Effect of gestational ethanol exposure on parvalbumin and calretinin expressing hippocampal neurons in a chick model of fetal alcohol syndrome Audrey G. Marshall a , Molly M. McCarthy a , Kirk M. Brishnehan a , Venugopal Rao b , Lyn M. Batia a , Madhul Gupta c , Srijit Das d , Nilesh K. Mitra e , Joydeep D. Chaudhuri a, * a Department of Anatomy, Midwestern University, 19555N, 59th Avenue, Glendale, AZ 85308, USA b Department of Anatomy, University Malaysia Sarawak, Sarawak, Malaysia c Department of Biology, University of Toronto, Toronto, ON, Canada d Department of Anatomy, University Kebangsaan Malaysia, Kuala Lumpur, Malaysia e Department of Anatomy, International Medical University, Kuala Lumpur, Malaysia Received 25 October 2007; received in revised form 29 October 2008; accepted 9 December 2008 Abstract Fetal alcohol syndrome (FAS), a condition occurring in some children of mothers who have consumed alcohol during pregnancy, is characterized by physical deformities and learning and memory deficits. The chick hippocampus, whose functions are controlled by inter- neurons expressing calcium-binding proteins parvalbumin (PV) and calretinin (CR), is involved in learning and memory mechanisms. Effects on growth and development and hippocampal morphology were studied in chick embryos exposed to 5% and 10% ethanol volume/volume (vol/vol) for 2 or 8 days of gestation. There was a significant dose-dependent reduction (P ! .05) in body weight and mean number per section of PV and CR expressing hippocampal neurons in ethanol-exposed chicks, without alterations in neuronal nuclear size or hippocampal volume, compared appropriate controls. Moreover, when chicks exposed to 5% ethanol for 2 and 8 days of gestation were compared, no significant differences were found in body parameters or neuronal counts. Similarly, exposure to 10% ethanol did not induce any significant changes in chicks exposed for 2 or 8 gestational days. Thus, these results suggest that gestational ethanol exposure induces a reduction in the mean number per section of PV and CR expressing hippocampal neurons, and could be a possible mechanism responsible for learning and memory disorders in FAS. Ó 2009 Elsevier Inc. All rights reserved. Keywords: FAS; Chicks; Hippocampus; Learning; Memory; Parvalbumin; Calretinin Introduction Fetal alcohol spectrum disorder (FASD) is a broad term describing a range of effects that can occur in an individual following maternal alcohol consumption during pregnancy. The most serious complication of FASD is an irreversible condition termed fetal alcohol syndrome (FAS) by Jones and Smith (1973). Children affected with FAS exhibit char- acteristic craniofacial anomalies and physical and mental growth retardation. They also demonstrate learning and memory impairments without apparent physical deformities (Rasmussen, 2005) and with a relatively unaffected overall intelligence quotient (IQ) performance (Mattson et al., 1996). The reported neurological deficits include impairment in verbal information acquisition and word comprehension, and defects in visuospatial processing (Eckardt et al., 1998; Riley and McGee, 2005; Sokol et al., 2003). Deficits in spatial learning have also been demonstrated in animal models of FAS following gestational ethanol exposure in early pregnancy (Matthews and Simson, 1998; Summers et al., 2006) and throughout the entire pregnancy (Gianou- lakis, 1990). Impairment in long-term memory formation has also been reported in chicks following ethanol exposure during the first 3 days of gestation (Rao and Chaudhuri, 2007). The hippocampus in both mammals and chicks is an essential functional component of the learning and memory mechanism (Horner et al., 1995; Kullmann and Lamsa, 2007) and is vulnerable to damage by neurotoxic agents (Korbo et al., 1996; Walsh and Emerich, 1988). The chick hippocampus is homologous to the mammalian hippo- campus based on topography, development, and role in * Corresponding author. Tel.: þ1-623-572-3332; fax: þ1-623-572- 3679. E-mail address: joydeep.chaudhuri@rediffmail.com (J.D. Chaudhuri). 0741-8329/09/$ e see front matter Ó 2009 Elsevier Inc. All rights reserved. doi: 10.1016/j.alcohol.2008.12.004 Alcohol 43 (2009) 147e161