Voltage-Sensitive Dye Optical Imaging Vassiliy Tsytsarev Department of Anatomy and Neurobiology University of Maryland School of Medicine

Principles of Voltage-Sensitive Dye Optical Imaging RH-795 chemical structure Dye interactions with the lipid bi- layer Unbound to the lipid bilayer dye molecules are not fluorescent Bound dye molecules are fluorescent (after: Grinvald et al In-vivo Optical imaging of cortical architecture and dynamics)

Main Parts of the VSD Imaging Setup Figure: Schematic of measurement CCD camera Excitation Filter ( nm) Emission Filter (695nmLP) Dichroic mirror LFP E-stim 4AP Figure 2 Schematics of experimental setup Emission Filter (695 nm LP) CCD camera Dichroic mirror Electrical stimulation LFP recording Intracortical Injection Excitation Filter (620± 20 nm Light Shutter

How VSD works? (After: Tsytsarev et al, 2008 Imaging cortical electrical stimulation in vivo: fast intrinsic optical signal versus voltage-sensitive dyes) Electrode 1 mm Frame = 10 ms VSD vs fast IOS

VSD Imaging of Tonotopicity 1 kHz 5 kHz 7 kHz Color-coded tonotopic map of sound frequencies showing the organization of subfields of the auditory cortex (After: Tsytsarev et al, Optical imaging of interaural time difference representation in rat auditory cortex) 1 kHz 5 kHz 7 kHz

Voltage-Sensitive Dye Optical Imaging of the Auditory Cortex Where is in the brain? “Where”

Interaural Time Difference (ITD) Binaural click produces the sensation of a sound source to the left, in front of, or to the right of the animal. The angle is determined by the Interaural Time Difference (ITD) (After: Tsytsarev et al, Optical imaging of interaural time difference representation in rat auditory cortex)

Imaging of the ITD Representation sound stimulus onset Click leading: Red: contralateral Green: ipsilateral Blue: (ITD = 0) (After: Tsytsarev et al, Optical imaging of interaural time difference representation in rat auditory cortex)

Conclusion l The virtual map of auditory space in rat auditory cortex does not exist but l Patterns of neural activity recorded in response to binaural click presentations can encode ITD

Vibrissae System: Optical Imaging Research The main goal: to create a functional map of the directional sensitivity of the barrel field using Voltage-Sensitive Dye Optical Imaging

Vibrissae System: Angular Selectivity in the Barrel Field? 1 mm Stimuli Arrangement (color coded) Whisker’s Stimulator VSD optical patterns evoked by different stimuli

Studying of the Cortical Representation of Whisker Directional Deflection Using Voltage-Sensitive Dye Optical Imaging Barrel Field VSD Pseudocolor Pattern in the Region of Interest Direction of the Deflection Magnetized Whisker inside the Stimulator

Barrel Field Voltage-Sensitive Dye Optical Imaging Histology (Cytochrome-c oxidase) Extracted brain Barrel field Brain opening and area of recording Δ F/F(%) 0 20 time, ms Integrated signal in response to different stimuli

Activity Centers in the Single Coordinates System Borders of the Activity Patterns after thresholding 0.5 mm

Disposition of Centers of Mass of Activity Patterns Stimulus 1 Stimulus 2 Stimulus 3 Stimulus 4

Acknowledgments  Drs. Shigeru Tanaka, Hidenao Fukuyama, Kazuyuki Imamura, Ayako Ajima, Hisayuki Ojima, Minoru Kimura and Jerom Ribot – Brain Science Institute of RIKEN and Kyoto University, Japan  Dr. Sonya Bahar, Director, Center for Neurodynamics, University of Missouri at St. Louis  Daisuke Takeshita Douglas Joseph Brumm and Dr. Micheal Hoffman – Center for Neurodynamics, University of Missouri at St. Louis  Song Hu, Junjie Yao, Li Li – Ph.D. student of the Washington University in St. Louis  Drs Konstantin Maslov and Lihong Wang – Department of Biomedical Engineering, Washington University in St. Louis

Thank you very much for your attention