Decreased Interleukin-4 Release from the Neurons of the Locus Coeruleus in Response to Immobilization Stress

Abstract

It has been demonstrated that immobilization (IMO) stress affects neuroimmune systems followed by alterations of physiology and behavior. Interleukin-4 (IL-4), an anti-inflammatory cytokine, is known to regulate inflammation caused by immune challenge but the effect of IMO on modulation of IL-4 expression in the brain has not been assessed yet. Here, it was demonstrated that IL-4 was produced by noradrenergic neurons in the locus coeruleus (LC) of the brain and release of IL-4 was reduced in response to IMO. It was observed that IMO groups were more anxious than nontreated groups. Acute IMO (2 h/day, once) stimulated secretion of plasma corticosterone and tyrosine hydroxylase (TH) in the LC whereas these increments were diminished in exposure to chronic stress (2 h/day, 21 consecutive days). Glucocorticoid receptor (GR), TH, and IL-4-expressing cells were localized in identical neurons of the LC, indicating that hypothalamic-pituitary-adrenal- (HPA-) axis and sympathetic-adrenal-medullary- (SAM-) axis might be involved in IL-4 secretion in the stress response. Accordingly, it was concluded that stress-induced decline of IL-4 concentration from LC neurons may be related to anxiety-like behavior and an inverse relationship exists between IL-4 secretion and HPA/SAM-axes activation.

Figure 1

Mean (± SEM) time spent in the closed arms and the open arms during EPM test for the acutely and chronically stressed and control rats. (a) Chronic IMO induced a significant increase in time spent on the closed arms. (b) Acute IMO led animals to stay for significantly less amount of time in the open arms. (n = 6 per group) ^∗ P < 0.05 versus control; ANOVA.

Figure 2

Mean (± SEM) IL-4 protein level in the LC of the acutely and chronically stressed and control rats. Expression of IL-4 was significantly decreased in the acutely stressed group. (n = 6 per group) ^∗ P < 0.05 versus control; ANOVA.

Figure 3

Mean (± SEM) CORT level in the plasma of the control, acute, and chronic IMO-submitted rats. Acute IMO significantly increased plasma CORT whereas chronic IMO induced reduction of CORT release. (n = 8 per group) ^∗ P < 0.05 versus control; ANOVA.

Figure 4

Characterization of IL-4-producing cells in the LC. (a–c) Representative fluorescent images showed that IL-4-secreting cells (a) were colocalized (c) with TH-releasing cells (b). Photomicrographs were taken at ×200 magnification. Scale bars = 50 μm. (d–f) High power (taken at ×400 magnification) photographs of (a), (b), and (c), respectively, are shown. Scale bars = 20 μm. IL-4-releasing cells (g and j) were not merged (i and l) with GFAP-positive cells (h) or Iba-1-immunoreactive cells (k). Photomicrographs were taken at ×400 magnification. Scale bars = 20 μm.

Figure 5

Localization of IL-4 and GR in the LC. (a–c) Representative fluorescent images showed that IL-4 (a) and GR (b) were expressed in the neurons but not in the identical part (c). Photomicrographs were taken at ×200 magnification. Scale bars = 50 μm. (d–f) High power (taken at ×400 magnification) photographs of (a), (b), and (c), respectively, are shown. Scale bars = 20 μm.

Figure 6

Mean (± SEM) quantification of TH expression in the LC of the control group and groups subjected acute and chronic IMO. Acutely stressed rat expressed significantly increase in TH density whereas chronically stressed animals showed no change. (n = 4-5 per group) ^∗∗∗ P < 0.001 versus control; ANOVA.