Recent studies have shown that cholinergic amacrine cells possess unique membrane properties. However, voltage-gated ionic channels in cholinergic amacrine cells have not been characterized systematically. In this study, using electrophysiological and immunohistochemical techniques, we examined voltage-gated ionic channels in a transgenic mouse line the cholinergic amacrine cells of which were selectively labeled with green fluorescent protein (GFP). Voltage-gated K⁺ currents contained a 4-aminopyridine-sensitive current (A current) and a tetraethylammonium-sensitive current (delayed rectifier K⁺ current). Voltage-gated Ca²⁺ currents contained a ω-conotoxin GVIA-sensitive component (N-type) and a ω-Aga IVA-sensitive component (P/Q-type). Tetrodotoxin-sensitive Na⁺ currents and dihydropyridine-sensitive Ca²⁺ currents (L-type) were not observed. Immunoreactivity for the Na channel subunit (Pan Nav), the K channel subunits (the A-current subunits [Kv. 3.3 and Kv 3.4]) and the Ca channel subunits (α1_A [P/Q-type], α1_B [N-type] and α1_C [L-type]) was detected in the membrane fraction of the mouse retina by Western blot analysis. Immunoreactivity for the Kv. 3.3, Kv 3.4, α1_A [P/Q-type], and α1_B [N-type] was colocalized with the GFP signals. Immunoreactivity for α1_C [L-type] was not colocalized with the GFP signals. Immunoreactivity for Pan Nav did not exist on the membrane surface of the GFP-positive cells. Our findings indicate that signal propagation in cholinergic amacrine cells is mediated by a combination of two types of voltage-gated K⁺ currents (the A current and the delayed rectifier K⁺ current) and two types of voltage-gated Ca²⁺ currents (the P/Q-type and the N-type) in the mouse retina.