1. Marilyn Zúñiga Advisor: Dr. Roorda Supervisor: Dr. Grieve Site: University of California-Berkeley Imaging Intrinsic Signals in the Retina Using Different Powers of Red Light as a Stimulus

2. Outline Background –Anatomy of Eye –References Experiment –Adaptive Optics Scanning Laser Ophthalmoscope (AOSLO) –Methods: Dual Frame Imaging Results –Systematic Trend Observed with Higher Energies –Limitations –Future Experiments

3. Background: The Retina Fovea

4. Motivation In brain imaging, brain function is assessed by looking for changes in scattered light in response to a stimulus Researchers have recently observed similar reflectance changes in the retina in response to a visible light stimulus We want to look for these intrinsic signals in the retina using AOSLO

5. Previous Research Detected IR Changes On Subjects After Stimulus Exposure Author Imaging MethodSubject Flash TypeFlash Duration Flash PowerIR Change TsunodaFundusMacaqueWhite1 ms 41.9 cd- s/m 2 Decrease AbramoffFundusHumanGreen3 sec10 cd/m 2 Decrease GrinvaldFundusCatGreen1 secN/A Increase dim stimulus; decrease bright stimulus BizhevaOCT Rabbit in vitroWhite200 ms 2300 photons/ rod/flashIncrease SrinivasanOCTRat in vivoWhite1.3 s 1400 cd/m 2 Increase Roorda and GrieveAOSLOHumanRed0.5-5 sVarious

6. Horizontal Scanner Wavefront Correction Vertical Scanner Eye Laser Confocal Pinhole PMT Wavefront Sensing Light Detection Light Delivery Raster Scanning Deformable Mirror Experimental Methods: The AOSLO red on 1.2 degrees (~ 360 microns)

7. Method: Dual Frame Imaging Frame 1: Red LaserFrame 2: Infrared Laser red on IR on red on IR on

8. Procedure: Intrinsic Imaging of Retina Frame 1: Red LaserFrame 2: Infrared Laser Dual Imaging Sets of 5 Randomized Trials (30 seconds) Constant infrared light (840 nm) Control (no red flash stimulus) Flashes of Red Light (658 nm; 0.5-5 s)

9. Image Analysis 1 1)Compensation for Eye Movements –Signals may be detected from different parts of the retina as the eye moves Solution –Stabilization of movies to better extract the signal

10. Image Analysis 2 2)Compensation for Eye Blinking - Intensity changes due to blinking are larger than the desired signal Solution - Compare stimulated vs. unstimulated region of the retina (ratio of upper to lower half of IR image) Frame 1: Red Laser Frame 2: Infrared Laser

11. Intensity Ratio of Stimulated to Unstimulated Retinal Areas 051015202530 Preliminary Testing Standard Deviation Mean of 5 Trials Legend Intensity Ratio Time (seconds) 0.92 0.96 1.00 1.04 1.08 Two Second Red Flash Energy: 2.00E-05 Joules Visible Light Stimulus

12. Experiment One: Half Second Red Flash with Designated Powers Time (seconds) Energy (Joules) 1.00E-08 2.14E-082.04E-06 9.55E-07 4.47E-07 2.09E-07 9.77E-08 4.57E-08 Control 9.33E-06 4.37E-06 Intensity Ratio 051015202530 0.92 0.96 1.00 1.04 1.08 051015202530 Standard Deviation Mean of 5 Trials Legend

13. Energy (Joules) 1.02E-05 3.12E-05 8.40E-05 7.26E-05 5.16E-05 Control Experiment Two: Three Second Red Flash with Designated Powers Time (seconds) Intensity Ratio 0.92 0.96 1.00 1.04 1.08 510152025300510152025300 Standard Deviation Mean of 5 Trials Legend

14. Image of Retina Before Stimulation Image of Retina After Stimulation

15. Summary/Conclusion Systematic change (increase in scattered lR light in response to stimulation) observed with higher flash energies and a longer flash duration Future Plans More testing with different stimuli and possibly patients with retinal disease

16. Acknowledgements Austin Roorda; Advisor Kate Grieve; Supervisor Kaccie Li Ethan Rossi Pavan Tiruveedhula Pinky Weiser Yuhua Zhang Funding Provided by NSF and the Center for Adaptive Optics Grant No. AST- 9876783

17. Thank you! Questions?