si-RNA inhibition of brain insulin or insulin-like growth factor receptors causes developmental cerebellar abnormalities: relevance to fetal alcohol spectrum disorder

BACKGROUND: In experimental models of fetal alcohol spectrum disorder (FASD), cerebellar hypoplasia and hypofoliation are associated with insulin and insulin-like growth factor (IGF) resistance with impaired signaling through pathways that mediate growth, survival, plasticity, metabolism, and neurotransmitter function. To more directly assess the roles of impaired insulin and IGF signaling during brain development, we administered intracerebroventricular (ICV) injections of si-RNA targeting the insulin receptor, (InR), IGF-1 receptor (IGF-1R), or IGF-2R into postnatal day 2 (P2) Long Evans rat pups and examined the sustained effects on cerebellar function, structure, and neurotransmitter-related gene expression (P20). RESULTS: Rotarod tests on P20 demonstrated significant impairments in motor function, and histological studies revealed pronounced cerebellar hypotrophy, hypoplasia, and hypofoliation in si-InR, si-IGF-1R, and si-IGF-2R treated rats. Quantitative RT-PCR analysis showed that si-InR, and to a lesser extent si-IGF-2R, broadly inhibited expression of insulin and IGF-2 polypeptides, and insulin, IGF-1, and IGF-2 receptors in the brain. ELISA studies showed that si-InR increased cerebellar levels of tau, phospho-tau and β-actin, and inhibited GAPDH. In addition, si-InR, si-IGF-1R, and si-IGF-2R inhibited expression of choline acetyltransferase, which mediates motor function. Although the ICV si-RNA treatments generally spared the neurotrophin and neurotrophin receptor expression, si-InR and si-IGF-1R inhibited NT3, while si-IGF-1R suppressed BDNF. CONCLUSIONS: early postnatal inhibition of brain InR expression, and to lesser extents, IGF-R, causes structural and functional abnormalities that resemble effects of FASD. The findings suggest that major abnormalities in brains with FASD are mediated by impairments in insulin/IGF signaling. Potential therapeutic strategies to reduce the long-term impact of prenatal alcohol exposure may include treatment with agents that restore brain insulin and IGF responsiveness.