Kevin A. O'Connor

Spinal Glia and Proinflammatory Cytokines Mediate Mirror-Image Neuropathic Pain in Rats

Mirror-image allodynia is a mysterious phenomenon that occurs in association with many clinical pain syndromes. Allodynia refers to pain in response to light touch/pressure stimuli, which normally are perceived as innocuous. Mirror-image allodynia arises from the healthy body region contralateral to the actual site of trauma/inflammation. Virtually nothing is known about the mechanisms underlying such pain. A recently developed animal model of inflammatory neuropathy reliably produces mirror-image allodynia, thus allowing this pain phenomenon to be analyzed. In this sciatic inflammatory neuropathy (SIN) model, decreased response threshold to tactile stimuli (mechanical allodynia) develops in rats after microinjection of immune activators around one healthy sciatic nerve at mid-thigh level. Low level immune activation produces unilateral allodynia ipsilateral to the site of sciatic inflammation; more intense immune activation produces bilateral (ipsilateral + mirror image) allodynia. The present studies demonstrate that both ipsilateral and mirror-image SIN-induced allodynias are (1) reversed by intrathecal (peri-spinal) delivery of fluorocitrate, a glial metabolic inhibitor; (2) prevented and reversed by intrathecal CNI-1493, an inhibitor of p38 mitogen-activated kinases implicated in proinflammatory cytokine production and signaling; and (3) prevented or reversed by intrathecal proinflammatory cytokine antagonists specific for interleukin-1, tumor necrosis factor, or interleukin-6. Reversal of ipsilateral and mirror-image allodynias was rapid and complete even when SIN was maintained constantly for 2 weeks before proinflammatory cytokine antagonist administration. These results provide the first evidence that ipsilateral and mirror-image inflammatory neuropathy pain are created both acutely and chronically through glial and proinflammatory cytokine actions.

Evidence that exogenous and endogenous fractalkine can induce spinal nociceptive facilitation in rats

Recent evidence suggests that spinal cord glia can contribute to enhanced nociceptive responses. However, the signals that cause glial activation are unknown. Fractalkine (CX3C ligand‐1; CX3CL1) is a unique chemokine expressed on the extracellular surface of spinal neurons and spinal sensory afferents. In the dorsal spinal cord, fractalkine receptors are primarily expressed by microglia. As fractalkine can be released from neurons upon strong activation, it has previously been suggested to be a neuron‐to‐glial signal that induces glial activation. The present series of experiments provide an initial investigation of the spinal pain modulatory effects of fractalkine. Intrathecal fractalkine produced dose‐dependent mechanical allodynia and thermal hyperalgesia. In addition, a single injection of fractalkine receptor antagonist (neutralizing antibody against rat CX3C receptor‐1; CX3CR1) delayed the development of mechanical allodynia and/or thermal hyperalgesia in two neuropathic pain models: chronic constriction injury (CCI) and sciatic inflammatory neuropathy. Intriguingly, anti‐CX3CR1 reduced nociceptive responses when administered 5–7u2003days after CCI, suggesting that prolonged release of fractalkine may contribute to the maintenance of neuropathic pain. Taken together, these initial investigations of spinal fractalkine effects suggest that exogenous and endogenous fractalkine are involved in spinal sensitization, including that induced by peripheral neuropathy.

Prior Stressor Exposure Sensitizes LPS-Induced Cytokine Production

Exposure to stressors often alters the subsequent responsiveness of many systems. The present study tested whether prior exposure to inescapable tailshock (IS) alters the interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, or IL-6 response to an injection of bacterial endotoxin (lipopolysaccharide; LPS). Rats were exposed to IS or remained as home cage controls (HCC); 24 h later animals were injected i.p. with either 10 microg/kg LPS or equilvolume sterile saline. IS significantly increased plasma TNF-alpha, IL-1beta, and pituitary, hypothalamus, hippocampus, cerebellum IL-1beta 1 h, but not 2 h, after LPS, compared to controls. Additional animals were injected with LPS or saline 4, 10, or 21 days after exposure to IS and tail vein blood was collected and assayed for IL-1beta. An enhanced plasma IL-1beta response occurred 4 days after IS, but was gone by 10 days. These results suggest that exposure to IS sensitizes the innate immune response to LPS by resulting in either a larger or a more rapid induction of proinflammatory cytokines.

Peripheral and central proinflammatory cytokine response to a severe acute stressor.

The role of proinflammatory cytokines in the response to acute stressor exposure has received recent attention. Exposure to a single session of inescapable shock (IS) induces peripheral and central proinflammatory cytokines. Other stressors also increase expression of proinflammatory cytokine mRNA and/or protein in various tissues. However, the induction of central and peripheral proinflammatory cytokines by stressors remains controversial and the pattern of cytokine induction is not consistent across stressors. The present experiments sought to examine the pattern of the proinflammatory cytokine response to a stressor known to cause elevations of IL-1beta protein. mRNA expression for three proinflammatory cytokines, IL-1beta, TNF-alpha and IL-6, and IL-1beta protein was examined after IS. IS increases IL-1beta mRNA and/or protein in a variety of tissues, including hypothalamus, hippocampus, pituitary and spleen. Furthermore, IS concomitantly alters IL-1beta mRNA and protein in hypothalamus and spleen, while the IL-1beta mRNA increase in pituitary lags behind the increase of IL-1beta protein. Interestingly, IL-1beta mRNA is elevated in hippocampus 4 h after IS, but an increase of IL-1beta protein in hippocampus is not detected. Expression of TNF-alpha and IL-6 mRNA do not increase in response to IS. Indeed, TNF-alpha mRNA expression decreases in cortex, pituitary and liver immediately after IS. These findings suggest that alterations of proinflammatory cytokine expression by stressors, and IS in particular, are region- and cytokine-specific.

Controlling pathological pain by adenovirally driven spinal production of the anti-inflammatory cytokine, interleukin-10.

Gene therapy for the control of pain has, to date, targeted neurons. However, recent evidence supports that spinal cord glia are critical to the creation and maintenance of pain facilitation through the release of proinflammatory cytokines. Because of the ability of interleukin‐10 (IL‐10) to suppress proinflammatory cytokines, we tested whether an adenoviral vector encoding human IL‐10 (AD‐h‐IL10) would block and reverse pain facilitation. Three pain models were examined, all of which are mediated by spinal pro‐inflammatory cytokines. Acute intrathecal administration of rat IL‐10 protein itself briefly reversed chronic constriction injury‐induced mechanical allodynia and thermal hyperalgesia. The transient reversal caused by IL‐10 protein paralleled the half‐life of human IL‐10 protein in the intrathecal space (t1/2u2003∼u20032u2003h). IL‐10 gene therapy both prevented and reversed thermal hyperalgesia and mechanical allodynia, without affecting basal responses to thermal or mechanical stimuli. Extra‐territorial, as well as territorial, pain changes were reversed by this treatment. Intrathecal AD‐h‐IL10 injected over lumbosacral spinal cord led to elevated lumbosacral cerebrospinal fluid (CSF) levels of human IL‐10, with far less human IL‐10 observed in cervical CSF. In keeping with IL‐10s known anti‐inflammatory actions, AD‐h‐IL10 lowered CSF levels of IL‐1, relative to control AD. These studies support that this gene therapy approach provides an alternative to neuronally focused drug and gene therapies for clinical pain control.

Prior stressor exposure primes the HPA axis

Exposure to stressors often alters the subsequent responsiveness of many systems. The present study tested whether prior exposure to inescapable tailshock (IS) alters the corticosterone (CORT) or adrenocorticotropin hormone (ACTH) response to either an injection of bacterial endotoxin (lipopolysaccharide; LPS) or subsequent placement on a pedestal. Rats were exposed to IS or remained as home cage controls (HCC). 1, 4, 10, or 21 days later animals were injected i.p. with either 10 microg/kg LPS or equivolume sterile saline. Prior IS significantly increased plasma CORT 1 h, but not 2 or 5 h after LPS, compared to controls 1, 4, and 10 days, but not 21 days after IS. Exposure to IS 24 h earlier also significantly increased plasma ACTH 1 h after LPS. Additional animals were placed on a pedestal 24 h after IS, and plasma CORT was measured 15, 30, and 60 min later. IS significantly increased plasma CORT 15 min after pedestal exposure, but not after 30 or 60 min. These results suggest that exposure to IS sensitizes the CORT and ACTH response to subsequent HPA activation.

The role of IL-1β in stress-induced sensitization of proinflammatory cytokine and corticosterone responses

Proinflammatory cytokines often sensitize neuronal, hormonal, and behavioral responses to subsequent challenge. Recently, it was observed that exposure to inescapable tailshock enhances peripheral and central proinflammatory cytokine and corticosterone (CORT) responses to subsequent immune challenge up to 4 days later. Thus, we examined the role of central interleukin-1beta (IL-1beta) in stress-induced sensitization of proinflammatory cytokine and CORT responses to a subsequent immune challenge. Rats were administered IL-1 receptor antagonist (IL-1ra) or vehicle into the intra-cisterna magna 1 h prior to tailshock (100, 1.6 mA 5 s shocks) exposure. Twenty-four hours later, rats were challenged i.p. with 10 microg/kg lipopolysaccharide (LPS) and killed 1 h later. IL-1ra had no effect on basal proinflammatory cytokines, but completely blocked the stress-induced enhancement in central and pituitary IL-1beta and plasma IL-6 release following LPS challenge. IL-1ra had no effect on stress-induced enhancement in CORT responses following LPS challenge. Additional rats were administered i.c.v. hrIL-1beta or vehicle and returned to their home cage. Twenty-four hours later, rats were challenged i.p. with either saline or 10 microg/kg LPS and killed 1 h later. Central hrIL-1beta administration significantly elevated central IL-1beta levels and plasma CORT following LPS challenge compared with vehicle-injected controls. These data demonstrate that elevations in central IL-1beta, whether stress-induced or exogenously administered, are sufficient for sensitizing central IL-1beta and CORT responses to subsequent immune challenge. However, during times of stress, exogenous central IL-1ra administration only blocked sensitization of subsequent central IL-1beta responses, not CORT responses, suggesting other factors during the stress response can sensitize CORT responses.

HIV-1 gp120 Stimulates proinflammatory cytokine-mediated pain facilitation via activation of nitric oxide synthase-I (nNOS)

&NA; It has become clear that spinal cord glia (microglia and astrocytes) importantly contribute to the creation of exaggerated pain responses. One model used to study this is peri‐spinal (intrathecal, i.t.) administration of gp120, an envelope protein of HIV‐1 known to activate glia. Previous studies demonstrated that i.t. gp120 produces pain facilitation via the release of glial proinflammatory cytokines. The present series of studies tested whether spinal nitric oxide (NO) contributes to i.t. gp120‐induced mechanical allodynia and, if so, what effect NO has on spinal proinflammatory cytokines. gp120 stimulation of acutely isolated lumbar dorsal spinal cords released NO as well as proinflammatory cytokines (tumor necrosis factor‐alpha, interleukin‐1beta (IL1), interleukin‐6 (IL6)), thus identifying NO as a candidate mediator of gp120‐induced behavioral effects. Behaviorally, identical effects were observed when gp120‐induced mechanical allodynia was challenged by i.t. pre‐treatment with either a broad‐spectrum nitric oxide synthase (NOS) inhibitor (L‐NAME) or 7‐NINA, a selective inhibitor of NOS type‐I (nNOS). Both abolished gp120‐induced mechanical allodynia. While the literature pre‐dominantly documents that proinflammatory cytokines stimulate the production of NO rather than the reverse, here we show that gp120‐induced NO increases proinflammatory cytokine mRNA levels (RT‐PCR) and both protein expression and protein release (serial ELISA). Furthermore, gp120 increases mRNA for IL1 converting enzyme and matrix metalloproteinase‐9, enzymes responsible for activation and release of proinflammatory cytokines.

A new function of the leptin receptor: mediation of the recovery from lipopolysaccharide-induced hypothermia

Obese (f/f) Koletsky rats lack the leptin receptor (LR), whereas their lean (F/?) counterparts bear a fully functional LR. By using f/f and F/? rats, we studied whether the LR is involved in lipopolysaccharide (LPS)‐induced fever and hypothermia. The body temperature responses to LPS (10 or 100 µg/kg iv) were measured in Koletsky rats exposed to a thermoneutral (28°C) or cool (22°C) environment. Rats of both genotypes responded to LPS with fever at 28°C and with dose‐dependent hypothermia at 22°C. The fever responses of the f/f and F/? rats were identical. The hypothermic response of the f/f rats was markedly prolonged compared with that of the F/? rats. The prolonged hypothermic response to LPS in the f/f rats was accompanied by enhanced NF‐κB signaling in the hypothalamus and an exaggerated rise in the plasma concentration of tumor necrosis factor (TNF)‐α. The f/f rats did not respond to LPS with an increase in the plasma concentration of corticosterone or adrenocorticotropic hormone, whereas their F/? counterparts did. The hypothermic response to TNF‐α (80 μg/kg iv) was markedly prolonged in the f/f rats. These data show that the LR is essential for the recovery from LPS hypothermia. LR‐dependent mechanisms of the recovery from LPS hypothermia include activation of the anti‐inflammatory hypothalamo‐pituitary‐adrenal axis, inhibition of both the production and hypothermic action of TNF‐α, and suppression of inflammatory (via NF‐κB) signaling in the hypothalamus.

A Dialogical Approach to Conceptualizing Resident Participation in Community Organizing

This article considers “concept formation in the wild” among community organizers aiming to initiate change through processes involving resident participation and dialogue with community members. While this approach is thought to be more effective than “top down” processes, there is little clear understanding of how conceptualizations of resident participation translate into authentic dialogue or substantive change. Our dialogical approach examines how a planning committee worked through differing interpretations of resident participation, as a concept and a way of being, and suggests that genuine dialogicality can point toward experimental forms of relating and increase possibilities for conceptualizing and organizing social futures.