Perfusion of 100 microM 2-amino-4-phosphonobutyric acid (APB) into the in vivo rabbit eye-cup selectively and reversibly blocked the b-wave of the ERG and all On responses from retinal ganglion cells. In contrast, Off responses were occasionally enhanced, sometimes dramatically. The antagonistic surround inputs to Off ganglion cells, identified by their latency to light stimulation and magnesium sensitivity, were unchanged by APB. These observations suggest that APB selectively blocks depolarizing bipolar cells in rabbit retina in close agreement with the results of Slaughter and Miller (1981) from mudpuppy retina. We conclude that APB may be useful as a pharmacological tool to differentiate On and Off pathways in the rabbit visual system.