Rod bipolar cells experience PDE-dependent loss of response reproducibility near threshold

Abstract

Purpose : We have demonstrated that a single-site D167A mutation of the gene for the α subunit of rod phosphodiesterase (PDEA) alters the rod response to single photons: rod responses were larger, more sensitive, slower, and less reproducible from trial-to-trial than those of WT rods indicating that response reproducibility depends on PDE concentration. Given the presumed importance of rod response reproducibility in signaling near threshold, we sought to understand the synaptic consequences of reduced rod PDE expression on synaptic transmission between rods and their postsynaptic rod bipolar cells (RBCs).

Methods : Slices of mouse retina were perfused with bicarbonate-buffered Ames' media at 35-37^oC and light response characteristics of individual RBCs from D167A (n=6) and wild-type (WT) (n=6) were measured with whole-cell patch-clamp recordings. Sensitivity was determined by the light intensity required to generate a half-maximal response (I_1/2) obtained from Hill equation fits to the response-intensity relationship. Single-photon responses were isolated in response to a series of consecutive dim flashes using weak light stimuli. Light-independent current fluctuations (dark noise) were recorded in the absence and in the presence of GTP-δ-S in the recording pipette. Two-tailed Student's t-tests was used for statistical analysis.

Results : Interestingly, little difference was observed in the sensitivity of D167A RBCs (I_1/2=1.6 R*/rod) compared to WT (I_1/2=1.3 R*/rod). However, single-photon responses from D167A RBCs (t_peak=270 ms) were significantly slower compared to WT (t_peak=150 ms) (p<0.001), roughly double in amplitude (WT=3.4 versus D167A=6.0 pA), and displayed ~3-fold more variability in peak amplitudes. Lastly, cellular dark noise was 10-fold greater at frequencies between 0 and 1 Hz in D167A compared to WT RBCs.

Conclusions : These data reveal the reductions in the response reproducibility of D167A rods are conveyed to downstream RBCs near their threshold. We found that compared to D167A rods, D167A RBCs are less variable from trial-to-trial. In addition, we found that D167A RBC responses are temporally slower than comparable responses in WT RBCs, presumably due to the summation of responses from multiple rods. We speculate that rod-to-RBC synaptic transmission is partially resilient to variations in the rod response characteristics.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.