Determination of pioglitazone hydrochloride by flow injection chemiluminescence tris(2,2'-bipyridyl)ruthenium(II)-silver(III) complex system.

A novel flow injection-chemiluminescence (FI-CL) approach is proposed for the assay of pioglitazone hydrochloride (PG-HCl) based on its enhancing influence on the tris(2,2'-bipyridyl)ruthenium(II)-silver(III) complex (Ru(bipy)3 2+ -DPA) CL system in sulfuric acid medium. The possible CL reaction mechanism is discussed with CL and ultraviolet (UV) spectra. The optimum experimental conditions were found as: Ru(bipy)3 2+ , 5.0 × 10-5  M; sulfuric acid, 1.0 × 10-3  M; diperiodatoargentate(III) (DPA), 1.0 × 10-4  M; potassium hydroxide, 1.0 × 10-3  M; flow rate 4.0 ml min-1 for each flow stream and sample loop volume, 180 µl. The CL intensity of PG-HCl was linear in the range of 1.0 × 10-3 to 5.0 mg L-1 (R2 = 0.9998, n = 10) with limit of detection [LOD, signal-to-noise ratio (S/N) = 3] of 2.2 × 10-4  mg L-1 , limit of quantification (LOQ, S/N = 10) of 6.7 × 10-4  mg L-1 , relative standard deviation (RSD) of 1.0 to 3.3% and sampling rate of 106 h-1 . The methodology was satisfactorily used to quantify PG-HCl in pharmaceutical tablets with recoveries ranging from 93.17 to 102.77 and RSD from 1.9 to 2.8%.

Determination of lansoprazole in pharmaceuticals using flow injection with rhodamine 6G-diperiodatoargentate(III) chemiluminescence detection.

A chemiluminescence (CL) method based on rhodamine 6G (R6G)-diperiodatoargentate(III) (silver(III) complex) reaction in acid solution is reported for the determination of lansoprazole (LNP) combined with a flow injection (FI) technique. The most likely mechanism for CL reaction was elucidated considering reported data, spectrophotometric and spectrofluorimetric studies. The weak CL reaction between R6G and silver(III) complex could be magnanimously increased in the presence of LNP with a limit of detection (LOD) of 0.002 mg L-1 (S/N = 3), a linear range of 0.01 to 10 mg L-1 (R2 = 0.9997, n = 7), a relative standard deviation (RSD) of 1.2 to 3.2% (n = 4) and an injection throughput of 140 h-1 . No interference activity of commonly found excipients in LNP was detected. After LNP extraction from pharmaceutical samples, the recovery rate ranging from 93 to 110% (RSD, 1.4-3.3%, n = 4) was calculated. The results of the proposed flow CL method were assessed with a spectrophotometric approach applying paired Student's t-test and the calculated value (0.178) was lower than the distributed value (2.20) at a 95% confidence limit.

Patients' Perceptions of Healthcare Quality at Hospitals Measured by the Revised Humane Caring Scale.

The patients' perceptions are central to quality improvement of the healthcare system worldwide. This study aimed to examine patients' perceptions of quality care and investigate the demographic factors related to the overall patients' perceptions. The Revised Humane Caring Scale was distributed to 367 adult patients who were admitted at medical, surgical, and obstetrics and gynecology departments in 2 tertiary hospitals in Oman. Overall patients' perceptions of quality of care were high, with professionalism being rated the highest, and cognition of physical needs and human resources rated the lowest. Significant differences in patients' perceptions between hospitals as well as in the subscales of interdisciplinary collaboration and outcome variables, between planned- and emergency-admitted patients were found. The linear regression analysis indicated a relationship between gender and overall quality care where male patients reported higher satisfaction compared to counterparts. This study suggested the need to improve the cognition of physical needs (food quality and environmental sanitation) and human resources (staff to patient ratio) as well as pay attention to the individual patients' needs especially for emergency-admitted patients.

A systematic review of healthcare professionals' core competency instruments.

While technical and profession-specific competencies are paramount in the delivery of healthcare services, the cross-cutting core competencies of healthcare professionals play an important role in healthcare transformation, innovation, and the integration of roles. This systematic review describes the characteristics and psychometric properties of existing instruments for assessing healthcare professionals' core competencies in clinical settings. It was guided by the JBI methodology and used the COSMIN checklist (Mokkink et al., User manual, 2018, 78, 1) to evaluate the methodological quality of the included studies. A database search (CINAHL, Scopus, and PubMed) and additional manual search were undertaken for peer-reviewed papers with abstracts, published in English between 2008 and 2019. The search identified nine studies that were included in the synthesis demonstrating core competencies in professionalism, ethical and legal issues, research and evidence-based practice, personal and professional development, teamwork and collaboration, leadership and management, and patient-centered care. Few instruments addressed competencies in quality improvement, safety, communication, or health information technology. The findings demonstrate the reviewed tools' validity and reliability and pave the way for a comprehensive evaluation and assessment of core competencies into clinical practice.

Flow injection-chemiluminescence determination of cetirizine dihydrochloride in pharmaceuticals using tris(2,2'-bipyridyl)ruthenium (II)-Ag(III) complex reaction.

A simple and sensitive flow injection-chemiluminescence (FI-CL) method was developed for determination of cetirizine dihydrochloride (CTZH) in pharmaceuticals. The method is primarily based on the enhancement effect of CTZH on the tris(2,2'-bipyridyl)ruthenium (II)-diperiodatoargentate (III) ([Ru(bpy)3 ]2+ -Ag(III) complex) CL system in an acidic medium. The optimum investigated variables of the CL reaction were: [Ru(bpy)3 ]2+ , 50 × 10-6 mol/L; sulfuric acid, 1.0 × 10-3 mol/L; Ag(III) complex, 100 × 10-6 mol/L; potassium hydroxide, 1.0 × 10-3 mol/L; flow rate, 3.0 ml/min and sample loop volume, 300 µl. The detection and quantification limits were 2.0 × 10-4 and 5.0 × 10-4 mg/L (S/N of 3 and 10) respectively with a linear calibration range of 5.0 × 10-4 to 7.5 mg/L (R2 = 0.9999, n = 11), injection throughput of 110/h and the relative standard deviations of 1.5-3.5% over the range studied. The methodology was successfully applied to determine CTZH in different pharmaceutical samples and validated with a high-performance liquid chromatography method, and resulted in the recovery of 94.6-108.6%. The probable CL reaction mechanism is described in brief.

Determination of Nalbuphine Hydrochloride in Pharmaceutical Formulations Using Diperiodatoargentate(III)-Rhodamine-B Chemiluminescence System by Flow Injection Analysis.

A novel method for the analysis of nalbuphine hydrochloride (NAL) is reported based on its enhancement effect on a diperiodatoargentate(III)-rhodamine-B (Ag(III) complex-Rh-B) chemiluminescence (CL) system in an aqueous sulfuric acid medium using flow-injection analysis (FIA). The optimal conditions of the CL reaction were: sulfuric acid, 10-2 M; Ag(III) complex, 2.0 × 10-4 M; Rh-B, 2.0 × 10-5 M; Brij-35, 0.01%; sample loop volume, 300 µL; and flow rate, 3.0 mL/min/stream. The limit of detection (LOD) and limit of quantification (LOQ) were 0.001 and 0.003 mg/L (S/N = 3 and 10); linear calibration range, 5 × 10-3 - 5.0 mg/L (R2 = 0.9999) and injection throughput, 150/h. The relative standard deviation (RSD) was from 0.8 - 3.2% over the range studied. The suggested technique was applied for the determination of NAL in pharmaceutical injections, compared with a reported spectrophotometric method, and obtained results were found to be satisfactory. Based on spectrophotometric studies, the most probable mechanism of the CL reaction has been briefly described and drawn schematically.

Lansoprazole Determination in Pharmaceutical Formulations by Flow Injection Coupled with Acidic KMnO4-Quinine Chemiluminescence System.

A simple and rapid flow injection chemiluminescence (FI-CL) method based on the reaction of potassium permanganate (KMnO4) and quinine was established for the determination of lansoprazole in pharmaceutical formulations. A linear calibration curve was achieved over the range from 0.01 to 20.0 mg L-1 LNP (R2 = 0.9997 (n = 8); RSD = 1.1 - 3.7% (n = 4)) with a limit of detection of 3.0 × 10-3 mg L-1 (S/N = 3) and injection throughput of 150 h-1. By applying the Student t-test (calculated t-test value: t = 1.059907664, and tabulated t-distributed (95%) = 2.200985) it was found that the proposed method and reported spectrophotometric method were not significantly different. The LNP was efficiently extracted and the recovery of LNP from the spiked pharmaceutical formulations was in the range of 91.0 - 105.9% (%RSD = 1.6 - 3.6, n = 4). No significant interference activity was detected from the excipients commonly found in the drug samples analyzed. The possible chemiluminescence emission mechanism is discussed briefly.

Eye and foot checks in patients with diabetes on haemodialysis: Are they done, and who does them?

AIM: To determine if retinal and foot checks are carried out on patients with diabetes receiving haemodialysis. METHODS: Eighty-four patients with diabetes receiving haemodialysis were asked if they recalled having eye and foot screening in the last year, and if so, by whom was the check done. RESULTS: Seventy-seven (91.7%) patients recalled having an eye check in the preceding 12 mo. Of these, 52 (67.5%) did so in an ophthalmology clinic, 17 (22%) in retinal screening, three (3.9%) in an optician clinic. Three patients (3.9%) went to both ophthalmology and retinal screening, and two (2.6%) attended an ophthalmology and optician. Seventy (83.3%) patients recalled having a foot check in the preceding 12 mo. Of these, 33 (47.1%) were done by practice nurse, 14 (20%) by a diabetes nurse, 11 (15.7%) by a general practitioner, eight (11.4%) by a chiropodist, and four (5.7%) were each checked by renal nurse, diabetes consultant, junior doctor, or unknown person at a foot clinic. CONCLUSION: Most patients with diabetes on haemodialysis are able to recall having an eye check in the last year, although 8.3% could not. A significant proportion of patients could not recall having a foot check (16.7%) in the last year. This baseline audit suggests that an improvement in the rate of foot screening is important to achieve in patients with diabetes on haemodialysis in our unit.

Anandamide produced by Ca(2+)-insensitive enzymes induces excitation in primary sensory neurons.

The endogenous lipid agent N-arachidonoylethanolamine (anandamide), among other effects, has been shown to be involved in nociceptive processing both in the central and peripheral nervous systems. Anandamide is thought to be synthesised by several enzymatic pathways both in a Ca(2+)-sensitive and Ca(2+)-insensitive manner, and rat primary sensory neurons produce anandamide. Here, we show for the first time, that cultured rat primary sensory neurons express at least four of the five known Ca(2+)-insensitive enzymes implicated in the synthesis of anandamide, and that application of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-arachidonoyl, the common substrate of the anandamide-synthesising pathways, results in anandamide production which is not changed by the removal of extracellular Ca(2+). We also show that anandamide, which has been synthesised in primary sensory neurons following the application of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-arachidonoyl induces a transient receptor potential vanilloid type 1 ion channel-mediated excitatory effect that is not inhibited by concomitant activation of the cannabinoid type 1 receptor. Finally, we show that sub-populations of transient receptor potential vanilloid type 1 ion channel-expressing primary sensory neurons also express some of the putative Ca(2+)-insensitive anandamide-synthesising enzymes. Together, these findings indicate that anandamide synthesised by primary sensory neuron via a Ca(2+)-insensitive manner has an excitatory rather than an inhibitory role in primary sensory neurons and that excitation is mediated predominantly through autocrine signalling. Regulation of the activity of the Ca(2+)-insensitive anandamide-synthesising enzymes in these neurons may be capable of regulating the activity of these cells, with potential relevance to controlling nociceptive processing.

Will adoption of the 2010 WHO ART guidelines for HIV-infected TB patients increase the demand for ART services in India?

BACKGROUND: In 2010, WHO expanded previously-recommended indications for anti-retroviral treatment to include all HIV-infected TB patients irrespective of CD4 count. India, however, still limits ART to those TB patients with CD4 counts <350/mm(3) or with extrapulmonary TB manifestations. We sought to evaluate the additional number of patients that would be initiated on ART if India adopted the current 2010 WHO ART guidelines for HIV-infected TB patients. METHODS: We evaluated all TB patients recorded in treatment registers of the Revised National TB Control Programme in June 2010 in the high-HIV prevalence state of Karnataka, and cross-matched HIV-infected TB patients with ART programme records. RESULTS: Of 6182 TB patients registered, HIV status was ascertained for 5761(93%) and 710(12%) were HIV-infected. 146(21%) HIV-infected TB patients were on ART prior to TB diagnosis. Of the remaining 564, 497(88%) were assessed for ART eligibility; of these, 436(88%) were eligible for ART according to 2006 WHO ART guidelines. Altogether, 487(69%) HIV-infected TB patients received ART during TB treatment. About 80% started ART within 8 weeks of TB treatment and 95% received an efavirenz based regimen. CONCLUSION: In Karnataka, India, about nine out of ten HIV-infected TB patients were eligible for ART according to 2006 WHO ART guidelines. The efficiency of HIV case finding, ART evaluation, and ART initiation was relatively high, with 78% of eligible HIV-infected patients actually initiated on ART, and 80% within 8 weeks of diagnosis. ART could be extended to all HIV-infected TB patients irrespective of CD4 count with relatively little additional burden on the national ART programme.

Bone marrow mononuclear cells reduce myocardial reperfusion injury by activating the PI3K/Akt survival pathway.

OBJECTIVE: Adult bone marrow mononuclear cells (BMMNCs) can restore cardiac function following myocardial necrosis. Protocols used to date have administered cells relatively late after ischaemia/reperfusion injury, but there is the opportunity with elective procedures to infuse cells shortly after restoration of blood flow, for example after angioplasty. Our aim was therefore to try and quantify protection from myocardial injury by early infusion of BMMNCs in a rat ischaemia reperfusion (I/R) model. METHODS AND RESULTS: Male Wistar rats underwent 25 min of ischaemia followed by 2 h reperfusion of the left anterior descending coronary artery. Ten million BMMNCs were injected i.v. at reperfusion. We found BMMNCs caused a significant reduction in infarct size at 2 h when assessed by staining the area at risk with p-nitro blue tetrazolium (42% reduction, P<0.01). Apoptosis and necrosis of isolated cardiomyocytes was significantly reduced in the area at risk. Functional assessment at 7 days using echocardiography and left ventricular catheterisation showed improved systolic and diastolic function in the BMMNC treatment group (LVEF: BMMNC 71 ± 3% vs. PBS 48 ± 4%, P<0.0001). In functional studies BMMNC injected animals showed increased activation of Akt, inhibition of GSK-3ß, amelioration of p38 MAP kinase phosphorylation and NF-κB activity compared to control myocardium. Inhibition of PI3K with LY294002 abolished all beneficial effects of BMMNC treatment. Proteomic analysis also demonstrated that BMMNC treatment induced alterations in proteins within known cardioprotective pathways, e.g., heat shock proteins, stress-70 protein as well as the chaperone protein 14-3-3 epsilon. CONCLUSIONS: Early BMMNC injection during reperfusion preserves the myocardium, with evidence of reduced apoptosis, necrosis, and activation of survival pathways.

Overcoming hERG issues for brain-penetrating cathepsin S inhibitors: 2-cyanopyrimidines. Part 2.

We describe here orally active and brain-penetrant cathepsin S selective inhibitors, which are virtually devoid of hERG K(+) channel affinity, yet exhibit nanomolar potency against cathepsin S and over 100-fold selectivity to cathepsin L. The new non-peptidic inhibitors are based on a 2-cyanopyrimidine scaffold bearing a spiro[3.5]non-6-yl-methyl amine at the 4-position. The brain-penetrating cathepsin S inhibitors demonstrate potential clinical utility for the treatment of multiple sclerosis and neuropathic pain.

Generation of endogenous hydrogen sulfide by cystathionine gamma-lyase limits renal ischemia/reperfusion injury and dysfunction.

The generation of endogenous hydrogen sulfide may either limit or contribute to the degree of tissue injury caused by ischemia/reperfusion. A total of 74 male Wistar rats were used to investigate the effects of endogenous and exogenous hydrogen sulfide in renal ischemia/reperfusion. Administration of the irreversible cystathionine gamma-lyase (CSE) inhibitor, dL-propargylglycine, prevented the recovery of renal function after 45 min ischemia and 72 h reperfusion. The hydrogen sulfide donor sodium hydrosulfide attenuated the (renal, tubular, and glomerular) dysfunction and injury caused by 45 min ischemia and 6 h reperfusion. Western blot analysis of kidneys taken at 30 min reperfusion showed that sodium hydrosulfide significantly attenuated phosphorylation of mitogen-activated protein kinases (p-38, c-JUN N-terminal protein kinase 1/2, and extracellular signal-regulated kinase 1/2) and activation of nuclear factor-kappaB. At 6 h reperfusion, sodium hydrosulfide significantly attenuated the histological score for acute tubular necrosis, the activation of caspase-3 and Bid, the decline in the expression of anti-apoptotic Bcl-2, and the expression of nuclear factor-kappaB-dependent proteins (inducible nitric oxide synthase, cyclo-oxygenase-2, and intercellular adhesion molecule-1). These findings suggest that (1) the synthesis of endogenous hydrogen sulfide by CSE is essential to protect the kidney against ischemia/reperfusion injury and dysfunction and aids in the recovery of renal function following ischemia/reperfusion, (2) hydrogen sulfide generated by sodium hydrosulfide reduces ischemia/reperfusion injury and dysfunction, and morphological changes of the kidney, and (3) the observed protective effects of hydrogen sulfide are due to both anti-apoptotic and anti-inflammatory effects.

Discovery of orally bioavailable cathepsin S inhibitors for the reversal of neuropathic pain.

Cathepsin S inhibitors are well-known to be an attractive target as immunological therapeutic agents. Recently, our gene expression analysis identified that cathepsin S inhibitors could also be effective for neuropathic pain. Herein, we describe the efficacy of selective cathepsin S inhibitors as antihyperalgesics in a model of neuropathic pain in rats after oral administration.

Renal artery stenosis in chronic renal failure: caution is advised for percutaneous revascularization.

INTRODUCTION: Percutaneous intervention for renal artery stenosis may lead to acute deterioration of renal function and, consequently, of a patient's well-being. The purpose of this study was to determine whether selection by indication for renal artery stenosis was predictive of outcome. METHODS: All patients who underwent intervention for renal artery stenosis were selected to participate in the study and their indication for intervention was determined. Patient characteristics, i.e., renal function and clearance by modified diet in renal disease (MDRD), blood pressure and its treatment, kidney size, proteinuria, and cardiovascular events, were recorded before intervention, at 1 year, and at the end of follow-up. An intervention was classified as a success, no change, or a failure with respect to the indication for intervention. Successful interventions were compared to failures with respect to indication and patient characteristics. RESULTS: Twenty-four patients were included in the study: 11 for renal failure (RF), 9 for hypertension (HT), and 4 due to flash pulmonary edema (FPE). One patient with RF, four with HT, and one with FPE benefited from intervention. Nine patients with RF and two with HT were classified as failures. Failure was most prevalent in the RF group (p<0.05). Other predictors of failure were older age (p<0.02), worse renal function (p<0.02), smaller kidneys (p<0.03), and previous cardiovascular events (p<0.05). CONCLUSIONS: Renal failure must be considered a contraindication for intervention in renal artery stenosis. Intervention can be considered in FPE and hypertension, provided other predictive factors for failure are absent.

Naphthalen-1-yl-(4-pentyloxynaphthalen-1-yl)methanone: a potent, orally bioavailable human CB1/CB2 dual agonist with antihyperalgesic properties and restricted central nervous system penetration.

Selective activation of peripheral cannabinoid CB1 receptors has the potential to become a valuable therapy for chronic pain conditions as long as central nervous system effects are attenuated. A new class of cannabinoid ligands was rationally designed from known aminoalkylindole agonists and showed good binding and functional activities at human CB1 and CB2 receptors. This has led to the discovery of a novel CB1/CB2 dual agonist, naphthalen-1-yl-(4-pentyloxynaphthalen-1-yl)methanone (13), which displays good oral bioavailability, potent antihyperalgesic activity in animal models, and limited brain penetration.

Nitrite-derived nitric oxide protects the rat kidney against ischemia/reperfusion injury in vivo: role for xanthine oxidoreductase.

In normal conditions, nitric oxide (NO) is oxidized to the anion nitrite, but in hypoxia, this nitrite may be reduced back to NO by the nitrite reductase action of deoxygenated hemoglobin, acidic disproportionation, or xanthine oxidoreductase (XOR). Herein, is investigated the effects of topical sodium nitrite administration in a rat model of renal ischemia/reperfusion (I/R) injury. Rats were subjected to 60 min of bilateral renal ischemia and 6 h of reperfusion in the absence or presence of sodium nitrite (30 nmol) administered topically 1 min before reperfusion. Serum creatinine, serum aspartate aminotransferase, creatinine clearance, fractional excretion of Na(+), and plasma nitrite/nitrate concentrations were measured. The nitrite-derived NO-generating capacity of renal tissue was determined under acidic and hypoxic conditions by ozone chemiluminescence in homogenates of kidneys that were subjected to sham, ischemia-only, and I/R conditions. Nitrite significantly attenuated renal dysfunction and injury, an effect that was abolished by previous treatment of rats with the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazole-1-oxyl-3-oxide (2.5 mumol intravenously 5 min before ischemia and 50 nmol topically 6 min before reperfusion). Renal tissue homogenates produced significant amounts of NO from nitrite, an effect that was attenuated significantly by the xanthine oxidoreductase inhibitor allopurinol. Taken together, these findings demonstrate that topically administered sodium nitrite protects the rat kidney against I/R injury and dysfunction in vivo via the generation, in part, of xanthine oxidoreductase-catalyzed NO production. These observations suggest that nitrite therapy might prove beneficial in protecting kidney function and integrity during periods of I/R such as those encountered in renal transplantation.

Identification and biological characterization of 6-aryl-7-isopropylquinazolinones as novel TRPV1 antagonists that are effective in models of chronic pain.

Vanilloid receptor 1 (VR1, TRPV1) is a cation-selective ion channel that is expressed on primary afferent neurons and is upregulated following inflammation and nerve damage. Blockers of this channel may have utility in the treatment of chronic nociceptive and neuropathic pain. Here, we describe the optimization from a high throughput screening hit, of a series of 6-aryl-7-isopropylquinazolinones that are TRPV1 antagonists in vitro. We also demonstrate that one compound is active in vivo against capsaicin-induced hyperalgesia and in models of neuropathic and nociceptive pain in the rat.

Antihyperalgesic properties of the cannabinoid CT-3 in chronic neuropathic and inflammatory pain states in the rat.

CT-3 (ajulemic acid) is a synthetic analogue of a metabolite of Delta9-tetrahydrocannabinol that has reported analgesic efficacy in neuropathic pain states in man. Here we show that CT-3 binds to human cannabinoid receptors in vitro, with high affinity at hCB1 (Ki 6 nM) and hCB2 (Ki 56 nM) receptors. In a functional GTP-gamma-S assay CT-3 was an agonist at both hCB1 and hCB2 receptors (EC50 11 and 13.4 nM, respectively). In behavioural models of chronic neuropathic and inflammatory pain in the rat, oral administration of CT-3 (0.1-1 mg/kg) produced up to 60% reversal of mechanical hyperalgesia. In both models the antihyperalgesic activity was prevented by the CB1-antagonist SR141716A but not the CB2-antagonist SR144528. In the tetrad of tests for CNS activity, CT-3 (1-10 mg/kg, po) produced dose-related catalepsy, deficits in locomotor performance, hypothermia, and acute analgesia. Comparison of 50% maximal effects in the tetrad and chronic pain assays produced an approximate therapeutic index of 5-10. Pharmacokinetic analysis showed that CT-3 exhibits significant but limited brain penetration, with a brain/plasma ratio of 0.4 measured following oral administration, compared to ratios of 1.0-1.9 measured following subcutaneous administration of WIN55,212-2 or Delta9-THC. These data show that CT-3 is a cannabinoid receptor agonist and is efficacious in animal models of chronic pain by activation of the CB1 receptor. Whilst it shows significant cannabinoid-like CNS activity, it exhibits a superior therapeutic index compared to other cannabinoid compounds, which may reflect a relatively reduced CNS penetration.

Anandamide-evoked activation of vanilloid receptor 1 contributes to the development of bladder hyperreflexia and nociceptive transmission to spinal dorsal horn neurons in cystitis.

The role of anandamide in the development of inflammatory hyperalgesia and visceral hyperreflexia was studied in the rat urinary bladder. Animals were given intraperitoneal cyclophosphamide injection, which evokes painful hemorrhagic cystitis accompanied by increased bladder reflex activity. The vanilloid receptor 1 [transient receptor potential vanilloid 1 (TRPV1)] antagonist capsazepine, applied onto the serosal surface of bladders, significantly reduced the hyperreflexia. Mass spectrometric analysis revealed that cyclophosphamide injection significantly and persistently increased the anandamide content of bladder tissues. The increase in the anandamide content paralleled the development of reflex hyperactivity. Anandamide (1-100 microm), applied onto the serosal surface of naive bladders, increased the reflex activity in a concentration-dependent manner. Repeated anandamide applications did not produce desensitization of the response. The anandamide-evoked effect was blocked by capsazepine or by instillation of resiniferatoxin, the ultrapotent TRPV1 agonist, into the bladders 24 hr before the anandamide challenge. The cannabinoid 1 receptor antagonist SR141716A [N-piperidino-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methylpyrazole-3-carboxamide] significantly increased the potency of anandamide in enhancing bladder reflex activity in naive but not in cyclophosphamide-injected animals. Application of the fatty acid amide hydrolyze inhibitor palmitoylisopropylamine onto the serosal surface of bladders also increased the reflex activity both in naive and cyclophosphamide-injected rats. This latter effect in naive animals was blocked by capsazepine and by resiniferatoxin pretreatment. Finally, intravesical instillation of anandamide (50 microm) increased c-fos expression in the spinal cord, which was reduced by capsazepine or by resiniferatoxin pretreatment. These results suggest that anandamide, through activating TRPV1, contributes to the development of hyperreflexia and hyperalgesia during cystitis.