Heterothermic insectivorous bats are supposed to experience differential adverse effects of insecticidal pollutants depending on their seasonal and/or daily variation of metabolic and detoxification rates. Here, we investigated effects of imidacloprid, cypermethrin and fipronil on Nyctalus noctula bat-derived hepatocytes through cytotoxicity, cell inhibition and death at different concentrations (0.01, 0.1, 1, 10, 100, 1000μg/ml), exposure times (10, 24, 48 hrs), incubation temperatures simulating hibernation (8 °C), daily torpor (20 °C), normothermy (37 °C) and active flight (40 °C), and cytochrome P450 addition. Toxic effects were significantly influenced by temperature (p < 0.05), with strong cytotoxicity after 10hour exposure to fipronil or cypermethrin at 37 and 40 °C, cell replication inhibition (all insecticides at 8 °C) and cellular stimulation, with slight culture proliferation after 48hours (all insecticides at 40 °C). Replacing protected chiropterans with cell cultures is a way to assess and extrapolate risks of insecticides for bats.