Regional differences in cell loss associated with binge-like alcohol exposure during the first two trimesters equivalent in the rat

Abstract

Women who abuse alcohol during pregnancy may deliver offspring who could be diagnosed with fetal alcohol syndrome (FAS) or a less severe deficit involving cognitive and behavioral disorders. The severity of the deficits may involve the interaction of several known risk factors, such as alcohol consumption pattern or duration, the timing of alcohol consumption relative to critical windows of vulnerability, or the inherent differential vulnerability among the various brain regions to alcohol-induced brain injury. In this study, we explore the vulnerability of the different brain regions by making cell counts from multiple brain regions. Specifically, we used stereological cell-counting techniques to estimate the total cell numbers in the cerebellum (Purkinje and granule cells), olfactory bulb (mitral and granule cells), hippocampus (CA1 and CA3 cells), and dentate gyrus (granule cells). Groups of timed-pregnant Sprague–Dawley rats were assigned to one of five treatments: alcohol by intragastric intubation (2.25, 4.5, or 6.5 g/kg/day), nutritional control [pairfed and intubated=Pairfed) and intubated], and normal control (Chow). Treatments began on embryonic day 1 (E1) and continued through E20. On E33 (usually postnatal day 10), all offspring were perfused intracardially with saline followed by fixatives. Representative forebrains, cerebella, and olfactory bulb from each group were processed for cell counting. The optical dissector was used to obtain cell densities, while Cavalieri's principle was used to calculate the reference volume. The product of density and volume gave unbiased estimates of the total neuronal number within each brain region. Overall peak BACs (regardless of sampling day) for the three alcohol groups averaged 136, 290, and 422 mg/dl for the 2.25-, 4.5-, and 6.5-g/kg groups, respectively. The total number of cerebellar Purkinje cells was reduced in the 6.5-g/kg group relative to controls, while the total number of olfactory bulb mitral cells and hippocampal CA1 and CA3 pyramidal cells from all alcohol-treated groups was not different from controls. Total numbers of granule neurons were reduced in the cerebellum and olfactory bulb of offspring exposed to 4.5 or 6.5 g/kg/day, but granule cell numbers in the dentate gyrus were not affected by the prenatal alcohol treatment. Taken together with previous findings, these data demonstrate that prenatal alcohol exposure results in regional vulnerability of various brain structures and underscores the variability of deleterious effects of alcohol on brain development.