Multifocal Electroretinogram

Anatomy and Physiology

The retinal structural architecture includes 10 layers constituting various cell types and their synaptic connections for visual processing. The inner retina comprises nerve fiber layer axons, ganglion cells and their dendritic synaptic connections, and amacrine cells. The outer retina comprises the rod and cone photoreceptors, which transfer visual information to second-order bipolar cells in the middle retina. Cones are found in the highest concentrations in the central macula or fovea, the area of the retina responsible for visual acuity. Following phototransduction from the outer retina, the inner retinal ganglion cells transmit this electrical information to the brain via the optic nerve for visual information processing.[1]

Indications

The mfERG is a specialized test beyond standard ophthalmologic examination. Testing may be indicated in the following scenarios:

• The diagnostic and discriminatory ability between various types of retinal diseases
• Ruling out outer retinal disease
• Monitoring disease progression
• Differentiating organic from nonorganic disease entities[2]

Contraindications

There are no specific contraindications for the mfERG.

Equipment

The following equipment is required:

• Electrodes
• Amplification system
• Data recording and display system

Personnel

Appropriately trained technicians perform mfERG in large referral centers with an electrophysiology laboratory. Retina specialists and neuro-ophthalmologists are typically responsible for interpreting the electrophysiological results.

Preparation

Electrode Placement

Recording Electrodes

• Depending on the electrode type (contact lens, foil, fiber electrodes), the recording electrode is placed on the corneal surface, on the bulbar conjunctiva adjacent to the inferior limbus of the cornea.
• The electrode type can affect a response's signal-to-noise ratio. Bipolar corneal contact electrodes characteristically exhibit the highest signal-to-noise ratio. In contrast, foil or fiber electrodes require longer recording times, repeated measurements, and few stimulus elements to acquire comparable signal-to-noise ratios. 

Reference Electrodes

• Separate surface electrodes are placed on the skin close to the outer canthus of each ipsilateral eye.
• For monocular mfERG recordings, the contralateral occluded eye may place the reference electrode.
• The forehead, earlobe, or mastoid are not recommended for placement as these sites may contaminate the mfERG with potentials generated by the fellow eye. 

Ground Electrodes

• Typically placed on the forehead and connected to the “ground input" of the recording system.

Patient Preparation

Per ISCEV Guidelines

Pupils

• During contact lens electrode use, the pupils should be maximally dilated and centered within the ring of the corneal electrode.
• Patients unable to see the fixation point may be advised to look straight ahead while maintaining a steady gaze.
• Binocular recording is recommended as it improves fixation stability and reduces examination time.
• A monocular recording is recommended in individuals with ocular misalignment. 

Patient Positioning

• Encourage comfortable seating and relaxation of neck and facial muscles to reduce muscle-related artifacts. Consider using a head or chin rest as needed.

Fixation Monitoring

• Instructing patients to fixate on a target within the stimulator while minimizing eye movement is critical. Careful, direct monitoring of fixation is encouraged to ensure fixation stability. For patients exhibiting eccentric fixation, the fixation target may be moved.

Refraction

• Eyes should be refracted at the viewing distance for optimal acuity, and the same correction should consistently be used for a given patient to ensure comparable results. Alternatively, lenses can be placed in a holder before the eye. In the latter case, care must be taken to avoid blocking the view of the stimulus screen by the rim of the lens or the lens holder and thus creating an apparent scotoma.

Monocular Versus Binocular Recording

• The monocular recording is the standard for the mfERG.
• The binocular recording is only used when the eyes are aligned.

Adaptation

• Patients exposed to strong light stimuli from alternative imaging techniques (eg, fundus photography or fluorescein angiography) require a minimum of 15 minutes of recovery time in normal room lighting.

Room Illumination

• Background illumination beyond the checkerboard stimulus typically involves dim or ordinary room lighting. For all recordings, ambient lighting should be the same; reflections from lens surfaces and bright lights should be kept from a subject’s direct view.[3]

Technique or Treatment

The mfERG is traditionally recorded in photopic conditions. This excludes rod contributions to the signal and primarily ensures a cone-driven response. The mfERG waveform includes an initial negative deflection (N1), followed by a positive deflection (P1), and a second negative deflection (N2).[2][3] 

Waveform Components

N1

The N1-wave is the initial negative deflection corresponding to cone photoreceptor cell activity. This wave component largely measures outer retinal function.

P1

The P1-wave is the positive deflection following the N1-wave. It represents the depolarization of inner retinal Muller, bipolar, and amacrine cells and provides a measure of phototransduction activity.

Waveform Analysis 

The mfERG can be analyzed according to the amplitudes and implicit times of the wave components. The amplitude and implicit time of P1 are generally the standard measurements. The amplitude and timing of N1 may also be measured. However, these measurements are not part of the current standard.

Amplitude

The amplitude is the maximal light-induced electrical response (voltage) of the various retinal cells. The mfERG amplitude is the trough-to-peak amplitude, measured from the trough of N1 to the peak of P1.

Implicit Time

Implicit time (time-to-peak) refers to the time needed for the electrical response to reach maximum amplitude. Implicit time is measured from stimulus onset to the corresponding wave-component peak and reflects the signal conduction rate.

Protocols

The ISCEV standards for generating the mfERG response are designed to minimize variability between procedures, thus enabling data to be compared among laboratories.  ISCEV defines the following clinical protocols for mfERG stimulus parameters and recording:

Stimulus and Recording Parameters  

Stimulus Size and Number of Elements

The stimulus display should comprise an array of 61 or 103 black-and-white hexagons over 40-50 degrees (20-25-degree radius to display edge) of the central visual field, including a central fixation target. 

Duration of Recording

Recording the mfERG for a minimum duration of 4 minutes for 61-element arrays and 8 minutes for 103-element arrays is recommended. The recording time is typically divided into shorter segments of approximately 15–30 seconds to permit patient rest between runs. This helps minimize data loss secondary to movement, noise, or other potential artifacts.

Stimulation

Stimulus Source

Cathode-ray tube monitors have traditionally been used to display mfERG stimuli. However, they have recently been replaced with liquid crystal displays. Since alternative sources of stimulation can impact the mfERG waveform, it is important to specify the details of the model used.  

Frame Frequency

Cathod-ray tube frame frequencies of 75 Hz (most common) and 60 Hz may be used. Since variation in frequency may alter the mfERG response, normative values for healthy individuals must be separately determined for a given frequency. Therefore, it is important to note the frame frequency during data interpretation.  

Luminance and Contrast

• When using cathode-ray tubes, stimulus luminance should be at least 100 cd/m2 in the light state. In the dark state, luminance should be low enough to achieve a contrast (Michelson) of at least 90%.
• A higher luminance setting may be required to achieve clear waveforms with reasonable mfERG amplitudes in the light state when using liquid crystal displays.
• For all recordings, the mean stimulus luminance should match the background luminance.

Calibration

Since luminance and contrast affect signal recordings, it is important to calibrate the stimulus according to ISCEV guidelines. In particular, the light and dark elements are not always uniform for many monitors, and variations greater than 15% are unacceptable.

Stimulus Parameters  

Stimulus Pattern

The Standard display is a hexagonal stimulus with larger peripheral hexagons and smaller central hexagons. This scaled pattern produces mfERG responses of approximately equal amplitudes over the healthy retina.

Temporal Sequence

The m-sequence is the standard for routine testing in mfERG. This algorithm determines the rate at which hexagonal elements change between dark and light stages with every frame.

Stimulus Size and Number of Elements

The size of the contrast-reversing stimulus may comprise an array of either 64 or 103 black-and-white hexagons over 30-40 degrees of the central visual field. The width of the stimulus field must include the blind spot. The selection of 64 versus 103 elements depends on balancing good spatial resolution and a high signal-to-noise ratio while minimizing the recording time.

Fixation Targets

Stable fixation is critical for acquiring reliable mfERG recordings. To avoid a diminished response, fixation targets should cover the central stimulus element minimally. The examiner should also confirm that the patient properly visualizes the fixation target.

Signal Recording

Amplifiers and Filters:  amplifiers and filters produce recognizable signals and remove confounding electrical noise. A filter bandpass range of 5 to 200 Hz is acceptable for basic mfERG, with 3 to 10 Hz and 100 to 300 Hz ranges for the high and low pass cutoffs, respectively. Since filter settings within these ranges can variably alter the waveform, the same settings should be uniformly applied for all patients tested in a given study.

Signal Analysis  

Artifact Rejection: artifact rejection algorithms remove sources of signal distortion, such as artificially induced blinks or movement.

Spatial Averaging: Spatial averaging may eliminate noise and smoothen waveforms. The contribution from the averaged neighboring elements should not exceed 17% to ensure equal influence by each of the 6 neighbors for a given hexagon.  

Signal Extraction/Kernels.

The first-order kernel is the standard response. The second-order kernel and other higher-order kernels are used in special circumstances and are occasionally reported.  

Interpreting and Reporting the mfERG

• Carefully examine the mfERG trace array for areas of diminished or delayed signals.
• Assess mfERG normality based on the overall appearance of the waveform and comparison with locally available normative data.
• Evaluate 3-D representations and ring response plots to help further identify potentially damaged areas.

Displaying Results

 Trace arrays:

• Displays an array of the mfERG traces for visually inspecting topographic variability and the quality of recordings.
• Trace lengths of 100 ms or more should be used.
• It can be spatially compared to visual field tests to help evaluate potential relationships between retinal dysfunction and visual field defects.

Topographic (3-D) response density plots:

• Depicts the overall signal strength per unit area of the retina.
• Observing the location and depth of the blind spot can be useful for assessing the quality of fixation. The presence of a blind spot assures good fixation. The absence of a blind spot can be due to poor fixation or a generalized signal loss due to disease.
• Limitations of the 3-D plot include:
  • Loss of waveform information. Thus, large but abnormal or delayed responses can produce normal 3-D plots.
  • A central peak in the 3-D plot can be seen in some records without any retinal signal. The appearance of the 3-D plot from a given recording depends on how the local amplitude is measured. For these reasons, 3-D plots should not be used without a simultaneous display of the trace array.

Ring and other regional averages: regional responses can be averaged per hexagon and compared between affected and unaffected or control eyes. Averaged responses within successive rings may also be used to evaluate visual dysfunction in patients with radial asymmetry. Since the stimulus hexagons are scaled to provide approximately equal response amplitudes, responses are approximately constant across rings. Responses within a ring can further be calculated as amplitude/unit area, whereby the summed responses in each ring are divided by the total area of the hexagons in the ring and plotted as nV/ deg2. The largest response is observed in the central foveal region, given the high density of cone photoreceptors and bipolar cells. Measurements and calibration marks: all traces/graphs must include calibration marks for adequate patient data comparison.

Normal values

Each laboratory should develop its own age-adjusted, normative database. Median values rather than mean values should be reported, given electrophysiologic data are not always normally distributed.[2][3]

Complications

The mfERG is a non-invasive test with minimal risks. Patients may experience mild ocular discomfort during the procedure or, in sporadic cases, develop a corneal abrasion depending on the type of electrode used.

Interfering Factors

• Deviating from standardized testing conditions (ie, lighting, flash intensity, recording environment, duration of light or dark adaptation, and pupil size)
• Electrode-based artifacts, including poor contact with skin or cornea, incorrect placement, unstable position, and high electrical impedance
• Eye blinking or movement
• Defocus or uncorrected refractive error
• Reduced electrical response with aging
• Ocular media opacification
• Diurnal fluctuation
• Depressed response under anesthesia
• Variability in recordings between different device types and normative databases between laboratories[3]

Common Artifact Types

• Line frequency interference
• Movement errors
• Eccentric fixation
• Positioning errors and head tilt
• Erroneous central peak (weak signal artifact)
• Averaging and smoothing artifacts
• Blindspot[2]

Clinical Significance

The mfERG is a more recent electrophysiologic test that detects and localizes distinct areas of outer retinal damage in the macula and paramacular and discrete peripheral areas. This precision enables electrophysiologic findings to be correlated with visual field testing. The mfERG is valuable in evaluating patients with ambiguous retinal diseases and monitoring the disease's progression. An abnormal mfERG generally reflects foveal cone and/or bipolar cell dysfunction along with the source of vision loss.[2] Therefore, damage to the inner retina may have minimal effects on the mfERG waveform.[4][5][6] In turn, the mfERG is most applicable to patients with focal deficits in visual function and an otherwise normal-appearing fundus, as commonly seen in macular dystrophies.[2]

Retinitis Pigmentosa

Retinitis pigmentosa (RP), a generalized retinal degenerative disease, is the most commonly inherited disease of the outer retinal rod-cone photoreceptors. The mfERG shows strong central responses with weak or flat signals in the peripheral rings. The mfERG is especially useful in the late stages of retinitis RP, when standard ERG tests may not be recordable given the severity of the disease. In these cases, the mfERG exhibits a generalized diminished response. Intriguingly, the mfERG is also affected in asymptomatic RP carriers, demonstrating patchy areas of retinal dysfunction. 

Hydroxychloroquine Retinopathy and Bull’s Eye Maculopathy

Hydroxychloroquine (Plaquenil) is a commonly prescribed anti-inflammatory medication for treating rheumatologic and dermatologic conditions. Hydroxychloroquine has recently been suggested as a potential off-label therapy for coronavirus, COVID-19. Among its side effects, this medication has been associated with macular retinal toxicity. Specifically, the macular rod and cone cells are damaged, while the foveal cones are spared. In advanced disease stages, this pattern of retinal loss results in a bullseye appearance, also known as bull’s eye maculopathy. A pericentral loss in the mfERG response is the most characteristic of hydroxychloroquine toxicity. The mfERG has prognostic value by identifying patients who are more prone to retinal toxicity. Early cessation of medication is necessary to avoid irreversible vision loss. Hydroxychloroquine is generally continued in patients with a normal mfERG, with the test repeated annually, whereas individuals with significant mfERG loss are advised to stop the medication.[7]

Stargardt’s Macular Dystrophy

Stargardt's macular dystrophy is a recessive retinal disease caused by a mutation in the ABCA gene. The clinical exam typically reveals a normal fovea with mid-peripheral flecks and poor central vision. However, patients may also present atypically without visibly appreciable flecks or diminished central vision. In affected patients, the central and paracentral responses in the mfERG is significantly diminished. Conversely, a normal mfERG excludes Stargardt's disease. 

Occult Macular Dystrophy

Occult macular dystrophy (OMD) is a rare, inherited retinal degenerative disease. Patients may clinically present with unexplained, progressive central vision loss despite a normal-appearing fundus, decreased color vision, and normal fluorescein angiogram. Affected patients exhibit a decreased central response density on mfERG.

Branch Retinal Artery Occlusion

Branch Retinal Artery Occlusion (BRAO) is retinal ischemic damage caused by a blockage in a branch of the central retinal artery. The mfERG response is characteristically decreased in a “cookie-cutter” fashion corresponding to the pattern of the affected retinal arterial blood supply. This diminished mfERG response is notably observed despite a normal-appearing retina in chronic disease.[8] 

Multiple Evanescent White Dot Syndrome

Multiple Evanescent White Dot Syndrome (MEWDS) is an atypical retinal inflammatory disease that commonly affects otherwise healthy young to middle-aged women. Patients are clinically present with photopsia and an enlarged blind spot on visual field testing. The mfERG response is characteristically depressed in the retinal region corresponding to the blind spot. Since these ocular findings are self-limiting and frequently accompanied by flu-like symptoms, MEWDS is often called the “common cold” of the retina. Similarly, these mfERG abnormalities are generally reversible, and retinal responses return to normal after a few months.[9] 

Neurodegenerative Disease

Alzheimer Disease: the leading form of dementia pathologically hallmarked by fibrillar beta-amyloid and hyper-phosphorylated tau deposition in the central nervous system (McKhann G, Drachman D, et al, Neurology, 1984). Notably, abnormal protein deposition and degenerative changes have also been reported in the retina.[10][11] Recent mfERG studies have suggested retinal electrophysiologic dysfunction in AD. In particular, mfERG responses in Alzheimer Disease have shown significant amplitude reduction in the foveal and perifoveal outer retina.[12] Studies have also provided evidence of retinal dysfunction in early Alzheimer Disease. mfERG responses have shown decreased amplitudes along with implicit time prolongation.[13]

Parkinson Disease: a neurodegenerative disease involving abnormal accumulation of α-synuclein (α-syn) protein resulting in dopaminergic neuronal atrophy.[14] A diminished P1 amplitude density in the mfERG response has been shown as a significant non-invasive clinical biomarker for diagnosing Parkinson Disease.[15]

Enhancing Healthcare Team Outcomes

The retina is a complex neuronal structure, and patients with retinal disease frequently present with difficult-to-diagnose causes of vision loss. Given this challenge in identifying the etiology of disease within the retinal infrastructure, an interprofessional team approach is essential for providing sufficient patient care. Patients with acute onset vision loss commonly present in the emergency department, where nurses, as the first point of contact, triage the patients according to the severity and acuity of symptoms. Medical professionals, including physicians and nursing staff, routinely order extensive and expensive testing for conditions concerning indeterminate clinical diagnosis, most of which returns unremarkable. In turn, patients are instructed to follow up with an outpatient ophthalmologist. Typically, this involves a standard eye examination by a comprehensive ophthalmologist. However, these preliminary diagnostic tests largely detect structural abnormalities that are inconsistent with clinical presentation. In turn, patient diagnoses may be mistaken for a benign condition and, in some cases, presumed to be malingering. When the cause of vision impairment remains undefined, despite extensive medical workup, neuro-ophthalmology is the field to which clinicians often turn.[16]

Considering a broad differential is important to distinguish retinal dysfunction from similar-appearing causes since the choice of therapy depends on the underlying etiology of the disease process. Electroretinography, in conjunction with clinical findings, provides invaluable data for patient management while avoiding unnecessary testing. Transparent communication and care coordination between nurses, physicians, and ophthalmologists, including subspecialists, is essential for deriving a correct diagnosis and therapeutic decision-making with proper management.