1. Simultaneous, whole-cell recordings were obtained from synaptically coupled photoreceptor/bipolar cell pairs, by the use of direct visualization in a superfused, mudpuppy retinal slice preparation. 2. OFF-bipolar cells (BPs) generated sign-conserving responses when extrinsic current was injected into rods and cones, whereas ON-BPs generated a sign-reversing response. OFF- BPs (n = 24) responded faster than ON-BPs (n = 12), in terms of response latency (27.8 vs. 80.6 ms) and peak response times (50.5 vs. 159.8 ms) when current was injected into photoreceptors. We did not detect any significant difference between rod- versus cone-mediated latency or peak response times in the ON- and OFF-BP subtypes. 3. Rod and cone inputs to OFF-BPs were blocked by kynurenic acid (Kyn), but the doses required were significantly higher for rod inputs: the IC50 (the concentration at which an antagonist blocks 50% of the responses) for Kyn was 0.3 mM for cone inputs and 1 mM for rod inputs. 4. Rod inputs to OFF-BPs showed the same Kyn sensitivity as rod inputs to horizontal cells (HCs). However, cone inputs to HCs (IC50 <200 μM) were more sensitive to Kyn than those to OFF-BPs. 5. The pharmacological studies presented here, together with previous studies, suggest that the sign-conserving pathway in the outer plexiform layer of the mudpuppy retina involves at least three subtypes of glutamate receptors: 1) cone-activated receptors of HCs: 2) cone-activated receptors of OFF-BPs; and 3) rod- activated receptors found in HCs and BPs. 6. The sign-reversing rod and cone inputs into ON-BPs were blocked by 2-amino-4-phosphonobutyric acid (APB; <10 μM). The effect of APB was permanent: ON-BPs did not recover their sign- reversing responses after returning to control conditions. However, in the presence of APB, current injections into photoreceptors revealed sign- conserving inputs in ON-BPs, which persisted after removing the APB. Both rod and cone stimulation was associated with this 'response reversal.' 7. The long latency of ON-BP activation and the permanent loss of the sign-reversing response presumably due to the dialysis by the whole-cell recording method support the idea that a second messenger system is involved in ON-BPs' synaptic responses. However, it is not clear where the uncovered sign- conserving inputs originate.