Background Excessive fluoride can accumulate in brain tissues, causing nervous system damage. MicroRNA (miRNA) can affect the ability of learning and memory by regulating the expressions of related proteins.

Objective This experiment explores the effects of fluoride exposure from pregnancy to adulthood on spatial learning and memory of offspring and the expressions of miR-204 and miR-34b-5P in the hippocampus.

Methods Sixteen pregnant SD rats were randomly divided into four groups and exposed to NaF via drinking water at 0, 60, 120, and 240 mg·L^-1, respectively, with four rats in each group. The pregnant rats were treated from pregnant day 0 to postnatal day 21 (PND21) of the offspring rats. From PND22 to PND90 (adulthood), 8 offspring rats (4 males and 4 females, and litter sex ratio was 1:1) from each group were treated with the same concentrations of NaF as the dams through the same procedure. The offspring rats were tested in the Morris water maze. Before death, 24 h urine was collected, and blood was collected by heart puncture. Urinary, brain, and serum fluoride concentrations were determined; the pathological changes of hippocampus were observed under a light microscope; the separated hippocampus were stored at -80℃ and tested for miR-204 and miR-34b-5P expression levels by real-time PCR.

Results Compared with the control group, the body weight of each exposure group was significantly reduced (P < 0.05), except the low fluoride exposure group at postnatal week 6. The Morris water maze results showed that on the second, third, and fourth days of training, the escape latency was positively correlated with fluoride exposure concentration (r=0.443, 0.519, 0.840; P < 0.05); the first arriving time and the number of crossing the platform were positively and negatively correlated with fluoride exposure concentration respectively (r=0.828, -0.599; P < 0.001). The urinary fluoride concentrations of the 60, 120, and 240 mg·L^-1 exposure groups were (13.08±1.60), (14.49±1.17), and (25.92±2.38) mg·L^-1, respectively, significantly higher than that of the control group(3.89±0.52) mg·L^-1 (P < 0.001). The brain fluoride concentration of each exposure group was (8.20±0.68), (16.03±0.84), and (25.39±0.62) μg·g^-1, respectively, significantly increased compared with the control group(1.28±0.11) μg·g^-1 (P < 0.001). The serum fluoride concentration of each exposure group was (0.04±0.00), (0.06±0.01), and (0.16±0.12) mg·L^-1, respectively, significantly higher than that of the control group(0.02±0.00) mg·L^-1 (P < 0.001). The urinary, brain, and serum fluoride levels of the progeny rats in the exposure groups were positively correlated with NaF exposure concentration (r=0.948, 0.996, 0.914; P < 0.001). Compared with the control group, there were pathological changes in the hippocampal neurons, such as shrinkage of hippocampal neurons, red staining of cytoplasm, pyknosis of nuclei, hyperchromasia, blurred structure, and disappearance of nucleoli in the exposure group. Compared with the control group, the expression levels of miR-204 in the medium fluoride group and the high fluoride group and the expression levels of miR-34b-5P in each exposure group were increased (P < 0.001). The expression levels of miR-204 and miR-34b-5P were positively correlated with fluoride exposure concentration (r=0.984, 0.980; P < 0.001).

Conclusion Continuous exposure to fluoride from pregnancy to adulthood may impair the ability of learning and memory in pups, which may be related to increasing expressions of miR-204 and miR-34b-5P in the hippocampus.