Experiment 2: Sex differences and effects of acute stress on flavor neophobia and conditioned taste aversion

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This study investigates the interaction between biological sex and acute stress (forced swim) on flavor neophobia and the acquisition/extinction of Conditioned Taste Aversion (CTA). Hypotheses: H1 (Neophobia): Females will show higher neophobia (lower consumption of the novel flavor) than males. H2 (Stress on Neophobia): Acute stress (forced swim) will increase neophobia (reduce consumption) compared to controls. We predict an interaction where the stress effect might be attenuated in females due to a ceiling effect in their baseline neophobia. H3 (CTA Acquisition): Males will show stronger CTA acquisition (lower intake on the first test day) compared to females. H4 (CTA Extinction): Females will show faster extinction of the taste aversion (faster recovery of consumption) compared to males across extinction sessions. Design PlanStudy type: Experiment. Blinding: No blinding. Experimenters will be aware of group assignments during data collection and handling. Study design: Factorial 2 x 2 between-subjects design. Factor 1 (Sex): Male vs. Female. Factor 2 (Stress): Stress (Forced Swim) vs. Control (No Swim). Sampling PlanData collection: Registration prior to creation of data.Sample size: N = 32 rats (8 subjects per group). Randomization strategy: Subjects will be assigned to groups using a stratified randomization method based on body weight to ensure equal mean weights across experimental conditions at the start of the study. Rationale: Sample size is based on previous experiments in our laboratory using similar protocols and a power analysis using G*Power (Power = 0.80, alpha = 0.05). Variables: Independent Variables: Sex (Male/Female). Stress Condition: Forced Swim (5 min session) vs. Control (Handling). Dependent Variable: Fluid consumption (ml) measured to the nearest 0.1 ml. Stimuli:CS (Flavor): Citric acid solutiomn (3% per liter of water). US (Aversive agent): LiCl (0.15M), injected i.p. at 0.5% body weight. Dissolved in saline solution . Procedure Experimental Procedure: Behavioral Protocol The following timeline describes the water deprivation and conditioning schedule: Monday: Water deprivation (bottles removed). Tuesday: Baseline recording. 30-minute access to water; total intake is recorded. Wednesday (Stress Induction): 5-minute Forced Swim Test (FST), followed by 5 minutes of activity. Subsequently, 30-minute access to the citric acid solution (intake recorded), followed by 30 minutes of water access. Thursday (Body Weight & Stress): Animal weighing. 5-minute FST, followed by 5 minutes of activity. 30-minute access to the citric acid solution (intake recorded). Friday: 5 minutes of activity, followed by 30-minute access to the citric acid solution (intake recorded). Saturday (Conditioning & Stress): 5-minute FST, followed by 5 minutes of activity. 30-minute access to the citric acid solution (intake recorded), immediately followed by LiCl administration. 30-minute water access provided thereafter. Sunday: 5 minutes of activity, followed by 30-minute access to the citric acid solution (intake recorded) and 30-minute water access. Monday (Testing): 5 minutes of activity, followed by 30-minute access to the citric acid solution (intake recorded) and 30-minute water access. Tuesday (Testing): 5 minutes of activity, followed by 30-minute access to the citric acid solution (intake recorded) and 30-minute water access. Exclusion Criteria: Animals consuming less than 1 ml of fluid during any experimental session (neophobia, conditioning, or tests). Animals showing visible motor deficits or vestibular problems during the forced swim sessions. Analysis Plan Software: Data will be analyzed using IBM SPSS Statistics.Neophobia: Univariate ANOVA (2 Sex x 2 Stress) on Wednesday consumption data. CTA Acquisition & Extinction: Mixed ANOVA of Repeated Measures.Between-subject factors: Sex (2) and Stress (2). Within-subject factor: Sessions (Test 1, Extinction 1, Extinction 2, Extinction 3). Significant interactions will be analyzed using Bonferroni-corrected post-hoc comparisons.

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