Choline acetyltransferase-immunoreactive neurons in the retina of normal and dark-reared turtle

Visual deprivation alters retinal-ganglion-cell response properties through changes in spontaneous wave-like activity (Sernagor and Grzywacz [1996] Curr Biol 6:1503-1508). This activity depends on cholinergic synaptic transmission in the turtle retina (ibid; Sernagor and Mehta [ 2001] J Anat 199:375-383). We studied the expression of choline acetyltransferase (ChAT) by immunocytochemistry and Western blot in developing retinas of control and dark-reared turtles. At postnatal day 0 (P0), right after hatching, ChAT-immunoreactivity was present in the ganglion cell layer (GCL), in the inner nuclear layer (INL), and in two distinct bands of the inner plexiform layer (IPL). In P14- and P28-control, and P14- and P28-dark-reared retinas, ChAT-immunoreactivity showed similar patterns to those in P0. However, in P14- and P28-dark-reared retinas the density of ChAT-immunoreactive cells was higher in both the INL and GCL than in P14- and P28-control retinas, respectively. Moreover, Western blotting showed that ChAT protein levels were significantly increased in the dark-reared retina compared to those of the control. TUNEL studies indicated that the difference between normal and dark-reared conditions was not due to extra apoptosis in the former. In turn, proliferating-cell nuclear antigen immunocytochemistry showed no extra proliferating cells in the latter. Finally, nearest-neighbor analysis revealed that the denser population of cholinergic cells in dark-reared turtles formed a mosaic as regular as the normal ones in the GCL. Thus, light deprivation increases the expression of ChAT, increasing the apparent density of cholinergic neurons in the developing turtle retina.