Hippocampal TNF-α Signaling Mediates Heroin Withdrawal-Enhanced Fear Learning and Withdrawal-Induced Weight Loss

Abstract

There is significant comorbidity of opioid use disorder (OUD) and post-traumatic stress disorder (PTSD) in clinical populations. However, the neurobiological mechanisms underlying the relationship between chronic opioid use and withdrawal and development of PTSD are poorly understood. Our previous work identified that chronic escalating heroin administration and withdrawal can produce enhanced fear learning, an animal model of hyperarousal, and is associated with an increase in dorsal hippocampal (DH) interleukin-1β (IL-1β). However, other cytokines, such as TNF-α, work synergistically with IL-1β and may have a role in the development of enhanced fear learning. Based on both translational rodent and clinical studies, TNF-α has been implicated in hyperarousal states of PTSD, and has an established role in hippocampal-dependent learning and memory. The first set of experiments tested the hypothesis that chronic heroin administration followed by withdrawal is capable of inducing alterations in DH TNF-α expression. The second set of experiments examined whether DH TNF-α expression is functionally relevant to the development of enhanced fear learning. We identified an increase of TNF-α immunoreactivity and positive cells at 0, 24, and 48 h into withdrawal in the dentate gyrus DH subregion. Interestingly, intra-DH infusions of etanercept (TNF-α inhibitor) 0, 24, and 48 h into heroin withdrawal prevented the development of enhanced fear learning and mitigated withdrawal-induced weight loss. Overall, these findings provide insight into the role of TNF-α in opioid withdrawal and the development of anxiety disorders such as PTSD.

MDMA administration attenuates hippocampal IL-β immunoreactivity and subsequent stress-enhanced fear learning: An animal model of PTSD

“…Further, we have also shown a mechanistic role of IL-1β in the behavioral consequences of stress, as blockade of IL-1β attenuates enhanced fear learning, suggesting that IL-1β signaling is necessary for the development of enhanced fear learning ( Jones et al, 2015 ; Jones et al, 2018a ; Parekh et al, 2020 ). Remarkably, we have also demonstrated that TNF-a signaling plays a role in the development of an enhanced fear learning paradigm ( Parekh et al, 2021 ).…”

“…On Days 9, 10, 15, and 22 (Test Days 1, 2, 7, and 14), animals were placed into Context B for 8 min, 23 s and were video recorded to analyze freezing behavior, a measure of learned fear. Automated video tracking and analysis software was used to quantify percent freezing behavior in all sessions (Ethovision XT, Activity Feature with Threshold = 10, Noldus Information Technology Inc., Netherlands) as previously analyzed (; Jones et al, 2018b ; Parekh et al, 2020 ; Parekh et al, 2021 ). Ethovision XT is a video tracking software that tracks and analyzes behavior, movement, and activity of any animal.…”

“…Experiment 2 used fluorescent immunohistochemistry (IHC) to examine alterations in DH IL-1β, TNF-α, GFAP, and IBA-1 in the DG. The IHC protocol used here has been described previously ( Jones et al, 2018a ; Parekh et al, 2020 ; Parekh et al, 2021 ) . The following primary antibodies were used: rabbit anti-IL-1β (1:500, Abcam, Cambridge, MA, Cat# Ab9722, USA), rabbit anti-TNF alpha (1:1000, Abcam, Cambridge, MA, Cat# ab66579), mouse anti-GFAP (1:1000, ThermoFisher Scientific, Waltham, MA, Cat #MS-1376P, USA), and rabbit anti-IBA-1 (1:500, Wako, Richmond, VA, Cat# 019–19741, USA).…”

Chronic ethanol consumption exacerbates future stress‐enhanced fear learning, an effect mediated by dorsal hippocampal astrocytes

“…SEFL was performed after the completion of the chronic EtOH consumption paradigm in Experiment 2 ( n = 24). This protocol has been previously described at length (Jones et al, 2015; Jones, Lebonville, et al, 2018; Jones, Paniccia, et al, 2018; Parekh et al, 2020, 2021; Rau et al, 2005). Briefly, 24 h following the replacement of EtOH Diet with Control Diet, animals were placed into Context A (BRS/LVE, Laurel, MD; H 26.7 cm × D 24.8 cm × 30.7 cm) for 90 min, an environment with distinct olfactory, auditory, and tactile characteristics from the home cage.…”

“…Fluorescent immunohistochemistry was used to verify the specificity of G i ‐DREADD expression in DH astrocytes in Experiment 4. The immunohistochemistry protocol used here has been described previously (Jones, Lebonville, et al, 2018; Jones, Paniccia, et al, 2018; Paniccia et al, 2018; Parekh et al, 2020, 2021). Tissue was incubated with the following primary antibodies: rabbit anti‐GFAP (1:1000, Agilent Technologies, Cat# Z0334), mouse anti‐NeuN (1:1000, Millipore‐Sigma, Cat # MAB377), and the corresponding secondary antibodies: goat anti‐rabbit Alexa Fluor‐405 (1:1000, ThermoFisher Scientific, Cat #A11008), goat anti‐mouse Alexa Fluor‐488 (1:1000, ThermoFisher Scientific, Cat #35501BID).…”