Neurons in the intermediate gray layer (SGI) of mammalian superior colliculus (SC) receive cholinergic innervation from the brainstem parabrachial region, which appears to modulate the signal processing in the SC. To clarify its role particularly in orienting behaviors, we investigated cholinergic effects on the major output neuron group of the SGI, crossed tecto-reticular neurons (cTRNs), identified by retrograde labeling from the contralateral brainstem gaze center in SC slices obtained from rats (PND 17-22) by whole cell patch clamp techniques. Bath application of carbachol induced either (1) nicotinic inward (nIN) + muscarinic inward (mIN)(11/24) or (2) nIN + mIN + muscarinic outward (mOUT)(13/24) current responses. Transient pressure application of 1 mM acetylcholine elicited nIN in all neurons tested (n=58). In a majority of these neurons (52/58), the nIN was completely suppressed by dihydro--erythroidine, a specific antagonist for 42 nicotinic receptor subtype. The remaining 6/58 neurons exhibited not only the slower 42 receptor-mediated component but also a faster component which was inhibited by a specific antagonist for 7 nicotinic receptor, -bungarotoxin. cTRNs expressing 7 nicotinic receptors tended to be smaller in size than those lacking 7 receptors. Bath application of muscarine induced two response patterns, mIN only (17/38) and mIN+ mOUT (21/38). The mIN and mOUT were mediated by M3 (plus M1) and M2 muscarinic receptors, respectively. These results suggest that a major response to cholinergic inputs to cTRNs is excitatory. This would indicate the facilitatory role of the brainstem cholinergic system in the execution of orienting behaviors including saccadic eye movements.