The paired flash protocol unmasks the time course of the rod response in vivo. (A) A bright probe flash (7.6 × 105 R*/rod) elicited a maximal ERG response (green trace) with a large negative-going a-wave, whereas a dim test flash (284 R*/rod, cyan) elicited a response (cyan trace) with a small a-wave component. When the probe flash was presented 80 ms after the dim test flash (black arrows), the response to the probe flash was greatly reduced (black trace) relative to its amplitude in the dark-adapted state (green trace). The cyan trace was subtracted from the black trace to isolate the pure probe response at a delay of 80 ms (red trace). (B) Pairing the bright probe and dim test flashes at delay intervals t yielded probe responses that varied in amplitude over time (top traces; bottom trace is a flash monitor for the probe flash). (C) Symbols plot the complement of the amplitude of the probe a-wave responses normalized by the maximum probe flash amplitude (amax, from green trace) as a function of the time interval between the test and probe flashes. All traces were from individual trials of the same animal in the same recording session. The green and red traces in A and B are the same, and the colored symbols in C plot the corresponding photoresponse amplitudes.
