Electroretinogram and Clinical Applications

Electroretinogram and Clinical Applications by Dr. Vikas Khatri

1.   ELECTRORETINOGRAM Dr. Vikas Khatri

2. ERG is the corneal measure of an electrical potential generated by the retina in response to a change in illumination which is recorded in biphasic waveform ELECTRORETINOGRAPHY

3. HISTORY 1865 : First known recording of an ERG (amphibian retina) Swedish Physiologist Alarik Frithiof 1908 : Einthoven and Jolly separated the ERG response into three components: a-wave, b-wave and c-wave 1967 : Ragnar Granit Nobel Prize for Physiology and Medicine (demonstrated the physiology of the receptor potential of each component of the ERG) Ragnar Granit

4. The electrical basis of ERG recordings ERG responses are recorded with an Active extracellular electrode positioned Cornea(Humans) Vitreous (Experimental Animals) At different levels inside the retina (Experimental Animals)

5. + - + - + - + - Active Electrode Negative Electrode ECF The electrical basis of ERG recordings ERG responses are recorded with an Active extracellular electrode positioned Cornea(Humans) Vitreous (Experimental Animals) At different levels inside the retina (Experimental Animals)

6. ERG Photoreceptors , Bipolar Cells, Ganglion Cells and Muller Cells are arranged in parallel and therefore, their currents are in parallel and sum up, giving rise to a strong radial extracellular current Cells arranged horizontally cancel the current of each other and cell arranged obliquely have small currents. Therefore, when homogenous light stimulation is directed at the whole retina only radial extracellular currents are formed. Current Pathway A - Current flowing through local route remaining entirely within the retina Current Pathway B - leaves the retina through the vitreous and anterior ocular tissue and returns to the retina through sclera> choroid>pigment epithelium layer

7. Electrical current flows through a resistor, a gradient of electrical potential is formed Ohms Law, V = I x R V - Potential Difference I - Current R - Resistance Each tissue - Retina , Vitreous , Sclera , Choroid , Pigment epithelium - behave as an electrical resistor Change in one of the resistances will cause a change in the magnitude of the current in the extraocular pathway and the ERG can change irrespective of retinal function. Patient undergone vitrectomy surgery and injection of silicon oil into the vitreous, the resistance of the vitreous increases by several folds causing the current  to be so reduced that the ERG becomes very small in amplitude. So if V (Potential Differece by Retina) = Constant Current inversely proportional to Resistance

8. Types of Electroretinogram Full Field ERG Focal ERG Multifocal ERG Pattern ERG

9. FULL FIELD ERG If 20 % or less of the retina is affected with a diseased state the Full-field ERGs are usually normal Summed activity of all retinal cells, and consists of overlapping positive and negative component potentials that originate from different stages of retinal processing Amplifier Stimulator Electrodes

10. 1. Active electrode It’s the main electrode. Can be Placed on : Cornea Conjunctiva Sclera Skin Application of electrodes Types of Electrodes :

11. 2. Reference electrode S ilver chloride electrode. Placed on – Outer Canthi or Zygomatic fossa Serves as the Negative pole as it is placed closer to the electrically negative posterior pole of the eye. Refrence electrode 3. Ground electrode Placed on – Forehead or Earlobe.

12. Recording Protocols Dark room with non-reflecting walls Full pupillary dilatation 30 minutes of dark adaptation Rod response (Scotopic) Maximum combined response Oscillatory potentials 10 minutes of light adaptation S ingle flash cone response (Photopic Response) 30 Hz flicker .

13. Stimulus for ERG Stimulus and background light should be homogeneous and cover the entire retina. Strobe lamp and LEDs - mobile and can be easily placed in front of a person whether sitting or reclining The Ganzfeld bowl - large white bowl which is used to stimulate the retina during the recording of the ERG. It diffuses the light & allows equal stimulation of all parts of retina.

14. a-Wave Initial negative wave Photoreceptors Resting State ( No light) Depolarized - - ------ + + +++++ + ++ + + ++ ++ + + + + - --- ---- Glutamate Hyperpolarised Hyperpolarised Light Depolarised No Inhibition on Bipolar Cells ++ ++ ++ ++ - - - + + + + + + + Third order Neuron Na+ Na+ Na+ Na+

15. b-Wave Resting State ( No light) Depolarized Glutamate Hyperpolarised Hyperpolarised Light Depolarised No Inhibition on Bipolar Cells ++ ++ ++ ++ - - - + + + + + + + Third order Neuron Na+ Na+ Na+ Na+ K+ K+ K+ Muller Cell Permeable to K+ ++ + + ++ ++ + + + + - --- ---- - - ------ + + +++++ + + + +++++ + K + K + K + K+ + + +++ Positive wave from Bipolar and Muller Cells

16. c-Wave Retinal Pigment Epithelium + + + + + + + + + + Apical Membrane (Towards Retina) More Permeable to Potential K+ Basal Membraen (Towards Choroid ) Less Permeable to Potential K+ The standing potential of the eye - - - - - - + + + + + - - - - - - - - - - - - - + + + + + + + + + + - - - - - - - - - - - - - + + + + + + + + + + + + + + + + Resting State ( No light) Depolarized Light > Hyperpolarised Increased Extracellular Positive Charge Increase in Potential Difference due to light induced electrical activity in the photoreceptors Extracellular Space c -wave + + + + +

17. c-wave originates from the pigment epithelium, it depends upon the integrity of the photoreceptors ERG c-wave can be used to assess the functional integrity of the photoreceptors, the pigment epithelial cells and the interactions between them

18. Scotopic Threshold Response The effects of BaCl2 on the STR and extracellular potassium ion concentration recorded from the proximal retina of the dark-adapted cat. Barium ions eliminate the STR but have no effect on the light-induced increase in extracellular potassium concentration in the proximal When very dim light stimuli (below the b-wave threshold) are applied in the dark-adapted state, a slow corneal negative potential is recorded Muller Cell response to K+ ions which affect the membrane potential of Muller cells Corneal negative component that is sometimes followed by a positive component

19. Oscillatory Potentials (OPs) When a bright light stimulus is used to elicit the ERG in humans or in animals, low amplitude oscillating waves can be identified on the rising phase of the b-wave . Frequency - 100-150Hz   Can be extracted with a band pass filter with a low-frequency cutoff of 75 Hz . Origin - Neuronal Inhibitory Feedback Circuits in Inner Retina and Amacrine Cells Isolating the oscillatory potentials from the bright flash ERG response of the human eye (a) by applying a digital filter (b). An FFT procedure was applied to the isolated oscillatory potential in order to obtain the power spectrum ( c)

20. Oscillatory potentials are significantly attenuated in various retinal degenerations amongst them are the following : Retinitis pigmentosa Central serous retinopathy Retinoschisis Early stage of Diabetic retinopathy Hypertensive retinopathy CRVO and CRAO Takayasu’s (pulseless) disease Two patients with diabeticretinopathy ( DR ) Delayed implicit time (Case 1) Reduced amplitude (Case 2 )

21. Factors affecting the ERG State of adaptation Light intensity Colour of light stimulus Frequency of photo stimulation Rod-mediated vision is very sensitive to dim light stimuli in the dark-adapted state. In Constant background illumination, Rod Photoreceptor saturates and does not respond to light increment or decrement ERG is of considerably larger amplitude (about 4 folds) and is characterized by slow temporal properties; time to peak of the b-wave is about 60ms This ERG response is a mixed rod-cone response (the cone system is operational too) but mainly reflects the activity in the rod system since the cone system contribution is considerably smaller . Cone-mediated vision is not as sensitive but is characterized by the ability to adapt to bright lights: processes that allow vision to adapt to background illumination over a wide range of intensities . ERG under these conditions is of small amplitude but of very fast kinetics

22. Factors affecting the ERG State of adaptation Light intensity Colour of light stimulus Frequency of photo stimulation b-wave Amplitude increases Size of the pupil is the major determinant of light intensity to the retina 3 fold change in pupil diameter = 9 fold change in light intensity reaching the retina a-wave precedes b-wave a-wave and the b-wave increase in amplitude Oscillatory potentials seen on the rising phase of the b-wave b-wave Amplitude increases

23. State of adaptation Light intensity Colour of light stimulus Frequency of photo stimulation Scotopic Condition - STR (Scotopic Threshold Response) First wave at -8.2 log units - b-wave Appears at -5.8 log units Saturates at -3.4 log unit - a-wave Appears at -1.7 log unit - OP (Oscillatory Potential) Light intensity higher than -0.8 log units 0 log unit = is 44.2 candela/meter 2 /second 1 Factors affecting the ERG

24. Factors affecting the ERG State of adaptation Light intensity Colour of light stimulus Frequency of photo stimulation Photopic Condition Short-flash ERGs elicited by increasing stimulus intensities b-wave maximum amplitude at 3.0 log unit Photopic Hill Phenomenon – b-wave amplitude decreases after maximum flash intensity forming inverted U shape (cone photoreceptor desensitization and Pigment bleach)

25. Factors affecting the ERG State of adaptation Light intensity Colour of light stimulus Frequency of photo stimulation Rod curve peaks at the blue-green region of the visible spectrum ( around 500nm ) Spectral sensitivity curve of cone-mediated vision is the sum of all three spectral types of cone; long-wavelength sensitive ( red ), medium-wavelength sensitive ( green ), short-wavelength sensitive ( blue ), In net, Cone has peak sensitivity in the orange range of the visible spectrum (around 560nm)

26. Factors affecting the ERG State of adaptation Light intensity Colour of light stimulus Frequency of photo stimulation Dark-adapted state Dim Blue Stimuli Bright Red Stimuli The red light stimulus produces an ERG response composed of two parts; a fast wave (30ms) a slow wave ( 100ms) The blue stimulus elicits a slow positive ERG of the more sensitive rod system With this procedure, cone-mediated function can be isolated from the large amplitude rod ERG and allows analysis of cone vs rod system in the dark-adapted state Equality in the slow ERG component when  two light stimuli were balanced

27. Factors affecting the ERG State of adaptation Light intensity Colour of light stimulus Frequency of photo stimulation Separate rod-mediated vision from cone-mediated vision Maximum frequency of stimulation that can be perceived as flickering – Critical Fusion Frequency(CFF) Highest CFF - Rod vision – 15-20Hz Highest CFF - Cone vision – 30-50Hz Bright light stimuli at a frequency of 30Hz is applied in order to isolate the cone system from the rod system

28. International Society for Clinical Electrophysiology of Vision (ISCEV) Standardised the protocols for performing electrophysiological tests (1989 ) Ensures uniformity and thus comparability between labs Dark-adapted 0.01 ERG A rod-driven response of bipolar cells Dark-adapted 3.0 ERG Combined responses arising from photoreceptors and bipolar cells of both the rod and cone systems; rod dominated

29. Dark-adapted 3.0 ERG with Band-pass Filter Oscillatory potentials Responses primarily from Amacrine cells Dark- adapted 10 ERG Combined response with enhanced a-waves reflecting photoreceptor function

30. Light- adapted 30 Hz flicker ERG A sensitive cone-pathway-driven response Light-adapted 3.0 ERG A cone-driven response of bipolar cells Recorded with a stimulus intensity of 3.3 log units Background illumination - 40 candela/meter 2 /second (Sufficient to suppress all rod activity)

31. Measurement of ERG Components Amplitude a-Wave amplitude – baseline to trough of a-wave b-Wave amplitude – trough of a-wave to peak of b-wave Time Sequences Latency – Stimulus to onset of wave response Implicit Time – Stimulus to Maximum wave response

32. Ratio of b-wave to a-wave ERG analysis that is based only upon amplitude measurements may lead to erroneous conclusions if the : Pupil is not maximally dilated Different recordings conditions  E xchange of information between laboratories So this Ratio give us information about normal signal transmission between the photoreceptor and post-synaptic neurons

33. ELECTRORETINOGRAM CLINICAL APPLICATIONS

34. Normal ERG Normal ERG can be seen in patients of : Localized macular dysfunction Optic nerve diseases Central nervous system disease such as amblyopia.

35. Subnormal ERG Amplitudes of all components are reduced approximately to the same degree Early stages of rod–cone dystrophy PRP for Diabetic retinopathy ( ERG components are reduced by 40–45%) b/a ratio remain normal Vitreous Haemorrhage (Hazy Media) - ERG + USG can be used to differentiate between Total RD and Dense Vitreous Membrane Case 1 - ERG is recordable, even if the amplitude is small, the thick membrane in the vitreous cavity is not totally detached retina, but vitreous membrane Case 2 - When the ERG is unrecordable , the thick membrane is most likely totally detached retina

36. Localized damage of the photoreceptors (Partial RD or Sectoral RD) - Amplitude of the full-field ERG is proportional to the area of functioning retina. Reduction of the ERG amplitude corresponds proportionally to the extent of RD

37. Negative ERG Indicates that the amplitude of the b-wave is smaller than that of the a-wave (b/a ratio <1.0 ) Normal a-wave with a reduced b-wave localizes the defect to post-synaptic photo transduction processes Negative ERG can be of useful Prognostic or Diagnostic value in retinal diseases Endophthalmitis after intraocular lens implantation Endophthalmitis (<1 Week) b/a ratio < 1.0 Endophthalmitis (<1 Week) b/a ratio > 1.0 Late-onset endophthalmitis b/a ratio > 1.0 Good Prognosis Worst Prognosis Should Undergo Vitrectomy Urgently

38. Central Retinal Artery Occlusion No b-Wave a-Wave Recordable : Outer Retina - Choroidal perfusion Ophthalmic artery occlusions usually result in unrecordable ERGs

39. Central Retinal Vein Occlusion Ischemic CRVO shows Negative ERG more frequently than Non Ischemic type b/a ratio can be an important index for evaluating the prognosis of CRVO

40. Proliferative Diabetic Retinopathy If patient present with VH it is difficult to predict the surgical and visual outcome after vitrectomy Most diabetic patients with vitreous hemorrhage have already undergone PRP and PRP reduces the ERG amplitude without changing the b/a ratio, So it is difficult to arrive at a prognosis of the outcome after vitrectomy using only the amplitudes b/a ratio provides more useful information about the visual prognosis after vitrectomy

41. b/a ratio>1.0 and the OPs are clearly recordable - b/a ratio >1.0 but the OPs are absent - b/a ratio <1.0 with absent Ops - Hiraiwa T, Horio N, Terasaki H, et al. Preoperative electroretinogram and postoperative visual outcome in patients with diabetic vitreous hemorrhage. Jpn J Ophthalmol 2003;47:307–11 . The postoperative visual acuity for group C was significantly worse than for group A or group B This observation is important when we discuss the visual prognosis with patients before surgery

42. ERG recordings in a normal patient and one with retinitis pigmentosa with residual cone physiology Retinitis Pigmentosa ERG is usually normal or only slightly subnormal in these infectious diseases. Pigment in the retina is prominent in many infectious diseases. Early stages of Syphilis and Rubella can mimic the fundus appearance of RP

43. Full-field ERGs are best for quantifying cone dystrophy Scotopic ERGs fairly normal in appearance but with slow implicit times. Cone Dystrophy 30 Hz flicker ERG Photopic white ERG Dependent on Functioning of Cones

44. Stargardt’s disease Full-field ERGs in these disorders are normal except in very late stages where full-field ERGs may become slightly subnormal. Macular multifocal ERGs are dramatically abnormal.

45. Congenital Stationary Night Blindness CSNB show essentially normal fundi and most patients with CSNB have moderately low visual acuity, So, ERG can be used to differentiate them from other diseases with normal fundi, low visual acuity, and normal ERG . Normal a-wave b/a < 1.0 Normal a-wave b/a > 1.0 Small a-wave b/a < 1.0 Without Photopic Hill Phenomenon Small a-wave b/a < 1.0 With Photopic Hill Phenomenon Psychological eye problems Amblyopia Optic nerve disease CNS disease Congenital Stationary Night Blindness Retinitis Pigmentosa Oguchis disease

46. Foreign bodies and Trauma Estimate the extent of retinal dysfunction Small piece of stainless steel or plastic outside the macula may have a minor affect on the retina . Copper or Iron would likely have deleterious affects within a few weeks In general if b-wave amplitudes are reduced 50% or greater compared to the fellow eye, it is unlikely that the retinal physiology will recover unless the foreign body is removed. Fundus photo of a patient with a hole in the retina caused by a metallic foreign body

47. Multifocal electroretinography is better way to assess the Drug Toxicity and should be Screened every 5-6 months to see Functional Retinal Changes Chloroquine Retinopathy

48. Extinct ERG Advanced stage of rod–cone dystrophy Retinitis pigmentosa Gyrate atrophy Choroideremia Total Retinal Detachment Even when the macular area is preserved, ERG may become undetectable .

49. FOCAL ERG Evaluate Macular Function Change in macular OPs are the most sensitive indicator in variable macular diseases.

50. Selective reduction of macular OP amplitude is observed in : Macular Edema Epimacular membrane Central Serous Chorioretinopathy Pseudophakic Cystoid Macular Edema

51. Occult macular dystrophy (OMD) Focal ERG or Multifocal ERG - key for diagnosis Normal fundus, FFA, Full-field ERGs, but abnormal focal macular ERG, multifocal ERG, OCT show mild Photoreceptor abnormalities

52. MULTIFOCAL ERG Record many focal retinal responses simultaneously in a brief time period Array of hexagons 64 or 103 hexagons over 30–40 ° of central visual field Recorded under photopic condition (To asses Foveal response) Every frame change ( 13.3ms) of the monitor , each hexagon can be either white or black based on a pseudorandom probability sequence (called an m-sequence )

53. Photoreceptor layer Disruption Normal Occult Macular Dystrophy

54. Full-field ERG showing Normal Response Occult Macular Dystrophy

55. Age Related Macular Degeneration (AMD)

56.   Patient with Early Age Related Macular Degeneration (AMD)

57. Multifocal ERG recordings in a patient with Stargart’s disease

58. PATTERN ERG Measure of macular function and generalized bipolar cell function . Checkerboard stimulus composed of white and black squares. Reduction of PERG amplitude reflect the reduced activity of dysfunctional RGCs. Inner retinal activity under light-adaptation.

59. The Normal Pattern Electroretinogram : • N35- a small negative component with a peak time occurring around 35ms • P50- a prominent positive wave emerging around 50ms • N95- a wide negative wave around 95ms

60. Macular diseases: • The P50 component was shown to be altered in all patients with retinal and macular diseases . Optic nerve disease : • N95 component was abnormal in 81% of patients with diseases of the optic nerve. The P50 component remain normal.

61. Thank You