Neurons in the intermediate gray layer (SGI) of the mammalian superior colliculus (SC) receive dense cholinergic innervations from the brainstem parabrachial region. Such cholinergic inputs may influence execution of orienting behaviors. To obtain deeper insights into how the cholinergic inputs modulate the SC local circuits, we analysed the cholinergic responses in identified γ-aminobutyric acid (GABA)ergic and non-GABAergic neurons using SC slices obtained from GAD67-GFP knock-in mice. The responses of SGI neurons to cholinergic agonists were various combinations of fast inward currents mediated mainly via α4β2 and partly by α7 nicotinic receptors (nIN), slow inward currents caused by activation of M1 plus M3 muscarinic receptors (mIN), and slow outward currents caused by activation of M2 muscarinic receptors (mOUT). The most common cholinergic responses in non-GABAergic neurons was nIN + mIN + mOUT (38/68), followed by nIN + mIN (16/68), nIN + mOUT (11/68), nIN only (2/68), and no response (1/68). On the other hand, the major response pattern in GABAergic neurons was either nIN only (26/54) or nIN + mIN (21/54), followed by nIN + mOUT (4/54), mOUT only (2/54), and no response (1/54). Thus, major effects of cholinergic inputs to both SGI GABAergic and non-GABAergic neurons are excitatory, but the response patterns in these two types of SGI neurons are different. Thus, actions of the cholinergic inputs to non-GABAergic and GABAergic SGI neurons are not simple push–pull mechanisms, like excitation vs inhibition, but might cooperate to balance the level of excitation and inhibition for setting the state of the response property of the local circuit.