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J.S. George, X.–C. Yao; Optical Imaging of Neurophysiological Activation in Isolated Retina . Invest. Ophthalmol. Vis. Sci. 2006;47(13):5671.
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© ARVO (1962-2015); The Authors (2016-present)
Transient intrinsic optical changes tightly coupled to neural activation promise high–resolution methodology for investigation of dynamic visual processing, and for characterization of the consequences of electrical stimulation of retina. Here we demonstrate the feasibility of imaging fast intrinsic optical responses evoked by light flashes in isolated frog retina.
Frogs (rana pipiens) were sacrificed, eyes were removed, and retinas were isolated under dim red illumination. Retinas were transferred to recording chamber incorporating a multi–electrode array. Optical recording employed NIR illumination (750–950 nm) in transmitted light, darkfield or cross–polarized configurations. Physiological stimulation employed a white LED. Optical recording employed a photodiode or a high resolution CCD camera operated at high frame rates (>100fps). An average prestimulus baseline image was subtracted from each image in the sequence, and difference images were divided by the original images (dI/I).
We routinely measured dynamic transmitted optical responses closely tracking the integral of electrophysiological activity. Photodiode measurements typically disclosed a positive peak, at a level 10–4 of background near infrared (NIR) transmitted light intenity. By implementing improvements to the experimental system we were able to improve signal to allow capture of the signals with CCD cameras. CCD image sequences show evidence of multiple response components with both negative– and positive–going signals with different time courses and consistent spatial organization, initially limited to the region activated by the flash. Because of the close juxtaposition of responses of opposite polarity, high–resolution imaging disclosed larger fractional responses in individual pixels, in some cases exceeding 10%. Fast negative–going responses are related to a–wave of the retinal electrophysiological response, and may reflect the activation of photoreceptors. Positive–going responses are related to the b–wave, and may result from the activation of ON and OFF bipolar cells and other postsynaptic neurons. Spatial and temporal structure in the image sequences indicates that we captured images of neurophysiological activation in large populations of individual neurons in single passes.
Improved methods allow dynamic optical imaging of neural activation in the isolated retina based on fast light scattering changes. Our previous demonstrations of the feasibility of recording fast optical changes in reflected light suggest the possibility of noninvasive functional imaging of the retina in situ.
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