Opioid action was thought to exert reinforcing effects solely via the initial agonism of opioid receptors. Here, we present evidence for an additional novel contributor to opioid reward: the innate ...immune pattern-recognition receptor, toll-like receptor 4 (TLR4), and its MyD88-dependent signaling. Blockade of TLR4/MD2 by administration of the nonopioid, unnatural isomer of naloxone, (+)-naloxone (rats), or two independent genetic knock-outs of MyD88-TLR4-dependent signaling (mice), suppressed opioid-induced conditioned place preference. (+)-Naloxone also reduced opioid (remifentanil) self-administration (rats), another commonly used behavioral measure of drug reward. Moreover, pharmacological blockade of morphine-TLR4/MD2 activity potently reduced morphine-induced elevations of extracellular dopamine in rat nucleus accumbens, a region critical for opioid reinforcement. Importantly, opioid-TLR4 actions are not a unidirectional influence on opioid pharmacodynamics, since TLR4(-/-) mice had reduced oxycodone-induced p38 and JNK phosphorylation, while displaying potentiated analgesia. Similar to our recent reports of morphine-TLR4/MD2 binding, here we provide a combination of in silico and biophysical data to support (+)-naloxone and remifentanil binding to TLR4/MD2. Collectively, these data indicate that the actions of opioids at classical opioid receptors, together with their newly identified TLR4/MD2 actions, affect the mesolimbic dopamine system that amplifies opioid-induced elevations in extracellular dopamine levels, therefore possibly explaining altered opioid reward behaviors. Thus, the discovery of TLR4/MD2 recognition of opioids as foreign xenobiotic substances adds to the existing hypothesized neuronal reinforcement mechanisms, identifies a new drug target in TLR4/MD2 for the treatment of addictions, and provides further evidence supporting a role for central proinflammatory immune signaling in drug reward.
Aim
To identify objective factors that can predict future sensitized stress responses, thus allowing for effective intervention prior to developing sensitization and subsequent stress‐related ...disorders, including post‐traumatic stress disorder (PTSD).
Methods
Adult male F344 rats implanted with biotelemetry devices were exposed to repeated conditioned fear or control conditions for 22 days followed by exposure to either no, mild or severe acute stress on day 23. Diurnal rhythms of locomotor activity (LA), heart rate (HR) and core body temperature (CBT) were biotelemetrically monitored throughout the study. In a subset of rat not implanted, corticosterone and indices of chronic stress were measured immediately following stress.
Results
Rats exposed to repeated fear had fear‐evoked increases in behavioural freezing and HR/CBT during exposure to the fear environment and displayed indices of chronic stress. Repeated fear produced flattening of diurnal rhythms in LA, HR and CBT. Repeated fear did not sensitize the corticosterone response to acute stress, but produced sensitized HR/CBT responses following acute stress, relative to the effect of acute stress in the absence of a history of repeated fear. Greater diurnal rhythm disruptions during repeated fear predicted sensitized acute stress‐induced physiological responses. Rats exposed to repeated fear also displayed flattened diurnal LA and basal increases in HR.
Conclusions
Exposure to repeated fear produces outcomes consistent with those observed in PTSD. The results suggest that diurnal rhythm disruptions during chronic stressors may help predict sensitized physiological stress responses following traumatic events. Monitoring diurnal disruptions during repeated stress may thus help predict susceptibility to PTSD.
One component of the adaptive stress response is that innate immunity is primed by circulating endogenous danger associated molecular patterns (DAMPs). Extracellular heat shock protein 72 (eHsp72) is ...a DAMP that is upregulated intracellularly after acute stress, but its mechanism of release is unknown. Previous in vitro studies have demonstrated the presence of Hsp72 on extracellular vesicles known as exosomes, which are biologically active vesicles secreted from all known cells in the body. We hypothesize that stress-evoked eHsp72 is released into the blood via an exosome pathway and that these exosomes are a critical component of stress-enhanced immunity. Male Fisher 344 rats (8/group) were subjected to tail shock stress or no stress, and exosome isolation from plasma was verified by size (EM and Nanosight) and common exosome markers (acetylcholinesterase, CD63, and Rab5b). We report that exposure to an acute stressor increases exosome expression of eHsp72, but not other stress-inducible proteins (IL-1β and IL-6). Additionally, exosomes from stressed, but not control, rats facilitates in vivo bactericidal inflammatory response ( p < 0.05) and an in vitro LPS-evoked inflammatory responses ( p < 0.05). These data suggest that exposure to stress can alter the proteomic composition of circulating exosomes, thereby enhancing the innate immune response.
Acute stressor exposure induces a sterile inflammatory response marked by shifts in circulating and white adipose tissue (WAT) cytokines. Although the sterile inflammatory response occurs in ...healthy/non-obese WAT and varies in magnitude relative to the WAT depot, the mechanisms which regulate stress-induced WAT cytokines remain unclear. Because the gut microbiota modulates visceral WAT inflammation and stress-induced systemic cytokines, we hypothesized that changes to the normal microbiota environment would also affect stress-induced WAT cytokines. To test this hypothesis 8 week old male Fischer 344 rats were treated for four days with antibiotics (4 mg/mL streptomycin and 2 mg/mL penicillin G ) administered via drinking water prior to receiving 100 tail-shocks (1.5 mA, 5s, every 60 s ± 25 s). Immediately before or following stress rats were sacrificed and subcutaneous, retroperitoneal, epididymal, omental, and mesenteric WAT were harvested for cytokine analyses via ELISA. Stress differentially increased interleukin-1beta (IL-1beta) in all WAT depots, decreased interleukin-10 (IL-10) in subcutaneous and mesenteric WAT and tumor necrosis factor-alpha in omental WAT. Antibiotic treatment reduced fecal colony forming units, dampened the stress-induced increase in IL-1beta in subcutaneous and retroperitoneal WAT and the stress-induced decrease in IL-10 in subcutaneous and mesenteric WAT. These data demonstrate that the gut microbiota directly affects stress-induced shifts in WAT cytokine concentrations in a depot specific manner suggesting a novel mechanism through which pro-inflammatory cytokine concentrations are regulated in healthy, stressed non-obese WAT. NSF-1022451.
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