MTHFR DEFICIENCY OR REDUCED INTAKE OF FOLATE OR CHOLINE IN PREGNANT MICE RESULTS IN IMPAIRED SHORT-TERM MEMORY AND INCREASED APOPTOSIS IN THE HIPPOCAMPUS OF WILD-TYPE OFFSPRING N. M. JADAVJI, a L. DENG, a O. MALYSHEVA, b M. A. CAUDILL b AND R. ROZEN a * a Departments of Human Genetics and Pediatrics, McGill University, The Research Institute of the McGill University Health Centre, Montreal, Canada b Division of Nutritional Sciences, Cornell University, Ithaca, USA Abstract—Genetic or nutritional disturbances in one-carbon metabolism, with associated hyperhomocysteinemia, can result in complex disorders including pregnancy complica- tions and neuropsychiatric diseases. In earlier work, we showed that mice with a complete deficiency of methylenetetrahydrofolate reductase (MTHFR), a critical enzyme in folate and homocysteine metabolism, had cogni- tive impairment with disturbances in choline metabolism. Maternal demands for folate and choline are increased dur- ing pregnancy and deficiencies of these nutrients result in several negative outcomes including increased resorption and delayed development. The goal of this study was to investigate the behavioral and neurobiological impact of a maternal genetic deficiency in MTHFR or maternal nutri- tional deficiency of folate or choline during pregnancy on 3-week-old Mthfr +/+ offspring. Mthfr +/+ and Mthfr +/À females were placed on control diets (CD); and Mthfr +/+ females were placed on folate-deficient diets (FD) or choline-deficient diets (ChDD) throughout pregnancy and lactation until their offspring were 3 weeks of age. Short- term memory was assessed in offspring, and hippocampal tissue was evaluated for morphological changes, apoptosis, proliferation and choline metabolism. Maternal MTHFR defi- ciency resulted in short-term memory impairment in off- spring. These dams had elevated levels of plasma homocysteine when compared with wild-type dams. There were no differences in plasma homocysteine in offspring. Increased apoptosis and proliferation was observed in the hippocampus of offspring from Mthfr +/À mothers. In the maternal FD and ChDD study, offspring also showed short-term memory impairment with increased apoptosis in the hippocampus; increased neurogenesis was observed in ChDD offspring. Choline acetyltransferase protein was increased in the offspring hippocampus of both dietary groups and betaine was decreased in the hippocampus of FD offspring. Our results reveal short-term memory deficits in the offspring of dams with MTHFR deficiency or dietary deficiencies of critical methyl donors. We suggest that deficiencies in maternal one-carbon metabolism during pregnancy can contribute to hippocampal dysfunction in offspring through apoptosis or altered choline metabolism. Ó 2015 IBRO. Published by Elsevier Ltd. All rights reserved. Key words: MTHFR deficiency, maternal folate deficiency, maternal choline deficiency, short-term memory, apoptosis. INTRODUCTION Folates are required for cellular proliferation, methylation reactions, and maintenance of homocysteine at non-toxic levels (Donnelly, 2001; Carmel et al., 2003; Obeid and Herrmann, 2005). The demand for folate is increased during pregnancy to accommodate development of the embryo, as well as growth of maternal tissue (Obeid and Herrmann, 2005; Safi et al., 2012). Inadequate dietary folate is a well- known risk factor for neural tube defects although the exact mechanism is unclear. Elevated plasma homocys- teine is observed in genetic or nutritional disruptions of folate metabolism since folate provides the one-carbon donor for homocysteine remethylation to methionine and consequently to S-adenosylmethionine for methyla- tion reactions. Hyperhomocysteinemia can have nega- tive consequences on fetal brain development, with impact on functions later in life, including effects on anxiety (Ferguson et al., 2005) and learning (Whitley et al., 1951). Elevated homocysteine levels have been shown to increase oxidative stress (Koz et al., 2010) and apoptosis (Chen et al., 2005; Jadavji et al., 2012), and to inhibit methylation reactions (Wang et al., 1997). Choline, through its metabolite betaine, can also provide carbon units to remethylate homocysteine, predominantly in the liver and kidney (Caudill, 2009). Flux through betaine-dependent homocysteine remethy- lation is increased when folate-dependent remethylation http://dx.doi.org/10.1016/j.neuroscience.2015.04.067 0306-4522/Ó 2015 IBRO. Published by Elsevier Ltd. All rights reserved. * Corresponding author. Address: The Research Institute of the McGill University Health Centre, 1001 De´carie Boulevard, Bloc E, Room E M0.3211, Montreal H4A 3J1, Canada. Tel: +1-514-934-1934x23839; fax: +1-514-933-3129. E-mail addresses: nafisa.jadavji@mail.mcgill.ca (N. M. Jadavji), deng_liyuan@hotmail.com (L. Deng), ovm4@cornell.edu (O. Maly- sheva), mac379@cornell.edu (M. A. Caudill), rima.rozen@mcgill.ca (R. Rozen). Abbreviations: Apo A-I, apolipoprotein A-I; CD, control diet; ChAT, choline acetyltransferase; ChDD, choline-deficient diet; EDTA, ethylenediaminetetraacetic acid; FD, folic acid-deficient diet; hpf, high-power field; HRP, horseradish peroxidase; MTHFR, methylenetetrahydrofolate reductase; NOR, novel object recognition. Neuroscience 300 (2015) 1–9 1