The contribution of the locus coeruleus-norepinephrine system in the emergence of defeat-induced inflammatory priming

Abstract

Exposure to psychosocial stress is known to precipitate the emergence of stress related psychiatric disorders such as depression and anxiety. While mechanisms by which this occurs remain largely unclear, recent evidence points towards a causative role for inflammation. Neurotransmitters, such as norepinephrine (NE), are capable of regulating expression of proinflammatory cytokines and thus may contribute to the emergence of stress-related disorders. The locus coeruleus (LC) is the major source of norepinephrine (NE) to the brain and therefore the current study utilized N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4), an LC selective noradrenergic neurotoxin, to determine the discrete involvement of the LC-NE system in social defeat-induced inflammation in LC projection regions including the central amygdala (CeA), dorsal raphe (DR) and plasma. In the current study, rats were exposed to brief social defeat or control manipulations on 5 consecutive days. To determine whether a history of social defeat enhanced or "primed" the inflammatory response to a subsequent defeat exposure, all rats regardless of stress history were exposed to an acute social defeat challenge immediately preceeding tissue collection. As anticipated, prior history of social defeat primed inflammatory responses in the plasma and CeA while neuroinflammation in the DR was markedly reduced. Notably, DSP-4 treatment suppressed stress-induced circulating inflammatory cytokines independent of prior stress history. In contrast, neuroinflammation in the CeA and DR were greatly augmented selectively in DSP-4 treated rats with a history of social defeat. Together these data highlight the dichotomous nature of NE in stress-induced inflammatory priming in the periphery and the brain and directly implicate the LC-NE system in these processes.

Keywords: Central amygdala; DSP-4; Dorsal raphe; Inflammation; Locus coeruleus; Neuroinflammatory priming; Norepinephrine; Peripheral inflammatory priming; Resident intruder paradigm; Social defeat.

Figure 1.. Study design and timeline for animals with a history of social defeat (A) or control (B).

10 days prior to the onset of stress all rats were implanted with intracerebroventricular (ICV) cannulas. Seven days prior to stress, DSP-4 or vehicle were administered. DSP-4 and vehicle treated rats were subjected to either 5 daily exposures to social defeat or control (daily between 0900–1130, Vehicle/Control N=8; DSP-4/Control N=7; Vehicle/Defeat N=7; DSP-4/Defeat N=7). Six days after the final social defeat/control exposure, all rats, regardless of stress history, were exposed to an acute social defeat challenge immediately prior to euthanasia and tissue collection. Two separate subsets of rats were exposed to 5 daily control sessions and euthanized 6 days later under resting, non-stress conditions (subset not represented in figure). These additional subsets were utilized for verification of study design.

Figure 2.. DSP-4 treatment suppresses circulating cytokine release in response to an acute social defeat challenge regardless of stress history.

Following the acute social defeat challenge, vehicle treated animals with a history of social defeat exhibited enhanced (A) IL-6, (B) INF-γ, and (C) TNF-α, compared with vehicle treated controls consistent with stress-induced inflammatory priming. Strikingly, treatment with DSP-4 (panels A-F) suppressed stress-induced facilitation of IL-6, TNF-α, and INF-γ, as well as increases in (D) IL-1β, (E) IL-4 and (F) IL-13, regardless of stress history. *p<0.05 vs. vehicle control; ^αp<0.05, ^βp<0.001 vs. vehicle defeat.

Figure 3.. DSP-4 treatment inhibits acute defeat-induced plasma NE and corticosterone habituation-induced by a history of social defeat.

(A) Following the acute social defeat challenge, vehicle treated rats with a history of social defeat exhibited a sensitized peripheral NE response. This defeat-induced sensitization was inhibited in DSP4 treated rats. (B) Vehicle treated rats with a history of social defeat selectively exhibit a habituated corticosterone response following the acute defeat challenge. In contrast, treatment with DSP-4 inhibited corticosterone habituation in rats with a history of defeat. *p<0.05 vs. vehicle treated rats with a history of control; ^αp<0.05 vs. vehicle treated rats with a history of defeat.

Figure 4.. Treatment with DSP-4 exacerbates the acute defeat-induced neuroinflammation and priming in the CeA.

Following the acute social defeat challenge, vehicle treated rats with a history of social defeat exhibited moderate but consistent increases in pro-inflammatory cytokines (A) IL-1β, (B) IL-6, (C) INF-γ, (D) TNF-α and anti-inflammatory (E) IL-1RA and (F) IL-4 compared with vehicle treated rats with a history of control. Treatment with DSP-4 produced modest increases in stress-induced cytokines in rats with a history of control, while the LC-NE lesion robustly exacerbated the neuroinflammatory priming evident in rats with a history of social defeat exposure for all cytokines except TNF-α. *p<0.05 vs. vehicle treated rats with a history of control; ^ƒp=0.07, ^βp<0.001 vs. vehicle treated rats with a history of defeat; ^γp=0.07 ^#p<0.05, ^##p<0.01, ^###p<0.001 vs. DSP-4 treated rats with a history of control. Dotted horizontal line indicates 100% of vehicle control.

Figure 5.. A history of stress and DSP-4 treatment produce cytokine specific effects in the DR following acute defeat challenge.

Following the acute defeat challenge, vehicle treated rats generally exhibited a reduction of (A) IL-1β, (B) TNF-α, (C) IL-2, and (D) IL-4. However, the magnitude of these reductions were largely cytokine dependent. While treatment with DSP-4 modestly enhanced IL-1β independent of stress history, TNF-α, IL-2, and IL-4 were selectively enhanced in DSP-4 treated rats with a history of social defeat. ^#p<0.05 vs. DSP-4 treated rats with a history of control; ^βp<0.05 vs. vehicle treated rats with a history of defeat.

Figure 6.. The contribution of the LC-NE system in peripheral and central social defeat-induced inflammation.

The LC serves as a significant source of NE to the brain, including regions such as the CeA and DR, as well as the periphery by acting on sympathetic spinal nerves located in the dorsal horn. In the brain NE can alter inflammatory tone by acting on microglia. Peripheral NE can similarly act upon circulating macrophages but also impacts the spleen to increase inflammatory tone in the periphery. Social defeat exposure in rats with functional LC projections produced enhanced peripheral inflammation and region-specific alterations of inflammatory tone within the brain (A). In contrast, defeat exposure in rats with chemical lesion of the LC-NE system resulted in the inhibition of peripheral inflammation but also produced a robust facilitation of neuroinflammation in both the CeA and DR (B).