Programming of Distinct Chemokine-Dependent and -Independent Search Strategies for Th1 and Th2 Cells Optimizes Function at Inflamed Sites.

T-helper (Th) cell differentiation drives specialized gene programs that dictate effector T cell function at sites of infection. Here, we have shown Th cell differentiation also imposes discrete motility gene programs that shape Th1 and Th2 cell navigation of the inflamed dermis. Th1 cells scanned a smaller tissue area in a G protein-coupled receptor (GPCR) and chemokine-dependent fashion, while Th2 cells scanned a larger tissue area independent of GPCR signals. Differential chemokine reliance for interstitial migration was linked to STAT6 transcription-factor-dependent programming of integrin αVß3 expression: Th2 cell differentiation led to high αVß3 expression relative to Th1 cells. Th1 and Th2 cell modes of motility could be switched simply by manipulating the amount of αVß3 on the cell surface. Deviating motility modes from those established during differentiation impaired effector function. Thus, programmed expression of αVß3 tunes effector T cell reliance on environmental cues for optimal exploration of inflamed tissues.

The Integrin LFA-1 Controls T Follicular Helper Cell Generation and Maintenance.

T follicular helper (Tfh) cells are a CD4+ T cell subset critical for long-lived humoral immunity. We hypothesized that integrins play a decisive role in Tfh cell biology. Here we show that Tfh cells expressed a highly active form of leukocyte function-associated antigen-1 (LFA-1) that was required for their survival within the germinal center niche. In addition, LFA-1 promoted expression of Bcl-6, a transcriptional repressor critical for Tfh cell differentiation, and inhibition of LFA-1 abolished Tfh cell generation and prevented protective humoral immunity to intestinal helminth infection. Furthermore, we demonstrated that expression of Talin-1, an adaptor protein that regulates LFA-1 affinity, dictated Tfh versus Th2 effector cell differentiation. Collectively, our results define unique functions for LFA-1 in the Tfh cell effector program and suggest that integrin activity is important in lineage decision-making events in the adaptive immune system.

Differential Enantiomer-Specific Signaling of Cannabidiol at CB1 Receptors.

The two main constituents of cannabis are Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD). While Δ9-THC pharmacology has been studied extensively, CBD-long considered inactive-is now the subject of vigorous research related to epilepsy, pain, and inflammation and is popularly embraced as a virtual cure-all. However, our understanding of CBD pharmacology remains limited, although CBD inhibits cannabinoid CB1 receptor signaling, likely as a negative allosteric modulator. Cannabis synthesizes (-)-CBD, but CBD can also exist as an enantiomer, (+)-CBD. We enantioselectively synthesized both CBD enantiomers using established conditions and describe here a new, practical, and reliable, NMR-based method for confirming the enantiomeric purity of two CBD enantiomers. We also investigated the pharmacology of (+)-CBD in autaptic hippocampal neurons, a well-characterized neuronal model of endogenous cannabinoid signaling, and in CHO-K1 cells. We report the inhibition constant for displacing CP55,940 at CB1 by (+)-CBD, is 5-fold lower than (-)-CBD. We find that (+)-CBD is ∼10 times more potent at inhibiting depolarization-induced suppression of excitation (DSE), a form of endogenous cannabinoid-mediated retrograde synaptic plasticity. (+)-CBD also inhibits CB1 suppression of cAMP accumulation but with less potency, indicating that the signaling profiles of the enantiomers differ in a pathway-specific manner. In addition, we report that (+)-CBD stereoselectively and potently activates the sphingosine-1 phosphate (S1P) receptors, S1P1 and S1P3 These results provide an attractive method for synthesizing and distinguishing enantiomers of CBD and related phytocannabinoids and provide further evidence that these enantiomers have their own unique and interesting signaling properties. SIGNIFICANCE STATEMENT: Cannabidiol (CBD) is the subject of considerable scientific and popular interest, but we know little of the enantiomers of CBD. We find that the enantiomer (+)-CBD is substantially more potent inhibitor of cannabinoid CB1 receptors and that it activates sphingosine-1-phosphate receptors in an enantiomer-specific manner; we have additionally developed an improved method for the synthesis of enantiomers of CBD and related compounds.

Pivotal role for αV integrins in sustained Tfh support of the germinal center response for long-lived plasma cell generation.

CD4+ follicular helper T cells (Tfh) are essential for germinal center (GC) reactions in the lymph node that generate high-affinity, long-lived plasma cells (LLPCs). Temporal GC analysis suggests B memory cells (Bmem) are generated early, while LLPCs are generated late in the GC reaction. Distinct roles for Tfh at these temporally different stages are not yet clear. Tfh entry into the GC is highly dynamic and the signals that maintain Tfh within the GC for support of late LLPC production are poorly understood. The GC is marked by inflammation-induced presentation of specific ECM components. To determine if T cell recognition of these ECM components played a role in Tfh support of the GC, we immunized mice with a T cell-restricted deletion of the ECM-binding integrin αV (αV-CD4 cKO). T cell integrin αV deletion led to a striking defect in the number and size of the GCs following immunization with OVA protein in complete Freund's adjuvant. The GC defect was not due to integrin αV deficiency impeding Tfh generation or follicle entry or the ability of αV-CD4 cKO Tfh to contact and support B cell activation. Instead, integrin αV was essential for T cell-intrinsic accumulation within the GC. Altered Tfh positioning resulted in lower-affinity antibodies and a dramatic loss of LLPCs. Influenza A infection revealed that αV integrin was not required for Tfh support of Bmem but was essential for Tfh support of LLPCs. We highlight an αV integrin-ECM-guided mechanism of Tfh GC accumulation that selectively impacts GC output of LLPCs but not Bmem.

Non-shoulder Injuries in Swimming: A Systematic Review.

OBJECTIVE: The shoulder is the most common injury in swimming, followed by the knee and spine. The repetitive nature of swimming training may predispose a swimmer to injury. Several risk factors have been previously identified, but the level of evidence and level of certainty that these risk factors predispose a swimmer to injury risk has yet to be evaluated critically in a systematic review. DESIGN: Systematic review and meta-analysis following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. DATA SOURCES: Data were obtained through PubMed, Cochrane, and Google Scholar. The database search was limited to articles that were published between January 1980 and December 2019. ELIGIBILITY CRITERIA FOR SELECTING STUDIES: Level I, II, and III studies were included in this review. All studies must have been conducted in swimmers or swimming, a description of the specific pathology and provide at least one identified risk factor with an association (P < 0.05). RESULTS: A total of 19 critically appraised articles identified 28 potential risk factors for musculoskeletal injuries in swimmers. The risk factors were grouped by the anatomical region: neck and back, pelvis and hip, and the knee. Only 2 risk factors were appraised at a moderate level of certainty, both occurring in the knee. The remaining 26 identified risk factors were appraised at a low level of certainty. There is a clear lack of research surrounding nonshoulder injuries in swimmers.

Beneath the Surface: Mental Health and Harassment and Abuse of Athletes Participating in the FINA (Aquatics) World Championships, 2019.

OBJECTIVE: To assess the mental health and experience of sport-related harassment and abuse of elite aquatic athletes and to analyze it in relation to gender and discipline. DESIGN: Cross-sectional study using an anonymous survey. SETTING: The FINA World Championships 2019. PARTICIPANTS: Registered athletes in the disciplines of swimming, diving, high diving, water polo, artistic swimming, and open water swimming. INTERVENTIONS: Athletes completed an online or paper-based questionnaire. MAIN OUTCOME MEASURES: The main outcome measures included screening for depression (10-item version of the Center for Epidemiological Studies Depression Scale revised), eating disorders (Brief Eating Disorders in Athletes Questionnaire), the subjective need for psychotherapeutic support, and the experience of sport-related harassment and/or abuse. RESULTS: A quarter (24.6%) of the 377 responding athletes were classified as depressed and 2.5% as having an eating disorder. More than 40% of the athletes stated that they wanted or needed psychotherapeutic support. Fifty-one athletes (14.9%) had experienced harassment/abuse in sport themselves, and 31 (9%) had witnessed it in another athlete. The experiences of harassment and abuse ranged from unwanted comments about body or appearance (40.2%) to rewards in sport for sexual favors (2.5%) and rape (0.3%). Athletes who had experienced harassment/abuse in sport themselves had higher average scores for depression and eating disorders, and more of them felt they needed psychotherapeutic support. Up to a third would not talk or report to anybody if they saw or experienced harassment/abuse, and less than 20% would talk to an official for help. CONCLUSION: Targeted initiatives are required to address mental health issues and harassment and abuse in sport in the FINA aquatic disciplines.

An Uneven Playing Field: Athlete Injury, Illness, Load, and Daily Training Environment in the Year Before the FINA (Aquatics) World Championships, 2017.

OBJECTIVE: To assess athlete's training environment and health problems before the FINA World Championships (WC) (2017) and to analyze the differences between the 6 disciplines, gender, and countries with different medal rankings during the championships. DESIGN: Retrospective anonymous questionnaire. SETTING: FINA WC 2017. PARTICIPANTS: Registered aquatic athletes (swimmers, divers, high divers, water polo players, artistic swimmers, open water swimmers). MAIN OUTCOME MEASURES: The outcome measures included the following: training and competition load, availability of support staff, performance of injury prevention exercises, and prevalence of health problems. The independent variables included gender, aquatic discipline, and country group based on medal ranking. RESULTS: In the 12 months preceding the Championships, 67% of the athletes reported physical complaints and 41% trained or competed with a diagnosed injury. Only half of the athletes reported that injury prevention exercises were always (29.9%) or often (23.4%) a regular part of their daily training. In the daily training environment, support staff (excluding coach) was not available or available only if the athlete pays in 28.4% (specialized trainer) to 58.9% (sport scientist) of cases. About one-quarter of the athletes rated the support offered by their National Team program as poor or very poor. There were discrepancies in training loads and support staff among the 6 aquatic disciplines. Availability of support staff, athlete's satisfaction with their training environment/support, and regular use of injury prevention exercises were higher in countries that ranked higher in the medal list. CONCLUSIONS: Injury prevention strategies should be promoted in aquatic sports, with prospective surveillance for the early identification of physical complaints. The facilitation of access to sport-specific experts could improve athlete's health and performance, especially in countries with low medal ranking.

An Evaluation of Understudied Phytocannabinoids and Their Effects in Two Neuronal Models.

Cannabis contains more than 100 phytocannabinoids. Most of these remain poorly characterized, particularly in neurons. We tested a panel of five phytocannabinoids-cannabichromene (CBC), cannabidiolic acid (CBDA), cannabidivarin (CBDV), cannabidivarinic acid (CBDVA), and Δ9-tetrahydrocannabivarin (THCV) in two neuronal models, autaptic hippocampal neurons and dorsal root ganglion (DRG) neurons. Autaptic neurons expressed a form of CB1-dependent retrograde plasticity while DRGs expressed a variety of transient receptor potential (TRP) channels. CBC, CBDA, and CBDVA had little or no effect on neuronal cannabinoid signaling. CBDV and THCV differentially inhibited cannabinoid signaling. THCV inhibited CB1 receptors presynaptically while CBDV acted post-synaptically, perhaps by inhibiting 2-AG production. None of the compounds elicited a consistent DRG response. In summary, we find that two of five 'minor' phytocannabinoids tested antagonized CB1-based signaling in a neuronal model, but with very different mechanisms. Our findings highlight the diversity of potential actions of phytocannabinoids and the importance of fully evaluating these compounds in neuronal models.

CD4 T Cell Epitope Specificity and Cytokine Potential Are Preserved as Cells Transition from the Lung Vasculature to Lung Tissue following Influenza Virus Infection.

Pulmonary CD4 T cells are critical in respiratory virus control, both by delivering direct effector function and through coordinating responses of other immune cells. Recent studies have shown that following influenza virus infection, virus-specific CD4 T cells are partitioned between pulmonary vasculature and lung tissue. However, very little is known about the peptide specificity or functional differences of CD4 T cells within these two compartments. Using a mouse model of influenza virus infection in conjunction with intravascular labeling in vivo, the cell surface phenotype, epitope specificity, and functional potential of the endogenous polyclonal CD4 T cell response was examined by tracking nine independent CD4 T cell epitope specificities. These studies revealed that tissue-localized CD4 cells were globally distinct from vascular cells in expression of markers associated with transendothelial migration, residency, and micropositioning. Despite these differences, there was little evidence for remodeling of the viral epitope specificity or cytokine potential as cells transition from vasculature to the highly inflamed lung tissue. Our studies also distinguished cells in the pulmonary vasculature from peripheral circulating CD4 T cells, providing support for the concept that the pulmonary vasculature does not simply reflect circulating cells that are trapped within the narrow confines of capillary vessels but rather is enriched in transitional cells primed in the draining lymph node that have specialized potential to enter the lung tissue.IMPORTANCE CD4 T cells convey a multitude of functions in immunity to influenza, including those delivered in the lymph node and others conveyed by CD4 T cells that leave the lymph node, enter the blood, and extravasate into the lung tissue. Here, we show that the transition of recently primed CD4 cells detected in the lung vasculature undergo profound changes in expression of markers associated with tissue localization as they establish residence in the lung. However, this transition does not edit CD4 T cell epitope specificity or the cytokine potential of the CD4 T cells. Thus, CD4 T cells that enter the infected lung can convey diverse functions and have a sufficiently broad viral antigen specificity to detect the complex array of infected cells within the infected tissue, offering the potential for more effective protective function.

Analysis of water polo injuries during 8904 player matches at FINA World Championships and Olympic games to make the sport safer.

OBJECTIVE: To analyse injuries of water polo players reported during four Summer Olympic Games (2004, 2008, 2012, 2016) and four Federation Internationale de Natation World Championships (2009, 2013, 2015, 2017). METHODS: Injuries during training and matches were reported daily by the team physicians and the local medical staff at the sports venues using an established surveillance system. RESULTS: A total of 381 injuries were reported, equivalent to 14.1 injuries per 100 players (95% CI ±1.42). The most frequent diagnoses were laceration (12.7%) and contusion (10.9%) of head, followed by (sub-)luxation/sprain of hand (9.5%) and contusion of trunk (6.5%) or hand (6.2%). More than half of the injuries (57.0%) occurred due to contact with another player. A quarter of the injuries (25.4%) were expected to result in absence from training or match; 10 (2.9%) resulted in an estimated time-loss of 3 or more weeks. About three-quarters of injuries (75.6%) occurred during matches, 86 during training. The incidence of match injuries was on average 56.2 injuries per 1000 match hours (95% CI ±6.74). The incidence of time-loss match injuries (14.7; 95% CI ±3.44) was significantly higher in men than in women. CONCLUSIONS: A critical review of water polo in-competition rules and the implementation of a Fair Play programme may help to mitigate the high incidence of contact injuries incurred during matches. A water polo-specific concussion education programme including recognition, treatment and return to play is recommended. Finally, a prospective injury surveillance programme would help to better define water polo injuries outside of the competition period.

Regulatory T Cell Numbers in Inflamed Skin Are Controlled by Local Inflammatory Cues That Upregulate CD25 and Facilitate Antigen-Driven Local Proliferation.

CD4(+)Foxp3(+) regulatory T cells (Tregs) are key immune suppressors that regulate immunity in diverse tissues. The tissue and/or inflammatory signals that influence the magnitude of the Treg response remain unclear. To define signals that promote Treg accumulation, we developed a simple system of skin inflammation using defined Ags and adjuvants that induce distinct cytokine milieus: OVA protein in CFA, aluminum salts (Alum), and Schistosoma mansoni eggs (Sm Egg). Polyclonal and Ag-specific Treg accumulation in the skin differed significantly between adjuvants. CFA and Alum led to robust Treg accumulation, with >50% of all skin CD4(+) T cells being Foxp3(+) In contrast, Tregs accumulated poorly in the Sm Egg-inflamed skin. Surprisingly, we found no evidence of inflammation-specific changes to the Treg gene program between adjuvant-inflamed skin types, suggesting a lack of selective recruitment or adaptation to the inflammatory milieu. Instead, Treg accumulation patterns were linked to differences in CD80/CD86 expression by APC and the regulation of CD25 expression, specifically in the inflamed skin. Inflammatory cues alone, without cognate Ag, differentially supported CD25 upregulation (CFA and Alum > Sm Egg). Only in inflammatory milieus that upregulated CD25 did the provision of Ag enhance local Treg proliferation. Reduced IL-33 in the Sm Egg-inflamed environment was shown to contribute to the failure to upregulate CD25. Thus, the magnitude of the Treg response in inflamed tissues is controlled at two interdependent levels: inflammatory signals that support the upregulation of the important Treg survival factor CD25 and Ag signals that drive local expansion.

Injury and illness in aquatic sport: how high is the risk? A comparison of results from three FINA World Championships.

BACKGROUND: Epidemiological information on injury/illness is required to develop effective injury prevention strategies. AIM: To assess the frequency and characteristics of injuries/illnesses (1) in the 4 weeks prior to and (2) during the Fédération Internationale de Natation (FINA) World Championships 2015 compared with 2013 and 2009. METHOD: (1) Athletes answered a retrospective questionnaire, and (2) the medical staff reported injuries/illnesses prospectively during the championships. RESULTS: (1) A quarter of responding athletes reported symptoms in the 4 weeks prior to the championships. More than half of all affected athletes presented with substantial severity, 80% took medication, 70% had overuse injuries and 30% did not modify their training regime despite symptoms. At the start of the championships, 70% of affected participants were still symptomatic. (2) During the championships, injury and illness incidence was 12.9 per 100 athletes. The most common injuries were shoulder sprains (5.7%) and muscle cramps of the lower back (5.7%). The most common illnesses were infections of the respiratory (33.9%) and gastrointestinal tract (23.5%). Risk factors included discipline and age, but not gender. Incidence was highest in athletes competing in high diving (HD), water polo (WP) and diving (DIV) for injuries, and WP and swimming (SW) for illnesses. The significantly higher incidence of injuries and illnesses at the FINA World Championships 2015 compared with 2013 and 2009 was most probably due to a similarly improved response rate of the medical staff. CONCLUSIONS: In aquatic sports, surveillance and health promotion should focus on prevention of out-of-competition overuse injuries and athlete education.

Influence of Cattle Trails on Runoff Quantity and Quality.

Cattle trails in grazed pastures close to rivers may adversely affect surface water quality of the adjacent river by directing runoff to it. The objective of this 3-yr study (2013-2015) in southern Alberta, Canada, was to determine if cattle trails significantly increased the risk of runoff and contaminants (sediment, nutrients) compared with the adjacent grazed pasture (control). A portable rainfall simulator was used to generate artificial rainfall (140 mm h) and runoff. The runoff properties measured were time to runoff and initial abstraction (infiltration), total runoff depth and average runoff rates, as well as concentrations and mass loads of sediment, N, and P fractions. Cattle trails significantly ( ≤ 0.10) decreased time to runoff and initial abstraction (26-32%) in the 2 yr measured and increased total runoff depth, runoff coefficients, and average runoff rates (21-51%) in 2 of 3 yr. Concentrations of sediment, N, and P fractions in runoff were not significantly greater for cattle trails than for control areas. However, mass loads of total suspended solids (57-85% increase), NH-N (31-90%), and dissolved reactive P (DRP) (30-92%) were significantly greater because of increased runoff volumes. Overall, runoff quantity and loads of sediment, NH-N, and DRP were greater for cattle trails compared with the adjacent grazed pasture, and hydrologic connection with cattle-access sites on the riverbank suggests that this could adversely affect water quality in the adjacent river. Extrapolation of the study results should be tempered by the specific conditions represented by this rainfall simulation study.

Influence of Long-term Application of Feedlot Manure Amendments on Water Repellency of a Clay Loam Soil.

Long-term application of feedlot manure to cropland may increase the quantity of soil organic carbon (C) and change its quality, which may influence soil water repellency. The objective was to determine the influence of feedlot manure type (stockpiled vs. composted), bedding material (straw [ST] vs. woodchips [WD]), and application rate (13, 39, or 77 Mg ha) on repellency of a clay loam soil after 17 annual applications. The repellency was determined on all 14 treatments using the water repellency index ( index), the water drop penetration time (WDPT) method, and molarity of ethanol (MED) test. The C composition of particulate organic matter in soil of five selected treatments after 16 annual applications was also determined using C nuclear magnetic resonance-direct polarization with magic-angle spinning (NMR-DPMAS). Manure type had no significant ( > 0.05) effect on index and WDPT, and MED classification was similar. Mean index and WDPT values were significantly greater and MED classification more hydrophobic for WD than ST. Application rate had no effect on the index, but WDPT was significantly greater and MED classification more hydrophobic with increasing application rate. Strong ( > 0.7) but nonsignificant positive correlations were found between index and WDPT versus hydrophobic (alkyl + aromatic) C, lignin at 74 ppm (O-alkyl), and unspecified aromatic compounds at 144 ppm. Specific aromatic compounds also contributed more to repellency than alkyl, O-alkyl, and carbonyl compounds. Overall, all three methods consistently showed that repellency was greater for WD- than ST-amended clay loam soil, but manure type had no effect.

Response of Soil Mesofauna to Long-Term Application of Feedlot Manure on Irrigated Cropland.

Long-term application of feedlot manure to cropland may influence soil mesofauna. These organisms affect the health, structure, and fertility of soils, organic matter decomposition, and crop growth. The objective was to study the long-term (16-17 yr) influence of feedlot manure type and bedding on soil mesofauna over 2 yr (2014-2015). Stockpiled or composted feedlot manure with straw (ST) or wood-chip (WD) bedding (plus unamended control) was annually applied (13 Mg ha dry wt.) to an irrigated clay loam soil with continuous barley (). Intact cores were taken from surface (0-5 cm) soil in the fall, and the densities of Acari (mites) suborders and Collembola (springtails) families were determined. Manure type had no significant ( > 0.05) effect on soil mesofauna density. In contrast, there was a significant two- to sixfold increase in density with WD- compared with ST-amended soils of total Acari in 2014 and 2015, as well as total Collembola, total Acari and Collembola, oribatid mites, and entomobryid springtails in 2014. The bedding effect was attributed to significantly greater soil water content and lower bulk density for WD than ST. Density of soil mesofauna was not significantly greater in amended soils than in unamended soils. A shift by feedlot producers from stockpiled to composted feedlot manure application should have no effect on soil mesofauna density, whereas a shift from ST to WD bedding may increase the density of certain soil mesofauna, which may have a beneficial effect on soil.

Intraspecific phytochemical variation shapes community and population structure for specialist caterpillars.

Chemically mediated plant-herbivore interactions contribute to the diversity of terrestrial communities and the diversification of plants and insects. While our understanding of the processes affecting community structure and evolutionary diversification has grown, few studies have investigated how trait variation shapes genetic and species diversity simultaneously in a tropical ecosystem. We investigated secondary metabolite variation among subpopulations of a single plant species, Piper kelleyi (Piperaceae), using high-performance liquid chromatography (HPLC), to understand associations between plant phytochemistry and host-specialized caterpillars in the genus Eois (Geometridae: Larentiinae) and associated parasitoid wasps and flies. In addition, we used a genotyping-by-sequencing approach to examine the genetic structure of one abundant caterpillar species, Eois encina, in relation to host phytochemical variation. We found substantive concentration differences among three major secondary metabolites, and these differences in chemistry predicted caterpillar and parasitoid community structure among host plant populations. Furthermore, E. encina populations located at high elevations were genetically different from other populations. They fed on plants containing high concentrations of prenylated benzoic acid. Thus, phytochemistry potentially shapes caterpillar and wasp community composition and geographic variation in species interactions, both of which can contribute to diversification of plants and insects.

Mutation of putative GRK phosphorylation sites in the cannabinoid receptor 1 (CB1R) confers resistance to cannabinoid tolerance and hypersensitivity to cannabinoids in mice.

For many G-protein-coupled receptors (GPCRs), including cannabinoid receptor 1 (CB1R), desensitization has been proposed as a principal mechanism driving initial tolerance to agonists. GPCR desensitization typically requires phosphorylation by a G-protein-coupled receptor kinase (GRK) and interaction of the phosphorylated receptor with an arrestin. In simple model systems, CB1R is desensitized by GRK phosphorylation at two serine residues (S426 and S430). However, the role of these serine residues in tolerance and dependence for cannabinoids in vivo was unclear. Therefore, we generated mice where S426 and S430 were mutated to nonphosphorylatable alanines (S426A/S430A). S426A/S430A mutant mice were more sensitive to acutely administered delta-9-tetrahydrocannabinol (Δ(9)-THC), have delayed tolerance to Δ(9)-THC, and showed increased dependence for Δ(9)-THC. S426A/S430A mutants also showed increased responses to elevated levels of endogenous cannabinoids. CB1R desensitization in the periaqueductal gray and spinal cord following 7 d of treatment with Δ(9)-THC was absent in S426A/S430A mutants. Δ(9)-THC-induced downregulation of CB1R in the spinal cord was also absent in S426A/S430A mutants. Cultured autaptic hippocampal neurons from S426A/S430A mice showed enhanced endocannabinoid-mediated depolarization-induced suppression of excitation (DSE) and reduced agonist-mediated desensitization of DSE. These results indicate that S426 and S430 play major roles in the acute response to, tolerance to, and dependence on cannabinoids. Additionally, S426A/S430A mice are a novel model for studying pathophysiological processes thought to involve excessive endocannabinoid signaling such as drug addiction and metabolic disease. These mice also validate the approach of mutating GRK phosphorylation sites involved in desensitization as a general means to confer exaggerated signaling to GPCRs in vivo.

Competing with injuries: injuries prior to and during the 15th FINA World Championships 2013 (aquatics).

BACKGROUND: Injury and illness surveillance is the foundation for the development of prevention strategies. OBJECTIVE: To examine injuries among the aquatic disciplines in the 4 weeks prior to and during the 2013 FINA World Championships. METHODS: The study was comprised of two components: (1) a retrospective athlete survey recording injuries in the 4 weeks prior to the Championships and (2) a prospective recording of injuries and illnesses by the medical teams of the participating countries and the local host medical team. RESULTS: One-third of the 1116 responding athletes reported an injury/physical complaint in the 4 weeks prior to the Championships. Significantly more women (36.7%) than men (28.6%) reported injuries. Divers reported the highest rate of injury/physical complaints (55.7%). At the start of the Championships, 70% of injured respondents (n=258) were still symptomatic; however, full participation was expected by 76%. During the Championships, 186 new injuries were reported (8.3/100 registered athletes) with the highest injury incidence rate in water polo (15.3/100 registered athletes). The most common injured body part was the shoulder (21%). A total of 199 illnesses were reported during the Championships (9.0/100 registered athletes) with the most common diagnosis of illness being gastrointestinal infection. Environmental exposure (allergy, otitis and jellyfish stings) was responsible for 27% of all illnesses in open water swimming. CONCLUSIONS: Injuries pose a significant health risk for elite aquatic athletes. A prospective study would improve understanding of out-of-competition injuries. Future injury and illness surveillance at FINA World Championships is required to direct and measure the impact of prevention strategies.

Optimization of a cisplatin model of chemotherapy-induced peripheral neuropathy in mice: use of vitamin C and sodium bicarbonate pretreatments to reduce nephrotoxicity and improve animal health status.

BACKGROUND: Cisplatin, a platinum-derived chemotherapeutic agent, produces antineoplastic effects coupled with toxic neuropathic pain and impaired general health status. These side-effects complicate long term studies of neuropathy or analgesic interventions in animals. We recently demonstrated that pretreatment with sodium bicarbonate (4% NaHCO3) prior to cisplatin (3 mg/kg i.p. weekly up to 5 weeks) was associated with improved health status (i.e. normal weight gain, body temperature, creatinine and ketone levels, and kidney weight ratio) in rats (Neurosci Lett 544:41-46, 2013). To reduce the nephrotoxic effects of cisplatin treatment in mice, we compared effects of sodium bicarbonate (4% NaHCO3 s.c.), vitamin C (25 mg/kg s.c.), resveratrol (25 mg/kg s.c.) and saline (0.9% NaCl) pretreatment on cisplatin-induced changes in animal health status, neuropathic pain and proinflammatory cytokine levels in spinal cord and kidney. RESULTS: Cisplatin-treated mice receiving saline pretreatment exhibited elevated ketone, creatinine and kidney weight ratios, representative of nephrotoxicity. Vitamin C and sodium bicarbonate lowered creatinine/ketone levels and kidney weight ratio whereas resveratrol normalized creatinine levels and kidney weight ratios similar to saline pretreatment. All pretreatments were associated with decreased ketone levels compared to saline pretreatment. Cisplatin-induced neuropathy (i.e. mechanical and cold allodynia) developed equivalently in all pretreatment groups and was similarly reversed by either morphine (6 mg/kg i.p.) or ibuprofen (6 mg/kg i.p.) treatment. RT-PCR showed that mRNA levels for IL-1ß were increased in lumbar spinal cord of cisplatin-treated groups pretreated with either saline, NaHCO3 or resveratrol/cisplatin-treated groups. However, IL-6 and TNF-alpha were elevated in the kidneys in all cisplatin-treated groups. Our studies also demonstrate that 60 days after the last cisplatin treatment, body weight, body temperature, kidney functions and mRNA levels have returned to baseline although the neuropathic pain (mechanical and cold) is maintained. CONCLUSIONS: Studies employing cisplatin should include NaHCO3 or vitamin C pretreatment to improve animal health status and reduce nephrotoxicity (lower creatinine and kidney weight ratio) without affecting the development of chemotherapy-induced neuropathy or analgesic efficacy.

Nitrogen and phosphorus in runoff from cattle manure compost windrows of different maturities.

Manure composting has become commonplace in the beef cattle ( L.) feedlot industry in Alberta. However, the nitrogen (N) and phosphorus (P) characteristics of runoff from windrows subjected to heavy rainfall at different compost maturities are unknown. On Days 18, 26, 40, 54, 81, 109, and 224 of composting, a rainfall simulator generated runoff, which was collected in timed 5-L increments, creating the variable "time during runoff event" (TDRE). The volumetric runoff coefficient of windrows increased from 24% of incident rainfall on Day 0 to 69% by Day 90. Ammonium-nitrogen showed a significant maturity × TDRE interaction on Day 18, increasing from 46 mg L for the 0- to 5-L increment to 172 mg L for the 25- to 30-L increment, as did total dissolved phosphorus (TDP), increasing from 36 to 61 mg L. Nitrate-nitrogen had a runoff export coefficient of 19.5 mg m min on Day 224, which was significantly higher than 1.8 to 6.3 mg m min on Days 18 to 54. Across the 224-d composting period, compost NO-N concentration explained 87% of runoff NO-N, whereas compost water-soluble P explained 68% of runoff TDP. The occurrence and duration of rainfall events relative to the compost maturity spectrum has implications for the magnitude of N and P mobility and overall nutrient losses.