Posttraumatic stress disorder-like induction elevates β-amyloid levels, which directly activates corticotropin-releasing factor neurons to exacerbate stress responses

Abstract

Recent studies have found that those who suffer from posttraumatic stress disorder (PTSD) are more likely to experience dementia as they age, most often Alzheimer's disease (AD). These findings suggest that the symptoms of PTSD might have an exacerbating effect on AD progression. AD and PTSD might also share common susceptibility factors such that those who experience trauma-induced disease were already more likely to succumb to dementia with age. Here, we explored these two hypotheses using a mouse model of PTSD in wild-type and AD model animals. We found that expression of human familial AD mutations in amyloid precursor protein and presenilin 1 leads to sensitivity to trauma-induced PTSD-like changes in behavioral and endocrine stress responses. PTSD-like induction, in turn, chronically elevates levels of CSF β-amyloid (Aβ), exacerbating ongoing AD pathogenesis. We show that PTSD-like induction and Aβ elevation are dependent on corticotropin-releasing factor (CRF) receptor 1 signaling and an intact hypothalamic-pituitary-adrenal axis. Furthermore, we show that Aβ species can hyperexcite CRF neurons, providing a mechanism by which Aβ influences stress-related symptoms and PTSD-like phenotypes. Consistent with Aβ causing excitability of the stress circuitry, we attenuate PTSD-like phenotypes in vivo by lowering Aβ levels during PTSD-like trauma exposure. Together, these data demonstrate that exposure to PTSD-like trauma can drive AD pathogenesis, which directly perturbs CRF signaling, thereby enhancing chronic PTSD symptoms while increasing risk for AD-related dementia.

Keywords: Alzheimer's disease; PTSD; crf; crfr1; crh; stress.

Figure 1.

PTSD-like induction procedure and phenotype. A, Diagram of the PTSD-like induction and testing procedure. B, Blood from wild-type animals was sampled every 15–30 min during the induction procedure. Circulating Cort rises dramatically at the onset of stress and slowly increases during immobilization before falling after the animal is released (n = 10). C, Cort levels were measured at rest (left) and after 20 min of restraint (right; peak). Cort levels in both conditions were lower in animals that had been exposed to PTSD-like induction (control, n = 18; stress, n = 15). D, Animals were tested for anxiety-related behavior. In the LDT test (2 left graphs), time in the light side and transitions to the light side were reduced after PTSD-like induction. In the OFA (3rd graph), the ratio between distance traveled in the center versus total distance traveled was reduced. In startle testing (right graph), wild-type animals displayed a nonstatistically significant increase in startle amplitude after PTSD-like induction (control, n = 15; stress, n = 21).

Figure 2.

APP/hAβ/PS1 PTSD-like phenotype. A, Corticosteroid levels during immobilization of wild-type animals (blue line) and APP/hAβ/PS1 animals (red). APP/hAβ/PS1 animals display a blunted corticosteroid release during PTSD-like induction (n = 10 per genotype). B, Resting and peak corticosteroid levels in control (light red) and PTSD-like induced (dark red) animals. Resting Cort levels fall in stress-exposed APP/hAβ/PS1 animals. Peak Cort levels are elevated in PTSD-like induced animals (n = 29 per condition). APP/hAβ/PS1 animals displayed exaggerated dexamethasone (DST) suppression compared with wild-type animals after being exposed to trauma (right; n = 6 per genotype). C, D, Behavioral assessment of PTSD-like phenotype in APP/hAβ/PS1 animals. APP/hAβ/PS1 animals have a baseline anxiety-related phenotype. Increased anxiety levels after PTSD-like induction are evident on the LDT test (light entries) and acoustic startle (control, n = 19; stress, n = 34). E, The percentage of animals that display a PTSD-like phenotype. The APP/hAβ/PS1 genotype is associated with a higher frequency of animals displaying PTSD-like behavior on all three behavioral assays. F, Aβ₄₀ and Aβ₄₂ levels in young (2–3 months, left bars; control, n = 8; stress, n = 10) and old (6–12 months, right bars; control, n = 8; stress, n = 5) APP/hAβ/PS1 animals. Control animals (light red bars) did not experience trauma, whereas PTSD animals experienced trauma at 8–12 weeks of age.

Figure 3.

PTSD-like phenotype requires Crfr1. A, Wild-type (blue) and APP/hAβ/PS1 (red) animals were crossed into a Crfr1^−/− background. In resulting mice, PTSD-like induction (darker bars) did not alter anxiety-related behavior or acoustic startle in either genotype. Data have been normalized to scores for wild-type, unstressed animals for each test (wild-type Crfr1^−/− control, n = 6; wild-type Crfr1^−/− stress, n = 5; APP/hAβ/PS1;Crfr1^−/− control, n = 14; APP/hAβ/PS1;Crfr1^−/− stress, n = 18). B, CSF Aβ levels were measured in APP/hAβ/PS1;Crfr1^−/− animals at both a young (3–4 months) and old (6–12 months) time point. PTSD-like induction did not alter Aβ levels when animals lacked Crfr1 (young APP/hAβ/PS1;Crfr1^−/−, n = 5 per condition; old APP/hAβ/PS1;Crfr1^−/−, n = 5 control, n = 6 stress).

Figure 4.

Aβ elevates CRF neuron activity in culture. A, Conditioned media from CHO cells (control CM) or CHO cells stably transfected with APP695 (SV695; Aβ CM) was measured for Aβ₄₀ concentration. In Aβ CM, we detected significantly more Aβ than in control CM, in which Aβ levels were below the detection limit of the assay (dashed line). CHO-K1 or SV695 cells incubated with 1 μm SEMA, a γ-secretase inhibitor, also had undetectable Aβ levels. B, Neurons were prepared from P0 mouse pups of the genotype CRF–cre;lox–stop–lox tomato (CRF–tomato). CRF neurons from these pups are fluorescent in culture. C, Whole-cell patch of CRF neurons with the addition of 10% conditioned media. Aβ conditioned media increased the action potential firing rate of CRF neurons (n = 8 cells). Both CM from CHO-K1 or SV695 cells incubated with SEMA did not affect CRF neuron firing rate (n = 3 cells). D, Quantification of results in C, normalized to the average firing rate of CRF neurons in 10% control CM. Aβ CM induced cells to fire, on average, three to four times faster than control CM.

Figure 5.

PVN CRF neurons are excited by Aβ conditioned media. Crf–cre;lsl–^tdTomato mice were imaged for tomato (left) and stained for mGluR5 (middle). CRF neurons in the PVN express a moderate level of mGluR5. Scale bar, 100 μm. B, CRF neurons in the PVN were patched in slice and perfused with control CM (top trace), CM from APP-expressing cells incubated with 1 μm SEMA (Aβ CM + Sema; middle trace), and CM from APP cells (Aβ CM; bottom trace). PVN CRF neurons increase action potential firing when exposed to Aβ containing CM (quantified in the bottom graph; n = 7 cells). C, Slices were incubated in synaptic blockers to isolate individual neuron excitability. CRF neurons increase action potential firing rate in Aβ CM (bottom trace; n = 5 cells) but not in control CM (top trace) or Aβ CM collected in the presence of SEMA (middle trace). D, PVN CRF neuron excitation by Aβ in the conditioned media is blocked when coapplied with MPEP, an mGluR5 antagonist (n = 6 cells). E, Dose response of PVN CRF neurons to varying dilutions of CM in ACSF. One percent Aβ CM caused a small increase in action potential firing. Ten percent Aβ CM caused a maximal response equivalent to the response to 20% Aβ CM (n = 3 cells).

Figure 6.

γ-Secretase inhibitors reverse PTSD-like phenotypes in APP/hAβ/PS1 animals. A, Treatment with SEMA during PTSD-like trauma exposure did not significantly alter behavior on the LDT test. B, SEMA decreased acoustic startle to a 120 dB sound. C, SEMA increased center/total distance traveled in the OFA. D, SEMA treatment decreased peak corticosteroid levels observed in trauma-exposed animals 2 weeks after the stress (for A–D, vehicle, n = 17; SEMA, n = 14). E, CSF sampled from unstressed APP/hAβ/PS1 animals after receiving 3 d of SEMA injections contained less Aβ₄₀ compared with vehicle controls (n = 7 per treatment).