Our findings can be used to inform future studies on community pharmacy-based screening programmes. The Author(s) declare(s) that they have no conflicts of interest to disclose. This research was funded by an MSc programme at the University of Aberdeen (Health Services and Public Health Research),

with additional financial support from a fellowship awarded jointly by the Medical Research Council and the Economic and Social Research Council, UK, to Dr Terry Porteous (Interdisciplinary Postdoctoral Y 27632 Fellowship). We are grateful to Cynthia Fraser (Information Specialist) at the University of Aberdeen for her advice in the development of the search strategy. We thank Graham Mowatt (University of Aberdeen) and Michelle Fiander, Trials Search Coordinator/Information

Specialist (University of Ottawa, Canada) for their help with the search on EPOC databases. Table S1 Characteristics of included studies (n = 50) Table S2 Quality assessment table for randomised controlled trial and cluster randomised studies[15] (n = 3 out of 50 included studies) Figure S1a Chart of quality assessment of comparative studies (n = 5) Figure S1b Chart of quality assessment of uncontrolled studies (n = 42) “
“This is the first of two papers which explore Roxadustat purchase the use of mixed-methods research in pharmacy practice. In an era of evidence-based medicine and policy, high-quality research evidence is essential for the development of effective pharmacist-led services. Over the DOK2 past decade, the use of mixed-methods research has become increasingly common in healthcare, although to date its use has been relatively limited in pharmacy practice research.

In this article, the basic concepts of mixed-methods research including its definition, typologies and advantages in relation to pharmacy practice research are discussed. Mixed-methods research brings together qualitative and quantitative methodologies within a single study to answer or understand a research problem. There are a number of mixed-methods designs available, but the selection of an appropriate design must always be dictated by the research question. Importantly, mixed-methods research should not be seen as a ‘tool’ to collect qualitative and quantitative data, rather there should be some degree of ‘integration’ between the two data sets. If conducted appropriately, mixed-methods research has the potential to generate quality research evidence by combining strengths and overcoming the respective limitations of qualitative and quantitative methodologies.

01% w/v arabinose for E. coli clones, both solidified with 12% gelatin (Oxoid, Adelaide, Australia). Colonies were grown at 25 °C for 5 days and then cooled at 4 °C for 3 h before checking for liquefaction by adding 3 μL of the 6 × gel loading dye (Fermentas Inc., Glen Burnie, MD) to each well. Evidence of liquefaction was established if the dye diffused rapidly (within 5 s) through the well and sank to the bottom. The Pseudoalteromonas tunicata D2 wild-type strain

(Holmström et al., 1998) and a genomic library of P. tunicata DNA, which was constructed by Burke et al. (2007) and which used the same fosmid vector and host strain as the metagenomic library described above, were used as positive controls. Cultures exhibiting activities on the solidified gelatin were subjected to a further assay EMD 1214063 in vivo using Azocoll, an insoluble, ground collagen, to which an azo-dye is attached. The assay was conducted in triplicates. Strains were grown for approximately 48 h at room temperature in MB and bacterial cells were harvested by centrifugation GPCR Compound Library at 8000 g for 10 min. Cell pellets were resuspended in the Azocoll substrate at a concentration of 5 × 108 CFU mL−1, supplemented

with a final concentration of 1 mM CaCl2. To prepare the substrate, 2.0 mg mL−1 of Azocoll (Sigma, St. Louis, MO) was washed twice using 0.01 M phosphate-buffered saline (pH 7.4) as described in Jiang et al. (2007). The tubes were incubated at room temperature with shaking at 90 r.p.m. for 24 h before centrifugation for 5 min to remove the undegraded Azocoll. Supernatants were taken for the measurement of OD520 nm. Escherichia coli Epi 300 pCC1FOS and Pseudomonas aeruginosa PAO1 strain were used as a negative and a positive control, respectively. The shotgun metagenome-sequencing data (92.6 Mbp of unique sequence) of the bacterial community associated with two C. concentrica specimen (BBAY04 and BBAY15) described in Thomas

et al. (2010) were searched for genes that were annotated as collagenase/matrix proteinase-related genes. Searches were performed on KEGG (Kanehisa & Goto, Cyclin-dependent kinase 3 2000), COG (Tatusov et al., 2003), Swiss-Prot (Boeckmann et al., 2003) and TIGRFAM (Haft et al., 2003) annotations using the keywords: ‘collagenase’, ‘Zn-dependent aminopeptidase’, ‘metalloproteinase’, ‘matrixin’ and ‘matrix proteinase’. The results were checked manually and matches that had an e-value lower than 1 × 10−20 in at least one annotation were regarded as putative collagenase protein sequences. In addition, collagenase-related proteins were retrieved from NCBI’s protein sequence database and the curated Swiss-Prot database (Boeckmann et al., 2003) using the keywords: ‘gelatinase’, ‘microbial collagenase’ and ‘matrix proteinase’ as well as proteins with the M9 peptidase and peptidase U32 conserved domains (which are domains in collagenases). Those database sequences were searched against the C. concentrica protein dataset with blastp (Altschul et al., 1990).

These results suggest the predominance of uncultured Treponema that appear to have distinct members related to the digestion of either Wee1 inhibitor hay or concentrate diet. The distribution of spiral-shaped bacteria (spirochetes) and their role in the degradation

of plant material in the rumen have been reported by different workers (Bryant, 1952; Stanton & Canale-Parola, 1979; Ziolecki & Wojciechowicz, 1980). Direct microscopic enumeration of spirochetes showed up to 2.0 × 108 cells mL−1 of bovine rumen fluid (Stanton & Canale-Parola, 1979), which is comparable to the population density of common rumen bacterial species (Bryant & Burkey, 1953). All strains of spirochetes isolated from the rumen have been classified in the genus Treponema, comprised of three described species: Treponema bryantii (Stanton & Canale-Parola, 1980), Treponema saccharophilum (Paster & Canale-Parola, 1985) and Treponema zioleckii (Piknova et al., 2008). Rumen Treponema strains are able to degrade plant polysaccharides (Ziolecki, 1979), and in vitro studies have shown a beneficial interaction of T. bryantii with the cellulolytic

learn more bacterium Fibrobacter succinogenes (Stanton & Canale-Parola, 1980). Recent application of molecular techniques in the study of microbial ecology demonstrated the existence of a considerable proportion of diverse uncultivated spirochetes involved in chronic disease in the human oral cavity and in degradation of lignocellulose materials in the termite gut (Paster et al., 1996, 2001; Dewhirst et al., 2000). For example, 16S rRNA gene-based clone library analysis of samples from the oral cavity of a human subject and from the hindgut of a single

termite species, respectively, suggested some 20 and 23 new species of spirochetes (Choi et al., 1994; Lilburn et al., 1999). Considering the individuality of human microbiota and the existence of ∼280 termite genera, these observations suggest the presence of a Teicoplanin considerable diversity of spirochetes, particularly uncultured members. In contrast to the above digestive tract environments, our knowledge of the uncultured Treponema community in the rumen is very limited. The current understanding of the rumen Treponema diversity is mainly based on earlier cultivation-based studies that showed morphological and physiological variations in rumen spirochetes (Paster et al., 1991; Piknova et al., 2008). A comprehensive analysis of 16S rRNA gene sequences derived from the rumen showed that rumen Treponema were not detected frequently (Edwards et al., 2004; Yang et al., 2010). However, we had previously retrieved a number of Treponema clones related to both cultured and uncultured members from a fiber-associated community (Koike et al., 2003; Shinkai et al., 2010). Based on these data, we speculated that rumen Treponema diversity has been underestimated and members of this group may play a metabolic role in fiber degradation.

Sparkle (Lee & La Rue, 1992). In Trifolium repens roots, ethylene inhibits cortical cell division, a process that is indispensable for nodule primordia formation (Goodlass & Smith, 1979). To obviate some of the inhibitory effects of ethylene in nodule formation, development and function, some rhizobial strains utilize different mechanisms for lowering ethylene levels such as the production of the BAY 57-1293 nmr enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase; this enzyme is responsible

for the cleavage of ACC (the immediate precursor of ethylene in plants) to ammonia and α-ketobutyrate (Honma & Shimomura, 1978), contributing to increase the competitiveness of the strains because of advantages in the processes of nodule formation and occupancy www.selleckchem.com/products/z-vad-fmk.html (Ma et al., 2003b, 2004). Other rhizobial strains lower ethylene levels by producing the compound rhizobitoxine, an inhibitor of the plant enzyme ACC synthase (Sugawara et al., 2006). The prevalence of ACC deaminase genes in rhizobia has been studied primarily in Rhizobium spp. (Ma et al., 2003a; Duan et al., 2009). In these studies, many Rhizobium spp. have been found to possess an acdS gene and produce ACC deaminase under free-living conditions. For example, in a rhizobia collection of isolates from Saskatchewan (Canada), 27 Rhizobium isolates possessed an acdS gene and were able to produce

ACC deaminase, thus, showing that acdS genes are present throughout Rhizobium isolates (Duan et al., 2009). On the other hand, notwithstanding reports documenting the presence of ACC deaminase in Mesorhizobium spp., not much is known about the environmental distribution of acdS genes in this bacterial genus. The first report on acdS gene presence in Mesorhizobium was obtained following the complete sequencing of Mesorhizobium sp. MAFF303099 (Kaneko et al., 2000). Subsequently, the presence of an acdS

gene in the symbiosis island of Mesorhizobium loti R7A was also reported (Sullivan et al., 2002). However, when Mesorhizobium sp. MAFF303099 and Mesorhizobium ciceri UPM Ca-7 were tested for ACC deaminase activity and the presence of an acdS gene, no activity was detected and the acdS gene was not found in M. ciceri (Ma et al., 2003b). Recently, the genome sequences of Mesorhizobium Reverse transcriptase opportunistum WSM2075T (Lucas et al., 2011a), Mesorhizobium australicum WSM2073T (Lucas et al., 2011b), and Mesorhizobium ciceri bv. biserrulae WSM1271 (Lucas et al., 2011c), revealed the presence of an acdS gene in these strains. In some strains of Mesorhizobium, the production of ACC deaminase has been shown to be an important mechanism to promote nodule formation. When compared to the wild-type strain, Mesorhizobium sp. MAFF303099 acdS knockout mutant has a decreased ability to form and occupy nodules, losing both its effectiveness and competitiveness (Uchiumi et al., 2004).

sedicola. The genus Stemphylium, anamorphic Pleospora (Dothideomycetes), was proposed by Wallroth in 1833 with Stemphylium botryosum Wallr. as the type species. More than 33 species are recognized in this check details genus (Câmara et al., 2002), many of which are saprophytic, growing on dead plants and cellulose materials (Simmons, 1969), but several species, including S. botryosum, S. solani G.F. Weber, and S. vesicarium (Wallr.) E.G. Simmons, are plant pathogens that cause diseases in important agricultural crops and fruit trees. This is the first report to show that S. sedicola, a member of the genus Stemphylium isolated from T. baccata, has the potential

to produce anticancer compound taxol. Taxol is the best known and most studied member of the taxane diterpenoids, or taxoids, and has been used in chemotherapy for many types of cancers since the 1970s. Many endophytic fungi, which are widely

found in almost all kinds of plants including Taxus species, can produce physiologically active compounds, such as taxanes, which are the same or analogous with those obtained from their hosts (Lu et al., 2000; Glienke-Blanco et al., 2002; Strobel et al., 2004). This constitutes a selleck new approach to resolving resource limitation and an alternative taxol source. Indeed, it represents a great opportunity to find new and interesting endophytic microorganisms among Taxus species in different settings and ecosystems. Taxol is known to be produced by a number of endophytic fungi, including the following families reported in the literature (Zhao et al.,

2008; Zhou et al., 2010): Hypocreaceae, Nectriaceae, Amphisphaeriaceae, Pleosporaceae, Chaetomiaceae, Kickxellaceae, Trichocomaceae, Clavicipitaceae, Thyridiaceae, and Xylariaceae. The genus Stemphylium has not been previously reported PRKACG from Taxus species, although both saprotrophic and pathogenic forms of Stemphylium occur in a wide range of plants and many species are economically damaging pathogens of agricultural crops (Câmara et al., 2002). Additionally, Stemphylium species are known sources of bioactive compounds, including the cytotoxic and protein kinase-inhibiting alterporriols G and H (Debbab et al., 2009), the antibacterial perylenequinones, stemphyltoxins I-IV (Arnone et al., 1986), as well as the phytotoxic chromone glucoside, stemphylin (Barash et al., 1975). In this study, based on morphological and molecular data as well as phytochemical analysis, S. sedicola SBU-16 was determined as a new strain of taxol-producing endophytic fungi. Quantitative HPLC analysis showed that the taxol content of S. sedicola SBU-16 was in the range of previously reported fungi (Stierle et al., 1993; Guo et al., 2006; Gangadevi et al., 2008), indicating its promising potential for taxol production.

However, relapse is described even among patients with successful treatment and CD4 T-cell counts >200 cells/μL [50]. Cases of leishmaniasis IRIS are described with new or worsening skin lesions including ulcers, mucocutaneous ulcers in the mouth or penis, post-kala-azar dermal leishmaniasis or uveitis [51,52]. There are also reports of visceral leishmaniasis presenting as an immune reconstitution Barasertib supplier phenomenon after the start of antiretroviral therapy [53]. There are

multiple overlapping toxicities with HIV medication and treatment for leishmaniasis and liaison with an HIV pharmacist is recommended. Chagas disease or American trypanosomiasis is caused by a parasite, which is a member of the genus Trypanosoma; Trypanosoma cruzi. It is confined to Central and South America and its distribution extends from Mexico in the north to the northern half of Argentina and Chile in the south. T. cruzi is spread by bloodsucking triatomine insects, also known as kissing bugs, found in particular in rural areas [54]. There is increasing recognition that reactivation of Trypanosoma cruzi infection can cause disease in patients with advanced immunosuppression, including HIV-seropositive individuals who have lived or travelled to endemic areas. T. cruzi causes two main types of disease in people with HIV: Neurological disease; space-occupying lesions or meningoencephalitis Clinical

syndromes are most common in individuals with CD4 T-cell counts <200 cells/μL selleck products [55]. Neurological syndromes are the commonest presentation, comprising 75% of presentations of Chagas disease in untreated HIV-seropositive patients. Patients can present with features of a space-occupying lesion, encephalitis, or meningoencephalitis [56]. Clinical symptoms are typically of fever, headaches,

seizures, vomiting and focal neurological signs and mimic toxoplasma encephalitis [54]. Myocarditis is the second most common presentation seen in approximately a third of cases, often with concomitant neurological disease. Myocarditis is often asymptomatic and only detected at autopsy Interleukin-3 receptor but can present with arrhythmias or heart failure [54,57]. Chagas disease may also affect the digestive tract and cause megaoesophagus or megacolon. Chagas disease should be suspected in patients from the endemic areas of Central and South America or with a history of blood transfusions or intravenous drug use with contacts from these areas. Diagnosis of Chagas disease requires a combination of imaging, serology, PCR and if available histological confirmation (category III recommendation). For neurological disease, imaging studies characteristically report space-occupying lesions similar to those described for toxoplasma encephalitis [58]. CSF examination typically describes lymphocytic pleocytosis and elevated protein with possibly low glucose [54]. Serological tests are generally not diagnostic for reactivation, indicating only previous exposure.

Both HIV-1 and HIV-2 are associated with similar opportunistic infections and AIDS. Natural history studies indicate Vorinostat clinical trial that HIV-2 is less pathogenic than HIV-1 [16–18]. Although the mortality rate in individuals infected with HIV-2 is two-to-three times that seen in HIV-negative populations, this compares with a 10-fold higher mortality rate in those

infected with HIV-1 than in those who are HIV negative. HIV-2 infection has a longer asymptomatic phase than HIV-1 infection and some patients with HIV-2 may never develop AIDS [19]. A cohort study of seroconverter women in Senegal found that the incidence of AIDS-defining illness was 0.95 [95% confidence interval (CI) 0.2–3.8] per

Akt inhibitor 100 person-years among HIV-2-infected women as compared with 5.6 (95% CI 3.3–9.8) in HIV-1-infected women [16]. In practice, it is not unusual to see patients who remain asymptomatic for 10–20 years without treatment [20]. There are, however, patients in whom disease progresses as rapidly as in those who have HIV-1. AIDS-defining illnesses have been noted to occur at higher CD4 cell counts in individuals infected with HIV-2 than in those infected with HIV-1, although this is unusual [21]. Plasma viral loads are lower in HIV-2-infected individuals, suggesting that HIV-2 replication is restricted in comparison to that of HIV-1. An in vivo study has clearly demonstrated that, like HIV-1, HIV-2 can establish a stable, integrated proviral infection but that HIV-2 produces less mRNA, which may attenuate HIV-2 replication and pathogenesis [22]. HIV-2 is less infectious than HIV-1 early in the course of infection and, although infectivity increases as the disease advances, in general HIV-2 has significantly lower infectivity than HIV-1 [23]. HIV-2 infection does not protect against HIV-1 infection and dual

infection is well documented [24–26] although it is still uncommon in the United Kingdom. Studies from West Africa demonstrate that dual infection is more common in older women [25]. Dually infected patients tend to present at a more advanced stage of disease than those with HIV-2 only. Levetiracetam Infection with both HIV-1 and HIV-2 generally carries the same prognosis as HIV-1 monoinfection [19]. It is important to note that HIV-2 has a different capsid antigen from the HIV-1 p24 antigen and that this capsid antigen may result in a prolonged seroconversion window period for HIV-2, but there is no current evidence from human studies that it is longer than the 3-month period described for HIV-1. Detection of HIV-2 infection is based on the demonstration of virus-specific antibodies using enzyme-linked immunosorbent assay-based techniques.

72; 95% CI Nivolumab ic50 0.26, 1.99), CD4 T-cell count

was positively associated with incident HTN (HR 1.15 per 100 cells/μL; 95% CI 1.03, 1.28). Among physically active HIV-infected men, exposure to ARVs was negatively associated with incident HTN (HR 0.15; 95% CI 0.03, 0.78). HIV infection was not associated with incident HTN in older men or women. This study provides additional evidence supporting a causal relationship between immune function and incident HTN, which warrants further study. “
“The aim of the study was to assess the significance of low-level viraemia (LLV) and the timing of treatment change in low/middle-income country (L/MIC) compared with high-income country (HIC) settings. Patients with virological control following commencement of combination antiretroviral therapy (cART) were included in the study. LLV was defined as undetectable viral load (<50 HIV-1 RNA copies/mL) followed by confirmed detectable viral load < 1000 copies/mL. Virological failure was defined as viral load > 1000 copies/mL. Kaplan−Meier plots of time to virological failure by prior LLV and income category were generated. Regimen changes in

the setting of LLV were compared between sites. Sensitivity analysis of rates of LLV and virological failure by person-years and number of tests was conducted for differing VX-770 definitions of LLV and virological failure. A total of 1748 patients from HICs and 823 patients from L/MICs were included in the study. One hundred and ninety-six (11.2%) HIC participants Fenbendazole and 36 (4.4%) L/MIC participants experienced at least one episode of LLV. Of the patients who underwent regimen switch in HIC settings, the majority changed from a nucleoside reverse transcriptase inhibitor (NRTI)/protease inhibitor (PI) regimen to an NRTI/nonnucleoside reverse transcriptase inhibitor (NNRTI) regimen (26.8%). Very few switches were made in L/MIC settings. Rates of LLV were significantly higher for HICs compared with L/MICs per 1000 person-years (28.6 and 9.9 per 1000 person-years,

respectively), but not in terms of the number of tests (9.4 and 7.2 per 1000 tests, respectively). Rates of virological failure per test were significantly higher for L/MICs compared with HICs (30.7 vs. 19.6 per 1000 tests, respectively; P < 0.001). LLV was a significant predictor of virological failure at 2 years in L/MICs [0.25; 95% confidence interval (CI) 0.11–0.50; P = 0.043] but not in HICs (0.13; 95% CI 0.08-0.22; P = 0.523). LLV is weakly predictive of virological failure at 2 years in L/MICs but not in HICs. This suggests that interventions targeted at subjects with LLV in L/MICs would help to improve treatment outcomes. "
“For the last 10 years there has been an epidemic of hepatitis C virus (HCV) infection in men who have sex with men (MSM) in Europe, North America and Australia. The majority of those infected are also HIV-positive and it is unclear to what extent HIV-negative MSM are also at increased risk of infection with HCV.

72; 95% CI Talazoparib molecular weight 0.26, 1.99), CD4 T-cell count

was positively associated with incident HTN (HR 1.15 per 100 cells/μL; 95% CI 1.03, 1.28). Among physically active HIV-infected men, exposure to ARVs was negatively associated with incident HTN (HR 0.15; 95% CI 0.03, 0.78). HIV infection was not associated with incident HTN in older men or women. This study provides additional evidence supporting a causal relationship between immune function and incident HTN, which warrants further study. “
“The aim of the study was to assess the significance of low-level viraemia (LLV) and the timing of treatment change in low/middle-income country (L/MIC) compared with high-income country (HIC) settings. Patients with virological control following commencement of combination antiretroviral therapy (cART) were included in the study. LLV was defined as undetectable viral load (<50 HIV-1 RNA copies/mL) followed by confirmed detectable viral load < 1000 copies/mL. Virological failure was defined as viral load > 1000 copies/mL. Kaplan−Meier plots of time to virological failure by prior LLV and income category were generated. Regimen changes in

the setting of LLV were compared between sites. Sensitivity analysis of rates of LLV and virological failure by person-years and number of tests was conducted for differing selleck kinase inhibitor definitions of LLV and virological failure. A total of 1748 patients from HICs and 823 patients from L/MICs were included in the study. One hundred and ninety-six (11.2%) HIC participants not and 36 (4.4%) L/MIC participants experienced at least one episode of LLV. Of the patients who underwent regimen switch in HIC settings, the majority changed from a nucleoside reverse transcriptase inhibitor (NRTI)/protease inhibitor (PI) regimen to an NRTI/nonnucleoside reverse transcriptase inhibitor (NNRTI) regimen (26.8%). Very few switches were made in L/MIC settings. Rates of LLV were significantly higher for HICs compared with L/MICs per 1000 person-years (28.6 and 9.9 per 1000 person-years,

respectively), but not in terms of the number of tests (9.4 and 7.2 per 1000 tests, respectively). Rates of virological failure per test were significantly higher for L/MICs compared with HICs (30.7 vs. 19.6 per 1000 tests, respectively; P < 0.001). LLV was a significant predictor of virological failure at 2 years in L/MICs [0.25; 95% confidence interval (CI) 0.11–0.50; P = 0.043] but not in HICs (0.13; 95% CI 0.08-0.22; P = 0.523). LLV is weakly predictive of virological failure at 2 years in L/MICs but not in HICs. This suggests that interventions targeted at subjects with LLV in L/MICs would help to improve treatment outcomes. "
“For the last 10 years there has been an epidemic of hepatitis C virus (HCV) infection in men who have sex with men (MSM) in Europe, North America and Australia. The majority of those infected are also HIV-positive and it is unclear to what extent HIV-negative MSM are also at increased risk of infection with HCV.

Spinal cords were obtained from 3- to 5-week-old male Sprague–Dawley rats by dorsal laminectomy. The rats were anesthetized with 3% isoflurane in an induction box and kept under isoflurane anesthesia during the extraction of the spinal cord, which took < 2 min and included euthanasia by bilateral thoracotomy. Coronal slices (400 μm) were cut with a vibratome (Integraslice 7550PSDS; Campden Instruments USA, Lafayette, IN, USA) from a lumbar ABT 199 spinal cord segment (L2–L4), as described (Marvizon et al., 2003a; Lao & Marvizon, 2005; Adelson et al., 2009). The spinal cord segment was glued vertically

to a block of agar on the stage of the vibratome and immersed in ice-cold sucrose-aCSF. Slices were cut using minimum forward speed and maximum vibration while Selumetinib chemical structure under observation with a stereo microscope mounted over the vibratome. Slices were prepared either without roots or with

one dorsal root, which was used for electrical stimulation. In the later case, fiber continuity between the dorsal root and the dorsal horn was assessed by examining the dorsal root and the dorsal surface of the slice with the stereo microscope. Slices were discarded if they did not meet the following criteria: (i) at least 80% of the dorsal funiculus had to be continuous with the dorsal root, and (ii) the dorsal root had no cuts or compression damage. Slices were kept for 1 h in K+-aCSF at 35°C, and then in regular aCSF at 35°C. The dorsal root attached to the slice was electrically stimulated using a custom-made chamber, as previously described (Marvizon et al., 2003b; Adelson et al., 2009). The root was placed on a bipolar stimulation electrode (platinum wire of 0.5 mm diameter, 1 mm pole separation) in a compartment separated from the superfusion chamber by a grease bridge. The root and the electrodes were covered with mineral oil, and

any excess aCSF was suctioned away. This ensured that electrical current circulated through the root and that the stimulus was consistent between preparations. Electrical stimulation was provided by a Master-8 stimulator and SIU5A stimulus isolating unit (A.M.P. Liothyronine Sodium Instruments, Jerusalem, Israel), and consisted of 1000 square pulses of 20 V and 0.4 ms (C-fiber intensity) delivered at 1 Hz or 100 Hz. In some experiments, the root was chemically stimulated by incubating it for 10 min with 1 μm capsaicin in aCSF in the side compartment of the chamber, as described (Lao et al., 2003). Slices were superfused at 3–6 mL/min with aCSF at 35°C. Drugs were present in the superfusate continuously starting 5 or 10 min before root stimulation. Ten minutes after the stimulus slices were fixed by immersion in ice-cold fixative (4% paraformaldehyde and 0.18% picric acid in 0.1 m sodium phosphate buffer). A round hole was punched in the ventral horn of the slice ipsilateral to the stimulus in order to identify it in the histological sections after immunohistochemistry.