There are increasing concerns about disruption of brain function due to exposure to environmental chemical substances, especially in the early stages of development. We aimed to develop a method to assess such effects on neuronal circuit activity. Bisphenol A (BPA), which is widely used in the production of polycarbonate plastics and epoxy resins, is a potentially hazardous chemical substance. BPA can bind estrogen receptors and affect the signal cascade of endogenous estradiol. A number of reports have indicated that estrogen affects synaptic plasticity in the hippocampal (HP) circuit in rats and mice. Pairs of male and female mice were transferred for mating into cages where we supplied BPA-containing drinking water at four different concentrations (0, 0.1, 1, and 10 ppm). On confirmation of pregnancy, BPA-containing drinking water was supplied until weaning. Male offspring were randomly chosen from mothers maintained under the same conditions. Hippocampal slices were made by standard procedures and analyzed after staining with voltage-sensitive dye (VSD: Di-4-ANEPPS) after at least 8 weeks. Stimulation of three main pathways of the HP, the perforant (PP), mossy fiber (MF), and Schaffer collateral pathways (Sch), was studied using the VSD imaging assay. The neuronal responses of the control (0 ppm) and BPA-exposed mice (0.1, 1 and 10 ppm BPA) were tested in normal and Gabazine (10 μM)-containing ACSF. Gabazine increased the peak amplitude and duration of the response to strong stimulation in both control and BPA-exposed mice. In control mice, even very weak stimulation was sufficient to induce large and long responses in the entire CA3-CA1, whereas BPA-exposed mice did not exhibit such responses. The clear difference may indicate a difference in the threshold of excitability of the cells. These results highlight the use of VSD imaging assay to assess environmental risk factors.