Ethanol-related teratogenicity and neurobehavioural impairments: influence of dietary zinc supplementation during pregnancy.

Abstract

Ethanol consumption during pregnancy can result in wide range of negative outcomes, including pre-and post-natal mortality, growth retardation, physical abnormalities and brain deficits, manifested as behavioural impairments. These outcomes can result from “binge-drinking” (generally defined as >5 standard drinks on a single occasion) or chronic ethanol consumption. Ethanol-induced zinc (Zn) deficiency is one of the mechanisms proposed as a cause of ethanol teratogenicity. We have previously demonstrated in mice that ethanol exposure on gestational day (GD)8 (during organogenesis) can alter Zn homeostasis by inducing the Zn-binding protein metallothionein (MT) in the maternal liver. This causes plasma Zn concentrations to decrease as Zn redistributes into the liver, and consequently decreases the fetal Zn supply and increases the risk of teratogenicity. Subcutaneous Zn treatment with ethanol on GD8 can prevent the deleterious effects of ethanol on the fetus (i.e. physical abnormalities and spatial memory impairments). The main objective of this thesis was to investigate whether a less invasive approach of giving dietary Zn supplementation throughout pregnancy could provide similar protective benefits against a range of adverse outcomes caused by prenatal binge or chronic ethanol exposure. Binge ethanol exposure in early pregnancy (i.e. where mice are injected with 25% ethanol (0.015 ml/g) intraperitoneally at 0 and 4 hours on GD8) significantly increased the incidence of birth abnormalities measured on GD18. These included craniofacial abnormalities (microphthalmia, anophthalmia) and limb defects. Ethanol also increased postnatal mortality between birth and postnatal day (PD)60. In a separate study, offspring from dams given ethanol on GD8 were subjected to a physical and behavioural screening protocol (including tests for vision, olfactory, exploratory, anxiety and motor impairments) and subsequently a cohort of phenotypically-normal offspring were randomly selected for testing in a cross-maze escape task (for spatial learning and memory) and an object recognition test (for short-term non-spatial memory). While ethanol did not affect behaviour measured during screening, it resulted in spatial memory and object recognition memory impairments in adult offspring. The most important finding was that dietary Zn supplementation throughout pregnancy significantly increased plasma Zn concentrations at the time of ethanol exposure (avoiding the “typical” ethanol-induced decrease in plasma Zn) and prevented all negative outcomes resulting from early ethanol exposure (birth abnormalities, mortality, spatial and object recognition memory impairments). In the chronic ethanol mouse model (i.e. where mice were fed a liquid diet containing 27 % v/v ethanol-derived calories from GD6-18), ethanol did not affect offspring growth between birth and PD21 or spatial memory in adult offspring, thus, the influence of Zn supplementation could not be examined for these parameters. While ethanol decreased offspring weight at PD50 and increased mortality between birth and PD40, they were not prevented by Zn supplementation throughout pregnancy. The findings from this thesis emphasise that organogenesis is a particularly vulnerable period to ethanol exposure and even a binge of ethanol during this time can result in dysmorphology, mortality and spatial and object memory impairments in adulthood. In addition, dietary Zn supplementation is protective against the deleterious effects of binge ethanol exposure in early pregnancy.