Objective: In Parkinson disease (PD), the onset of motor symptoms is linked to a substantial loss of dopaminergic substantia nigra (DA SN) neurons. While the pattern of DA SN cell loss has been extensively documented in animal PD models, little is known about electrophysiological changes of the remaining viable nigrostriatal DA neurons. To explore the functional changes of these neurons, intrastriatal infusions of 6-hydroxydopamine were combined with retrograde tracings and in vitro patch clamp analyses. Methods: To induce a partial degeneration of DA SN neurons, a single dose of 6ml (12mg) 6-hydroxydopamine was infused into the right dorsolateral striatum of adult C57Bl6 mice (12 weeks old) under stereotactic control. Loss of DA SN neurons was quantified 3 weeks after infusion by unbiased stereology and compared to control animals infused with 6ml ACSF. For in vitro patch clamp analyses, surviving nigrostriatal DA neurons were identified by injection of fluorescent tracer beads into the right striatum 2 weeks post lesion. Patch clamp recordings of retrogradely labeled DA SN neurons were performed 3 weeks after lesion. After recording, neurobiotin-filled cells were immunostained for tyrosine hydroxylase (TH) to verify their neurochemical identity. Results: Intrastriatal infusion of 6-hydroxydopamine resulted in a substantial loss of >50 % of TH-positive SN neurons. In the dorsal striatum, >50 % of DA fibers were lost. In combination with selective retrograde tracing, individual remaining DA SN neurons with intact axonal connectivity were identified and characterized electrophysiologically. Conclusion: The combination of intrastriatal infusions of 6-hydroxydopamine with retrograde tracings and in vitro patch clamp techniques provides a novel approach to investigate the functional properties of surviving DA SN neurons in the chronically dopamine depleted brain.