ORIGINAL RESEARCH ARTICLE

Light- and dark-adapted electroretinograms (ERGs)

and ocular pigmentation: comparison

of brown- and blue-eyed cohorts

Abdlsaed Al Abdlseaed • Yvonne McTaggart •

Thomas Ramage • Ruth Hamilton •

Daphne L. McCulloch

Received: 12 November 2009 / Accepted: 5 July 2010 / Published online: 28 July 2010

Ó Springer-Verlag 2010

Abstract This study characterizes differences in

human ERGs based on ocular pigmentation. Light-

and dark-adapted luminance-response (LR) series

for a-, b- and i-waves and light-adapted oscillatory

potentials (OPs) were recorded in 14 healthy volun-

teers (7 blue-eyed Caucasians; 7 brown-eyed Asians,

aged 20–22 years). Amplitude interpolations were by

logistic growth (Naka-Rushton), Gaussian or the

combined ‘photopic hill’ functions. Implicit times

(IT) for dark-adapted a- and b-waves, and for light-

adapted a-, b- and i-waves were earlier in the blue-

eyed group than in the brown-eyed group across all

flash strengths (P\ 0.05). For dark-adapted ERGs,

saturated a-wave amplitude was larger for blue eyes

(397 vs. 318 lV, P\ 0.05) as was the a-wave to

strong flash (10 cd�s/m2; 357 vs. 293 lV, P\ 0.05)

and the b-wave to ISCEV standard 0.01 (354 vs.

238 lV, P\ 0.05). Light-adapted b-waves for mid-

range flash stimuli were much larger for the blue-

eyed group (photopic hill, Gaussian peak: 155 vs.

82 lV, P\ 0.001) with no difference in saturated

amplitudes. Similarly, interpolated i-wave amplitudes

were larger (48 vs. 18 lV, P\ 0.01). For a light-

adapted 2.6 stimulus, a- and b-waves were larger for

the blue-eyed group (52 vs. 39 lV; 209 vs. 133 lV,

P\ 0.01) as were OP4 and OP5 (37.2 vs. 15.6 lV;

47.5 vs. 22.2 lV, P\ 0.01), but OP1-OP3 did not

differ. ERGs have shorter ITs in people with blue

irides than in those with dark pigmentation. Ampli-

tude differences are highly non-linear and substan-

tially larger from eyes with light pigmentation for

components thought to be associated with the OFF

retinal pathways.

Keywords Electroretinogram (ERG) �Ocular pigmentation � Luminance–response

function � OFF response � Photopic hill

Introduction

In all applications of clinical electroretinography,

results must be compared with suitable normative data

[1]. It has been reported that human eyes with lighter

fundi demonstrate larger full-field ERGs under both

light- and dark-adapted conditions with normal or

supranormal amplitudes in those with albinism [2–7].

Fundus pigmentation corresponds closely to iris pig-

mentation, iris colour and skin pigmentation [8–10].

Therefore, iris colour is a reasonable proxy for fundus

pigmentation. Although dark- and light-skinned

humans have similar amounts of melanin pigment in

A. Al Abdlseaed (&) � Y. McTaggart �T. Ramage � D. L. McCulloch

Department of Vision Sciences, Glasgow Caledonian

University, Glasgow G4 0BA, UK

e-mail: saed.alabdlseaed@gcal.ac.uk

R. Hamilton

Department of Clinical Physics, Royal Hospital for Sick

Children and University of Glasgow, Glasgow, UK

123

Doc Ophthalmol (2010) 121:135–146

DOI 10.1007/s10633-010-9240-3

their retinal pigment epithelium (RPE), humans with

dark skin have, on average, twice as much melanin

pigment in their choroid as those with light skin [9].

Although it seems intuitive that greater light absorption

by the denser choroidal pigment should reduce the

effective retinal illumination and hence the ERG

amplitude, this alone does not adequately explain the

larger ERGs from lighter eyes [3, 7].

To investigate the effect of pigmentation, the

present study compared ERGs between two distinct

groups—Caucasians with blue irides and Asians

with brown irides—using the light-adapted and the

dark-adapted luminance–response (LR) functions of

a-, b- and i-wave amplitudes. This included some of the

ISCEV standard ERGs (dark-adapted 0.01, 3.0 and 10;

light-adapted 3.0) and light-adapted oscillatory poten-

tials (OPs). The implications for clinical ERG record-

ings at the relevant ISCEV standards were considered.

Methods

Participants

The study included 14 adult participants, seven blue-

eyed Caucasians and seven brown-eyed Asians. All

were aged 20–22 years to limit any age effects

[11–13]. All were free of ocular disorders, had

corrected visual acuities of 6/6 or better in each eye;

refractive errors, if present, were in the range of-4.00

to?3.50 dioptres to limit effects associated with axial

length [14, 15]. Each gave informed consent to

participate in the study, as approved by the Human

Research Ethics Committee for Life Sciences at

Glasgow Caledonian University, which adheres to

the tenets of the declaration of Helsinki.

Protocols and recording

Visual acuity, intraocular pressure and eye examina-

tion including anterior chamber angle assessment were

conducted prior to pupil dilation (tropicamide 0.5%,

repeated after 5 min for the brown group) to achieve

maximal dilatation in all participants (C8 mm). A

ground electrode was placed on the contralateral

mastoid, and participants were then dark-adapted for

20 min. After instilling anaesthesia (oxybuprocaine

0.4%) and lubricant (CelluviscÒ), a Burian-Allen

bipolar contact-lens electrode was inserted into the

chosen eye using dim red light. Full-field, 4-ms white

flashes were generated by LEDs (colour temperature

6,500 K) or a xenon strobe as required. For the dark-

adapted series, 5–10 flashes at each of 17 flash levels

(ranging from -3.3 to 2.3 log cd�s/m2 in 0.3 log unit

steps) were delivered using an Espion E2 systemwith a

Colordome stimulator (Diagnosys, UK). The inter-

stimulus interval was 2 s for weaker flashes and up to

10 s for stronger flashes to avoid light adaptation

during testing [1]. Participants were then light-adapted

for 5 min to a 25.5 cd/m2 (1.39 log cd/m2) white

background. A second LR series was then recorded,

consisting of 16 white flashes from -2.0 to 3.0 log

cd�s/m2 in 0.3 log unit steps. The first 5 min of this

protocol consists of sub-threshold strength flashes and

acted as further light adaptation time. For the flash of

2.55 cd�s/m2 in the photopic series (within 0.07 log

units of current ISCEV standard [1]), OPs were

recorded with a bandpass filter of 75–300 Hz. Follow-

ing the ERG recordings, a fundus photograph was

taken, the correlation between iris colour and fundus

pigment concentration was confirmed by visual

inspection [8, 16], and the cornea and intraocular

pressure were examined.

Data analysis

Individual ERGs were examined off-line to remove

manually traces contaminated with blink or eye

movement artefacts. Amplitudes and implicit times

(IT) were measured from the remaining averaged

data as illustrated in Fig. 1.

A-wave amplitudes for both light- and dark-adapted

series and b-wave amplitudes for dark-adapted series

were plotted versus time-integrated luminance for each

individual, and a logistic growth function [17] (Naka-

Rushton) was fitted using Sigmaplot (v.8):

V ¼VmaxI

I þ rð1Þ

where V is amplitude (lV), I is time-integrated lumi-

nance (cd�s/m2), Vmax is the maximum saturated ampli-

tude (lV), and r is the semi-saturated time-integrated

luminance (the flash strength that evokes a half-

maximal response) (cd�s/m2). Only points up to the first

plateauwere included in thefit [18] to avoid distortion of

the logistic function by the secondary rising phase.

B-wave amplitudes for the light-adapted series

were plotted versus time-integrated luminance for

136 Doc Ophthalmol (2010) 121:135–146

123

each individual, and a photopic hill function com-

prising the sum of a logistic growth function and a

Gaussian function was fitted [19]:

Vb ¼ Gb

I

l

� �

lnðl=IÞ

B2

" #

þVbmaxI

I þ rb

ð2Þ

where Vb is b-wave amplitude (lV), Gb is the

maximum amplitude at the peak of the Gaussian

component (lV), B is a measure of the width of the

Gaussian component (cd�s/m2), and l is the flash

strength at the peak of the Gaussian component, Gb

(cd�s/m2). The variables in remaining portion of the

equation are as defined for Eq. 1.

Photopic i-waves were plotted versus time-inte-

grated luminance for each individual and fitted with a

Gaussian function. Up to five photopic OPs were

measured from the preceding trough to each peak.

Logarithms of all amplitudeswere used for statistical

purposes in order to normalise their distributions. Mean

values of the two groups were tested for differences

using two-sample, two-tailed t-tests. To assess ITs over

multiple flash strengths, the differences between group

mean values at each flash strength were tested using a

one-sample t-test relative to zero difference.

Results

Dark-adapted ERG series

All participants showed the expected dark-adapted

ERG waveforms with an increase in amplitude and

decrease in IT as the flash strength was increased.

Figure 2 illustrates typical data for one participant

from each pigmentation group. In all individuals, the

logistic curve provided a good fit to a-wave amplitudes

(Fig. 3, left). Average Vmax for a-waves from blue

eyes was 397 lV, significantly larger than from brown

eyes (318 lV) (P\ 0.05). Semi-saturated luminance

(log r), a measure of a-wave sensitivity, did not differ

between groups. Dark-adapted b-wave amplitudes

were also fitted with logistic growth curves: all showed

saturation by around 2 cd�s/m2 (Fig. 3, right). Neither

Vmax nor semi-saturated luminance, log r, for the

b-waves differed between groups (Table 1).

The a-wave IT shortened for stronger flashes as

expected (Fig. 4). ITs were marginally but signifi-

cantly shorter for blue eyes, by 1 ms on average across

all flash strengths (one-sample t-test, P\ 0.001). The

ITs of the dark-adapted b-wave decreased with

increasing flash strength, reaching a plateau of around

45 ms at around 1 cd�s/m2. Again, the blue-eyed group

had slightly shorter ITs, by 3 ms on average across all

flash strengths (one-sample t-test, P\ 0.01).

Dark-adapted ISCEV standard and recommended

strong flash ERGs

At the ISCEV standard flash [1] (dark-adapted 0.01),

only two (blue) eyes showed an a-wave. Blue eyes

had an average b-wave amplitude of 354 lV, signif-

icantly larger than the average from brown eyes of

238 lV (P\ 0.05). Mean b-wave IT did not differ

between groups (Table 1).

Fig. 1 A typical averaged

ERG from the light-adapted

series at 2.55 cd�s/m2 is

illustrated from one blue-

eyed participant. ITs are

measured from the flash,

a-wave amplitudes from the

pre-stimulus baseline and

the b- and i-waves

amplitudes are measured

from the preceding trough

Doc Ophthalmol (2010) 121:135–146 137

123

An ERG was not recorded at the ISCEV dark-

adapted 3.0 flash, but due to the linear nature of the

amplitude and IT response between the neighbouring

flashes of the LR series (2.0 and 5.0 cd�s/m2), a linear

interpolation of results was used to quantify results

[20] at the mid-point (3.16 cd�s/m2), which meets the

current ERG standard [1]. For this interpolated flash

strength, a-wave IT was 17.2 ms on average for

the blue-eyed group, significantly shorter than for the

brown-eyed group (19.1 ms) (P\ 0.05). For the

ISCEV strong flash (dark-adapted 10), only average

a-wave amplitudes differed between the groups,

being 357 lV for blue and 293 lV for brown eyes

(P\ 0.05) (Table 1).

Fig. 2 The dark-adapted

ERG LR series is illustrated

for two individuals, one

with brown eyes (left) and

the other with blue eyes

(right). Numbers indicate

the time-integrated

luminance of the full-field

flash (0.0005–200 cd�s/m2)

Fig. 3 Dark-adapted a-wave (left) and b-wave (right) LR

functions. Blue symbols (square) and solid line: blue eyes.

Brown symbols (filled circle) and dashed line: brown eyes. The

lines are based on the average values of the parameters of each

individual’s fit. Points beyond about 2 cd�s/m2 of the b-wave LR

function (right) reflect the secondary rising phase and were

excluded from the curve fitting [18]

138 Doc Ophthalmol (2010) 121:135–146

123

Light-adapted ERG series

Typical ERGs for the light-adapted LR series are

shown in Fig. 5 for a blue-eyed and for a brown-eyed

participant.

The LR function of light-adapted a-wave ampli-

tudes was well described by a logistic growth

function (Eq. 1; Fig. 6) in all participants. Neither

Vmax nor log r for the a-waves differed significantly

between blue and brown eyes (Table 2).

The photopic hill model [19] fitted well to each

individual’s data for b-wave amplitudes (Fig. 7, left).

The curve fitting parameters of the photopic hill model

showed no differences in log r of the logistic growth

function component (Fig. 7, lower right) or in the peak

luminance or width of the Gaussian components

(Fig. 7, upper right; Table 2). The b-wave amplitudes

for the strongest flashes, Vmax, of the logistic compo-

nent did not differ between groups (Fig. 7, lower

right). However, the Gaussian component, which

describes the amplitude increment in the mid-lumi-

nance portion of the photoic hill, was significantly

larger for blue eyes than for brown eyes (Fig. 7, upper

right) with mean values of 155 and 82 lV, respec-

tively (P = 0.0001).

Each individual’s i-wave amplitudes were well

fitted with a Gaussian curve (Fig. 8). The peak

amplitude was again substantially larger for blue eyes

than for brown eyes: 48 versus 18 lV, P = 0.005.

Neither the luminance at the amplitude peak nor the

Table 1 Summary of dark-adapted ERG results

Brown eyes Blue eyes P-value

from t-testMean 95% confidence

interval of mean

Mean 95% confidence

interval of mean

Luminance–response

functions

a-wave Vmax (lV) 318 274–367 397 348–454 0.046*

a-wave log r (log cd�s/m2) -0.12 -0.28–0.03 -0.04 -0.12–0.04 0.36

b-wave Vmax (lV) 403 344–472 448 394–510 0.33

b-wave log r (log cd�s/m2) -2.40 -2.55–-2.26 -2.49 -2.61–-2.36 0.10

ISCEV standard and

additional ERGs

0.01 b-wave amp (lV) 238 188–301 354 313–400 0.03*

0.01 b-wave IT (ms) 91.1 82.6–99.7 92.4 88.5–96.3 0.82

3.0 a-wave amp (lV) 236 199–280 287 253–326 0.09

3.0 a-wave IT (ms) 19.1 17.6–20.7 17.2 16.4–18.0 0.048*

3.0 b-wave amp (lV) 464 395–546 541 471–620 0.19

3.0 b-wave IT (ms) 48.1 45.7–50.4 46.6 44.6–48.5 0.35

10 a-wave amp (lV) 293 258–333 357 323–395 0.04*

10 a-wave IT (ms) 14.1 13.4–14.9 13.3 12.7–13.8 0.11

10 b-wave amp (lV) 462 377–566 565 486–658 0.14

10 b-wave IT (ms) 45.0 39.7–50.3 45.6 42.3–48.9 0.86

Amp amplitude

* P\ 0.05

Fig. 4 ITs for the dark-adapted ERG a-waves (lower) and b-

waves (upper) are shown for the blue-eyed group (blue) and the

brown-eyed group (brown). Error bars indicate the 95%

confidence interval for the mean of each group at each flash

strength

Doc Ophthalmol (2010) 121:135–146 139

123

width of Gaussian curves fitted to i-wave amplitudes

demonstrated significant differences between the

groups (Table 2).

The photopic a-wave IT shortened slightly for

stronger flashes; b-wave IT lengthened then short-

ened again and i-wave IT lengthened (Fig. 9). Across

Fig. 5 Light-adapted LR

series recorded for flashes

from 0.01 (bottom traces) to

1,000 cd�s/m2 (top traces)

flashes for two participants,

one with brown eyes (left)

and one with blue eyes

(right)

Fig. 6 Light-adapted

luminance–response

functions for a-wave

amplitude. Blue symbols

(open square) and solid

line: blue eyes. Brown

symbols (filled circle) and

dashed line: brown eyes.

The lines are based on the

average values of the

parameters of each

individual’s fit to Eq. 1

140 Doc Ophthalmol (2010) 121:135–146

123

all flash strengths, a-wave ITs were 0.7 ms shorter for

blue eyes (one-sample t-test, P = 0.0003); b-wave

ITs were 0.8 ms shorter for blue eyes (one-sample

t-test, P = 0.02) and i-wave ITs were 2 ms shorter

for blue eyes (one-sample t-test, P\ 0.05).

Light-adapted ISCEV standard ERGs

and oscillatory potentials

The 2.55 cd�s/m2 flash used here is close to the current

ISCEV standard flash of 3 cd�s/m2 [1] and within the

2004 standard range [21]. For this stimulus, blue eyes

showed an average a-wave amplitude of 52 lV, 33%

larger than the average amplitude from the brown eyes

(39 lV; P = 0.01). Blue eyes also showed a larger

average b-wave amplitude of 209 lV, 57% larger than

the average from brown eyes (133 lV, P = 0.001).

A-wave average IT was shorter for blue eyes than

brown eyes (14.0 vs. 14.6 ms, P = 0.02), but b-wave

average IT did not differ between the groups (Table 2).

The earlier oscillatory potentials, OP1, OP2 and

OP3, did not differ in amplitude between the groups.

However, OP4 and OP5 were more than twice as

large from blue eyes than from brown eyes,

P\ 0.0001 and P = 0.001 (Fig. 10, Table 2). These

later OPs were sufficiently different in amplitude to

cause the overall summed amplitude (SOPs) to be

significantly larger for blue eyes than for brown eyes:

106 lV for the blue-eyed group and 56 lV for the

brown-eyed group, P = 0.001. OP2 IT was shorter

for blue eyes (although there was zero variability),

and no other OP IT differences were found (Table 2).

Table 2 Summary of light-adapted ERG results

Brown eyes Blue eyes P-values

from t-testMean 95% confidence

interval of mean

Mean 95% confidence

interval of mean

Luminance–response

functions

a-wave amp (lV) 95 81–111 111 98–125 0.15

a-wave sensitivity log r

(log cd�s/m2)

0.54 0.44–0.65 0.44 0.27–0.52 0.30

b-wave amp (lV): Gaussian height 82 68–98 155 135–177 0.0001*

b-wave sensitivity: log Gaussian

position (log cd�s/m2)

0.71 0.53–0.88 0.66 0.56–0.77 0.69

b-wave amp (lV): logistic Vmax 78 62–99 74 57–96 0.79

b-wave sensitivity: logistic log r

(log cd�s/m2)

-0.08 -0.27–0.12 -0.01 -0.15–0.12 0.62

i-wave amp (lV): Gaussian height 18 12–28 48 34–67 0.005*

i-wave sensitivity (log cd�s/m2) 0.36 0.13–0.58 0.54 0.33–0.76 0.26

ISCEV standard

ERGs

2.55 a-wave amp (lV) 39 34–46 52 47–58 0.01*

2.55 a-wave IT (ms) 14.6 14.2–15.0 14.0 14.0–14.0 0.02*

2.55 b-wave amp (lV) 133 115–153 209 179–245 0.001*

2.55 b-wave IT (ms) 29.7 29.4–30.1 29.1 28.6–29.7 0.10

Light-adapted

oscillatory potentials

OP1 amp (lV) 3.3 2.7–4.0 4.9 3.9–6.0 0.11

OP2 amp (lV) 11.2 9.2–14.0 14.1 12.0–17.0 0.10

OP3 amp (lV) 13.2 10.0–18.0 11.0 11.0–11.0 0.43

OP4 amp (lV) 15.6 13.0–18.0 37.2 30.0–46 0.00003*

OP5 amp (lV) 22.2 16.0–30.0 47.5 39.0–58.0 0.001*

OP1 IT (ms) 13.7 13.2–14.3 13.3 12.9–13.6 0.23

OP2 IT (ms) 18.1 17.5–18.8 17.0 All 17.0 0.006*

OP3 IT (ms) 24.7 24.4–25.1 24.7 24.2–25.3 1.0

OP4 IT (ms) 28.3 27.9–28.6 28.1 27.9–28.4 0.55

OP5 IT (ms) 39.1 38.2–40.0 38.3 37.4–39.2 0.22

* P\ 0.05

Doc Ophthalmol (2010) 121:135–146 141

123

Fig. 7 Light-adapted LR functions (‘‘photopic hills’’) for

b-wave amplitudes. Blue symbols (open square) and solid

line: blue eyes. Brown symbols (filled circle) and dashed line:

brown eyes. On the right are the Gaussian components (top)

and logistic growth components (bottom). All lines are based

on the average values of the parameters of each individual’s fit

to Eq. 2

Fig. 8 Light-adapted LR functions for i-wave amplitude. Blue

symbols (open square) and solid line: blue eyes. Brown

symbols (filled circle) and dashed line: brown eyes. The lines

are based on the average values of the parameters of a Gaussian

curve fitted to each individual’s data

0

10

20

30

40

50

60

70

0.001 0.01 0.1 1 10 100 1000 10000

time-integrated luminance (cd·s/m2)

photo

pic

IT

(m

s)

Fig. 9 ITs of the light-adapted a-wave (lower), b-wave

(middle) and i-wave (upper). Error bars indicate the 95%

confidence interval for the mean of each group at each flash.

Where only one data point is available, that data point is

shown. Brown symbols: brown eyes. blue symbols: blue eyes

142 Doc Ophthalmol (2010) 121:135–146

123

The testing did not induce substantial elevation of

IOP, and there were no incidents of corneal staining.

The fundus photographs confirmed that all of the

brown-eyed Asian participants had typical brown–

pink fundus appearance, and the blue-eyed Cauca-

sians had light pink fundi with transparency of the

RPE and visible choroidal vessels.

Discussion

Some studies have reported ERG differences in

animal models based on fundus pigmentation levels

[22–28], and others have compared human partici-

pants [2–6, 29, 30]. Our findings that some ERGs are

larger from those with lighter eyes are in keeping

with previous studies [2–7]. The ITs were earlier for

the blue group compared with the brown group for

most of ERG components, both dark- and light-

adapted, which may suggest that a basic process, such

as the phototransduction cascade, may be slightly

accelerated in those with blue eyes.

Our LR curves have demonstrated that some ERG

components do not differ at all with eye colour, while

others differ markedly. For the light-adapted ERG

series, there were very marked amplitude differences

(blue[ brown) in the amplitude of the b-wave in the

mid-luminance range (photopic hill) (P = 0.0001)

and in the amplitude of the Gaussian-shaped i-wave

LR function (P = 0.005). These aspects of the ERG

are particularly associated with the OFF retinal

pathways [19, 31–37]. For the light-adapted OPs,

very marked amplitude differences (blue[ brown)

were noted for OP4 (P\ 0.0001) and OP5 (P =

0.001). These later OPs are also particularly associated

with the OFF retinal pathway [27, 38, 39]. Those

aspects of the light-adapted ERG associated with ON

retinal pathways, the saturated b-wave amplitude (as

described by the logistic growth component of the

photopic hill model) and the earlier OPs, showed no

difference between blue and brown eyes. However, the

light-adapted 2.55 ERG, which reflects a combination

of ON and OFF retinal pathway contributions, showed

larger and earlier a-waves (P = 0.01, P\ 0.02) and

larger b-wave amplitudes (P = 0.001) for blue eyes

than for brown eyes. To clarify these findings, Table 3

presents the significance or otherwise of group

differences.

The combination of findings presented in Table 3

suggests a selective association between some aspect

of fundus pigmentation and the OFF retinal pathway

with markedly larger responses in blue eyes than in

brown eyes.

Russell-Eggitt et al. [5] postulated the light scatter

theory as the cause of larger ERGs, as less pigmented

people will have more light reaching the photorecep-

tors via their more translucent iris and their reduced

light absorption in the choroid. Similarly, in 1992,

Wali and Leguire [2] hypothesised that light absorp-

tion by choroidal melanin in dark fundi could behave

as a 0.15 log unit neutral density filter. Slightly smaller

pupil diameters after dilation would have a similar

Fig. 10 Photopic OPs are

superimposed from all

brown-eyed participants

(left) and blue-eyed

participants (right).

Numbers indicate OPs.

Inset: amplitude

comparison for OPs 1–5,

and for summed OPs

(SOPs). Error bars are 95%

CI of the mean

Doc Ophthalmol (2010) 121:135–146 143

123

effect of reducing luminance for those with more

pigment. The small but significant increase in ERG

ITs in the brown-eyed group could indeed be

accounted for by light absorption: a 0.15 log unit

reduction in flash brightness would predict a 1.0-ms

slower a-wave IT and a 3.4-ms slower b-wave IT

under dark-adapted conditions; this matches very

closely the 1.0 and 3.0 ms differences actually mea-

sured. Wali and Leguire also postulated that reduced

light scatter would shift the LR function for darker

eyes to the right (less sensitive) without reducing Vmax.

However, as in the present study, they showed no such

shift and in fact recorded smaller Vmax values for

darker eyes [2, 3] and thus rejected reduced light

scatter as the cause for smaller ERGs in dark-eyed

participants.

An alternative explanation lies in the high resis-

tance and semi-conductor-like physical properties of

melanin [3, 46] which is found in greater concentra-

tions in the choroid of darker-skinned individuals [9,

47]. The choroid (with the sclera) forms a resistive

component in the ERG circuit [48], and it is therefore

plausible that the effective resistance of the brown-

eyed ERG circuit is greater than for blue-eyed people,

causing a relative reduction in ERG voltage [3].

However, this resistance theory does not explain the

findings in the present study, which suggest that

increased pigmentation has little effect on the ampli-

tude of ERG components generated by the ON retinal

pathways.

One further possibility is that there is a selective

effect on the OFF pathways of the higher calcium

binding found in the more pigmented tissues of animal

eyes [47]. Although calcium modulates a variety of

retinal functions [48, 49], the ERG signals seen in the

relatively calcium-rich blue eyes are unknown.

Implications for laboratory norms

In clinical electrophysiology, it is vital to be able to

compare individual patient data with comparable

normative ranges. Age [11–13], gender [50] and race

should be considered. In the present study, the groups

were selected to have distinctly different levels of

ocular pigmentation with iris colour and skin tone used

to define the groups. Iris colour has been shown to

closely correlate with fundus pigmentation [8, 9].

Other methods of categorisation, such as evaluation of

Table 3 ERG amplitudes for brown- versus blue-eyed participants with associated retinal sources

ERG component Difference Associated retinal source

LA OP4 **** OFF pathway [27, 38, 39]

Photopic b-wave peak (Gaussian) **** OFF response [19, 34]

LA 2.55 b-wave *** ON and OFF bipolar cells [33]

LA OP5 *** OFF pathway [27, 38, 39]

i-wave amplitude (Gaussian) ** OFF response [36, 37]

LA 2.55 a-wave * Photoreceptors; postreceptoral contributions, primarily OFF pathway [40]

DA 0.01 b-wave * Predominantly rod bipolar cells; ON pathway [41, 42]

DA 10 a-wave * Photoreceptors; postreceptoral contributions, primarily ON pathway [40]

DA a-wave LRF, Vmax * Photoreceptors; postreceptoral contributions, primarily ON pathway [40]

DA 3.0 a-wave – Photoreceptors; postreceptoral contributions, primarily ON pathway [40]

DA 3.0 b-wave – Predominantly rod bipolar cells; ON pathway [41, 42]

LA OP1 – Triggered through ON pathway [43]

LA OP2 – Triggered through ON pathway [43]

LA OP3 – Triggered through ON pathway [43]

DA 10 b-wave – Predominantly rod bipolar cells; ON pathway [41, 42]

LA a-wave LRF, Vmax – Photoreceptors & postreceptoral ON and OFF pathway [44]

DA b-wave LRF, Vmax – Predominantly mass activity of ON bipolar cells [45]

Photopic saturated b-wave (logistic) – Presumptive ON response [19]

LA light-adapted, OP oscillatory potential, LRF luminance–response function, DA dark-adapted

**** P\ 0.001; *** P\ 0.005; ** P\ 0.01; * P\ 0.05; – P[ 0.05

144 Doc Ophthalmol (2010) 121:135–146

123

calibrated digital fundus photographs, fundus reflec-

tometry or infrared imaging, can be used to provide

graded levels of pigmentation [2, 3, 8, 51–53].

We conclude that the major difference in ERGs

between groups with different levels of ocular pig-

mentation is larger photopic ERG components thought

to be associated with the OFF retinal pathways for blue

eyes. In addition, ERGs are slightly faster under most

test conditions, a result that can be accounted for by

reduced light absorption.

Future studies could include the EOG and long flash

duration ERGs to further understand the physiological

differences associated with pigmentation. Given the

higher prevalence of diseases associated with light-

induced damage in Caucasians with blue irides,

particularly age-related macular degeneration, it is

important to understand the underlying differences in

retinal physiology in this group. To this end, it would

seem important to understand fully the electrophysi-

ological consequences of denser retinal pigmentation.

References

1. Marmor MF, Fulton AB, Holder GE, Miyake Y, Brigell M,

Bach M (2009) ISCEV standard for full-field clinical

electroretinography (2008 update). Doc Ophthalmol 118

(1):69–77 (www.iscev.org)

2. Wali N, Leguire LE (1992) Fundus pigmentation and the

dark-adapted electroretinogram. Doc Ophthalmol 80(1):

1–11

3. Wali N, Leguire LE (1993) Fundus pigmentation and the

electroretinographic luminance-response function. Doc

Ophthalmol 84(1):61–69

4. Krill AE, Lee GB (1963) The electroretinogram in albinos

and carriers of the ocular albino trait. Arch Ophthalmol

69:32–38

5. Russell-Eggitt I, Kriss A, Taylor DS (1990) Albinism in

childhood: a flash VEP and ERG study. Br J Ophthalmol

74(3):136–140

6. Nusinowitz S, Sarraf D (2008) Retinal function in X-linked

ocular albinism (OA1). Curr Eye Res 33:789–803

7. Tomei F, Wirth A (1978) The electroretinogram of albinos.

Vision Res 18(10):1465–1466

8. Weiter JJ, Delori FC, Wing GL, Fitch KA (1985) Rela-

tionship of senile macular degeneration to ocular pig-

mentation. Am J Ophthalmol 99(2):185–187

9. Weiter JJ, Delori FC, Wing GL, Fitch KA (1986) Retinal

pigment epithelial lipofuscin and melanin and choroidal

melanin in human eyes. Invest Ophthalmol Vis Sci

27(2):145–152

10. Robins AH (1973) Skin melanin content in blue-eyed and

brown-eyed subjects. Hum Hered 23(1):13–18

11. Birch DG, Anderson JL (1992) Standardized full-field

electroretinography. Normal values and their variation with

age. Arch Ophthalmol 110(11):1571–1576

12. Westall CA, Panton CM, Levin AV (1998) Time courses

for maturation of electroretinogram responses from infancy

to adulthood. Doc Ophthalmol 96(4):355–379

13. Weleber RG (1981) The effect of age on human cone and

rod ganzfeld electroretinograms. Invest Ophthalmol Vis

Sci 20(3):392–399

14. Perlman I, Meyer E, Haim T, Zonis S (1984) Retinal

function in high refractive error assessed electroretino-

graphically. Br J Ophthalmol 68(2):79–84

15. Westall CA, Dhaliwal HS, Panton CM, Sigesmun D, Levin

AV, Nischal KK, Heon E (2001) Values of electroretino-

gram responses according to axial length. Doc Ophthalmol

102(2):115–130

16. Seddon JM, Sahagian CR et al (1990) Evaluation of an iris

color classification system. The Eye Disorders Case-Con-

trol Study Group. Invest Ophthalmol Vis Sci 31(8):

1592–1598

17. Naka KI, Rushton WA (1966) S-potentials from colour

units in the retina of fish (cyprinidae). J Physiol 185(3):

536–555

18. Severns ML, Johnson MA (1993) The care and fitting of

Naka-Rushton functions to electroretinographic intensity-

response data. Doc Ophthalmol 85(2):135–150

19. Hamilton R, Bees MA, Chaplin CA, McCulloch DL (2007)

The luminance-response function of the human photopic

electroretinogram: a mathematical model. Vision Res

47(23):2968–2972

20. Bach M, Poloschek CM, Wozniak S (2009) Moving from

non-standard to standard stimuli in the ERG [abstract]. Doc

Ophthalmol 119:17

21. Marmor MF, Holder GE, Seeliger MW, Yamamoto S

(2004) Standard for clinical electroretinography (2004

update). Doc Ophthalmol 108:107–114

22. Zemel E, Loewenstein A, Lei B, Lazar M, Perlman I

(1995) Ocular pigmentation protects the rabbit retina from

gentamicin-induced toxicity. Invest Ophthalmol Vis Sci

36(9):1875–1884

23. Bui BV, Sinclair AJ, Vingrys AJ (1998) Electroretino-

grams of albino and pigmented guinea-pigs (cavia porcel-

lus). Aust N Z J Ophthalmol 26(Suppl 1):S98–S100

24. Bui BV, Vingrys AJ (1999) Development of receptoral

responses in pigmented and albino guinea-pigs (cavia

porcellus). Doc Ophthalmol 99(2):151–170

25. Vingrys AJ, Bui BV (2001) Development of postreceptoral

function in pigmented and albino guinea pigs. Vis Neurosci

18(4):605–613

26. Behn D, Doke A, Racine J, Casanova C, Chemtob S, La-

chapelle P (2003) Dark adaptation is faster in pigmented

than albino rats. Doc Ophthalmol 106(2):153–159

27. Racine J, Joly S, Rufiange M, Rosolen S, Casanova C,

Lachapelle P (2005) The photopic ERG of the albino

guinea pigs (Cavia porcellus): a model of the human

photopic ERG. Documenta Ophthalmologica 110(1):67–77

28. Heiduschka P, Schraermeyer U (2008) Comparison of

visual function in pigmented and albino rats by electro-

retinography and visual evoked potentials. Graefes Arch

Clin Exp Ophthalmol 246(11):1559–1573

Doc Ophthalmol (2010) 121:135–146 145

123

29. Dodt E, Copenhaver RM, Gunkel RD (1959) Electroreti-

nographic measurement of the spectral sensitivity in albi-

nos, caucasians, and negroes. Arch Ophthalmol 62:

795–803

30. Schmidt B (1965) Sensorial examination in an African

albino child. Acta Fac Med Univ Brumen 25:83–89

31. Kondo M, Piao CH, Tanikawa A, Horiguchi M, Terasaki

H, Miyake Y (2000) Amplitude decrease of photopic erg

b-wave at higher stimulus intensities in humans. Jpn J

Ophthalmol 44(1):20–28

32. Sieving PA (1993) Photopic on- and off-pathway abnor-

malities in retinal dystrophies. Trans Am Ophthalmol Soc

91:701–773

33. Sieving PA, Murayama K, Naarendorp F (1994) Push-pull

model of the primate photopic electroretinogram: a role for

hyperpolarizing neurons in shaping the b-wave. Vis Neu-

rosci 11(3):519–532

34. Ueno S, Kondo M, Niwa Y, Terasaki H, Miyake Y (2004)

Luminance dependence of neural components that under-

lies the primate photopic electroretinogram. Invest Oph-

thalmol Vis Sci 45(3):1033–1040

35. Rufiange M, Dassa J, Dembinska O, Koenekoop RK, Little

JM, Polomeno RC, Dumont M, Chemtob S, Lachapelle P

(2003) The photopic ERG luminance-response function

(photopic hill): method of analysis and clinical application.

Vision Res 43(12):1405–1412

36. Miyake Y (2006) Electrodiagnosis of retinal diseases.

Springer, Tokyo, pp 1–41

37. Rangaswamy NV, Frishman LJ, Dorotheo EU, Schiffman

JS, Bahrani HM, Tang RA (2004) Photopic ERGs in

patients with optic neuropathies: comparison with primate

ERGs after pharmacologic blockade of inner retina. Invest

Ophthalmol Vis Sci 45(10):3827–3837

38. Lachapelle P, Rousseau S, McKerral M, Benoit J, Polo-

meno RC, Koenekoop RK, Little JM (1998) Evidence

supportive of a functional discrimination between photopic

oscillatory potentials as revealed with cone and rod med-

iated retinopathies. Doc Ophthalmol 95(1):35–54

39. Wachtmeister L (1998) Oscillatory potentials in the ret-

ina: what do they reveal. Prog Retin Eye Res 17(4):

485–521

40. Frishman LJ (2006) Origins of the electroretinogram. In:

Heckenlively JR, Arden GB (eds) Principles and practice

of clinical electrophysiology of vision, 2nd edn. MIT press,

Cambridge, p 150

41. Robson JG, Frishman LJ (1999) Dissecting the dark

adapted electroretinogram. Doc Ophthalmol 95:187–215

42. Robson JG, Saszik SM, Jameel A, Frishman LJ (2003) Rod

and Cone contributions to the a-wave of the electroreti-

nogram of the macaque. J Physiol 547(2):509–530

43. Kojima M, Zrenner E (1978) Off-components in response

to brief light flashes in the oscillatory potential of the

human electroretinogram. Albrecht Von Graefes Arch Klin

Exp Ophthalmol 206(2):107–120

44. Hood DC, Birch DG (1996) Beta wave of the scotopic

(rod) electroretinogram as a measure of the activity of

human on-bipolar cells. J Opt Soc Am A Opt Image Sci

Vis 13(3):623–633

45. Fulton AB, Hansen RM (2006) Stimulus-response func-

tions for the scotopic b-wave. In: Heckenlively JR, Arden

GB (eds) Principles and practice of clinical electrophysi-

ology of vision, 2nd edn. MIT press, Cambridge, p 473

46. Abbas M, D’Amico F et al (2009) Structural, electrical,

electronic and optical properties of melanin films. Eur Phys

J E Soft Matter 28(3):285–291

47. Drager UC (1985) Calcium binding in pigmented and

albino eyes. Proc Natl Acad Sci U S A 82(19):6716–6720

48. Yau KW (1994) Phototransduction mechanism in retinal

rods and cones. The Friedenwald Lecture. Invest Oph-

thalmol Vis Sci 35(1):9–32

49. Nakatani K, Yau KW (1988) Calcium and light adaptation

in retinal rods and cones. Nature 334(6177):69–71

50. Brule J, Lavoie MP, Casanova C, Lachapelle P, Hebert M

(2007) Evidence of a possible impact of the menstrual

cycle on the reproducibility of scotopic ergs in women.

Doc Ophthalmol 114(3):125–134

51. Elsner AE, Burns SA, Weiter JJ et al (1996) Infrared

imaging of sub-retinal structures in the human ocular

fundus. Vision Res 36(1):191–205

52. Bone RA, Brener B, Gibert JC (2007) Macular pigment,

photopigments, and melanin: distributions in young sub-

jects determined by four-wavelength reflectometry. Vision

Res 47(26):3259–3268

53. Kilbride PE, Alexander KR, Fishman M, Fishman GA

(1989) Human macular pigment assessed by imaging

fundus reflectometry. Vision Res 29(6):663–674

146 Doc Ophthalmol (2010) 121:135–146

123