Chromaticity type horizontal cells (C-type HCs) are the first retinal neurons exhibiting spectral information processing in cold-blooded vertebrates. The simple input of hyperpolarizing responses of cone photoreceptors is transformed in the C-type HCs into spectral opponent output. Nitric oxide (NO), a known background neuromodulator in the distal retina, was tested here for its effects upon spectral information processing by C-type HCs in the retina of turtle. Photoresponses were intracellularly recorded from C-type HCs, using light stimuli of different wavelength, applied over backgrounds of different wavelengths, and changing retinal NO level. Raising retinal level of NO in darkness by adding the precursor for its synthesis (l-Arginine) augmented the depolarizing photoresponses elicited by long-wavelength light stimuli, and reduced the hyperpolarizing photoresponses elicited by short-wavelength light stimuli. Lowering retinal level of NO by l-NAME, an inhibitor of NO synthesis, induced the opposite effects. However, the total voltage range of operation remained constant regardless of the level of NO. Qualitatively similar effects were observed under background illuminations regardless of background strength and wavelength. Altering retinal level of NO exerted a small effect upon the null wavelength. Our findings are consistent with the known effects of NO upon turtle distal retinal neurons, with the addition of NO strengthening the negative feedback pathway from L-type horizontal cells onto medium-wavelength cones.