Abstract Heterozygous mutations in the PINK1 gene are considered a susceptibility factor to develop early-onset Parkinson's disease (PD), as supported by dopamine hypometabolism in asymptomatic mutation carriers and subtle alterations of dopamine-dependent striatal synaptic plasticity in heterozygous PINK1 knockout (PINK1+/− ) mice. The aim of the present study was to investigate whether exposure to low-dose rotenone of heterozygous PINK1+/− mice, compared to their wild-type PINK1+/+ littermates, could impact on dopamine-dependent striatal synaptic plasticity, in the absence of apparent structural alterations. Mice were exposed to a range of concentrations of rotenone (0.01–1 mg/kg). Chronic treatment with concentrations of rotenone up to 0.8 mg/kg did not cause manifest neuronal loss or changes in ATP levels both in the striatum or substantia nigra of PINK1+/− and PINK1+/+ mice. Moreover, rotenone (up to 0.8 mg/kg) treatment did not induce mislocalization of the mitochondrial membrane protein Tom20 and release of cytochrome c in PINK1+/− striata. Accordingly, basic electrophysiological properties of nigral dopaminergic and striatal medium spiny neurons (MSNs) were normal. Despite the lack of gross alterations in neuronal viability in chronically-treated PINK1+/− , a complete loss of both long-term depression (LTD) and long-term potentiation (LTP) was recorded in MSNs from PINK1+/− mice treated with a low rotenone (0.1 mg/kg) concentration. Even lower concentrations (0.01 mg/kg) blocked LTP induction in heterozygous PINK1+/− MSNs compared to PINK1+/+ mice. Of interest, chronic pretreatment with the antioxidants alpha-tocopherol and Trolox, a water-soluble analog of vitamin E and powerful antioxidant, rescued synaptic plasticity impairment, confirming that, at the doses we utilized, rotenone did not induce irreversible alterations. In this model, chronic exposure to low-doses of rotenone was not sufficient to alter mitochondrial integrity and ATP production, but profoundly impaired the expression of long-term plasticity at corticostriatal synapses in PINK1 heterozygous knockout mice, suggesting that disruption of synaptic plasticity may represent an early feature of a pre-manifesting state of the disease, and a potential tool to test novel neuroprotective agents.