Gilly Wolf

Interleukin-1 antagonizes morphine analgesia and underlies morphine tolerance.

Pain sensitivity reflects a balance between pain facilitatory and inhibitory systems. To characterize the relationships between these systems we examined the interactions between the analgesic effects of morphine and the anti‐analgesic effects of the pro‐inflammatory cytokine interleukin‐1 (IL‐1). We report that administration of a neutral dose of IL‐1β abolished morphine analgesia in mice, whereas acute or chronic blockade of IL‐1 signaling by various IL‐1 blockers (IL‐1 receptor antagonist (IL‐1ra), α‐melanocyte‐stimulating hormone, or IL‐1 tri‐peptide antagonist) significantly prolonged and potentiated morphine analgesia. Morphine‐induced analgesia was also extended in strains of mice genetically impaired in IL‐1 signaling (mice with transgenic over‐expression of IL‐1 receptor antagonist, deletion of the IL‐1 receptor type I, or deletion of the IL‐1 receptor accessory protein). The finding that IL‐1 produces a marked anti‐analgesic effect, suggests that it may also be involved in the development of opiate tolerance. Indeed, genetic or pharmacological blockade of IL‐1 signaling prevented the development of tolerance following repeated morphine administration. Moreover, acute administration of IL‐1ra in wild type mice, either immediately following the cessation of acute morphine analgesia, or following the development of chronic morphine tolerance, re‐instated the analgesia, suggesting that blockade of the IL‐1 system unmasks the analgesic effect of morphine. These findings suggest that morphine produces an IL‐1‐mediated homeostatic response, which serves to limit the duration and extent of morphine analgesia and which underlies the development of tolerance.

Genetic impairment of interleukin-1 signaling attenuates neuropathic pain, autotomy, and spontaneous ectopic neuronal activity, following nerve injury in mice.

Abstract Peripheral nerve injury may lead to neuropathic pain, which is often associated with mechanical and thermal allodynia, ectopic discharge of from injured nerves and from the dorsal root ganglion neurons, and elevated levels of proinflammatory cytokines, particularly interleukin‐1 (IL‐1). In the present study, we tested the role of IL‐1 in neuropathic pain models using two mouse strains impaired in IL‐1 signaling: Deletion of the IL‐1 receptor type I (IL‐1rKO) and transgenic over‐expression of the IL‐1 receptor antagonist (IL‐1raTG). Neuropathy was induced by cutting the L5 spinal nerve on one side, following which mechanical and thermal pain sensitivity was measured. Wild‐type (WT) mice and the parent strains developed significant allodynia and hyperalgesia in the hind‐paw ipsilateral to the injury compared with the contralateral hind‐paw. The mutant strains, however, did not display decreased pain threshold in either hind‐paw. Pain behavior was also assessed by cutting the sciatic and saphenous nerves and measuring autotomy scores. WT mice developed progressive autotomy, beginning at 7 days post‐injury, whereas the mutant strains displayed delayed onset of autotomy and markedly reduced severity of the autotomy score. Electrophysiological assessment revealed that in WT mice a significant proportion of the dorsal root axons exhibited spontaneous ectopic activity at 1, 3, and 7 days following spinal nerve injury, whereas in IL‐1rKO and IL‐1raTG mice only a minimal number of axons exhibited such activity. Taken together, these results suggest that IL‐1 signaling plays an important role in neuropathic pain and in the altered neuronal activity that underlies its development.

Impairment of interleukin-1 (IL-1) signaling reduces basal pain sensitivity in mice: genetic, pharmacological and developmental aspects

The cytokine interleukin‐1 (IL‐1) has been implicated in modulation of pain perception under various inflammatory conditions. The present study examined the hypothesis that IL‐1 signaling is also involved in pain sensitivity under normal, non‐inflammatory states, using three mouse models of impaired IL‐1 signaling: targeted deletion of the IL‐1 receptor type I or the IL‐1 receptor accessory protein, and transgenic over‐expression of IL‐1 receptor antagonist within the brain and spinal cord. Thermal and mechanical pain sensitivity was assessed using the paw‐flick, hot‐plate, and von Frey tests. All mutant strains displayed significantly lower pain sensitivity, compared with their respective wild‐type control strains, and with their parent strains (C57BL/6, CBA and 129), in all tests. In contrast, mice with targeted deletion of the p55 or p75 TNF receptor, or of interleukin‐18, displayed normal or higher pain sensitivity compared to their respective controls. To differentiate between developmental vs. on‐going effects of IL‐1, mice were chronically treated with IL‐1 receptor antagonist (IL‐1ra) via osmotic micropumps, either in adulthood or prenatally (throughout the last 2 weeks of gestation). Adult mice that were treated with IL‐1ra either in adulthood or in utero, displayed lower pain sensitivity, similar to mice with impaired IL‐1 signaling. These findings suggest that basal pain sensitivity is genetically, developmentally and tonically influenced by IL‐1 signaling.

IL-1β signaling is required for mechanical allodynia induced by nerve injury and for the ensuing reduction in spinal cord neuronal GRK2

Many neurotransmitters involved in pain perception transmit signals via G protein-coupled receptors (GPCRs). GPCR kinase 2 (GRK2) regulates agonist-induced desensitization and signaling of multiple GPCRs and interacts with downstream molecules with consequences for signaling. In general, low GRK2 levels are associated with increased responses to agonist stimulation of GPCRs. Recently, we reported that in mice with reduced GRK2 levels, inflammation-induced mechanical allodynia was increased. In addition, mice with impaired interleukin (IL)-1 beta signaling did not develop mechanical allodynia after L5 spinal nerve transection (SNT). We hypothesized that in the L5 SNT model mechanical allodynia would be associated with reduced neuronal GRK2 levels in the spinal cord dorsal horn and that IL-1 beta signaling would be required to induce both the decrease in GRK2 and mechanical allodynia. We show here that in wild type (WT) mice L5 SNT induces a bilateral decrease in neuronal GRK2 expression in the lumbar spinal cord dorsal horn, 1 and 2 weeks after L5 SNT. No changes in GRK2 were observed in the thoracic segments. Moreover, spinal cord GRK2 expression was not decreased in IL-1R(-/-) mice after L5 SNT. These data show that IL-1 beta signaling is not only required for the development of mechanical allodynia, but also to reduce neuronal GRK2 expression. These results suggest a functional relation between the L5 SNT-induced IL-1 beta-mediated decrease in GRK2 and development of mechanical allodynia.

Interleukin-1 signaling is required for induction and maintenance of postoperative incisional pain: genetic and pharmacological studies in mice.

Postoperative incisional pain is characterized by persistent acute pain in the area of the cut, and is associated with release of proinflammatory cytokines, including interleukin-1 (IL-1), which play important hyperalgesic and allodynic roles in various inflammatory conditions. In the present study, we tested the role of IL-1 signaling in postoperative incisional pain using three mouse strains impaired in IL-1 signaling due to deletion of the IL-1 type I receptor on a mixed genetic background (IL-1rKO) or congenic background (IL-1rKOCog), or due to transgenic over-expression of IL-1 receptor antagonist (IL-1raTG). We used the relevant wild-type (WT) mice both as controls for the mutant strains, and for assessing the effects of pharmacological blockade of IL-1-signaling. Mechanosensitivity was assessed using the von-Frey filament test before, and up to 4 days following plantar incision, an animal model of postoperative pain. WT mice developed significant allodynia in the incised, compared with the intact, hind-paw beginning 3h after the incision and lasting up to 48h postoperatively. In contrast, IL-1rKO, IL-1rKOCog, and IL-1raTG mice, as well as WT mice chronically treated with IL-1ra, did not display increased mechanical pain sensitivity in either hind-paw. To test the hypothesis that IL-1-signaling is also involved in the maintenance of postoperative pain, WT mice were acutely treated with IL-1ra 24h following the incision, when allodynia was already evident. This treatment reversed the allodynic response throughout the observation period. Together, these findings suggest that IL-1 plays a critical role in the development and maintenance of postoperative incisional pain.

Effects of surgical stress on brain prostaglandin E2 production and on the pituitary–adrenal axis: Attenuation by preemptive analgesia and by central amygdala lesion

Surgical stress is the combined result of tissue injury, anesthesia, and postoperative pain. It is characterized by elevated levels of adrenocorticotropin (ACTH), corticosterone (CS), and elevated levels of prostaglandin E2 (PGE2) in the periphery and in the spinal cord. The present study examined the effects of perioperative pain management in rats undergoing laparotomy on serum levels of ACTH, CS, and on the production of PGE2 in several brain regions, including the amygdala. The amygdala is known to modulate the pituitary-adrenal axis response to stress. We, therefore, also examined the effects of bilateral lesions in the central amygdala (CeA) on laparotomy-induced activation of the pituitary-adrenal axis in rats. In the first experiment, rats either underwent laparotomy or were not operated upon. Half the rats received preemptive analgesia extended postoperatively, the other received saline. ACTH, CS serum levels, and ex vivo brain production of PGE2 were determined. In the second experiment, rats underwent bilateral lesions of the CeA. Ten days later, rats underwent laparotomy, and ACTH and CS serum levels were determined. Laparotomy significantly increased amygdala PGE2 production, and CS and ACTH serum levels. This elevation was markedly attenuated by perioperative analgesia. Bilateral CeA lesions also attenuated the pituitary-adrenal response to surgical stress. The present findings suggest that the amygdala plays a regulatory role in mediating the neuroendocrine response to surgical stress. Effective perioperative analgesia attenuated the surgery-induced activation of pituitary-adrenal axis and PGE2 elevation. The diminished elevation of PGE2 may suggest a mechanism by which pain relief mitigates pituitary-adrenal axis activation.

The Effects of Perioperative Pain Management Techniques on Food Consumption and Body Weight After Laparotomy in Rats

We examined the effects of two perioperative pain management techniques on recovery after laparotomy, as assessed by body weight (BW) and food consumption (FC). All rats received a preoperative intrathecal mixture of morphine plus bupivacaine combined with one of two treatments: (a) injection of slow-release morphine at the end of the surgery or (b) an antiinflammatory drug, interleukin-1 receptor antagonist (IL-1ra), combined with the preoperative mixture. Laparotomy significantly decreased FC and BW. Both analgesic treatments resulted in a faster recovery of FC and BW. This beneficial effect was more pronounced in the group receiving preoperative analgesics combined with IL-1ra.

Effects of prenatal morphine exposure on NK cytotoxicity and responsiveness to LPS in rats

Prenatal exposure to opiates can adversely affect fetal development, resulting in long-term growth retardation and impairments in physiological and behavioral functions. In the present study we studied long-term effects of prenatal morphine exposure on immune functions, including the activity of natural killer (NK) cells and the febrile and behavioral responses to lipopolysaccharide (LPS). Pregnant Fischer 344 rats were given increasing doses of morphine in slow release emulsion during gestational days 12-18. Control rats were injected with vehicle and were either pair fed to morphine rats or fed ad lib. Postnatal experiments were conducted when offspring were 10-12 weeks old. Compared to both control groups, rats prenatally exposed to morphine exhibited: 1) suppressed cytotoxic activity of NK cells; 2) reduced LPS-induced fever measured by a biotelemetric system; 3) reduced hyperalgesia measured by the hot-plate test at 30 min, and augmented hypoalgesia at 2-6 h post-LPS; 4) higher open-field activity in saline-treated animals, and more pronounced suppression of activity in LPS-injected animals; 5) LPS-induced reduction of food consumption, body weight, and social exploration, which did not differ from the reduction observed in control animals. These findings indicate that prenatal exposure to morphine induces long-term impairment of host-defense mechanisms, which may render the offspring more susceptible to infectious diseases.

Differentially Severe Cognitive Effects of Compromised Cerebral Blood Flow in Aged Mice: Association with Myelin Degradation and Microglia Activation

Bilateral common carotid artery stenosis (BCAS) models the effects of compromised cerebral blood flow on brain structure and function in mice. We compared the effects of BCAS in aged (21 month) and young adult (3 month) female mice, anticipating a differentially more severe effect in the older mice. Four weeks after surgery there was a significant age by time by treatment interaction on the radial-arm water maze (RAWM; p = 0.014): on the first day of the test, latencies of old mice were longer compared to the latencies of young adult mice, independent of BCAS. However, on the second day of the test, latencies of old BCAS mice were significantly longer than old control mice (p = 0.049), while latencies of old controls were similar to those of the young adult mice, indicating more severe impairment of hippocampal dependent learning and working memory by BCAS in the older mice. Fluorescence staining of myelin basic protein (MBP) showed that old age and BCAS both induced a significant decrease in fluorescence intensity. Evaluation of the number oligodendrocyte precursor cells demonstrated augmented myelin replacement in old BCAS mice (p < 0.05) compared with young adult BCAS and old control mice. While microglia morphology was assessed as normal in young adult control and young adult BCAS mice, microglia of old BCAS mice exhibited striking activation in the area of degraded myelin compared to young adult BCAS (p < 0.01) and old control mice (p < 0.05). These findings show a differentially more severe effect of cerebral hypoperfusion on cognitive function, myelin integrity and inflammatory processes in aged mice. Hypoperfusion may exacerbate degradation initiated by aging, which may induce more severe neuronal and cognitive phenotypes.

Effect of chronic unpredictable stress on mice with developmental under-expression of the Ahi1 gene: behavioral manifestations and neurobiological correlates

The Abelson helper integration site 1 (Ahi1) gene plays a pivotal role in brain development and is associated with genetic susceptibility to schizophrenia, and other neuropsychiatric disorders. Translational research in genetically modified mice may reveal the neurobiological mechanisms of such associations. Previous studies of mice heterozygous for Ahi1 knockout (Ahi1+/−) revealed an attenuated anxiety response on various relevant paradigms, in the context of a normal glucocorticoid response to caffeine and pentylenetetrazole. Resting-state fMRI showed decreased amygdalar connectivity with various limbic brain regions and altered network topology. However, it was not clear from previous studies whether stress-hyporesponsiveness reflected resilience or, conversely, a cognitive-emotional deficit. The present studies were designed to investigate the response of Ahi1+/− mice to chronic unpredictable stress (CUS) applied over 9 weeks. Wild type (Ahi1+/+) mice were significantly affected by CUS, manifesting decreased sucrose preference (p < 0.05); reduced anxiety on the elevated plus maze and light dark box and decreased thigmotaxis in the open field (p < 0.01 0.05); decreased hyperthermic response to acute stress (p < 0.05); attenuated contextual fear conditioning (p < 0.01) and increased neurogenesis (p < 0.05). In contrast, Ahi1+/− mice were indifferent to the effects of CUS assessed with the same parameters. Our findings suggest that Ahi1 under-expression during neurodevelopment, as manifested by Ahi1+/− mice, renders these mice stress hyporesponsive. Ahi1 deficiency during development may attenuate the perception and/or integration of environmental stressors as a result of impaired corticolimbic connectivity or aberrant functional wiring. These neural mechanisms may provide initial clues as to the role Ahi1 in schizophrenia and other neuropsychiatric disorders.