Anti-nociceptive and desensitizing effects of olvanil on capsaicin-induced thermal hyperalgesia in the rat

Background: Olvanil (NE 19550) is a non-pungent synthetic analogue of capsaicin, the natural pungent ingredient of capsicum which activates the transient receptor potential vanilloid type-1 (TRPV1) channel and was developed as a potential analgesic compound. Olvanil has potent anti-hyperalgesic effects in several experimental models of chronic pain. Here we report the inhibitory effects of olvanil on nociceptive processing using cultured dorsal root ganglion (DRG) neurons and compare the effects of capsaicin and olvanil on thermal nociceptive processing in vivo; potential contributions of the cannabinoid CB1 receptor to olvanil’s anti-hyperalgesic effects were also investigated. Methods: A hot plate analgesia meter was used to evaluate the anti-nociceptive effects of olvanil on capsaicin-induced thermal hyperalgesia and the role played by CB1 receptors in mediating these effects. Single cell calcium imaging studies of DRG neurons were employed to determine the desensitizing effects of olvanil on capsaicin-evoked calcium responses. Statistical analysis used Student’s t test or one way ANOVA followed by Dunnett’s post-hoctest as appropriate. Results: Both olvanil (100 nM) and capsaicin (100 nM) produced significant increases in intracellular calcium concentrations [Ca2+]I in cultured DRG neurons. Olvanil was able to des ensitise TRPV1 responses to further capsaicin exposure more effectively than capsaicin. Intra plantar injection of capsaicin (0.1, 0.3 and 1μg) produced a robust TRPV1-dependant thermal hyperalgesia in rats, whilst olvanil (0.1, 0.3 and 1μg) produced no hyperalgesia, emphasizing its lack of pungency. The highest dose of olvanil significantly reduced the hyperalgesic effects of capsaicin in vivo. Intraplantar injection of the selective cannabinoid CB1 receptor antagonist rimonabant (1μg) altered neither capsaicin-induced thermal hyperalgesia nor the desensitizing properties of olvanil, indicating a lack of involvement of CB1receptors. Conclusions: Olvanil is effective in reducing capsaicin-induced thermal hyperalgesia, probably via directly desensitizingTRPV1 channels in a CB 1 receptor-independent fashion. The results presented clearly support the potential for olvanil in the development of new topical analgesic preparations for treating chronic pain conditions while avoiding the unwanted side effects of capsaicin treatments

Dental management considerations for the patient with an acquired coagulopathy. Part 1: Coagulopathies from systemic disease

Current teaching suggests that many patients are at risk for prolonged bleeding during and following invasive dental procedures, due to an acquired coagulopathy from systemic disease and/or from medications. However, treatment standards for these patients often are the result of long-standing dogma with little or no scientific basis. The medical history is critical for the identification of patients potentially at risk for prolonged bleeding from dental treatment. Some time-honoured laboratory tests have little or no use in community dental practice. Loss of functioning hepatic, renal, or bone marrow tissue predisposes to acquired coagulopathies through different mechanisms, but the relationship to oral haemostasis is poorly understood. Given the lack of established, science-based standards, proper dental management requires an understanding of certain principles of pathophysiology for these medical conditions and a few standard laboratory tests. Making changes in anticoagulant drug regimens are often unwarranted and/or expensive, and can put patients at far greater risk for morbidity and mortality than the unlikely outcome of postoperative bleeding. It should be recognised that prolonged bleeding is a rare event following invasive dental procedures, and therefore the vast majority of patients with suspected acquired coagulopathies are best managed in the community practice setting

The web-based ASSO-food frequency questionnaire for adolescents: relative and absolute reproducibility assessment

Background: A new food frequency questionnaire (FFQ) has been recently developed within the Italian Adolescents and Surveillance System for the Obesity prevention (ASSO) Project; it was found to be appropriate for ranking adolescents in food and nutrient levels of intake. The aim of this study was to assess the relative and absolute reproducibility of the ASSO-FFQ for 24 food groups, energy and 52 nutrients. Methods: A test-retest study was performed on two ASSO-FFQs administered one month apart of each other to 185 adolescents, aged 14–17 and attending secondary schools in Palermo (Italy). Wilcoxon test assessed differences in median daily intakes between the two FFQs. Agreement was evaluated by quintiles comparison and weighted kappa. Intraclass Correlation Coefficients (ICC) and Bland-Altman method assessed the relative and absolute reliability respectively. Results: Significant difference (p < 0.05) in median intakes was found only for bread substitutes, savoury food, water, soft drinks, carbohydrates and sugar. The subjects classified into the same or adjacent quintiles for food groups ranged from 62% (white bread) to 91% (soft drinks); for energy and nutrients from 64% (polyunsaturated fatty acids) to 90% (ethanol). Mean values of weighted kappa were 0.47 and 0.48, respectively for food groups and nutrients. Fair to good ICC values (>0.40) were assessed for thirteen food groups, energy and forty-three nutrients. Limits of Agreement were narrow for almost all food groups and all nutrients. Conclusions: The ASSO-FFQ is a reliable instrument for estimating food groups, energy and nutrients intake in adolescents

Human keratinocytes are vanilloid resistant

BACKGROUND: Use of capsaicin or resiniferatoxin (RTX) as analgesics is an attractive therapeutic option. RTX opens the cation channel inflammatory pain/vanilloid receptor type 1 (TRPV1) permanently and selectively removes nociceptive neurons by Ca(2+)-cytotoxicity. Paradoxically, not only nociceptors, but non-neuronal cells, including keratinocytes express full length TRPV1 mRNA, while patient dogs and experimental animals that underwent topical treatment or anatomically targeted molecular surgery have shown neither obvious behavioral, nor pathological side effects. METHODS: To address this paradox, we assessed the vanilloid sensitivity of the HaCaT human keratinocyte cell line and primary keratinocytes from skin biopsies. RESULTS: Although both cell types express TRPV1 mRNA, neither responded to vanilloids with Ca(2+)-cytotoxicity. Only ectopic overproduction of TRPV1 rendered HaCaT cells sensitive to low doses (1-50 nM) of vanilloids. The TRPV1-mediated and non-receptor specific Ca(2+)-cytotoxicity ([RTX]>15 microM) could clearly be distinguished, thus keratinocytes were indeed resistant to vanilloid-induced, TRPV1-mediated Ca(2+)-entry. Having a wider therapeutic window than capsaicin, RTX was effective in subnanomolar range, but even micromolar concentrations could not kill human keratinocytes. Keratinocytes showed orders of magnitudes lower TRPV1 mRNA level than sensory ganglions, the bona fide therapeutic targets in human pain management. In addition to TRPV1, TRPV1b, a dominant negative splice variant was also noted in keratinocytes. CONCLUSION: TRPV1B expression, together with low TRPV1 expression, may explain the vanilloid paradox: even genuinely TRPV1 mRNA positive cells can be spared with therapeutic (up to micromolar) doses of RTX. This additional safety information might be useful for planning future human clinical trials

TRPV1 enhances the afferent response to P2X receptor activation in the mouse urinary bladder

Both TRPV1 and P2X receptors present on bladder sensory nerve fibres have been implicated in mechanosensation during bladder filling. The aim of this study was to determine possible interactions between these receptors in modulating afferent nerve activity. In wildtype (TRPV1+/+) and TRPV1 knockout (TRPV1−/−) mice, bladder afferent nerve activity, intravesical pressure, and luminal ATP and acetylcholine levels were determined and also intracellular calcium responses of dissociated pelvic DRG neurones and primary mouse urothelial cells (PMUCs). Bladder afferent nerve responses to the purinergic agonist αβMethylene-ATP were depressed in TRPV1−/− mice (p ≤ 0.001) and also in TRPV1+/+ mice treated with the TRPV1-antagonist capsazepine (10 µM; p ≤ 0.001). These effects were independent of changes in bladder compliance or contractility. Responses of DRG neuron to αβMethylene-ATP (30 µM) were unchanged in the TRPV1−/− mice, but the proportion of responsive neurones was reduced (p ≤ 0.01). Although the TRPV1 agonist capsaicin (1 µM) did not evoke intracellular responses in PMUCs from TRPV1+/+ mice, luminal ATP levels were reduced in the TRPV1−/− mice (p ≤ 0.001) compared to wildtype. TRPV1 modulates P2X mediated afferent responses and provides a mechanistic basis for the decrease in sensory symptoms observed following resiniferatoxin and capsaicin treatment for lower urinary tract symptoms

Nasal Chemosensory-Stimulation Evoked Activity Patterns in the Rat Trigeminal Ganglion Visualized by In Vivo Voltage-Sensitive Dye Imaging

Mammalian nasal chemosensation is predominantly mediated by two independent neuronal pathways, the olfactory and the trigeminal system. Within the early olfactory system, spatiotemporal responses of the olfactory bulb to various odorants have been mapped in great detail. In contrast, far less is known about the representation of volatile chemical stimuli at an early stage in the trigeminal system, the trigeminal ganglion (TG), which contains neurons directly projecting to the nasal cavity. We have established an in vivo preparation that allows high-resolution imaging of neuronal population activity from a large region of the rat TG using voltage-sensitive dyes (VSDs). Application of different chemical stimuli to the nasal cavity elicited distinct, stimulus-category specific, spatiotemporal activation patterns that comprised activated as well as suppressed areas. Thus, our results provide the first direct insights into the spatial representation of nasal chemosensory information within the trigeminal ganglion imaged at high temporal resolution

Phagocytosis depends on TRPV2-mediated calcium influx and requires TRPV2 in lipids rafts: alteration in macrophages from patients with cystic fibrosis.

Whereas many phagocytosis steps involve ionic fluxes, the underlying ion channels remain poorly defined. As reported in mice, the calcium conducting TRPV2 channel impacts the phagocytic process. Macrophage phagocytosis is critical for defense against pathogens. In cystic fibrosis (CF), macrophages have lost their capacity to act as suppressor cells and thus play a significant role in the initiating stages leading to chronic inflammation/infection. In a previous study, we demonstrated that impaired function of CF macrophages is due to a deficient phagocytosis. The aim of the present study was to investigate TRPV2 role in the phagocytosis capacity of healthy primary human macrophage by studying its activity, its membrane localization and its recruitment in lipid rafts. In primary human macrophages, we showed that P. aeruginosa recruits TRPV2 channels at the cell surface and induced a calcium influx required for bacterial phagocytosis. We presently demonstrate that to be functional and play a role in phagocytosis, TRPV2 might require a preferential localization in lipid rafts. Furthermore, CF macrophage displays a perturbed calcium homeostasis due to a defect in TRPV2. In this context, deregulated TRPV2-signaling in CF macrophages could explain their defective phagocytosis capacity that contribute to the maintenance of chronic infection

Chitosan encapsulation modulates the effect of capsaicin on the tight junctions of MDCK cells

Capsaicin has known pharmacological effects including the ability to reversibly open cellular tight junctions, among others. The aim of this study was to develop a strategy to enhance the paracellular transport of a substance with low permeability (FITC-dextran) across an epithelial cell monolayer via reversible opening of cellular tight junctions using a nanosystem comprised by capsaicin and of chitosan. We compared the biophysical properties of free capsaicin and capsaicin-loaded chitosan nanocapsules, including their cytotoxicity towards epithelial MDCK-C7 cells and their effect on the integrity of tight junctions, membrane permeability and cellular uptake. The cytotoxic response of MDCK-C7 cells to capsaicin at a concentration of 500 μM, which was evident for the free compound, is not observable following its encapsulation. The interaction between nanocapsules and the tight junctions of MDCK-C7 cells was investigated by impedance spectroscopy, digital holographic microscopy and structured illumination fluorescence microscopy. The nanocapsules modulated the interaction between capsaicin and tight junctions as shown by the different time profile of trans-epithelial electrical resistance and the enhanced permeability of monolayers incubated with FITC-dextran. Structured illumination fluorescence microscopy showed that the nanocapsules were internalized by MDCK-C7 cells. The capsaicin-loaded nanocapsules could be further developed as drug nanocarriers with enhanced epithelial permeability

Circum-Mediterranean cultural heritage and medicial plant uses in traditional animal healthcare: a field survey in eight selected areas within the RUBIA project

During the years 2003¿2005, a comparative ethnobotanical field survey was conducted on remedies used in traditional animal healthcare in eight Mediterranean areas. The study sites were selected within the EU-funded RUBIA project, and were as follows: the upper Kelmend Province of Albania; the Capannori area in Eastern Tuscany and the Bagnocavallo area of Romagna, Italy; Cercle de Ouezanne, Morocco; Sierra de Aracena y Picos de Aroche Natural Park in the province of Huelva, Spain; the St. Catherine area of the Sinai Peninsula, Egypt; Eastern and Western Crete, Greece; the Paphos and Larnaca areas of Cyprus; and the Mitidja area of Algeria. One hundred and thirty-six veterinary preparations and 110 plant taxa were recorded in the survey, with Asteraceae and Lamiaceae being the most quoted botanical families. For certain plant species the survey uncovered veterinary phytotherapeutical indications that were very uncommon, and to our knowledge never recorded before. These include Anabasis articulata (Chenopodiaceae), Cardopatium corymbosum (Asteraceae), Lilium martagon (Liliaceae), Dorycnium rectum (Fabaceae), Oenanthe pimpinelloides (Apiaceae), Origanum floribundum (Lamiaceae), Tuberaria lignosa (Cistaceae), and Dittrichia graveolens (Asteraceae). These phytotherapeutical indications are briefly discussed in this report, taking into account modern phytopharmacology and phytochemistry. The percentage of overall botanical veterinary taxa recorded in all the study areas was extremely low (8%), however when all taxa belonging to the same botanical genus are considered, this portion increases to 17%. Nevertheless, very few plant uses were found to be part of a presumed "Mediterranean" cultural heritage in veterinary practices, which raises critical questions about the concept of Mediterraneanism in ethnobotany and suggests that further discussion is required. Nearly the half of the recorded veterinary plant uses for mammals uncovered in this survey have also been recorded in the same areas in human folk medicine, suggesting a strong link between human and veterinary medical practices, and perhaps also suggesting the adaptive origins of a few medical practices. Since most of the recorded data concern remedies for treating cattle, sheep, goats, and camels, it would be interesting to test a few of the recorded phytotherapeuticals in the future, to see if they are indeed able to improve animal healthcare in breeding environments, or to raise the quality of dairy and meat products in the absence of classical, industrial, veterinary pharmaceuticals

Construction of a Global Pain Systems Network Highlights Phospholipid Signaling as a Regulator of Heat Nociception

The ability to perceive noxious stimuli is critical for an animal's survival in the face of environmental danger, and thus pain perception is likely to be under stringent evolutionary pressure. Using a neuronal-specific RNAi knock-down strategy in adult Drosophila, we recently completed a genome-wide functional annotation of heat nociception that allowed us to identify α2δ3 as a novel pain gene. Here we report construction of an evolutionary-conserved, system-level, global molecular pain network map. Our systems map is markedly enriched for multiple genes associated with human pain and predicts a plethora of novel candidate pain pathways. One central node of this pain network is phospholipid signaling, which has been implicated before in pain processing. To further investigate the role of phospholipid signaling in mammalian heat pain perception, we analysed the phenotype of PIP5Kα and PI3Kγ mutant mice. Intriguingly, both of these mice exhibit pronounced hypersensitivity to noxious heat and capsaicin-induced pain, which directly mapped through PI3Kγ kinase-dead knock-in mice to PI3Kγ lipid kinase activity. Using single primary sensory neuron recording, PI3Kγ function was mechanistically linked to a negative regulation of TRPV1 channel transduction. Our data provide a systems map for heat nociception and reinforces the extraordinary conservation of molecular mechanisms of nociception across different species. © 2012 Neely et al