The Interplay of Nutrition, Physical Activity, Severity of Illness, and Mortality in Critically Ill Burn Patients: Is There a Connection?

The purpose of this project was to evaluate the relationships between nutrition, physical activity levels (PALs), severity of illness (SOI), and survival in critically ill burn patients. We conducted a retrospective evaluation of consecutively admitted adult patients who had an intensive care unit stay ≥8 days after ≥20% TBSA burns. Linear regression was used to assess the association between SOI (sequential organ failure assessment scores) and PALs as well as between SOI and nutritional intake. After univariate analysis comparing survivors and nonsurvivors, factors with P < .10 were analyzed with multiple logistic regression. Characteristics of the 45 included patients were: 42 ± 15 years old, 37 ± 17% TBSA burns, 22% mortality. Factors independently associated with survival were burn size (negatively) (P = .018), height (positively) (P = .006), highest PAL during the first eight intensive care unit days (positively) (P = .016), and kcal balance during the fifth through the eighth intensive care unit days (positively) (P = .012). Sequential organ failure assessment scores had a significant (P < .001) but weak association with nutrition intake (R2 = 0.05) and PALs (R2 = 0.25). Higher nutritional intake and activity were significantly associated with lower mortality in critically ill burn patients. Given the weak associations between both nutritional intake and PALs with SOI, the primary barrier in achieving nutrition and activity goals was not SOI. We recommend that physical rehabilitation and nutritional intake be optimized in an effort to improve outcomes in critically ill burn patients.

Predicting wound healing rates and survival with the use of automated serial evaluations of burn wounds.

Healing of burn wounds is necessary for survival; however tracking progression or healing of burns is an inexact science. Recently, the relationship of mortality and wound healing has been documented with a software termed WoundFlow. The objective of the current study was to confirm various factors that impact burn wound healing, as well as to establish a timeline and rate of successful healing. A retrospective analysis was performed on adults (n=115) with at least 20% TBSA burn that had at least two computer-based wound mappings. The % open wound (%OW) was calculated over time to document healing trajectory until successful healing or death. Only 2% of patients in the group with successful wound healing died. A decrease in the %OW of 0.8 (IQR: 0.7-1.1) was associated with survival. Disparities in wound healing trajectories between survivors and non-survivors were distinguishable by 2weeks post-injury (P<0.05). When %TBSA was stratified by decile, the 40-49% TBSA group had the highest healing rate. Taken together, the data indicate that wound healing trajectory (%OW) varies with injury severity and survival. As such, automated mapping of wound healing trajectory may provide valuable information concerning patient/prognosis, and may recommend early interventions to optimize wound healing.

Comparison of military and civilian burn patients admitted to a single center during 12 years of war.

OBJECTIVE: The current conflicts in Iraq and Afghanistan resulted in an increased incidence of burn injury in the military population. We sought to compare the characteristics and outcomes of this population to a civilian cohort cared for at the same burn center over the same time-period. METHODS: A retrospective review was performed to examine differences in the demographics, etiology, mortality, and functional status over a 12-year period. Descriptive analyses were performed. Logistic regression was used to calculate the likelihood of mortality. RESULTS: A total of 3814 patients were included in this analysis; 1069 were military casualties. When compared to civilians, military patients were younger, had a higher incidence of flame-induced burn injury, mean total body surface area burned (% TBSA), rate of inhalation injury, and lower mortality. Civilian patients presented with a higher Baux score. Although most military patients had a full functional recovery, they had a greater incidence of severe disability. In a univariate model, likelihood of mortality was higher in civilians. No difference in mortality between the two cohorts was found after adjusting for age, inhalation injury, gender, % TBSA and percent full-thickness burn. CONCLUSIONS: Military patients exhibited improved survival and functional recovery over their civilian counterparts. However, mortality did not differ between civilian and military patients after controlling for known covariates. Further studies are needed to improve functional outcomes in civilian patients, who may not have the inherent advantages of younger age and healthier physical status found in military patients.

A Survey of Temperature Management Practices Among Burn Centers in North America.

Maintaining body temperature is a unique challenge with burn care. We sought to describe core temperature goals in the operating room (OR) and the methods used to achieve and maintain these goals, along with current methods of warming in the intensive care unit (ICU), the perception of effect of increased ambient temperature on work performance, and concerns with contamination of sterile fields due to increased ambient temperature. A 24 question survey was disseminated to burn centers in the United States and Canada. The questions included demographics, target core and ambient temperatures, warming methods, and beliefs on ambient temperature's effects. Of 121 burn centers, 52 questionnaires were completed (43% response rate). The majority of centers targeted a core temperature between 36 and 38°C in the OR and an ambient temperature between 75 and 95°F in the ICU. The most common methods for maintaining core temperature included warmed ambient temperature, forced air devices, and intravenous fluids. Although the majority of centers reported the belief that increased ambient temperature benefits patients, many also reported that there is a negative impact on staff performance and risk of staff perspiration contaminating sterile fields. Burn centers reported a range of target core temperatures and methods to reach target temperatures. More than a third of respondents perceived a negative impact work performance while more than half acknowledged the potential for contamination of sterile fields. A prospective observational study is needed to determine actual temperature regulation practice patterns and its impact on outcomes.

Thermal stability of mafenide and amphotericin B topical solution.

OBJECTIVE: Fungal infections remain a major cause of mortality in the burned population. Mafenide acetate/amphotericin B solution (SMAT) has been used topically for prophylaxis and treatment of these infections. Current manufacturer guidelines only guarantee the stability of mafenide solution and amphotericin B at room temperature. Additionally, the recommended maximum storage time for mafenide solution is 48h, leading to significant financial and material loss when unused solutions are discarded. The purpose of this study was to characterize the chemical stability, structure and bioactivity of SMAT stored at 2°C, 25°C, and 40°C for up to 90 days. METHODS: Stability analyses of SMAT solutions containing 2.5% or 5% mafenide plus 2µg/mL amphotericin B were performed using high performance liquid chromatography. Chemical structure was assessed using Fourier-transform infrared spectroscopy. Bioactivity against clinically relevant species was examined. RESULTS: The chemical structure and stability of mafenide did not change over 90days at all temperatures. Amphotericin B was undetectable in SMAT solutions after two days at high temperatures, which was slowed by refrigerated storage. Against Staphylococcus aureus, SMAT activity began to decrease generally between two and seven days. Against Pseudomonas aeruginosa, activity slowly tapered and was gone by day 90. SMAT retained high bioactivity against Candida albicans for over 40days and was not affected by temperature. CONCLUSIONS: The amphotericin B component of SMAT is degraded within 2days under warm storage. While mafenide was stable over 90 days, the bioactivity of SMAT solution may be lost within 2days as well.

Wound Penetration of Cefazolin, Ciprofloxacin, Piperacillin, Tazobactam, and Vancomycin During Negative Pressure Wound Therapy.

Objective: Negative pressure wound therapy (NPWT) uses subatmospheric pressure as a noninvasive adjunct to treat wounds and has demonstrated clinical efficacy by accelerating healing of a variety of acute and chronic wounds. NPWT may also play a role in preventing or treating wound infections, possibly by increasing wound penetration of antibiotics. However, clinical data in patients undergoing antibiotic and NPWT treatment are limited. Approach: To evaluate the wound penetration of antibiotics in NPWT patients, we conducted a prospective, observational study of burn and trauma patients treated with NPWT and systemic antibiotics. We evaluated the plasma pharmacokinetic profile of systemic vancomycin, ciprofloxacin, cefazolin, and piperacillin/tazobactam, as well as total and unbound antibiotic concentrations in wound exudate from the same patients. Results: Data from 32 patients with 37 wounds undergoing NPWT demonstrated that vancomycin, ciprofloxacin, and piperacillin/tazobactam all penetrated wounds with exudate to plasma concentration ratios more than 0.8. Cefazolin did not penetrate wounds in patients undergoing NPWT as effectively, with an average exudate to plasma concentration ratio of 0.51. Innovation: Clinical data on the wound penetration of antibiotics in patients undergoing NPWT are limited, but these data suggest that antibiotics have different capacities for wound penetration during NPWT that should be considered when making clinical decisions. Conclusion: This initial report suggests that (1) vancomycin, ciprofloxacin, and piperacillin/tazobactam effectively penetrate wounds during NPWT and (2) cefazolin as well as other antibiotics may not penetrate wounds during NPWT.

Perioperative Temperature Management During Burn Care.

Major physiologic alterations following a severe thermal injury disrupt thermal homeostasis and predispose burn patients to hypothermia. An important recommendation in many clinical practice guidelines is to increase the ambient temperature during the care of severely burned patients in the operating room and intensive care unit to mitigate the loss of thermoregulation, prevent hypothermia, and minimize the impact of hypermetabolism. However, the scientific support for this recommendation remains unclear. This review summarizes the current knowledge regarding the pathophysiology and treatment of thermal injury-induced hypermetabolism and hypothermia, with special emphasis on alterations in ambient temperature. Current evidence on the value of increasing ambient temperature during the care of severely burned patients in the operating room or intensive care unit is limited, with minimal human studies investigating physiologic benefit or potential adverse effects.

Elevations in growth hormone and glucagon-like peptide-2 levels on admission are associated with increased mortality in trauma patients.

BACKGROUND: Burn and trauma patients present a clinical challenge due to metabolic derangements and hypermetabolism that result in a prolonged catabolic state with impaired healing and secondary complications, including ventilator dependence. Previous work has shown that circulating levels of growth hormone (GH) are predictive of mortality in critically ill adults, but few studies have examined the prognostic potential of GH levels in adult trauma patients. METHODS: To investigate the utility of GH and other endocrine responses in the prediction of outcomes, we conducted a prospective, observational study of adult burn and trauma patients. We evaluated the serum concentration of GH, insulin-like growth factor 1 (IGF-1), IGF binding protein 3 (IGFBP-3), and glucagon-like peptide-2 (GLP-2) weekly for up to 6 weeks in 36 adult burn and trauma patients admitted between 2010 and 2013. RESULTS: Non-survivors had significantly higher levels of GH and GLP-2 on admission than survivors. DISCUSSION: This study demonstrates that GH has potential as a predictor of mortality in critically ill trauma and burn patients. Future studies will focus on not only the role of GH, but also GLP-2, which was shown to correlate with mortality in this study with a goal of offering early, targeted therapeutic interventions aimed at decreasing mortality in the critically injured. CONCLUSIONS: GH and GLP-2 may have clinical utility for outcome prediction in adult trauma patients.

Vitamin C in Burn Resuscitation.

The inflammatory state after burn injury is characterized by an increase in capillary permeability that results in protein and fluid leakage into the interstitial space, increasing resuscitative requirements. Although the mechanisms underlying increased capillary permeability are complex, damage from reactive oxygen species plays a major role and has been successfully attenuated with antioxidant therapy in several disease processes. However, the utility of antioxidants in burn treatment remains unclear. Vitamin C is a promising antioxidant candidate that has been examined in burn resuscitation studies and shows efficacy in reducing the fluid requirements in the acute phase after burn injury.

Intravenous Antibiotic and Antifungal Agent Pharmacokinetic-Pharmacodynamic Dosing in Adults with Severe Burn Injury.

PURPOSE: Despite advances in the care of patients with severe burn injury, infection-related morbidity and mortality remain high and can potentially be reduced with antimicrobial dosing optimized for the infecting pathogen. However, anti-infective dose selection is difficult because of the highly abnormal physiologic features of burn patients, which can greatly affect the pharmacokinetic (PK) disposition of these agents. We review published PK data from burn patients and offer evidence-based dosing recommendations for antimicrobial agents in burn-injured patients. METHODS: Because most infections occur at least 48 hours after initial burn injury and anti-infective therapy often lasts ≥10 days, we reviewed published data informing PK-pharmacodynamic (PD) dosing of anti-infectives administered during the second, hypermetabolic stage of burn injury, in those with >20% total body surface area burns, and in those with normal or augmented renal clearance (estimated creatinine clearance ≥130 mL/min). Analyses were performed using 10,000-patient Monte Carlo simulations, which uses PK variability observed in burn patients and MIC data to determine the probability of reaching predefined PK-PD targets. The probability of target attainment, defined as the likelihood that an anti-infective dosing regimen would achieve a specific PK-PD target at the single highest susceptible MIC, and the cumulative fraction of response, defined as the population probability of target attainment given a specific dose and a distribution of MICs, were calculated for each recommended anti-infective dosing regimen. FINDINGS: Evidence-based doses were derived for burn-injured patients for 15 antibiotics and 2 antifungal agents. Published data were unavailable or insufficient for several agents important to the care of burn patients, including newer antifungal and antipseudomonal agents. Furthermore, available data suggest that antimicrobial PK properties in burned patients is highly variable. We recommend that, where possible, therapeutic drug monitoring be performed to optimize PK-PD parameter achievement in individual patients. IMPLICATIONS: Given the high variability in PK disposition observed in burn patients, doses recommended in the package insert may not achieve PK-PD parameters associated with optimal infectious outcomes. Our study is limited by the necessity for fixed assumptions in depicting this highly variable patient population. New rapid-turnaround analytical technology is needed to expand the menu of antimicrobial agents for which therapeutic drug monitoring is available to guide dose modification within a clinically actionable time frame.

Elevations in inflammatory cytokines are associated with poor outcomes in mechanically ventilated burn patients.

BACKGROUND: The treatment of burn patients who undergo mechanical ventilation is complicated by many factors; patient outcomes and mortality could potentially be improved with predictive biomarkers. Severe burn provokes a systemic inflammatory response characterized by the release of a host of cytokines. Recent studies evaluated the prognostic value of temporal changes in cytokine levels in several patient populations, but few have compared differences in the cytokine profiles of survivors and nonsurvivors following severe burn. We previously compared high-frequency percussive ventilation and low-tidal-volume ventilation and found no difference in mortality or cytokine levels between the two treatments. Since it is unknown whether cytokine levels are predictive of mortality in these patients, we performed a post hoc analysis comparing cytokine levels in survivors and nonsurvivors. METHODS: We evaluated plasma levels of several cytokines (interleukin 1ß [IL-1ß], IL-6, IL-8, granulocyte-macrophage colony-stimulating factor, and tumor necrosis factor α) for their prognostic biomarker potential related to mortality at 0, 3, and 7 days in survivors and nonsurvivors of burns. RESULTS: While the majority of values for IL-1ß, granulocyte-macrophage colony-stimulating factor, and tumor necrosis factor α fell below the limit of quantification, univariate analysis demonstrated higher plasma levels of IL-6 and IL-8 in nonsurvivors on Day 7. Logistic regression revealed that elevated plasma IL-8 was independently associated with an increased likelihood of the composite end point of death or ventilator-associated pneumonia with odds ratios of 7.9, 26, and 7.3 on Days 0, 3, and 7, respectively. CONCLUSION: Early increases in plasma IL-8 are associated with a multifold increase in death or ventilator-associated pneumonia in mechanically ventilated burn patients. LEVEL OF EVIDENCE: Prognostic/epidemiologic study, level IV; therapeutic study, level IV.

A-kinase anchoring protein 79/150 coordinates metabotropic glutamate receptor sensitization of peripheral sensory neurons.

Glutamate serves as the primary excitatory neurotransmitter in the nervous system. Previous studies have identified a role for glutamate and group I metabotropic receptors as targets for study in peripheral inflammatory pain. However, the coordination of signaling events that transpire from receptor activation to afferent neuronal sensitization has not been explored. Herein, we identify that scaffolding protein A-kinase anchoring protein 79/150 (AKAP150) coordinates increased peripheral thermal sensitivity after group I metabotropic receptor (mGluR5) activation. In both acute and persistent models of thermal somatosensory behavior, we report that mGluR5 sensitization requires AKAP150 expression. Furthermore, electrophysiological approaches designed to record afferent neuronal activity reveal that mGluR5 sensitization also requires functional AKAP150 expression. In dissociated primary afferent neurons, mGluR5 activation increases TRPV1 responses in an AKAP-dependent manner through a mechanism that induces AKAP association with TRPV1. Experimental results presented herein identify a mechanism of receptor-driven scaffolding association with ion channel targets. Importantly, this mechanism could prove significant in the search for therapeutic targets that repress episodes of acute pain from becoming chronic in nature.

Burn wound healing and treatment: review and advancements.

Burns are a prevalent and burdensome critical care problem. The priorities of specialized facilities focus on stabilizing the patient, preventing infection, and optimizing functional recovery. Research on burns has generated sustained interest over the past few decades, and several important advancements have resulted in more effective patient stabilization and decreased mortality, especially among young patients and those with burns of intermediate extent. However, for the intensivist, challenges often exist that complicate patient support and stabilization. Furthermore, burn wounds are complex and can present unique difficulties that require late intervention or life-long rehabilitation. In addition to improvements in patient stabilization and care, research in burn wound care has yielded advancements that will continue to improve functional recovery. This article reviews recent advancements in the care of burn patients with a focus on the pathophysiology and treatment of burn wounds.

Antifungal wound penetration of amphotericin and voriconazole in combat-related injuries: case report.

BACKGROUND: Survivors of combat trauma can have long and challenging recoveries, which may be complicated by infection. Invasive fungal infections are a rare but serious complication with limited treatment options. Currently, aggressive surgical debridement is the standard of care, with antifungal agents used adjunctively with uncertain efficacy. Anecdotal evidence suggests that antifungal agents may be ineffective in the absence of surgical debridement, and studies have yet to correlate antifungal concentrations in plasma and wounds. CASE PRESENTATION: Here we report the systemic pharmacokinetics and wound effluent antifungal concentrations of five wounds from two male patients, aged 28 and 30 years old who sustained combat-related blast injuries in southern Afghanistan, with proven or possible invasive fungal infection. Our data demonstrate that while voriconazole sufficiently penetrated the wound resulting in detectable effluent levels, free amphotericin B (unbound to plasma) was not present in wound effluent despite sufficient concentrations in circulating plasma. In addition, considerable between-patient and within-patient variability was observed in antifungal pharmacokinetic parameters. CONCLUSION: These data highlight the need for further studies evaluating wound penetration of commonly used antifungals and the role for therapeutic drug monitoring in providing optimal care for critically ill and injured war fighters.

Colistin pharmacokinetics in burn patients during continuous venovenous hemofiltration.

While colistin is considered a last resort for the treatment of multidrug-resistant Gram-negative bacterial infections, there has been an increase in its use due to the increasing prevalence of drug-resistant infections worldwide. The pharmacology of colistin is complex, and pharmacokinetic data are limited, especially in patients requiring renal replacement therapy. As a result, dosing for patients who require renal replacement remains a challenge. Here, we present pharmacokinetic data for colistin from two burn patients (37 and 68 years old) infected with colistin-susceptible isoclonal Acinetobacter baumannii and receiving continuous venovenous hemofiltration (CVVH). To our knowledge, we are the first to examine data from before and during CVVH (for one patient), allowing analysis of the effect of CVVH on colistin pharmacokinetics. Pharmacokinetic/pharmacodynamic analysis indicated that a dose increase from 1.5 to 2.2 mg/kg of body weight colistin base activity on CVVH was insufficient to satisfy the target parameter of an AUC24/MIC (area under the concentration-time curve over 24 h in the steady state divided by the MIC) of ≥ 60 at an MIC of ≥ 1 µg/ml in one patient with residual endogenous renal function. Plasma concentrations of colistin ranged from 0 to 15 µg/ml, with free colistin levels ranging from 0.4 to 2.2 µg/ml. While both patients resolved their clinical infections and survived to discharge, colistin-resistant colonizing isolates resulted from therapy in one patient. The variabilities observed in colistin concentrations and pharmacokinetic characteristics highlight the importance of pharmacokinetic monitoring of antibiotics in patients undergoing renal replacement therapy.

ß-arrestin-2-biased agonism of delta opioid receptors sensitizes transient receptor potential vanilloid type 1 (TRPV1) in primary sensory neurons.

Despite advances in understanding the signaling mechanisms involved in the development and maintenance of chronic pain, the pharmacologic treatment of chronic pain has seen little advancement. Agonists at the mu opioid receptor (MOPr) continue to be vital in the treatment of many forms of chronic pain, but side-effects limit their clinical utility and range from relatively mild, such as constipation, to major, such as addiction and dependence. Additionally, chronic activation of MOPr results in pain hypersensitivity known as opioid-induced hyperalgesia (OIH), and we have shown recently that recruitment of ß-arrestin2 to MOPr, away from transient potential vanilloid eceptor type 1 (TRPV1) in primary sensory neurons contributes to this phenomenon. The delta opioid receptor (DOPr) has become a promising target for the treatment of chronic pain, but little is known about the effects of chronic activation of DOPr on nociceptor sensitivity and OIH. Here we report that chronic activation of DOPr by the DOPr-selective agonist, SNC80, results in the sensitization of TRPV1 and behavioral signs of OIH via ß-arrestin2 recruitment to DOPr and away from TRPV1. Conversely, chronic treatment with ARM390, a DOPr-selective agonist that does not recruit ß-arrestin2, neither sensitized TRPV1 nor produced OIH. Interestingly, the effect of SNC80 to sensitize TRPV1 is species-dependent, as rats developed OIH but mice did not. Taken together, the reported data identify a novel side-effect of chronic administration of ß-arrestin2-biased DOPr agonists and highlight the importance of potential species-specific effects of DOPr agonists.

Activation of estrogen receptor α enhances bradykinin signaling in peripheral sensory neurons of female rats.

Numerous studies have demonstrated that females have a higher risk of experiencing several pain disorders with either greater frequency or severity than males. Although the mechanisms that underlie this sex disparity remain unclear, several studies have shown an important role for sex steroids, such as estrogen, in the modulation of nociception. Receptors for estrogen are present in primary afferent neurons in the trigeminal and dorsal root ganglia, and brief exposure to estrogen increases responses to the inflammatory mediator bradykinin (BK). However, the mechanism for estrogen-mediated enhancement of BK signaling is not fully understood. The aim of the present study was to evaluate the relative contributions of estrogen receptor α (ERα), ERß, and G protein-coupled estrogen receptor 1 (GPER) to the enhanced signaling of the inflammatory mediator BK by 17ß-estradiol (17ß-E2) in primary sensory neurons from female rats in culture (ex vivo) and in behavioral assays of nociception in vivo. The effects of 17ß-E2 on BK responses were mimicked by ERα-selective agonists and blocked by ERα-selective antagonists and by small interfering RNA knockdown of ERα. The data indicate that ERα is required for 17ß-E2-mediated enhancement of BK signaling in peripheral sensory neurons in female rats.

Activation of mu opioid receptors sensitizes transient receptor potential vanilloid type 1 (TRPV1) via ß-arrestin-2-mediated cross-talk.

The transient receptor potential family V1 channel (TRPV1) is activated by multiple stimuli, including capsaicin, acid, endovanilloids, and heat (>42C). Post-translational modifications to TRPV1 result in dynamic changes to the sensitivity of receptor activation. We have previously demonstrated that ß-arrestin2 actively participates in a scaffolding mechanism to inhibit TRPV1 phosphorylation, thereby reducing TRPV1 sensitivity. In this study, we evaluated the effect of ß-arrestin2 sequestration by G-protein coupled receptors (GPCRs) on thermal and chemical activation of TRPV1. Here we report that activation of mu opioid receptor by either morphine or DAMGO results in ß-arrestin2 recruitment to mu opioid receptor in sensory neurons, while activation by herkinorin does not. Furthermore, treatment of sensory neurons with morphine or DAMGO stimulates ß-arrestin2 dissociation from TRPV1 and increased sensitivity of the receptor. Conversely, herkinorin treatment has no effect on TRPV1 sensitivity. Additional behavioral studies indicate that GPCR-driven ß-arrestin2 sequestration plays an important peripheral role in the development of thermal sensitivity. Taken together, the reported data identify a novel cross-talk mechanism between GPCRs and TRPV1 that may contribute to multiple clinical conditions.

Allosteric interactions between δ and κ opioid receptors in peripheral sensory neurons.

The peripheral δ opioid receptor (DOR) is an attractive target for analgesic drug development. There is evidence that DOR can form heteromers with the κ-opioid receptor (KOR). As drug targets, heteromeric receptors offer an additional level of selectivity and, because of allosteric interactions between protomers, functionality. Here we report that selective KOR antagonists differentially altered the potency and/or efficacy of DOR agonists in primary cultures of adult rat peripheral sensory neurons and in a rat behavioral model of thermal allodynia. In vitro, the KOR antagonist nor-binaltorphimine (nor-BNI) enhanced the potency of [D-Pen(2,5)]-enkephalin (DPDPE), decreased the potency of [D-Ala(2),D-Leu(5)]-enkephalin (DADLE), and decreased the potency and efficacy of 4-[(R)-[(2S,5R)-4-allyl-2,5-dimethylpiperazin-1-yl](3-methoxyphenyl)methyl]-N,N-diethylbenzamide (SNC80) to inhibit prostaglandin E(2) (PGE(2))-stimulated adenylyl cyclase activity. In vivo, nor-BNI enhanced the effect of DPDPE and decreased the effect of SNC80 to inhibit PGE(2)-stimulated thermal allodynia. In contrast to nor-BNI, the KOR antagonist 5'-guanidinonaltrindole (5'-GNTI) reduced the response of DPDPE both in cultured neurons and in vivo. Evidence for DOR-KOR heteromers in peripheral sensory neurons included coimmunoprecipitation of DOR with KOR, a DOR-KOR heteromer selective antibody augmented the antinociceptive effect of DPDPE in vivo, and the DOR-KOR heteromer agonist 6'-GNTI inhibited adenylyl cyclase activity in vitro as well as PGE(2)-stimulated thermal allodynia in vivo. Taken together, these data suggest that DOR-KOR heteromers exist in rat primary sensory neurons and that KOR antagonists can act as modulators of DOR agonist responses most likely through allosteric interactions between the protomers of the DOR-KOR heteromer.

Regulation of κ-opioid receptor signaling in peripheral sensory neurons in vitro and in vivo.

There is considerable interest in understanding the regulation of peripheral opioid receptors to avoid central nervous system side effects associated with systemically administered opioid analgesics. Here, we investigated the regulation of the κ-opioid receptor (KOR) on rat primary sensory neurons in vitro and in a rat model of thermal allodynia. Under basal conditions, application of the KOR agonist trans-(1S,2S)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]benzeneacetamide hydrochloride hydrate (U50488) did not inhibit adenylyl cyclase (AC) activity nor release of calcitonin gene-related peptide (CGRP) in vitro and did not inhibit thermal allodynia in vivo. However, after 15-min pretreatment with bradykinin (BK), U50488 became capable of inhibiting AC activity, CGRP release, and thermal allodynia. Inhibition of AC by 5-hydroxytryptamine 1 or neuropeptide Y(1) receptor agonists and stimulation of extracellular signal-regulated kinase activity by U50488 did not require BK pretreatment. The effect of U50488 in BK-primed tissue was blocked by the KOR antagonist nor-binaltorphimine both in vitro and in vivo. The effect of BK in vitro was blocked by either indomethacin or bisindolylmaleimide, suggesting that an arachidonic acid (AA) metabolite and protein kinase C (PKC) activation mediate BK-induced regulation of the KOR system. Furthermore, the effect of U50488 in BK-treated tissue was blocked by a soluble integrin-blocking peptide (GRGDSP), but not the inactive reverse sequence peptide (GDGRSP), suggesting that, in addition to AA and PKC, RGD-binding integrins participate in the regulation of KOR signaling in response to U50488. Understanding the mechanisms by which peripheral KOR agonist efficacy is regulated may lead to improved pharmacotherapy for the treatment of pain with reduced adverse effects.