Fibroblast Growth Factor 2 Reverses the Effects of Anxious Phenotype on the Early Emergence of Threat Learning in Infancy

Hider J, White A, Boulanger-Bertolus J, Sullivan R, Akil H, Debiec J
American College of Neuropsychopharmacology. 2017.


Background: Anxiety disorders are the most prevalent psychiatric disorders in childhood affecting around 10% of youth (Kessler et al., 2005). Maladaptive anxiety emerges early in life and disturbs child's psychosocial functioning and development. It is generally accepted that anxiety disorders are caused by a combination of inborn/hereditary and environmental factors. One of the best studied inborn risk factors for anxiety disorders in youth is anxious temperament whereas one of the best known environmental risk factors for anxiety disorders is a history of emotional trauma in childhood. Threat conditioning (TC) is the most commonly used experimental model of emotional trauma. In TC a neutral stimulus (conditioned stimulus, CS), such as neutral sound or odor is paired with an aversive unconditioned stimulus (US), typically a mild electric shock. As a result, an organism expresses threat responses such as freezing upon subsequent exposures to the CS. Previous studies show that fear conditioning in rodents is naturally attenuated until postnatal day (PN) 10 (for review: Debiec & Sullivan, 2016). However, the ontogeny of fear learning in phenotypically anxious organisms is mostly unknown. To address this issue, we used a selectively-bred anxiety-prone rat model, in which spontaneous anxiety-like behaviors emerge as early as at PN 11 (Maras et al., SFN Abstracts, 2014). We also investigated effects of the fibroblast growth factor 2 (FGF 2), which in known to have anxiolytic effects in developing rats (Graham & Richardson, 2009), on TC in anxiety-prone infant pups.
Methods: Experiment 1: Sprague-Dawley (SD) PN 4 rat pups selectively-bred for anxiety received 11 US electric shocks (0.4 mA, 1 s) to the tail, either paired with a CS peppermint odor (30 s) (Paired), or unpaired (Unpaired). Another group included pups that received 11 CS presentations (CS Only). Additional controls included wild-type SD pups matched for age and experimental conditions. OnPN 11, all pups were reexposed to 3 CSs and their behavior was video recorded and scored for freezing. Experiment 2: SD PN 2 pups received s.c. FGF2 (Sigma-Aldrich, F0291; 20ng/g in 50μl in 0.1M PBS w/ 1% BSA) or vehicle (0.1M PBS w/1% BSA, s.c.) injections. On PN 4, all pups received TC training (and on PN 11 testing) as described above. Data were analyzed with ANOVA followed by post hoc test (Experiment 1) or student t-test (Experiment 2).
Results: Experiment 1: Statistical analysis revealed no significant differences in freezing behavior among the wild type animals (p40.05), a finding consistent with previous studies using similar training parameters. However, the anxiety-prone Paired pups showed significantly higher levels of freezing as compared to the control groups (Unpaired and CS Only pups expressed comparable levels of freezing) (ANOVA: F(3,46) =16.51; po0.0001; post hoc: po0.05). Experiment 2: Statistical analysis revealed that pups injected with FGF 2 display significantly lower levels of freezing than vehicle controls. In other experiments, we found that early emergence of TC in anxiety-prone pups is associated with elevated levels of early expression c Fos gene expression, an effect moderated by FGF 2 neonatal administration.
Conclusions: Our results demonstrate that threat learning in phenotypically anxious pups occurs very early in life, before they spontaneously express anxious behaviors at PN 11, and before the emergence of adult-like threat conditioning in wild-type rats at PN 10. These results suggest that the early emergence of threat learning in the anxious phenotype affects early-life experiences and may contribute to the development of maladaptive threat responses, such as these seen in anxiety disorders. The fact that early postnatal administration of FGF 2 reversed the effects of anxious phenotype on TC in infancy suggests a potential role for FGF 2 in prevention of childhood anxiety disorders.