Forebrain-Specific CRF Overproduction During Development is Sufficient to Induce Enduring Anxiety and Startle Abnormalities in Adult Mice

Abstract

Corticotropin releasing factor (CRF) regulates physiological and behavioral responses to stress. Trauma in early life or adulthood is associated with increased CRF in the cerebrospinal fluid and heightened anxiety. Genetic variance in CRF receptors is linked to altered risk for stress disorders. Thus, both heritable differences and environmentally induced changes in CRF neurotransmission across the lifespan may modulate anxiety traits. To test the hypothesis that CRF hypersignaling is sufficient to modify anxiety-related phenotypes (avoidance, startle, and conditioned fear), we induced transient forebrain-specific overexpression of CRF (CRFOE) in mice (1) during development to model early-life stress, (2) in adulthood to model adult-onset stress, or (3) across the entire postnatal lifespan to model heritable increases in CRF signaling. The consequences of these manipulations on CRF peptide levels and behavioral responses were examined in adulthood. We found that transient CRFOE during development decreased startle habituation and prepulse inhibition, and increased avoidance (particularly in females) recapitulating the behavioral effects of lifetime CRFOE despite lower CRF peptide levels at testing. In contrast, CRFOE limited to adulthood reduced contextual fear learning in females and increased startle reactivity in males but did not change avoidance or startle plasticity. These findings suggest that forebrain CRFOE limited to development is sufficient to induce enduring alterations in startle plasticity and anxiety, while forebrain CRFOE during adulthood results in a different phenotype profile. These findings suggest that startle circuits are particularly sensitive to forebrain CRFOE, and that the impact of CRFOE may be dependent on the time of exposure.

Overexpression of Forebrain CRH During Early Life Increases Trauma Susceptibility in Adulthood

“…These data are consistent with our and others' previous reports showing reduced PPI and habituation following developmental or lifetime CRHOE (Dirks et al, 2002;Groenink et al, 2008;Toth et al, 2014). Pharmacological and genetic manipulation studies reported increased startle and reduced PPI following CRHR1 receptor hypersignaling, whereas CRHR2 receptor stimulation increased PPI (Risbrough et al, 2003(Risbrough et al, , 2004.…”

“…In contrast, predator stress increased avoidance of the light chamber in females regardless of CRHOE dev exposure ( Figure 1 and Table 2). CRHOE dev females also exhibited a trend for increased avoidance in non-stressed groups (duration: F stress × CRHOE (1,32) = 3.49, p = 0.071, post hoc: p = 0.098; Table 2) as described previously (Toth et al, 2014).…”

“…To induce CRHOE in spatio-temporally restricted manner, we used double-mutant mice carrying CamkIIα promoterdriven rtta2 transgene (Michalon et al, 2005) and doxycycline (DOX)-regulated tetO promoter fused to the Crh gene (Vicentini et al, 2009) on a C57BL/6J background as previously described (Toth et al, 2014). The Crh transgene was turned on by DOX administration in breeder chow (Harlan Laboratories, Indianapolis, IN) to 'single-mutant' dams from postnatal day 2 for 3 weeks (PND2-PND23).…”

“…Typical litter sizes were four or five pups, producing one double-mutant male and one double-mutant female on average for testing. The DOX dose administered to the dam (6 mg/g food) induces forebrain-specific expression of Crh or Lacz reporter genes in the forebrain as early as PND0, with detectable levels after 4 days, reaching its maximum after 1 week and returning to baseline levels 14 days after DOX treatment is terminated (Michalon et al, 2005;Toth et al, 2014). We and others have previously established that DOX alone (administration between PND2-23) does not affect startle reactivity and avoidance behavior in wild-type mice (Kolber et al, 2010;Toth et al, 2014), therefore control subjects were double-mutant mice without DOX treatment.…”

“…Startle reactivity was assessed in Plexiglas chambers (San Diego Instruments, San Diego, CA) as previously described (Adamec et al, 2010;Toth et al, 2014). Briefly, 1 week before stress exposure, baseline startle was assessed over 3 consecutive days using Session 1, which presented 10 105 dB pulses over 50 dB background in dark chambers.…”