical motion asymmetry in specific relation to Supported by Grant OR2001-99a from The Guide Dogs for the 37 Blind Association, United Kingdom, and The Smith Kettlewell Eye the steady state VEP. In these studies in which horizontal and Research Institute, San Francisco, California. vertical motion asymmetry were each assessed, there is the Submitted for publication June 22, 2005; revised July 26, 2005; consistent finding that, when asymmetric responses to vertical accepted September 26, 2005. motion are present, they are consistently less robust than Disclosure: S.I. Chen, None; A.M. Norcia, None; M.W. Pettet, asymmetric responses to horizontal motion. None; A. Chandna, None This evidence suggests that a horizontally oriented carrier The publication costs of this article were defrayed in part by page grating with vertically introduced vernier offsets offers a better charge payment. This article must therefore be marked “advertise- ment” in accordance with 18 U.S.C. §1734 solely to indicate this fact. choice of stimulus design. In this study, we examine the levels Corresponding author: Sean I. Chen, Department of Ophthalmol- of motion contamination and confound of the steady state ogy. Royal Liverpool Children’s Hospital, Liverpool L12 2AP, UK; vernier VEP by recordings of patients with early-onset esotro- [email protected]. pia who first had a high level of motion asymmetry docu-

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Bloomington, IL). Nine of the youngest children (six amblyopes and three normal) were not testable with the Lea chart, and the presence of amblyopia was assessed by fixation preference (FP), as described in A the next section. In the EOE group, mean right eye acuity was 0.08 Ϯ 0.14 logMAR (SD; range, Ϫ0.10–0.30). Mean left eye acuity was 0.00 Ϯ 0.14 logMAR (Ϫ0.20–0.30). The group mean interocular acuity difference (IAD) was 0.09 Ϯ 0.15 logMAR (0.00–0.50). Only three participants from 0 this group had amblyopia, with IADs of 0.20, 0.20, and 0.50 logMAR. Amblyopia. Thirty children with untreated amblyopia aged between 0.5 and 8 years were recruited consecutively. The group comprised 13 male and 17 female patients (10 with anisometropia, 10 with strabismus and, 10 with mixed disease) with a mean age of 4.76 Ϯ 1.63 10 seconds years (SD; range, 1.02–7.10). Children were considered to have ambly- Ն Vernier Acuity Sweep opia if the IAD was 0.2 logMAR. Those unable to perform this test reliably were assessed for amblyopia by the state of FP based on previously described techniques.54–56 Briefly, in this study, grade 1 FP referred to patients consistently demonstrating equal FP between eyes (in three or all four of four trials). Grade 2 referred to patients able to B hold fixation with the nonpreferred eye through a blink. Grade 3 patients were able to hold fixation with the nonpreferred eye only up to a blink and, grade 4 patients demonstrated an immediate switch back to the preferred eye when it was uncovered. Only Grade 3 and 4 0 patients were recruited as having amblyopia. (This equated to the grades used by Sener et al.54 grades 0 to 2 and grades 1 and 2 of Zipf et al.55) Participants with amblyopia were prescribed their full optical correction after cycloplegic retinoscopy, adjusting only for working distance. In participants able to perform logMAR chart acuity reliably, 10 seconds VEP testing began on the first occasion after spectacles were prescribed when the fellow-eye acuity became equal to or better than its Vernier Control Sweep previously recorded threshold in the unaided state. In those unable to perform chart acuity, VEP testing was performed when spectacle- FIGURE 1. Schematic of swept vernier acuity and vernier control stim- wearing time was greater than 50% of waking hours. uli. The vertically introduced offsets are modulated between alignment The group mean acuity of the amblyopic eye was 0.59 Ϯ 0.38 and misalignment (A, vernier acuity) and misalignment and misalign- logMAR (range, 0.20–1.77), and the group mean IAD was 0.46 Ϯ 0.36 ment (B, vernier control) at 3.76 Hz. The magnitude of the offset is logMAR (0.20–1.52). Six patients were assessed by FP, and their grades increased in 10 logarithmic steps toward the visible direction. The were all four of four (immediate switch back to preferred eye). range of offset and amount of symmetric motion are matched between Normal Subjects. Fifteen normally developing children without (A) and (B). strabismus or amblyopia (seven male and eight female) with a mean age of 5.37 Ϯ 2.28 years (SD; range, 0.60–8.58) were recruited. They mented with oscillating gratings.32,34,35 In this group, the pres- were required to have a normal birth history without being more than ence of a motion asymmetry was investigated in response to 2 weeks premature or having low birth weight, to possess no manifest both horizontally and vertically oriented gratings. After this, strabismus, and to have IAD Յ 0.1 logMAR and/or FP of no worse than the level of motion confound of the steady state vernier VEP 1 (i.e., to prefer each eye equally). Anterior and undilated posterior was also tested for both vertically and horizontally introduced segment examination of normal subjects, performed by one or both of vernier offsets. two fully qualified pediatric ophthalmologists (SC or AC), also had to The level of motion confound of the steady state vernier yield clinically normal findings. Potential observers were subjected to VEP was also tested in a group of children with untreated manifest retinoscopy with fogging control. Normal recruits found to amblyopia (vertically introduced offsets only), since this group have spherical and/or cylindrical refraction outside acceptable limits is a likely target for application of vernier acuity measurements for their age57 were to be subjected to cycloplegic refraction; but none in the clinical setting. A group of normal children served as of the normal children met this requirement, and all were therefore control subjects. tested in the unaided state. Group mean acuity was 0.03 logMAR in right and 0.05 in left eyes, with the same ranges (Ϫ0.10–0.20) and similar standard deviations METHODS (right ϭ 0.10; left ϭ 0.11). Group mean IAD was 0.01 Ϯ 0.03 (0.00– Observers 0.10). Three children were assessed according to FP, as they were too young to be tested with the Lea chart. Early-Onset Esotropia. Twelve participants with a history of This work was performed with IRB approval. Informed written EOE (mean onset, at 5 months) by history and/or medical records, consent was obtained from the participant or guardian, and the pro- were recruited for the study. The mean age at testing was 18.04 Ϯ tocol conformed to the tenets of the Declaration of Helsinki. 12.94 (SD) years (range, 5–42). Each had a documented VEP motion asymmetry, as determined by a preliminary experimental protocol of VEP Recording and Analysis oscillating gratings.30–35 Participants in this group were tested with their habitual spectacle The EEG was amplified by 50,000 for adults and 20,000 for children correction (if one was worn), which had been prescribed within the under 2 years of age over an amplifier pass-band of 1 to 100 Hz (Ϫ6 preceding year. In this as well as other study groups, clinical acuity was dB), using Grass amplifiers (model P511; Grass Telefactor, West War- measured in logMAR (minimum angle of resolution) units, using the wick, RI). The sampling rate was 600 Hz (16 bits), analog to digital crowded Lea symbols, Early Treatment Diabetic Retinopathy Study conversion. The electrode montage consisted of Oz, O1, and O2, each (ETDRS)–style distance chart (Catalog no. 2503; Precision Vision, referenced to Cz with additional bipolar derivations (O1–Oz, O2–Oz)

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into which vernier offsets were introduced. The display comprised equal height regions of moving and static pattern, with the distance between offsets being 0.50°. In all groups tested, vernier offsets were introduced vertically within a horizontally oriented square-wave grating (Fig. 1A). Figure 1A depicts a schematic of the vernier acuity stimulus. The offsets were presented in alignment–misalignment mode with the offset parameter swept from small to large (invisible to visible). In the vernier control condition (Fig. 1B) the offsets were presented symmetrically about the static reference panels to produce a misalignment–misalignment mode. All vernier acuity and vernier control conditions were swept-parameter conditions. The offset was swept in logarithmic steps over a 10-second recording period divided into 1-second epochs termed bins. The sweep range was 0.25 and 4 arcmin in observers aged 8 years or more (0.5–8 arc min under 8 years).27 This sweep range was altered toward the larger offset direction in patients with amblyopia who failed to produce a vernier acuity threshold for the starting sweep FIGURE 2. Motion asymmetry in a patient shown by spectrum analysis range. The sweep range for the control stimulus always matched that of the response to 10-Hz horizontal oscillatory motion. Temporal fre- of the vernier acuity condition. Between 3 and 6 trials were averaged quency and response amplitude are plotted on the x-axis and y-axis, for each stimulus condition. respectively. Compared with other frequencies and the second harmonic (2F1 ϭ 1.3 ␮V), the first harmonic (1F1 ϭ 4.8 ␮V) dominates, Threshold Determination in ϭ making this patient’s response highly asymmetric (asymmetry index Swept-Parameter Conditions 0.79). For each swept-stimulus condition, response thresholds are estimated by regression of amplitudes from the trial-average bins where the and one Laplacian derivation Oz Ϫ [(O1 ϩ O2)/2] being calculated response increased monotonically with stimulus visibility. The range of off-line. Spectrum analysis was used to extract the amplitude and phase bins eligible for regression depended on the statistical significance and of the evoked response at the first six harmonics of the stimulus phase consistency of the response, according to an algorithm adapted frequency. Specially written software (PowerDIVA, software, ver. 2.0; from Norcia et al.60 The regression range was limited to those bins in developed by one of the authors [AMN], Smith Kettlewell Institute, San which the following criteria were met: (1) The response probability in Francisco, CA) was used for recording and analysis of evoked potential each bin was at most 0.16; (2) the difference in response phase for data. Details of the spectrum analysis58 have been presented else- each pair of consecutive bins was between 80° and Ϫ100°, where where, and information related to the software’s algorithm for com- phase increased with response latency; (3) at least one pair of consec- puting threshold data is presented later in this article. utive bins had a response P Յ 0.077; and (4) to exclude spike artifacts, the amplitude of the bin immediately before and the bin immediately Oscillatory Motion VEP Test for after any given bin in the range could not both be Ͻ0.3 times the Response Asymmetry amplitude of that given bin. Once the regression range was established, The symmetry of the monocular motion VEP was assessed in the EOE the threshold stimulus value was determined by extrapolating the group by presenting the participants with high-contrast horizontal and regression line to the zero response amplitude. When applied to vertical sine-wave gratings, each oscillating at 6 Hz and also at 10 Hz, spectral data from background EEG, these criteria yielded a 5% false- hereafter termed motion VEP. The spatial frequency of the gratings was positive rate (data not shown). 2 cyc/deg and the oscillation was through 90° of spatial phase. The gratings were presented at 80% Michelson contrast at a space-averaged Motion Artifact Categorization luminance of 150 candelas per square meter (cd/m2). The field size Our criterion for detecting the intrusion of a motion artifact (penetra- was 8° ϫ 8° throughout. Stimuli were generated at a resolution of tion) into the measurement of the vernier VEP was the rate at which 1600 ϫ 1200 on a high-bandwidth video monitor (model MR2000-HB; the vernier acuity scoring algorithm was triggered at the first harmonic Richardson Electronics, LaFox, IL). The frame rate was 60 Hz, and the (1F1) in the vernier control stimulus. The level of penetration is monitor contrast was linearized through a gamma table correction. reported herein as both the total (relevant ϩ irrelevant) number of The symmetry of the motion VEP was quantified with an asymme- penetrations and the number of relevant penetrations (when the 1F1 try index.30,31,33,34 Patients with asymmetric motion responses such as threshold of the control was less that that of the vernier condition). EOE display significant first-harmonic responses to oscillatory motion The notion of relevance takes into account clinically relevant versus that are smaller or nonexistent in normal patients. Figure 2 shows the irrelevant penetrations. A threshold at the first harmonic due to motion spectral analysis of the VEP response to 10-Hz horizontal oscillatory asymmetry is problematic (clinically relevant) if that threshold is better motion in one of the EOE observers. The temporal frequency spectrum (lower) than the true vernier acuity and one would erroneously at- is profiled on the x-axis and response amplitude on the y-axis. The first tribute too good an acuity to the patient. If the vernier onset–offset (1F1, 10 Hz) and second (2F1, 20 Hz) harmonic responses are in black, stimulus produces a lower threshold than the control condition (irrel- and the responses of neighboring frequencies are shaded. It is clear evant penetration), it can only have resulted from mechanisms sensi- that the 1F1 (4.8 ␮V) dominates the response and is greater than the tive to the offset, per se, since the motion is the same in test and 2F1 (1.3 ␮V), making the asymmetry index in this case equal to 0.79 control conditions and the threshold measure is likely to be valid. [1F1/(1F1 ϩ 2F1)]. In this study, only EOE patients with an asymmetry index Ն0.35 to horizontal motion were included as motion asymmet- Procedure 31,59 ric. Early Onset Esotropia. Monocular motion VEPs were mea- Swept-Parameter Measurement of VEP sured in each eye under four viewing conditions (horizontal and vertical gratings, 6 and 10 Hz temporal frequency). The order of Vernier Acuity presentation of conditions was randomized within an eye and the Vernier acuity thresholds were measured using vertical (EOE group order of eye tested first was randomized across observers. Observers only) and horizontal (all observers) 2-cyc/deg square-wave gratings from this group were then presented with an additional eight swept-

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FIGURE 3. Results of group mean (ϮSEM) asymmetry indices for EOEs by temporal frequency, eye, and recording channel. Responses to horizontal induced motion was consistently more asymmetric than those to vertical induced motion, and 10-Hz motion induced greater response asymmetry than 6 Hz in this group.

parameter conditions (vernier acuity stimulus: alignment and misalign- The mean asymmetry index across channels was deter- ment; and vernier control stimulus: misalignment–misalignment con- mined for each patient with EOE, in each of the eight different trol, in both horizontal and vertical orientation, right and left eyes). conditions tested. These eight conditions represented the pos- The order of presentation was randomized within eye, and the order of sible combinations of three two-level factors: temporal fre- testing of eyes was randomized across observers. quency (6 or 10 Hz), grating orientation (horizontal or verti- Children with Amblyopia and Normal Children. VEP cal), and eye (right or left). Because three observers were data were recorded in each eye for vernier acuity and vernier control amblyopic, however, data from them were excluded from a stimuli in horizontal orientation with vertically introduced vernier repeated-measures ANOVA (Statistica GLM module; Statsoft offsets. These younger participants were part of a longer experimental Inc., Tulsa, OK) testing for interactions between experimental protocol, and for this reason only a horizontally oriented stimulus with conditions. Two main effects were revealed: temporal fre- ϭ ϭ vertical offsets was used. The order of presentation was randomized quency (F(1,8) 16.075, P 0.004) and grating orientation ϭ ϭ within eye, and the order of testing of eyes was randomized across (F(1,8) 7.315, P 0.027). The effect of eye was not signifi- ϭ ϭ observers. cant (F(1,8) 0.963, P 0.355), nor were there any interactions between any of the factors. This group of patients thus manifests a high level of motion asymmetry that was signifi- RESULTS cantly greater for horizontal than vertical oscillatory motion and was also greater at higher temporal frequencies (10 Hz vs. Motion Asymmetry in the EOE Group 6 Hz). The EOE group demonstrated horizontal asymmetry indices that were higher than previously reported in normal observers Vernier Penetration in the EOE Group (Fig. 3, Table 1).31,33,34 Figure 3 charts the mean asymmetry indices for the EOEs, grouped by temporal frequency, eye, and Having established abnormal levels of motion asymmetry in the recording channel. It is clear that in every category, the indices EOE group, we then proceeded to determine whether these for horizontal oscillatory motion (mean, 0.41 Ϯ 0.14) were asymmetries were likely to interfere with the measurement of greater than for vertical (mean, 0.30 Ϯ 0.10). Also across all vernier acuity. channels and eyes, asymmetry indices were greater for 10-Hz Figure 4 shows first harmonic response functions from an (mean, 0.47 Ϯ 0.14) than for 6-Hz motion (mean, 0.25 Ϯ 0.14). EOE observer in the vernier condition and in the control

TABLE 1. Early-Onset Esotropes: Mean Asymmetry Index for Horizontal and Vertical Induced Motion

6Hz 10Hz

RE LE Both RE LE Both

Horizontal 0.26 (0.14) 0.31 (0.20) 0.29 (0.17) 0.50 (0.22) 0.53 (0.19) 0.52 (0.17) Vertical 0.19 (0.13) 0.22 (0.14) 0.21 (0.13) 0.41 (0.16) 0.37 (0.19) 0.40 (0.16)

Data are the mean Ϯ SD group asymmetry index in response to induced motion in either eye or both eyes.

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FIGURE 5. Patient data showing irrelevant penetration. The vernier acuity response triggered a threshold (0.30 arcmin) by regression to zero amplitude. The vernier control response also triggered a threshold (0.57 arcmin) by regression that was worse, making this an irrelevant penetration.

FIGURE 4. Patient data showing the specificity of the first-harmonic (1F1) response to the vernier stimulus (no penetration). The 1F1 (9/97), but the relevant penetration rate was 0% (0/97; not response for the control condition was at noise level, whereas the shown). vernier acuity response rose monotonically with increasing vernier offset. Acuity Results: EOE Group The logMAR acuity did not differ significantly between eyes of condition and average noise. Increasing vernier offset is plot- the EOE or normal group or between the right eye of EOE and ted in log terms on the x-axis and response amplitude in linear normal subjects. As in the case of logMAR acuity, there was no terms on the y-axis. This observer did not have a significant first-harmonic response in the control condition and thus does not demonstrate penetration. Figure 5 shows an example of irrelevant penetration. In this figure the line segments between points of each function have been removed for clarity, and the regression lines determining the thresholds are included. The 1F1 for the vernier stimulus and its control both triggered thresholds. The vernier acuity threshold (0.30 arcmin) was, in this case, better than the control threshold (0.57), making this penetration irrelevant. An example of a relevant penetration is shown in Figure 6, which has the same format as Figure 5. In this case, the control threshold (0.57) was better than the vernier acuity threshold (0.62). Note that the peak signal-to-noise ratio in the nonpen- etrating case (Fig. 4, compare amplitude of solid lines to that of noise levels shown by dotted lines) was better than the cases in which penetration occurred (Figs. 5, 6), especially for the 1F1 of the control condition—a frequent observation. We report herein, the total penetration rate (relevant ϩ irrelevant) and the relevant penetration rate. In this study, the scoring algorithm was applied 288 times to the EOE control data (six channels, two orientations, 24 eyes of 12 observers) yielding 205 thresholds. The total penetration rate (all eyes both orientations) was 17 of 205 applications for an 8.29% penetration rate but the relevant penetration rate was 0.49% (1/205). For vertically introduced offsets (Table 2), the total penetration rate was 7.41% (8/108), but the relevant penetra- FIGURE 6. Patient data showing relevant penetration. The vernier tion rate was 0.93% (1/108) and therefore much less than the acuity response triggered a threshold (0.62 arcmin). The vernier con- false-positive rate of the scoring algorithm itself. For horizon- trol response triggered a threshold (0.57 arcmin) which was better tally introduced offsets, the total penetration rate was 9.28% (lower) than the vernier acuity threshold.

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TABLE 2. Total and Relevant Penetration Rates for Vertical Vernier Offsets

Penetration Index

Total Relevant Penetrations Penetrations Vernier Thresholds Study Group Eye n (%) n (%) n (100%)

EOE RE 2 (3.85) 0 (0.00) 52 LE 6 (10.71) 1 (1.79) 56 Amblyopes Amblyopic 6 (7.23) 4 (4.82) 83 Fellow 11 (10.48) 4 (3.81) 105 Normal Subjects RE 5 (7.46) 1 (1.49) 67 LE 9 (15.52) 2 (3.45) 58 All Groups Both 39 (9.26) 12 (2.85) 421

significant difference in VEP vernier acuity between the EOE penetration rate, however, was lower than the amblyopic and normal subjects for vertical vernier offsets. There was no group at 2.40% (3/125). significant (P ϭ 0.26) difference between the VEP vernier acuity thresholds generated by the two vernier orientations. Vernier Penetration in the Entire Study Group Vernier Penetration in the Amblyopic Group Considering vertically introduced vernier offsets only, which was tested across all study groups, the total penetration rate In this group, vernier offsets were introduced only in the was 9.26% (39/421), whereas the relevant penetration rate was vertical orientation. Four eyes in this group were untestable 2.85% (12/421). Concerning relevant penetrations, the mean because of reduced cooperation. The scoring algorithm was difference in thresholds between the vernier and control con- applied 336 times to the amblyopic group data (six channels, ditions was 0.52 arcmin (median, 0.36 Ϯ 0.52). The extremely one orientation; 56 eyes of 30 observers) yielding 188 thresh- low number of relevant penetrations between horizontal and olds. The total penetration rate was 17 of 188 applications for vertical vernier offset conditions tested in the EOE group pre- a 9.04% penetration rate (Table 2). The relevant penetration cludes meaningful comparison. rate, however, was 4.26% (8/188). It may appear from cursory inspection of the total penetration rates (Table 2) that left eyes seem more likely to penetrate, Acuity Results: Amblyopic and Normal Group at least in the normal and EOE groups. Overall, there were 21 There was a significant (P ϭ 2.2 ϫ 10Ϫ6) difference in logMAR penetrations in left eyes and 18 penetrations in right eyes. As 2 61 acuity between amblyopic and fellow eyes of the amblyopic expected therefore, ␹ analysis ruled out eye as a factor in group, but no difference between fellow eyes of the amblyopic total penetration rate (P ϭ 0.37). Furthermore, the conditions group and the right eyes of normal subjects. Findings were and eyes were randomized, making order effects much less similar for the VEP vernier acuity thresholds, in that a signifi- likely. cant (P ϭ 0.0001) difference was found only between amblyopic and fellow eyes of the amblyopic group (Table 3). DISCUSSION Vernier Penetration in the Normal Control Group The present study shows that monocular, position-specific VEP As in the amblyopic group, vernier offsets were introduced responses can be measured with low levels of contamination only in the vertical orientation. The scoring algorithm was by motion artifacts in patients with strabismus and/or ambly- applied 178 times to the normal group data (six channels, one opia. Acuity estimates based on regression of the first harmonic orientation; 30 eyes of 15 observers) yielding a total of 125 recorded from vernier onset–offset stimuli are better than thresholds. The total penetration rate was 14 of 125 applica- those recorded from a symmetric control condition in 97% of tions for an 11.20% penetration rate (Table 2). The relevant cases. Thus, in only 3% percent of cases did we find evidence

TABLE 3. Threshold Yield and Mean VEP Vernier Acuity Thresholds for Vertical Vernier Offsets

Threshold Count Mean Threshold Value Study Group Eye Possible (n) Actual (n) arcmin (SD)

EOE RE 72 52 0.71 (0.57) LE 72 56 0.56 (0.27) Both 144 108 Amblyopes Amblyopic 168* 83 1.65 (1.45) Fellow 168* 105 0.91 (0.67) Both 336* 188 Normal Subjects RE 89 67 0.86 (0.85) LE 89 58 0.86 (0.70) Both 178 125

Possible (n) is the number of thresholds that could be derived, given the number of observers, eyes, and recording channels. Actual (n) is the number of instances in which a threshold was actually returned. * Reduced number, since recording was not possible in four eyes in this group (30 observers).

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for a motion response that would lead to an erroneous deter- lead to a less robust preference than is the case in grating mination of the vernier threshold. These erroneous determina- acuity measurements in which only one salient stimulus is tions constitute false positives for the vernier acuity test pro- presented. It is commonly reported10,13,15,21,22 that many in- cedure (e.g., concluding position-specificity where there is fants were either unable to complete the testing protocol or none). This false-positive rate is similar to that of the scoring had psychometric functions that did not reach high levels of algorithm itself when applied multiple times across recording percent correct. It is well known that infants prefer to look at channels and stimulus conditions. a moving or flickering stimulus,20,64 rather than a stationary Previous work26,27 using a vernier stimulus such as the one one. Many studies of infant vernier acuity11,14–18,20–22 incor- used herein failed to detect evidence of motion penetration porated some element of motion in their vernier stimulus that into the first harmonic response of the alignment–misalign- otherwise maintained the magnitude of the offsets presented. ment stimulus, but these studies were on a smaller scale and Although attempts to counteract this introduced motion were tested only normal children and adults. More important, these made in some,15,16 none used adequate control for the intro- two studies used binocular recordings which may have ob- duced motion. The fact that motion can influence behaviorally scured motion asymmetries that would be expected to cancel measured vernier thresholds has been clearly documented.17,18 under binocular viewing conditions. We show here that the The stimuli used in these studies11,14–18,20–22 therefore could steady state vernier VEP is similarly immune to motion artifacts very well have been confounded by motion and as such may not both in untreated patients with amblyopia (age range tested reflect solely the measurement of a position-sensitive mechanism. here), as well as in patients with EOE who had documented high levels of motion asymmetry. When penetration did occur, Potential limitations of the VEP Vernier Paradigm the signal-to-noise ratio was usually lower, especially in the The results for high-frequency oscillation described earlier and control condition. in Figure 3 suggest that caution should be used at higher temporal frequencies. The VEP motion asymmetry was in fact Horizontal Versus Vertical Motion VEP first observed in a deeply amblyopic patient tested with the Asymmetries for Oscillating Gratings vernier onset stimulus at 10 Hz (Norcia AM, Wesemann W, As part of our effort to determine the extent to which the unpublished observations, 1985). vernier VEP paradigm can be contaminated, asymmetries in The group of patients with amblyopia tested in this study motion processing were studied in a group of patients with had no previous treatment and as such, our findings may not be high levels of horizontal motion asymmetry, as determined applicable at other stages of treatment. Changes in motion with oscillating gratings.32,34,35 Previous behavioral studies in asymmetry during treatment have been found after surgical 30,33 31 infants52 and in adults with strabismus43,45,49 or amblyopia45,49 realignment of strabismus and alternate occlusion. In and previous VEP work from our laboratory37 has found lower both cases, treatment resulted in decreases in motion asymme- levels of motion asymmetry for vertical as opposed to horizon- try, however, and thus would not be expected to increase tal motion. We also find the magnitude of VEP vertical motion motion penetration in the vernier task. asymmetry to be significantly less than that for horizontal. We If the vernier VEP technique we have described is to be may have observed low penetration rates for the vertical mo- applied in a novel situation, such as patients with a different tion condition because of this, but if motion orientation on its diagnoses, it would be advisable to include the motion-control own is the sole controlling factor, we would expect to have condition as a precaution. seen more penetration with horizontal motion. We did not. 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