Center/surround organization of retinal bipolar cells: High correlation of fundamental responses of center and surround to sinusoidal contrasts

Abstract

Receptive field organization of cone-driven bipolar cells was investigated by intracellular recording in the intact light-adapted retina of the tiger salamander (Ambystoma tigrinum). Centered spots and concentric annuli of optimum dimensions were used to selectively stimulate the receptive field center and surround with sinusoidal modulations of contrast at 3 Hz. At low contrasts, responses of both the center and surround of both ON and OFF bipolar cells were linear, showing high gain and thus contrast enhancement relative to cones. The contrast/response curves for the fundamental response, measured by a Fast Fourier Transform, reached half maximum amplitude quickly at 13% contrast followed by saturation at high contrasts. The variation of the normalized amplitude of the center and surround responses was remarkably similar, showing linear regression over the entire response range with very high correlations, r2 = 0.97 for both ON and OFF cells. The contrast/response curves of both center and surround for both ON and OFF cells were well fit (r2 = 0.98) by an equation for single-site binding. In about half the cells studied, the nonlinear waveforms of center and surround could be brought into coincidence by scaling and shifting the surround response in time. This implies that a nonlinearity, common to both center and surround, occurs after polarity inversion at the cone feedback synapse. Evidence from paired whole-cell recordings between single cones and OFF bipolar cells suggests that substantial nonlinearity is not due to transmission at the cone synapse but instead arises from intrinsic bipolar cell and network mechanisms. When sinusoidal contrast modulations were applied to the center and surround simultaneously, clear additivity was observed for small responses in both ON and OFF cells, whereas the interaction was strikingly nonadditive for large responses. The contribution of the surround was then greatly reduced, suggesting attenuation at the cone feedback synapse.

Fig. 1

Response of an ON bipolar cell to a contrast step of variable diameter (A) and variable inner diameter of a concentric annulus (B). Dimensions in microns are shown above each response. The 100-µm spot and all other stimuli are centered in the receptive field. The contrast step is 0.3 log unit, that is, two times greater than the steady background of 20 cd/m². (C) Response to a sinusoidal modulation of 80% at a frequency of 3 Hz and diameter of 240 µm.

Fig. 2

Signal averaged waveforms and Fourier spectra (FFT) for an ON bipolar cell to contrast modulations from 4 to 92%. The FFT is to the left of the average response waveform. The left column shows responses to a centered spot of 240 µm. The right column shows responses to a concentric annulus with an inner diameter of 382 µm and an outer diameter of 2000 µm.

Fig. 3

Signal averaged waveforms and Fourier spectra (FFT) for an OFF bipolar cell to contrast modulations from 4 to 92%. The FFT is to the left of the average response waveform. The left column shows responses to a centered spot of 240 µm. The right column shows responses to a concentric annulus with an inner diameter of 496 µm and an outer diameter of 2000 µm.

Fig. 4

Contrast modulation/response measurements for the fundamental component of the FFT for two OFF bipolar cells (A and B, left column) and two ON bipolar cells (C and D, right column). The open and filled circles show results for centered spots and concentric annuli, respectively. Reponses are normalized with respect to the maximum response evoked by the spot. The smooth curves show the best-fitting equation for the single-site binding equation (see text). The percent of variance explained (r²) is 98% for the cell in (B) and 97% for the other three cells. All stimuli were of optimal dimensions for stimulating the center or surround mechanism, as determined from the basic procedure described for Fig. 1.

Fig. 5

Plot of the response of the center and surround for all measurements of the normalized fundamental component of the FFT. Squares and diamonds are for ON and OFF bipolar cells, respectively. Many points are not evident due to overlap. The total number of measurements is n = 67 for OFF cells and n = 56 for ON cells. The percent of variance explained (r²) of the linear best-fit regression lines (not shown for clarity since they overlap closely) are 96.6 and 96.5% for ON and OFF cells, respectively.

Fig. 6

Plot of the observed amplitude of the fundamental component of the FFT versus the amplitude predicted from the best-fitting equation for single-site binding (see text for details). Many points are not evident due to overlap. The total number of measurements is n = 67 for OFF cells and n = 56 for ON cells. The percent of variance explained (r²) is: OFF center, 97.5; OFF surround, 97.2; ON center, 98.2; and OFF surround, 96.5.

Fig. 7

Plot of the amplitude of the fundamental response (3 Hz) of the FFT and the total amplitude of the harmonics at 6, 9, 12, and 15 Hz. (A) and (B) show results for the center and surround, respectively. The symbols shown as X and open circles are for OFF and ON cells, respectively.

Fig. 8

Response of an OFF bipolar cell for stimuli applied to the center (C) or the surround (S) at contrast modulation of 4% in (A) and 78% in (B). The response of the surround has been normalized and shifted laterally to yield a best fit with the response of the center. The bottom trace in (B) (S 78%) shows the response of the surround without a lateral shift. Note that the waveforms in (B), although distorted sine waves and in the nonlinear range, can still be brought into close registration. The diameter for the centered spot is 240 µm. The i.d. of the annulus used to stimulate the surround is 382 µm.

Fig. 9

Synaptic transmission does not contribute substantial nonlinearities to OFF bipolar cell responses. (A) Cone membrane currents evoked by sinusoidal modulation of the cone membrane potential using 3 Hz waveforms of 4 (−35 to −39 mV, top) and 20 mV (−30 to −50 mV, bottom). Current segments (right) and FFTs (left) are illustrated for both stimulus protocols. Note the very small harmonic evident at 6, 9, and 12 Hz in the responses to 20 mV sinusoids. (B) Postsynaptic currents evoked in OFF bipolar cells by sinusoidal modulation of cones by 3 Hz waveforms of 4 (top) and 20 mV (bottom). Synaptic current waveforms (right) and FFTs (left) are illustrated for both stimulus protocols. For both cone and OFF bipolar cells, responses with 20 mV waveforms were averaged from single trials across 12 cells. For the 4-mV waveform, responses were averaged from 16 trials across 12 cells.

Fig. 10

Injection of 3 Hz sinusoidal current (15 pA) into an OFF bipolar cell produced only modest nonlinearities in the resulting voltage change. Note the small harmonics at 6 and 9 Hz in the FFT (left). The FFT was calculated from the average of eight trials. A segment of the average voltage waveform is also shown (right). Mean membrane potential was −48.6 mV.

Fig. 11

Intracellular recordings of responses of an OFF cell to stimulation of the center (C) or surround (S) separately or together (C&S) at 4 (A) and 92% (B) modulations. ADD shows the results of the addition of the C and S traces. The diameter for the centered spot is 240 µm. The i.d. of the annulus used to stimulate the surround is 382 µm.