Doc Ophthalmol (2018) 136:207–211 https://doi.org/10.1007/s10633-018-9638-x ISCEV STANDARDS ISCEV extended protocol for the photopic negative response (PhNR) of the full-ﬁeld electroretinogram . . . . Laura Frishman Maja Sustar Jan Kremers J. Jason McAnany . . Marc Sarossy Radouil Tzekov Suresh Viswanathan Received: 7 May 2018 / Accepted: 9 May 2018 / Published online: 31 May 2018 The Author(s) 2018 Abstract The International Society for Clinical information about the function of retinal ganglion cells Electrophysiology of Vision (ISCEV) Standard for and their axons. The PhNR can be reduced in disorders full-ﬁeld electroretinography (ERG) describes a min- that affect the innermost retina, including glaucoma imum procedure, but encourages more extensive and other forms of optic neuropathy. This document, testing. This ISCEV extended protocol describes an based on existing literature, provides a protocol for extension to the ERG Standard, namely the photopic recording and analyzing the PhNR in response to a negative response (PhNR) of the light-adapted ﬂash brief ﬂash. The protocol includes full-ﬁeld stimula- ERG, as a well-established technique that is broadly tion, a frequency bandwidth of the recording in which accepted by experts in the ﬁeld. The PhNR is a slow the lower limit does not exceed 0.3 Hz, and a negative-going wave after the b-wave that provides spectrally narrowband stimulus, speciﬁcally, a red L. Frishman (&) R. Tzekov College of Optometry, University of Houston, Houston, Department of Ophthalmology, University of South TX, USA Florida, Tampa, FL, USA e-mail: Lfrishman@uh.edu S. Viswanathan M. Sustar College of Optometry, State University of New York, Eye Hospital, University Medical Centre Ljubljana, New York, NY, USA Ljubljana, Slovenia J. Kremers Department of Ophthalmology, University Hospital Erlangen, Erlangen, Germany J. J. McAnany Department of Ophthalmology and Visual Sciences, Department of Bioengineering, University of Illinois at Chicago, Chicago, IL, USA M. Sarossy Department of Ophthalmology, Centre for Eye Research Australia, The Royal Victorian Eye and Ear Hospital, University of Melbourne, Melbourne, VIC, Australia 123 208 Doc Ophthalmol (2018) 136:207–211 ﬂash on a rod saturating blue background. Suggested Patient population ﬂash strengths cover a range up to and including the minimum required to elicit a maximum amplitude This protocol for recording the PhNR can be used for PhNR. This extended protocol for recording the PhNR testing patients in whom inner retinal integrity, and provides a simple test of generalized retinal ganglion speciﬁcally signaling by retinal ganglion cells and cell function that could be added to standard ERG their axons, may be compromised due to ganglion cell testing. pathology or limitations in the input to the ganglion cells. For example, since 2000, reduced PhNR ampli- Keywords Clinical standards Electroretinogram tudes have been reported in patients with glaucoma (ERG) Full-ﬁeld ERG International Society of [3–6], optic atrophy [7, 8], central retinal artery Clinical Electrophysiology of Vision (ISCEV) occlusion [9, 10], ischemic optic neuropathy , diabetic retinopathy , and idiopathic intracranial Photopic negative response PhNR Optic neuropathy Glaucoma Retinal ganglion cells hypertension . In some cases, the protocol may be useful for monitoring treatment effects in eyes with ocular hypertension or glaucoma . Abnormal potassium (K ) channel activity or other dysfunction Introduction of retinal glia may also be reﬂected in PhNR recordings . This is because generation of the The International Society for Clinical Electrophysiol- PhNR, which has a slow time course (Fig. 1), is ogy of Vision (ISCEV) Standard for full-ﬁeld elec- thought to involve glial K currents that serve to troretinography (ERG) describes a minimum set of remove the excess K released into extracellular space tests, but encourages the use of additional ERG during activation of retinal ganglion cells . protocols for clinical ERG testing . This extended protocol describes the photopic negative response (PhNR) of the ﬂash ERG, as a specialized procedure Technical issues which is well established and broadly accepted by experts in the ﬁeld. The protocol was prepared by the The electrodes and electronic recording equipment for authors in accordance with ISCEV procedures (http:// this PhNR protocol are as described in the ISCEV www.iscev.org/standards/index.html.) and was Standard for full-ﬁeld ERG . The present protocol approved by the ISCEV Board of Directors on March assumes full-ﬁeld stimulation, while acknowledging 25, 2018. that focal stimulation has been shown to be effective in assessing inner retinal function . For the frequency bandwidth of the recording, the ISCEV Standard Scope and applications suggests a minimum range of 0.3–300 Hz. For PhNR recordings, the bottom limit of the ﬁltering could be The photopic negative response (PhNR) of the light- lower to minimize distortion and possible attenuation adapted (LA) electroretinogram (ERG) is a negative- of the slow negative wave. For spectral characteristics going wave that occurs after the b-wave in response to of the stimulus, whereas the ISCEV Standard recom- a brief ﬂash. The PhNR reﬂects generalized activity of mends ‘‘visibly white’’ (broadband) stimuli, narrow- retinal ganglion cells and their axons , and its band stimuli are recommended for recording the amplitude can be reduced early in diseases that affect PhNR. Speciﬁcally, a long-wavelength (red) ﬂash on the innermost retina. The PhNR also occurs in a rod saturating short-wavelength (blue) background response to long-duration ﬂashes, following the b- yields a larger amplitude PhNR than broadband wave at light onset and d-wave at light offset , but stimuli. LED-based stimulators typically provide a most publications to date have described brief ﬂashes. 20-nm half-height bandwidth for the red and blue Only the brief ﬂash PhNR will be addressed in this LEDs. The recommendation for narrowband stimuli is protocol. based on the outcome of studies that compared PhNR amplitudes using broad- vs narrowband stimuli in nonhuman primates  and in glaucoma patients 123 Doc Ophthalmol (2018) 136:207–211 209 Calibration The stimulus strength for the brief ﬂashes can be speciﬁed in photopic candela seconds per meter -2 squared (phot cd s m ); the background in phot -2 cd m . A spectroradiometer (or spectrometer) is required to determine the spectral characteristics of chromatic ﬂashes. Care should be taken to measure a range of ﬂash luminances as some Ganzfeld stimula- tors use different combinations and banks of LEDs for different luminance ranges, and these may have different wavelength speciﬁcations. It is useful also to conﬁrm that the background is strong enough to saturate rod photoreceptors, for example, about 100 -2 scot cd m . Blue backgrounds will saturate the rods while minimizing the photopic stimulus strength and hence the adapting effect of the background on cone- driven responses. Protocol speciﬁcations The procedures for patient preparation and recording are as speciﬁed by the ISCEV Standard for the light- adapted ERG, including pupil dilation and 10 min of light adaptation if the patient was dark adapted for other testing prior to recording the light-adapted ERG. Other speciﬁcations are listed below; (a) The chromatic characteristics of the stimuli. Fig. 1 Illustration of the light-adapted ERG of a healthy Background: steady, blue LED (450–485 nm); -2 subject (35 years.) in response to a brief red LED ﬂash (660 nm) 100 scot cd m ; equivalent to * 10 phot at each of four ﬂash strengths, on a blue background (460 nm) of -2 -2 cd m . Light ﬂash: red LED (630–660 nm). 10 cd m . Figure shows PhNR amplitude measurements from (b) Flash strengths and background luminance. baseline to PhNR trough (BT) and from b-wave peak to PhNR -2 trough (PT). Adapted from Ref.  (the Association for Flash: \ 5 ms; 1.0–2.5 phot cd s m , or the Research in Vision and Ophthalmology is the copyright holder) stimulus strength that produces the largest PhNR amplitude, but does not exceed the initial [6, 18, 19], and more generally on a review of the stimulus strength producing amplitude satura- literature which shows that most studies in patients tion, or lead to the decline in response amplitude have used red LED ﬂashes on blue LED backgrounds. associated with the photopic hill [21, 22]. The It should be noted that other narrowband combinations dynamic range of the stimulus response func- using blue ﬂashes on yellow or orange backgrounds tion generally ranges from * 0.01 to have also been reported to be effective for eliciting a -2 [ 2.0 phot cd s m . robust PhNR [19, 20]. (c) Frequency of ﬂash presentation. Inter-ﬂash interval: 1 s. Some studies have used an interval of 500 ms, but this may not allow enough time for PhNR to fully recover to baseline. 123 210 Doc Ophthalmol (2018) 136:207–211 (d) Recording bandwidth. The low-frequency ﬁlter PhNR amplitudes can be greater than that of a- and b- should be 0.3 Hz or lower; the high-frequency waves [21–24]. ﬁlter, a minimum of 300 Hz. (e) Signal averaging. There should be sufﬁcient repetitions to provide good signal-to-noise Reporting ratio, and many studies have used 20 trials or more. At least 8–10 trials or more are necessary Reporting of results of PhNR testing should include for lower stimulus strengths if a range of stimuli measurements of the a-wave, b-wave, and PhNR and a are used that include weak stimuli, fewer may computation of the PhNR: b-wave ratio. This helps to be necessary for saturated responses. Artifact determine whether the origin of any change in PhNR rejection should be used if available. If single amplitude is at the retinal ganglion cells themselves or responses are saved, noisy responses can be a more distal location in the retina. The choice of removed during off-line analysis before method for measuring PhNR amplitude is open to the averaging. study and the site, but for comparison with other studies, inclusion of the BT measure is advised. Some studies have compared the sensitivity of the ratio of PhNR to b-wave amplitude (i.e., PhNR normalized to Response evaluation b-wave) versus the simple BT measure for detecting glaucoma, and results were mixed [5, 25]. Caution is As shown in Fig. 1, the PhNR amplitude can be needed as the ratio measure could be misleading in measured from baseline to the minimum point in the diseases where the b-wave is abnormal. trough (BT). It also can be measured from the peak of the b-wave to the maximum amplitude in trough (PT). Acknowledgements We would like to thank Anthony G. Alternatively, PhNR amplitude can be measured at a Robson for his advice and careful editing, and the members of ISCEV in particular, Michael F. Marmor for their valuable ﬁxed time, for example, at 65–75 ms after the ﬂash in discussion during the consultation period. the trough of the response (not shown). Using a ﬁxed time could be helpful when responses in diseased eyes are small and the trough is difﬁcult to locate. Note that Compliance with ethical standards the PT measurement is largely dominated by the b- wave amplitude, and a change in b-wave amplitude Conﬂict of interest The authors declare that they have no conﬂict of interest. reﬂecting a change in bipolar cell function must be considered when interpreting a change in PhNR Statement of human rights This article does not contain any amplitude. When measuring the PhNR, it may also studies with human participants performed by any of the be necessary to take account of the i-wave, or i-waves, authors. positive deﬂection(s) of Off pathway origin  in the Statement on the welfare of animals This article does not falling limb of the b-wave, and/or later in the trough contain any studies with animals performed by any of the (Fig. 1). For responses to the suggested narrowband authors. stimuli, such as those used for responses in Fig. 1, the Informed consent This article does not contain any studies maximum trough amplitude generally occurs after the with human participants performed by any of the authors. initial i-wave. Given the slow nature of the response, and the variety of amplitude criteria that have been Open Access This article is distributed under the terms of the used, peak time of the PhNR is generally not reported. Creative Commons Attribution 4.0 International License (http:// creativecommons.org/licenses/by/4.0/), which permits unre- The PhNR is moderately affected by age, so, for the stricted use, distribution, and reproduction in any medium, particular measure(s) chosen, appropriate age- provided you give appropriate credit to the original matched normative data should be used [3, 22]. author(s) and the source, provide a link to the Creative Com- Comparisons of longitudinal ﬁndings in patients to mons license, and indicate if changes were made. normal test-retest repeatability of PhNR amplitudes are also important, as the test–retest variability of 123 Doc Ophthalmol (2018) 136:207–211 211 Appendix: Justiﬁcation for the protocol details optic neuropathies: comparison with primate ERGs after pharmacologic blockade of inner retina. Invest Ophthalmol Vis Sci 45:3827–3837 A systematic literature review was performed using 12. 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Documenta Ophthalmologica – Springer Journals
Published: May 31, 2018
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