The correlation between cholinergic sensitivity and the level of stratification for ganglion cells was examined in the rabbit retina. As examples, we have used ON or OFF α ganglion cells and ON/OFF directionally selective (DS) ganglion cells. Nicotine, a cholinergic agonist, depolarized ON/OFF DS ganglion cells and greatly enhanced their firing rates but it had modest excitatory effects on ON or OFF α ganglion cells. As previously reported, we conclude that DS ganglion cells are the most sensitive to cholinergic drugs. Confocal imaging showed that ON/OFF DS ganglion cells ramify precisely at the level of the cholinergic amacrine cell dendrites, and co-fasciculate with the cholinergic matrix of starburst amacrine cells. However, neither ON or OFF α ganglion cells have more than a chance association with the cholinergic matrix. Z-axis reconstruction showed that OFF α ganglion cells stratify just below the cholinergic band in sublamina a while ON α ganglion cells stratify just below cholinergic b. The latter is at the same level as the terminals of calbindin bipolar cells. Thus, the calbindin bipolar cell appears to be a prime candidate to provide the bipolar cell input to ON α ganglion cells in the rabbit retina. We conclude that the precise level of stratification is correlated with the strength of cholinergic input. Alpha ganglion cells receive a weak cholinergic input and they are narrowly stratified just below the cholinergic bands.