Beta adrenergic blockade decreases the immunomodulatory effects of social disruption stress

Abstract

During physiological or psychological stress, catecholamines produced by the sympathetic nervous system (SNS) regulate the immune system. Previous studies report that the activation of β-adrenergic receptors (βARs) mediates the actions of catecholamines and increases pro-inflammatory cytokine production in a number of different cell types. The impact of the SNS on the immune modulation of social defeat has not been examined. The following studies were designed to determine whether SNS activation during social disruption stress (SDR) influences anxiety-like behavior as well as the activation, priming, and glucocorticoid resistance of splenocytes after social stress. CD-1 mice were exposed to one, three, or six cycles of SDR and HPLC analysis of the plasma and spleen revealed an increase in catecholamines. After six cycles of SDR the open field test was used to measure behaviors characteristic of anxiety and indicated that the social defeat induced increase in anxiety-like behavior was blocked by pre-treatment with the β-adrenergic antagonist propranolol. Pre-treatment with the β-adrenergic antagonist propranolol did not significantly alter corticosterone levels indicating no difference in activation of the hypothalamic-pituitary-adrenal axis. In addition to anxiety-like behavior the SDR induced splenomegaly and increase in plasma IL-6, TNFα, and MCP-1 were each reversed by pre-treatment with propranolol. Furthermore, flow cytometric analysis of cells from propranolol pretreated mice reduced the SDR-induced increase in the percentage of CD11b(+) splenic macrophages and significantly decreased the expression of TLR2, TLR4, and CD86 on the surface of these cells. In addition, supernatants from 18h LPS-stimulated ex vivo cultures of splenocytes from propranolol-treated SDR mice contained less IL-6. Likewise propranolol pre-treatment abrogated the glucocorticoid insensitivity of CD11b(+) cells ex vivo when compared to splenocytes from SDR vehicle-treated mice. Together, this study demonstrates that the immune activation and priming effects of SDR result, in part, as a consequence of SNS activation.

Copyright © 2012 Elsevier Inc. All rights reserved.

Figures

Fig. 1

Repeated social defeat significantly increased plasma and splenic catecholamines. Separate groups of mice were subjected to one, three or six cycles of repeated social defeat (SDR; filled bars) or left undisturbed in their home cage (HCC; open bars). Immediately following the first, third or sixth stress cycle plasma and spleens were collected to determine catecholamine levels. Mice subjected to SDR had increased levels of (A) plasma norepinephrine and (B) plasma epinephrine as well as (C) increased splenic norepinephrine regardless of the number of SDR cycles experienced. In a follow up study separate groups of mice were subjected to six cycles of SDR in order to collect plasma and spleens at 20 min, 3, 12 or 24 h post termination of the stressor in order to assess catecholamine levels. (D and E) Plasma norepinephrine and epinephrine were elevated 20 min following termination of the stressor but returned to control levels at later time points. (F) Defeated mice had elevated splenic norepinephrine except at 3 h post termination of the stressor. Bars represent the mean ± SEM (n = 6). Means with different letters (a or b) are significantly different (p < 0.05) from each other. (tw = total tissue weight).

Fig. 2

β-adrenergic antagonism reduced anxiety-like behavior in socially defeated mice. CD-1 male mice were subjected to six days of repeated social defeat (SDR; filled bars) or left undisturbed in their home cage (HCC; open bars) after a s.c. injection of vehicle or propranolol prior to each cycle of defeat. Mice were assessed in the open field test the day after the last stress cycle. (A) Defeated mice spent less time in the center of the open field, propranolol prevents this behavior. (B) Defeated mice took longer to enter the center of the field than controls, propranolol blocks this behavior. (C) Distance traveled and (D) Time spent moving were not significantly different between any groups. Bars represent the mean ± SEM (n = 9). Means with different letters (a or b) are significantly different (p < 0.05) from each other.

Fig. 3

β-adrenergic antagonism treatment of socially defeated mice significantly altered spleen weight but not serum corticosterone. Male CD-1 mice were injected s.c. with vehicle or propranolol prior to defeat each day for 6 days. Corticosterone was measured via radioimmune assay from defeated (SDR; filled bars) and home cage control (HCC; open bars) mice in the serum obtained via retro orbital plexus (A) immediately following the last cycle of SDR (i.e., at 1900 EST) and at 18 h after the last cycle of SDR (i.e. at 0900 EST). (C) Spleen and body weight were measured the day after the last cycle of defeat and a relative spleen mass (spleen mass/body mass) was calculated. Bars represent the mean ± SEM (n = 9). Means with different letters (a or b) are significantly different (p < 0.05) from each other.

Fig. 4

β-adrenergic antagonism treatment of socially defeated mice significantly alters cell activation. Male CD-1 mice were injected s.c. with vehicle or propranolol prior to defeat each day for 6 days. (A) The day after the last cycle of defeat there was a significant increase in CD11b+ cells within the spleen of socially defeat (SDR; filled bars) mice compared to home cage control (HCC; open bars) mice as assessed via flow cytometry. (B, C, and D) The expression of Toll-like receptor (TLRs) 2 and 4 (TLR2 and TLR4) and CD86 on splenic CD11b+ monocytes/macrophages was increased as assessed via flow cytometry using fluorescent anti-TLR and anti-CD86 antibody staining. Bars represent the mean ± SEM (n = 9). Means with different letters (a or b) are significantly different (p < 0.05) from each other.

Fig. 5

β-adrenergic antagonism treatment prevented stress induced cytokines after social defeat in plasma. Male CD-1 mice were subjected to six cycles of repeated social defeat (SDR; filled bars) or left undisturbed in their home cage (HCC; open bars). Mice were injected s.c. with vehicle or propranolol prior to each cycle of defeat. The day after the last cycle of defeat (A) IL-6, (B) MCP-1, and (C) TNF-α protein levels were determined in plasma. Bars represent the mean ± SEM (n = 9). Means with different letters (a or b) are significantly different (p < 0.05) from each other.

Fig. 6

β-adrenergic antagonism prevented social defeat induced IL-6 production and attenuated social defeat-induced glucocorticoid insensitivity. Spleen cells were harvested from socially defeated (SDR) or home cage control (HCC) mice treated with vehicle or propranolol, stimulated with LPS, and 18 h later IL-6 was quantified from collected supernatants by ELISA. 48 h later GC insensitivity, defined as high cell viability in the presence of increased concentrations of corticosterone, was determined. SDR mice (closed square) demonstrate GC insensitivity compared to control groups (open square & circle). Cell viability was decreased in socially defeated mice treated with propranolol (closed circle). Lines represent the mean ± SEM (n = 9). *p < 0.05 vs. HCC vehicle treated mice; **p < 0.05 vs. SDR vehicle treated mice.