Objectives: It is unclear whether parental consumption of non-nutritive sweetener (NNS) can affect subsequent generations. The aim of this study was to determine whether chronic parental consumption of sucralose and stevia in mice affects body weight gain and liver and intestinal expression of histone deacetylase 3 (Hdac3) in these animals and in the subsequent first filial (F1) and second filial (F2) generations. Methods: Male and female mice (n = 47) were divided into three groups to receive water alone or supplemented with sucralose (0.1 mg/mL) or stevia (0.1 mg/mL) for 16 wk (parental [F0] generation). F0 mice were bred to produce the F1 generation; then, F1 mice were bred to produce the F2 generation. F1 and F2 animals did not receive NNSs. After euthanasia, hepatic and intestinal expression of Hdac3 was determined by quantitative reverse transcription polymerase chain reaction. Results: Body weight gain did not differ between the three groups in the F0 generation, but it was greater in the F1 sucralose and stevia groups than in the control group. Consumption of both NNSs in the F0 generation was associated with lower Hdac3 expression in the liver and higher in the intestine. Hepatic Hdac3 expression was normalized to the control values in the F1 and F2 animals of the sucralose and stevia groups. Intestinal expression was still higher in the F1 generations of the sucralose and stevia groups but was partially normalized in the F2 generation of these groups, compared with control. Conclusions: NNS consumption differentially affects hepatic and intestinal Hdac3 expression. Changes in hepatic expression are not transmitted to the F1 and F2 generations whereas those in intestinal expression are enhanced in the F1 and attenuated in the F2 generations.