Developmental alterations in olivary climbing fiber distribution following postnatal ethanol exposure in the rat

Abstract

Ethanol exposure during postnatal days (PN) 4-6 in rats alters cerebellar development resulting in significant loss of Purkinje cells. There is little knowledge, however, on what happens to the neurons that survive. In this study, rat pups were treated with a daily dose of ethanol (either 3.6 or 4.5 g/kg body weight) delivered by intragastric intubation on PN4, PN4-6, or PN7-9. Then the interactions between climbing fibers and Purkinje cell were examined on PN14 using confocal microscopy. Mid-vermal cerebellar sections were stained with antibodies to calbindin-D28k (to visualize Purkinje cells) and vesicular glutamate transporter 2 (VGluT2, to visualize climbing fibers). Confocal z-stack images were obtained from Lobule 1 and analyzed with Imaris software to quantify the staining of the two antibodies. The VGluT2 immunostaining was significantly reduced in the PN4 and PN4-6 ethanol groups for the 4.5 g/kg dose level, compared to controls, indicating that the cerebellar circuitry was significantly altered following developmental ethanol exposure. Not only were there fewer Purkinje cells following ethanol exposure, but the surviving neurons had significantly fewer VGluT2-labeled synapses. These alterations in the synaptic integrity were both dose dependent and temporally dependent.

Overexpression of Arabidopsis Translationally Controlled Tumor Protein Gene AtTCTP Enhances Drought Tolerance with Rapid ABA-Induced Stomatal Closure

“…The confocal acquired z-stacks were also reconstructed into 3D image projections using the IMARIS software (Bitplane AG) and converted the confocal images into surface mode images. This software allowed the construction of an isosurface for each fluorescent channel (GFP channel; microtubule arrays) and the investigation of microtubule arrays more precisely (Pierce et al, 2010). transgenic lines overexpressing AtTCTP (hereafter, OX lines) were generated in a Col-0 background.…”

Electrophysiological and Immunohistochemical Evidence for an Increase in GABAergic Inputs and HCN Channels in Purkinje Cells that Survive Developmental Ethanol Exposure

“…Thus, when PCs die during the postnatal period, the loss is permanent. Interestingly, the window of vulnerability for PCs has a precise temporal boundary in that there is very little PC loss from ethanol exposures on PN 7 or later [30, 33, 34]. Additionally, although a recent study reported significant ethanol-induced loss of PCs following exposure on PN 0 or PN 2 [35], the general findings have not identified ethanol-specific PC loss before PN 4 [30, 36].…”

“…The climbing fibers have a critical role in the proper functioning of the cerebellar circuitry, yet they comprise only 1–2 % of the synapses on the PC [45–47]. We hypothesized that developmental ethanol exposure would negatively impact the establishment of synaptic contacts by climbing fibers onto PCs [34, 48]. We tested this hypothesis by using immunostaining for vesicular glutamate transporter 2 (VGluT2) to identify climbing fiber synapses in Lobule I [45–47] and calbindin-D28k to identify PCs [37].…”

“…The VGluT2 immunostaining was significantly reduced by the PN 4 and PN 4–6 but not PN7–9 exposures when the peak blood ethanol concentration was 365±39 mg/dL [34, 48]. Additionally, we identified that the surviving neurons from these two exposure groups also had significantly fewer VGluT2-labeled synapses per PC when compared to controls [34, 48].…”

“…Additionally, we identified that the surviving neurons from these two exposure groups also had significantly fewer VGluT2-labeled synapses per PC when compared to controls [34, 48]. These effects were both dose- and temporally dependent, and a single ethanol exposure proved sufficient to cause alterations in the synaptic integrity.…”