Title : Effects of probiotic supplementation on behaviour and hippocampal neurometabolites in pregnant rats exposed to chronic stress
Abstract:
Introduction: Chronic stress during pregnancy increases the risk of anxiety and adverse outcomes in mothers and offspring [1]. Probiotics may improve mental health and modulate biochemical processes during pregnancy, as supported by clinical and preclinical studies [2–3]. In our previous work, Lacticaseibacillus rhamnosus JB?1 mitigated mood-related disturbances and restored neurochemical balance in a rat stress model [4]. Here, we investigated whether JB-1 influences neurometabolic and behavioural outcomes in a preclinical model of gestational stress.
Materials and methods: Female Wistar rats were mated and assigned to stressed or control conditions. Stressed rats (n = 12) underwent chronic unpredictable mild stress (CUMS) during gestation, receiving either JB-1 bacteria or placebo (phosphate-buffered saline), while control rats (n = 13) were supplemented the same but were not exposed to stress. At gestational day (GD) 18, anxiety-like behaviour was assessed using the Open Field (OFT) and Elevated Plus Maze (EPM) tests. On GD19, Magnetic Resonance Spectroscopy (MRS) was performed in the hippocampus with a 7T Bruker animal MRI system. Spectral data were processed with jMRUI software. Statistical analysis was performed using two-way ANOVA with Tukey’s multiple comparisons test.
Results: There was a significant effect of stress on the number of entries to the center arena (F(1,21) = 5.30, p = 0.03) and on moving time in the OFT (F(1,21) = 8.69, p = 0.008), indicating reduced exploratory behaviour and locomotor activity in stressed groups compared to controls. No significant effects were observed in the EPM test, and bacteria did not affect behaviour in either task. While stress effects were evident in OFT parameters, Tukey’s post hoc tests did not identify significant pairwise differences among groups. MRS analysis revealed a significant stress × bacteria interaction for GABA (F(1,21) = 6.02, p = 0.02), although no significant post hoc group differences were detected. For glutamate, both an interaction (F(1,21) = 5.89, p = 0.02) and a main effect of stress (F(1,21) = 4.51, p = 0.05) were found; post hoc analysis indicated that JB?1 bacteria reduced glutamate levels under stress (Control: JB-1 vs. Stress: JB-1, p = 0.02). For choline, a significant interaction was observed (F(1,21) = 9.99, p = 0.005), with post hoc tests showing higher choline levels in the Control: JB-1 group compared to Control: Placebo (p = 0.03) and Stress: JB-1 (p = 0.02). Finally, a main effect of stress was detected for N-acetylaspartate (F(1,21) = 4.87, p = 0.04), but without significant post hoc differences.
Conclusion: Chronic stress during pregnancy reduced exploratory behaviour and locomotor activity, confirming effective stress induction. While JB-1 supplementation did not alter behavioural outcomes, MRS analyses revealed metabolite-specific effects, including modulation of glutamate and choline under stress. These findings suggest that JB-1 may influence neurometabolic responses to gestational stress, even in the absence of overt behavioural changes.
