Response characteristics and receptive field widths of on-bipolar cells in the mouse retina

Abstract

Voltage-clamp and current-clamp recordings were made from bipolar cells in dark-adapted mouse retinal slices. Light-evoked responses fell into three groups corresponding to the rod bipolar cells, on-cone bipolar cells and off-cone bipolar cells. The morphology of the recorded cells confirmed this classification. Intensity-response relations were well fitted by a Michaelis saturation function with Hill coefficients of 1.15 +/- 0.11 (n = 6) for rod bipolar cells and 2.33 +/- 0.06 (n = 4) for cone inputs onto on-cone bipolar cells. In the absence of antagonists for GABA or glycine receptors, light-evoked synaptic currents for all cells displayed linear current-voltage relations that reversed near 0 mV, indicating that very little inhibition was activated under dark-adapted recording conditions. Saturating light stimuli evoked conductances of 0.81 +/- 0.56 nS (n = 4) in rod bipolar cells and 1.1 +/- 0.8 nS (n = 4) in on-cone bipolar cells. Receptive field widths were estimated by flashing a vertical light bar at various locations along the slice. Rod and on-cone bipolar cells had receptive field widths of 67 +/- 16 micrometer (n = 6) and 43 +/- 7 microm (n = 5), respectively. The maximum spatial resolution of an array of such cone bipolar cells was estimated to be 0.3 cycles deg-1, compared with a maximum resolution of 0.5 cycles deg-1 obtained from behavioural studies in mice. Our results suggest that this limit to spatial resolution could be imposed early in the visual system by the size of the bipolar cell receptive fields.

Figure 3. Normalised intensity-response plots for RBPs

For each cell the data have been normalised to have the same half-maximal light intensity and maximum amplitude. A, intensity-response relation obtained from the peak photocurrents in six cells, including those illustrated in Fig. 2A and C. The line though the points was fitted to eqn (1) where h = 1.15, L₀= 0.070 lm m⁻² and I_min=−115 pA. B, intensity-response relation obtained from the peak photovoltages in four cells, including the cell shown in Fig. 2B. The line though the points was fitted to eqn (1) where h = 1.46, L₀= 0.028 lm m⁻² and V_max= 16.1 mV.

Figure 1. Three classes of bipolar cell

Whole-cell voltage-clamp responses to flashes (upper traces) and steps (lower traces) of light in rod and cone bipolar cells. Holding potential, -70 mV. The light intensity produced saturating responses in all cases except the flash responses for the RBP, which were subsaturating. The light stimulus monitor traces above the records show the timing of the light flashes. A, RBP. Two flash responses have been superimposed. Stimulus intensity, 0.1 lm m⁻². Despite the characteristically high dark noise (and low signal:noise ratio of ≈3) the timing of the responses to the two flashes is very accurately registered. Note the strong suppression of current noise during the light step (stimulus intensity, 0.35 lm m⁻²). B, on-CBP. The dark noise was lower in on-CBPs than in RBPs. C, off-CBP. The dark noise was strongly suppressed by light. Stimulus intensity in B and C, 14.4 lm m⁻².

Figure 4. Intensity-response relations determined for on-CBPs

Responses in A and B, and C and D were obtained from two different cells. A, photocurrents recorded during light steps of increasing intensity (shown to the left of the records in lm m⁻²). The holding potential was set to -50 mV. The light stimulus timings for both A and C are shown by the stimulus monitor trace below the traces. B, the amplitudes of the photocurrents recorded in two separate runs are plotted as a function of the stimulus intensity. The line though the points was fitted to the sum of two eqn (1) with h₁= 1.15, I_max(1)=−33.5 pA and L₀₍₁₎= 0.053 lm m⁻²; and h₂= 2.28, I_max(2)=−125 pA and L₀₍₂₎= 0.91 lm m⁻². C, photovoltages during light steps of increasing intensity (shown to the left of the records in lm m⁻²). Potassium gluconate internal solution. The resting potential of the cell was -28 mV. The r.m.s. voltage noise was 0.74 mV in darkness, and dropped to 0.24 mV during the brightest steps. D, the peak photovoltages in C are plotted as a function of the stimulus intensity (•). Measurements were made just before the light step (▿), and at the end of the record (□) to illustrate the baseline variability. The line though the points was fitted to the sum of two eqn (1) with h₁= 1.15, V_max(1)= 2.2 mV and L₀₍₁₎= 0.035 lm m⁻²; and h₂= 2.28, V_max(2)= 6.5 mV and L₀₍₂₎= 2.9 lm m⁻².

Figure 2. Intensity-response relations determined for RBPs

The light stimulus monitor trace is shown above the records in each panel. A and B, whole-cell photocurrents and photovoltages, respectively, recorded in the same RBP during light steps of increasing intensity (shown to the left of the records as × 10⁻³ lm m⁻²). The internal solution contained potassium gluconate. The resting potential in current clamp was -47 mV. Note the appearance of the ‘rod tail’ for the strongest light steps in both A and B. For the nine records in B the r.m.s. voltage noise averaged 4.1 ± 0.6 mV in darkness and dropped to 0.5 ± 0.1 mV during the brightest three steps. C, intensity- response series for photocurrents in another cell plotted on an expanded time base to show the change in activation kinetics as the light intensity was increased. D, the bottom records from A and B (5700 × 10⁻³ lm m⁻²) have been scaled and superimposed to compare the activation time courses. The sag is less pronounced in the current-clamp recording.

Figure 5. Current-voltage relations recorded from RBPs and on-CBPs

The holding potential was -70 mV. The membrane potential was stepped from -109 to +41 mV and ganzfeld light stimuli (390 ms; open bar in A and C) were delivered 150 ms after establishing the new potential. The peak amplitude of the light-evoked current was measured at each potential. A and C, light-evoked currents in one RBP and one on-CBP, respectively. B and D, averaged peak current-voltage relations from light-evoked currents measured in four RBPs and four on-CBPs, respectively, including the cells shown in A and C.

Figure 6. Receptive field profiles of a RBP (A and B) and an on-CBP (C and D)

The width of the stimulus bar was 10 μm for A and B and 20 μm for C and D. A and C, the peaks of the light responses have been aligned to the relative spatial offset of the stimulus bar. At the central locations (0 μm) two separate responses are superimposed, one obtained at the start and one in the middle of the series. The close agreement in both cases indicates that the sensitivity did not change significantly during the data collection periods. B and D, response amplitudes plotted over a wider range. The continous line shows the best-fitting Gaussian spatial profiles (eqn (2)). Parameters were: w = 34μm, c =−4 μm and I_max=−57 pA for B; and w = 17.5μm, c = 10μm and I_max=−50 pA for D.