Kori L. Brewer

Neuroprotective Effects of Interleukin-10 Following Excitotoxic Spinal Cord Injury

Intraspinal injection of quisqualic acid (QUIS) produces excitotoxic injury with pathological characteristics similar to those associated with ischemic and traumatic spinal cord injury (SCI). Inflammatory responses appear to be a major component of the secondary neuronal injury initiated by SCI and play a role in the pathogenesis of QUIS-induced injury. IL-10 is a potent antiinflammatory cytokine that has been shown to reduce inflammation and improve functional outcome in human and animal models of inflammatory diseases. We propose the administration of IL-10 following excitotoxic SCI will attenuate the inflammatory response, thus resulting in increased neuronal survival. Female, Sprague-Dawley rats were given intraspinal injections of QUIS followed by either intraspinal (5 ng, n = 8) or systemic injections (5 microgram n = 14) of IL-10. Survival times were varied (2-3 days) in order to produce a range of injury states and inflammatory involvement. When administered intraspinally, IL-10 significantly exacerbated the QUIS damage (P < 0.05), resulting in an 11.2% increase in lesion volume. When given systemically, IL-10 significantly decreased lesion volume by 18.1% in the more advanced injury (P < 0.05), but did not effect the more acute injury. These divergent effects were attributed to the modest inflammatory response in the short-term injury compared to the more robust inflammatory response in the more chronic injury. In conclusion, reducing the inflammatory response to SCI by systemic administration of IL-10 resulted in a significant reduction in neuronal damage, suggesting that targeting injury-induced inflammation may be an effective treatment strategy for acute SCI.

Weight gain associated with chronic exposure to chlorpyrifos in rats.

ObjectiveThis work exposed rats to low levels of the organophosphate insecticide chlorpyrifos and monitored for toxic effects, including weight gain.MethodsRats received either a subcutaneous injection of chlorpyrifos, 5 mg/kg/day, or an equal volume of vehicle daily for 4 months. Subjects were observed for 30 minutes after injection for signs of acute toxicity. Body weights were recorded at baseline, 2 months, 3 months, and 4 months. At the end of the experiment, the weights of hearts, medial lobe of the livers, peri-nephric fat pads, and gastrocnemius muscles were recorded. Effects of chlorpyrifos on adipocyte differentiation in culture were studied. Results were compared using RMANOVA.ResultsNo signs of acute cholinergic toxicity were observed after injections in any subject. Rats in the 5 mg/kg group were significantly heavier than those in the control group by 2 months (335.7 ± 16.7 g vs. 318.6 ± 15.8 g; p = 0.034). This difference increased at 3 months (350.1 ± 16.4 g vs. 322.3 ± 21.3 g p = 0.006) and 4 months (374.4 ± 22.2 g vs. 340.2 ± 25.2 g p = 0.006). At 4 months, the weights of the perinephric fat pads were significantly increased in the chlorpyrifos group relative to controls (2.867 + 0.516 vs. 1.130 + 0.171, p = 0.0039). The two groups showed no weight differences between hearts, livers, and gastrocnemius muscles. Chlorpyrifos did not affect adipocyte differentiation in tissue culture.ConclusionsChronic exposure to chlorpyrifos at 5 mg/kg/day caused an increase in rat body weight when compared to controls. This increase was in adipose tissue. Chlorpyrifos did not induce differentiation of adipocytes in culture.

The effects of endogenous interleukin-10 on gray matter damage and the development of pain behaviors following excitotoxic spinal cord injury in the mouse.

Interleukin-10 (IL-10) has been utilized as a neuroprotective agent in experimental models of spinal cord injury because of its potent anti-inflammatory properties. Previous studies have delivered a single dose (5 microg) of IL-10 following experimental spinal cord injury in the rat, and demonstrated various degrees of neuroprotection. However, the role of endogenous production of IL-10 has not been considered. Therefore, the purpose of the current study was to establish the role of endogenous IL-10 and demonstrate the true potential of exogenous IL-10 administration through the use of IL-10((-/-)) mice. Using the quisqualic acid model of spinal cord injury, we examined the extent of gray matter damage and onset of injury-induced pain behaviors at various time points following injury in wild-type vs. IL-10((-/-)) mice. Additionally, IL-10 was reconstituted in IL-10 deficient mice by the intraperitoneal administration of 50 ng recombinant murine (rm) IL-10 30 min following quisqualic acid injection. Animals were observed daily following injury for the onset of pain-behaviors. At days 1, 7, and 14 following injection, lesion analysis revealed a greater extent of damage at early time points (1 day, 7 days) following injury in the IL-10((-/-)) animals as compared with wild-type animals. However, by 14 days post-experimental spinal cord injury, the extent of damage between the two groups was not significant. IL-10((-/-)) animals that received the single (50 ng) rmIL-10 injection following injury displayed gray matter damage patterns similar to wild-type animals. The pronounced early damage noted in the IL-10((-/-)) animals was associated with an approximately two-fold increase in peripheral neutrophils, an index of an innate immune response to injury, compared with wild-type mice. In addition, wild type and IL-10((-/-)) animals receiving rmIL-10 demonstrated a delay in the onset of injury-induced pain behaviors. However, by 14 days post-experimental spinal cord injury the overall incidence of pain behaviors was similar between all treatment groups. Therefore, the absence of IL-10 expression accelerates the kinetics of lesion expansion, the onset of pain behaviors, and the peripheral immune response to spinal cord injury. Endogenous IL-10 and low doses of exogenous IL-10 are neuroprotective at 1 and 7 days following injury. Therefore, the results of the current study suggest that low dose IL-10 administration acutely following spinal cord injury has potential as a therapeutic agent for limiting tissue loss following injury.

Chronic, Selective Forebrain Responses to Excitotoxic Dorsal Horn Injury

Intraspinal injection of the AMPA/metabotropic receptor agonist quisqualic acid (QUIS) results in excitotoxic injury which develops pathological characteristics similar to those associated with ischemic and traumatic spinal cord injury (SCI) (R. P. Yezierski et al., 1998, Pain 75: 141-155; R. P. Yezierski et al., 1993, J. Neurotrauma 10: 445-456). Since spinal injury can lead to partial or complete deafferentation of ascending supraspinal structures, it is likely that secondary to the disruption of spinal pathways these regions could undergo significant reorganization. Recently, T. J. Morrow et al. (Pain 75: 355-365) showed that autoradiographic estimates of regional cerebral blood flow (rCBF) can be used to simultaneously identify alterations in the activation of multiple forebrain structures responsive to noxious formalin stimulation. Accordingly, we examined whether excitotoxic SCI produced alterations in the activation of supraspinal structures using rCBF as a marker of neuronal activity. Twenty-four to 41 days after unilateral injection of QUIS into the T12 to L3 spinal segments, we found significant increases in the activation of 7 of 22 supraspinal structures examined. As compared to controls, unstimulated SCI rats exhibited a significant bilateral increase in rCBF within the arcuate nucleus (ARC), the hindlimb region of S1 cortex (HL), parietal cortex (PAR), and the thalamic posterior (PO), ventral lateral (VL), ventral posterior lateral (VPL), and ventral posterior medial (VPM) nuclei. All structures showing significantly altered rCBF are associated with the processing of somatosensory information. These changes constitute remote responses to injury and suggest that widespread functional changes occur within cortical and subcortical regions following injury to the spinal cord.

A prospective double-blind study of nasal sumatriptan versus IV ketorolac in migraine

We conducted a study to compare the efficacy in migraine headache of nasal sumatriptan and intravenous ketorolac. The study was a prospective, double-blind study done with a convenience sample of 29 patients presenting to the emergency department (ED) with acute migraine. Patients received either 20 mg of nasal sumatriptan or 30 mg of intravenous ketorolac. Patients scored the severity of their headache on a 100-mm visual analog scale (VAS) of pain prior to medication, and again 1 hour after medication. Differences between initial and 1-hour scores were analyzed. Before treatment, no difference existed between the groups in the intensity of headache. One hour after medication, the sumatriptan group had a decrease in pain score of 22.937 mm and the ketorolac group a decrease of 71.462 mm on the VAS. The decrease in pain score with ketorolac was significantly greater than that with sumatriptan (P < 0.001). The study therefore showed that both sumatriptan and ketorolac effectively reduced the pain associated with acute migraine headache, but that intravenous ketorolac produced a greater reduction in pain than did nasal sumatriptan.

The role of kainic acid/AMPA and metabotropic glutamate receptors in the regulation of opioid mRNA expression and the onset of pain-related behavior following excitotoxic spinal cord injury

Intraspinal injection of quisqualic acid, a mixed kainic acid/2-amino-3(3-hydroxy-5-methylisoxazol-4-yl)propionic acid and metabotropic glutamate receptor agonist, produces an excitotoxic injury that leads to the onset of both spontaneous and evoked pain behavior as well as changes in spinal and cortical expression of opioid peptide mRNA, preprodynorphin and preproenkephalin. What characteristics of the quisqualic acid-induced injury are attributable to activation of each receptor subtype is unknown. This study attempted to define the role of activation of the kainic acid/2-amino-3(3-hydroxy-5-methylisoxazol-4-yl)propionic acid (AMPA) and metabotropic glutamate receptor subtypes in the regulation of opioid peptide expression and the onset of spontaneous and evoked pain-related behavior following excitotoxic spinal cord injury by comparing quisqualic acid-induced changes with those created by co-injection of quisqualic acid and the kainic acid/AMPA antagonist, 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo[f]quinoxaline, (NBQX) or the metabotropic antagonist, (RS)-1-aminoindan-1,5-dicarboxylic acid (AIDA). Therefore, 42 male Long-Evans adult rats were divided into seven treatment groups and received intraspinal microinjections of saline (sham), 0.5% dimethylsulphoxide (sham), quisqualic acid (1.2 microl, 125 mM), NBQX (1.2 microl, 60 microM), AIDA (1.2 microl, 250 microM), quisqualic acid/NBQX (1.2 microl, 125 mM/60 microM), or quisqualic acid/AIDA (1.2 microl, 125 mM/250 microM) directed at spinal levels thoracic 12-lumbar 2. Behavioral observations of spontaneous and evoked pain responses were completed following surgery. After a 10-day survival period, animals were killed and brain and spinal cord tissues were removed and processed for histologic analysis and in situ hybridization. Both AIDA and NBQX affected the quisqualic acid-induced total lesion volume but only AIDA caused a decrease in the percent tissue damage at the lesion epicenter. Preprodynorphin and preproenkephalin expression is increased in both spinal and cortical areas in quisqualic acid-injected animals versus sham-, NBQX or AIDA-injected animals. NBQX did not affect quisqualic acid-induced spinal or cortical expression of preprodynorphin or preproenkephalin except for a significant decrease in preproenkephalin expression in the spinal cord. In contrast, AIDA significantly decreases quisqualic acid-induced preprodynorphin and preproenkephalin expression within the spinal cord and cortex. AIDA, but not NBQX, significantly reduced the frequency of, and delayed the onset of, quisqualic acid-induced spontaneous pain-related behavior. From these data we suggest that both the kainic acid/AMPA and metabotropic glutamate receptor subtypes are involved in the induction of the excitotoxic cascade responsible for quisqualic acid-induced neuronal damage and changes in opioid peptide mRNA expression, while metabotropic glutamate receptors may play a more significant role in the onset of post-injury pain-related behavior.

Spinal and supraspinal changes in opioid mRNA expression are related to the onset of pain behaviors following excitotoxic spinal cord injury.

&NA; Excitotoxic spinal cord injury (SCI) causes anatomic, physiologic and molecular changes within the spinal cord and brain. Intraspinal injection of quisqualic acid (QUIS) produces an excitotoxic injury that leads to the onset of behavioral syndromes, believed to be related to the clinical condition of chronic pain. The opioid system, classically involved in the suppression of pain transmission, has been associated with the onset of pain‐related behaviors and changes in spinal opioid peptide expression have been demonstrated in various models of SCI and chronic pain. Recently, changes in opioid peptide expression have been demonstrated in both spinal and supraspinal areas following excitotoxic SCI. Therefore, the purpose of this study was to examine changes in opioid peptide gene expression as they relate to the onset of pain behaviors following excitotoxic SCI. Male, Long–Evans rats were given an intraspinal injection of 1.2 &mgr;l of 125 mM QUIS and allowed to survive for 10 days, a duration sufficient for the development of pain‐related behaviors. Animals were assessed daily for the presence of excessive grooming behavior, i.e. self‐directed biting and scratching resulting in damage to superficial and deeper layers of the skin. Animals were also tested for thermal hypersensitivity using a cold plate apparatus on days 5, 7, and 10 following QUIS injection. After sacrifice, quantitative in situ hybridization was performed on regions of the spinal cord surrounding the lesion site as well as whole brain sections through various levels of the thalamus and cortex. Spinal preproenkephalin (PPE) and preprodynorphin (PPD) expression was significantly increased in animals that developed excessive grooming behaviors vs. those that did not. For PPE, this difference was seen bilaterally, in areas of cord caudal to the site of injury. For PPD, this difference was seen only ipsilateral to the site of injection, rostral to the site of injury. In addition, PPE expression in the anterior cingulate cortex and PPD expression in the contralateral parietal cortex were significantly higher in grooming vs. non‐grooming animals. These results support previous conclusions that both spinal and supraspinal regulation of endogenous opioid peptide expression plays a role in the response to or onset of post‐SCI pain. These results also suggest that the opioid peptides are regulated independently and serve different functions in response to SCI.

Opioid peptide messenger RNA expression is increased at spinal and supraspinal levels following excitotoxic spinal cord injury

Spinal cord injury in rats is known to cause anatomical, physiological and molecular changes within the spinal cord. These changes may account for behavioral syndromes that appear following spinal cord injury, syndromes believed to be related to the clinical condition of chronic pain. Intraspinal injection of quisqualic acid produces an excitotoxic injury with pathological characteristics similar to those associated with ischemic and traumatic spinal cord injury. In addition, recent studies have demonstrated changes in blood flow, neuronal excitability and gene expression in the brain following excitotoxic injury, indicating that behavioral changes may result from modification of neuronal substrates at supraspinal levels of the neuraxis. Because changes in spinal opioid peptide expression have been demonstrated in models of traumatic spinal cord injury and chronic pain, the present study investigated messenger RNA expression of the opioid peptides, preproenkephalin and preprodynorphin, at spinal and supraspinal levels following excitotoxic spinal cord injury. Male, Long-Evans rats were given three intraspinal injections of quisqualic acid (total 1.2 microl, 125mM). After one, three, five, seven or 10days, animals were killed and quantitative in situ hybridization performed on regions of the spinal cord surrounding the lesion site, as well as whole-brain sections through various levels of the thalamus. Preproenkephalin and preprodynorphin expression was increased in spinal cord areas adjacent to the site of quisqualic injection and in cortical regions associated with nociceptive function, preproenkephalin in the cingulate cortex and preprodynorphin in the parietal cortex, both ipsilaterally and contralaterally at various time-points following injury. These results further our knowledge of the secondary events that occur following spinal cord injury, specifically implicating supraspinal opioid systems in the CNS response to spinal cord injury.

Pilot studies of pressure-immobilization bandages for rattlesnake envenomations

Study objective. Pressure-immobilization bandages sequester venom in extremities and are recommended for snakebites without local toxicity. Pilot studies were performed to determine the time of onset of toxicity and efficacy of pressure-immobilizations bandages in a porcine model of rattlesnake envenomation. Methods. After IACUC approval, anesthetized pigs were injected subcutaneously in a distal hind leg with 200 mg of Crotalus atrox venom. After 1 min, pigs received either a pressure-immobilization bandage (N = 3) or no treatment (N = 3). At 24 h, surviving pigs received antivenin and then the pressure-immobilization bandages were removed. Surviving subjects were followed for 1 week. Chi-square analysis and paired t-test were used. Results. Pigs with pressure-immobilization bandages survived for 24 h, whereas untreated pigs died at 13.68 ± 3.42 h (p = 0.014). Surviving pigs walked on the extremity at 7 days. Potassium rose from 4.033 ± 0.252 at baseline to 17.767 ± 5.218 mEq/L (p < 0.0001) at time of death in untreated pigs but was normal at 24 h in treated subjects. Widespread tissue necrosis was seen in the untreated group but only local necrosis in the treatment group. Conclusions. Pressure-immobilization bandages prevented death from severe C. atrox envenomations with a 24 h delay to treatment. Surviving pigs had recovery of limb use at 1 week.

RVU Ready? Preparing Emergency Medicine Resident Physicians in Documentation for an Incentive‐based Work Environment

OBJECTIVES The emergency medicine (EM) job market is increasingly focused on incentive-based reimbursement, which is largely based on relative value units (RVUs) and is directly related to documentation of patient care. Previous studies have shown a need to improve resident education in documentation. The authors created a focused educational intervention on billing and documentation practices to meet this identified need. The hypothesis of this study was that this educational intervention would result in an increase in RVUs generated by EM resident physicians and the average amount billed per patient. METHODS The authors used a quasi-experimental study design. An educational intervention included a 1-hour lecture on documentation and billing, biweekly newsletters, and case-specific feedback from the billing department for EM resident physicians. RVUs and charges generated per patient were recorded for all second- and third-year resident physicians for a 3-month period prior to the educational intervention and for a 3-month period following the intervention. Pre- and postintervention data were compared using Students t-test and repeated-measures analysis of variance, as appropriate. RESULTS The evaluation and management (E/M) chart levels billed during each phase of the study were significantly different (p < 0.0001). The total number of RVUs generated per hour increased from 3.17 in the first phase to 3.71 in the second phase (p = 0.0001). During the initial 3-month phase, the average amount billed per patient seen by a second- or third-year resident was 282.82 USD, which increased to 301.94 USD in the second phase (p = 0.0004). CONCLUSIONS The educational intervention positively affected resident documentation resulting in greater RVUs/hour and greater billing performance in the study emergency department (ED).