Transgenerational modification of dopaminergic dysfunctions induced by maternal immune activation

Prenatal exposure to infectious and/or inflammatory insults is increasingly recognized to contribute to the etiology of psychiatric disorders with neurodevelopmental components. Recent research using animal models suggests that maternal immune activation (MIA) can induce transgenerational effects on brain and behavior, possibly through epigenetic mechanisms. Using a mouse model of MIA that is based on gestational treatment with the viral mimetic, poly(I:C) (= polryriboinosinic-polyribocytidilic acid), the present study explored whether the transgenerational effects of MIA are extendable to dopaminergic dysfunctions. We show that the direct descendants born to poly(I:C)-treated mothers display signs of hyperdopaminergia, as manifested by a potentiated sensitivity to the locomotor-stimulating effects of amphetamine (Amph) and increased expression of tyrosine hydroxylase (Th) in the adult ventral midbrain. In stark contrast, second- and third-generation offspring of MIA-exposed ancestors displayed blunted locomotor responses to Amph and reduced expression of Th. Furthermore, we found increased DNA methylation at the promoter region of the dopamine-specifying factor, nuclear receptor-related 1 protein (Nurr1), in the sperm of first-generation MIA offspring and in the ventral midbrain of second-generation offspring of MIA-exposed ancestors. The latter effect was further accompanied by reduced mRNA levels of Nurr1 in this brain region. Together, our results suggest that MIA has the potential to modify dopaminergic functions across multiple generations with opposite effects in the direct descendants and their progeny. The presence of altered DNA methylation in the sperm of MIA-exposed offspring highlights the possibility that epigenetic processes in the male germline play a role in the transgenerational effects of MIA.Fig. 1BREEDING SCHEME USED TO GENERATE F1, F2, AND F3 OFFSPRING WITH CONTROL AND POLY(I:C)-DERIVED ANCESTORS.: a Breeding scheme used to generate F1 and F2 generations with control and immune-challenged ancestors. Pregnant F0 mice were treated with vehicle control (CON) or poly(I:C) (POL) solution to obtain F1 CON and F1 POL offspring. To obtain F2 POL offspring via the paternal lineage (POL-PL), male F1 POL offspring were mated with female F1 CON offspring; and to generate F2 POL offspring via the maternal lineage (ML-POL), female F1 POL offspring were crossed with male F1 CON offspring. F1 CON males and females were crossed to obtain the F2 control lineage (F2 CON). b Breeding scheme used to obtain the F3 generation with control and immune-challenged ancestors deriving from the paternal lineage (PL). Pregnant F0 mice were treated with control (CON) or poly(I:C) (POL) solution, and F1 POL males were mated with F1 CON females to generate F2 POL-PL offspring. F2 POL-PL males were then mated with F2 CON females to obtain F3 POL-PL offspring. CON F3 offspring were generated by crossing F2 CON males and females.Fig. 2EFFECTS OF MATERNAL IMMUNE ACTIVATION ON THE LOCOMOTOR REACTION TO ACUTE SYSTEMIC AMPHETAMINE CHALLENGE IN F1, F2, AND F3 OFFSPRING.: All animals were first placed into the open field and allowed to habituate for 30 min, after which they were injected with amphetamine (Amph; 2.5 mg/kg) or saline (Sal) solution. The line plots show the distance moved as a function of 5-min-bins during the initial habituation and subsequent drug phase; the bar plots depict the mean distance moved during the drug phase. a Locomotor activity levels in F1 offspring of control (CON) and poly(I:C)-treated (POL) mothers. The data represent male and female offspring together, as there were no sex-specific effects of MIA on Amph sensitivity in F1 offspring. b Locomotor activity levels in male F2 CON offspring and male F2 POL offspring generated via the maternal (POL-ML) or paternal (POL-PL) lineages. c Locomotor activity levels in male F3 CON offspring and male F3 POL offspring generated via the paternal lineage (POL-PL). *p < 0.05, **p < 0.01, and ***p < 0.001, based on post-hoc comparisons. All values are means + SEM. The numbers of animals in each group are provided in Table 1.Fig. 3EFFECTS OF MATERNAL IMMUNE ACTIVATION ON THE EXPRESSION OF TYROSINE HYDROXYLASE (TH) AND NUCLEAR RECEPTOR-RELATED 1 PROTEIN (NURR1) IN THE VENTRAL MIDBRAIN OF MALE F1, F2 AND F3 OFFSPRING.: The graphs depict the normalized levels of Th and Nurr1 mRNA levels (fold changes) of individual animals measured by quantitative RT-PCR. a Th and Nurr1 mRNA levels in F1 offspring of control (CON) and poly(I:C)-treated (POL) mothers. b Th and Nurr1 mRNA levels in F2 CON offspring and F2 POL offspring generated via the paternal lineage (POL/PL). c Th and Nurr1 mRNA levels in F3 CON offspring and F3 POL offspring generated via the paternal lineage (POL/PL). *p < 0.05 and **p < 0.01, based on independent Student's t tests (two-tailed).Fig. 4EFFECTS OF MATERNAL IMMUNE ACTIVATION ON DNA METHYLATION AT PROMOTER REGIONS OF TYROSINE HYDROXYLASE (TH) AND NUCLEAR RECEPTOR-RELATED 1 PROTEIN (NURR1) IN THE VENTRAL MIDBRAIN (VMB) AND SPERM CELLS OF MALE OFFSPRING.: The line plots depict percent DNA methylation of specific CpGs in selected amplicons, and the scatter plots show the mean percent DNA methylation across all individual CpGs included in the corresponding amplicon. Amplicon and primer sequences and position of CpGs in each amplicon are provided in Supplementary Table S2 and S3. a DNA methylation levels of Th and Nurr1 in the vMB of F1 offspring of control (CON) and poly(I:C)-treated (POL) mothers. *p < 0.05 and #p = 0.059, based on ANOVA and post-hoc tests. b DNA methylation levels of Th and Nurr1 in the sperm of F1 CON and POL offspring. *p < 0.05, based on ANOVA. c DNA methylation levels of Th and Nurr1 in the vMB of F2 CON offspring and F2 POL offspring generated via the paternal lineage (POL/PL). *p < 0.05, based on ANOVA.