Cerebellar hypoplasia in experimental
fetal alcohol syndrome (FAS) is associated with impaired
insulin-stimulated survival signaling. In vitro studies demonstrated that
ethanol inhibition of neuronal survival is mediated by apoptosis and
mitochondrial dysfunction. Since
insulin and
insulin-like growth factors (IGFs) regulate energy metabolism, and
ethanol can exert its toxic effects by causing oxidative damage to
DNA and
proteins, we further characterized the effects of chronic gestational exposure to
ethanol on mitochondrial gene expression, and the degree to which
ethanol inhibition of mitochondrial function is mediated by impaired
insulin/IGF responsiveness. Pregnant Long-Evans rats were fed isocaloric liquid diets containing 0, 2, 4.5, 6.5, or 9.25% v/v
ethanol from gestation day 6 through delivery. Cerebella harvested on postnatal day 1 were examined for indices of oxidative stress, and
mRNA levels of mitochondrial,
pro-oxidant, and pro-apoptosis gene expression. Rat primary cerebellar neuron cultures were used to characterize the effects of
ethanol (50 mM for 96 h) on
insulin and IGF stimulated mitochondrial function and
ATP production.
Ethanol-exposed cerebella had significantly reduced
mRNA levels of mitochondrial genes encoding Complexes II-A, IV, and V, increased expression of p53 and
NADPH oxidase (NOX) 1 and 3, and increased immunoreactivity for
4-hydroxy-2,3-nonenal (HNE) and 8-OHdG in cerebellar granule cells. The activations of p53 and NOX genes were highest in cerebella from pups exposed to the 6.5 or 9.25%
ethanol containing diet, whereas the impairments in mitochondrial Complex IV and V expression were similar at low and high levels of
ethanol exposure. In vitro experiments confirmed that
ethanol treatment reduces neuronal expression of mitochondrial genes encoding Complexes IV and V, impairs mitochondrial function and
ATP production, and increases HNE and 8-OHdG immunoreactivity, but they also showed that these effects were not
insulin- or IGF-dependent. Together, the results suggest that
mitochondrial dysfunction, oxidative stress, and DNA damage in FAS may be largely due to the toxic effects of
ethanol rather than specific impairments in
insulin or IGF signaling.