Supplementation with Choline Chloride in a Liquid Rodent Diet: Recipe Development for Differing Caloric Intake in a Fetal Alcohol Model

Objective Choline supplementation of maternal diet during preconception and gestational periods has been shown to ameliorate fetal alcohol effects on offspring. Researchers generally provide alcohol and choline to pregnant dams via gavage, but this is not physiologically relevant. An alternative method of choline supplementation is to supplement saccharine‐sweetened drinking water with choline, but this still requires ethanol gavage. These limitations necessitate a palatable novel method of choline delivery to ethanol‐exposed dams. The objective of this study was to devise a physiologically relevant means of delivering choline and ethanol to rat dams throughout gestation by developing a novel choline‐supplemented liquid ethanol diet that replicated previous choline supplementation doses of 250 mg choline per kg body weight per day (250 mg/kg/day). Methods Twelve Wistar rats were purchased from Charles River and bred at 100–120 days. Dams consumed diets in a 2 × 2 model: (Ethanol, No Ethanol) x (Choline, No Choline). Ethanol‐exposed animals also had a pair‐fed, calorie‐restricted group. Thus, 5 recipes were designed: 3 with choline supplementation and 2 without. Each diet was developed by analyzing consumption relative to weight after different treatments and increasing the choline chloride content in recipes until approximately 250 mg/kg/day choline was supplemented across groups. Blood was taken at gestational day 18 (GD18) for assay of choline and its metabolites. ANOVAs were run to analyze group differences in consumption and plasma choline metabolites. Results A set of recipes was formulated that achieved a 258.38 mg/kg/day mean choline dose with no significant differences between groups (means = 269.85, 251.71, 237.35, p <0.05). Importantly, the supplemented dams consumed the same amount of diet as the unsupplemented ( p <0.05). Significantly higher concentrations of choline and its metabolites were found in blood taken at GD18 from choline‐supplemented dams compared to control dams ( p <0.05). Discussion We successfully devised recipes that allow us to achieve a 250 mg/kg/day choline dose in choline‐supplemented groups. Separate recipes are required for Ethanol versus No Ethanol choline‐supplemented groups, because dams consuming an ethanol diet consume fewer calories than dams on a No Ethanol diet. Findings in choline metabolite concentrations in blood demonstrate that we have provided biologically active choline to pregnant dams. Conclusion It is feasible to supplement rodent liquid diet with choline chloride to achieve a more physiologically relevant model while accounting for non‐isocaloric intake in a fetal alcohol model. Support or Funding Information NIAAA K01‐AA021134 to last author