Striatal dopamine (DA) release plays an essential role in many physiological functions including motor and non-motor behaviors (such as reward, motivation, and cognition). We have previously reported that, following a single electrical field stimulation, the amperometric recording of DA release from presynaptic terminals in striatal slices (both ventral and dorsal) contains two temporally separated phases. The first phase (direct DA transmission, direct DT) arises from DA terminal release following autologous action potentials (APs), while the second phase (cholinergic transmission-induced DA transmission, CTDT) arises from delayed DA release triggered by the activation of cholinergic interneurons to DA terminals (axon-axon transmission). The millisecond time-resolution of amperometry permits separation of an ∼7 ms latency difference from the single synapse (axon-axon) within the two-phase DA-release (2pDA) signal, and thus the 2pDA signal provides a novel method to study either direct DT, or CTDT, or both. Here, we describe the 2pDA method, including signal recording, processing, analysis, and troubleshooting (anti-artifact). Compared with other DA assays using different stimuli, recording methods, and preparations (such as high performance liquid chromatography or fast scan cyclic voltammetry), 2pDA recording is a novel and powerful physiological recording method for the study of DA transmissions in situ.