Chronic Ethanol Treatment and Withdrawal Alter ACPD-Evoked Calcium Signals in Developing Purkinje Neurons

Abstract

Background: Alcohol exposure during human fetal development can result in fetal alcohol syndrome, a condition characterized by central nervous system dysfunction. Detailed studies in animal models of fetal alcohol syndrome show that the cerebellar region is particularly sensitive to alcohol exposure during early development; however, the cellular mechanisms underlying the alcohol sensitivity of the immature cerebellum are poorly understood. Methods: Primary neuronal cultures of cerebellar cells were prepared from embryonic day 20 rat pups. Cultures were exposed to ethanol (33 mM; 150 mg/100 ml) during the main period of morphological development of the Purkinje neurons, from 6 to 17 days in vitro. After the ethanol treatment, the response of Purkinje neurons to the selective metabotropic glutamate receptor (mGluR) agonist (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD; 300 μM) was examined in parallel fura-2 Ca2+ imaging and current-clamp experiments. In an additional set of experiments, ethanol-treated cultures were allowed to withdraw from ethanol overnight before recordings were performed. Results: In Ca2+ imaging studies, the mean peak amplitude of ACPD-evoked Ca2+ signals was depressed in the dendritic region of chronic ethanol–treated Purkinje neurons compared with control neurons (p < 0.05, unpaired t test), whereas there was no apparent difference in the somatic region. In contrast, peak ACPD-evoked Ca2+ signals were enhanced in both the somatic and dendritic regions of withdrawn Purkinje neurons compared with control neurons. Parallel current-clamp studies showed no consistent effect of chronic ethanol treatment or ethanol withdrawal on the membrane response to ACPD. Conclusions: These results show that prolonged ethanol exposure and early withdrawal lead to alterations in mGluR-evoked Ca2+ signaling in cerebellar Purkinje neurons. Metabotropic GluRs in the Purkinje neuron play important roles in cerebellar development and function, suggesting that alterations of mGluR signaling pathways by ethanol may play a key role in the actions of ethanol on the developing cerebellum.