Metatranscriptomics sheds light on the links between the functional traits of fungal guilds and ecological processes in forest soil ecosystems.

Soil fungi belonging to different functional guilds, such as saprotrophs, pathogens, and mycorrhizal symbionts, play key roles in forest ecosystems. To date, no study has compared the actual gene expression of these guilds in different forest soils. We used metatranscriptomics to study the competition for organic resources by these fungal groups in boreal, temperate, and Mediterranean forest soils. Using a dedicated mRNA annotation pipeline combined with the JGI MycoCosm database, we compared the transcripts of these three fungal guilds, targeting enzymes involved in C- and N mobilization from plant and microbial cell walls. Genes encoding enzymes involved in the degradation of plant cell walls were expressed at a higher level in saprotrophic fungi than in ectomycorrhizal and pathogenic fungi. However, ectomycorrhizal and saprotrophic fungi showed similarly high expression levels of genes encoding enzymes involved in fungal cell wall degradation. Transcripts for N-related transporters were more highly expressed in ectomycorrhizal fungi than in other groups. We showed that ectomycorrhizal and saprotrophic fungi compete for N in soil organic matter, suggesting that their interactions could decelerate C cycling. Metatranscriptomics provides a unique tool to test controversial ecological hypotheses and to better understand the underlying ecological processes involved in soil functioning and carbon stabilization.

ENSO and NAO affect long-term leaf litter dynamics and stoichiometry of Scots pine and European beech mixedwoods.

Litterfall dynamics (production, seasonality and nutrient composition) are key factors influencing nutrient cycling. Leaf litter characteristics are modified by species composition, site conditions and water availability. However, significant evidence on how large-scale, global circulation patterns affect ecophysiological processes at tree and ecosystem level remains scarce due to the difficulty in separating the combined influence of different factors on local climate and tree phenology. To fill this gap, we studied links between leaf litter dynamics with climate and other forest processes, such as tree-ring width (TRW) and intrinsic water-use efficiency (iWUE) in two mixtures of Scots pine (Pinus sylvestris L.) and European beech (Fagus sylvatica L.) in the south-western Pyrenees. Temporal series (18 years) of litterfall production and elemental chemical composition were decomposed following the ensemble empirical mode decomposition method and relationships with local climate, large-scale climatic indices, TRW and Scots pine's iWUE were assessed. Temporal trends in N:P ratios indicated increasing P limitation of soil microbes, thus affecting nutrient availability, as the ecological succession from a pine-dominated to a beech-dominated forest took place. A significant influence of large-scale patterns on tree-level ecophysiology was explained through the impact of the North Atlantic Oscillation (NAO) and El Niño-Southern Oscillation (ENSO) on water availability. Positive NAO and negative ENSO were related to dry conditions and, consequently, to early needle shedding and increased N:P ratio of both species. Autumn storm activity appears to be related to premature leaf abscission of European beech. Significant cascading effects from large-scale patterns on local weather influenced pine TRW and iWUE. These variables also responded to leaf stoichiometry fallen 3 years prior to tree-ring formation. Our results provide evidence of the cascading effect that variability in global climate circulation patterns can have on ecophysiological processes and stand dynamics in mixed forests.

[Onychomycosis by yeast not common in diabetics of a health center]. / Onicomicosis por levaduras no comunes en diabéticos de un centro de salud.

BACKGROUND: Mexican diabetic population frequently presents mycosis under foot hyperkeratosis; however, in another type of onychomycosis as the ones that is assumed Candida albicans is the causal agent, it is unknown the frequency, the prevalence and if another Candida species or other yeasts are found. OBJECTIVE: Evaluate the frequency of yeasts causing onychomycosis in diabetic patients looked after in public institutions of health of the State of Hidalgo, Mexico, and its association with clinical epidemiological variables. MATERIALS AND METHODS: An observational, descriptive and transversal study was made on 261 patients, from which one nail sample of each one was obtained, used to isolate and identify dermatophytes and yeasts; the results were statistically correlated with 24 epidemiological parameters. The clinical study was done through interrogation and by medical exploration in order to evaluate Tinea pedis and onychomycosis. RESULTS: Onychomycosis were caused by Candida guilliermondii, Candida parapsilosis, Candida glabrata, Candida krusei, Candida spp., Kodamaea ohmeri, Prototheca wickerhamii and unidentified yeasts. The prevalence for general onychomycosis, by dermatophytes, mixed onychomycosis and by yeasts were: 24.1, 19.5, 2.3 and 14.6%, respectively. Patients with significant probability to be diagnosed as having onychomycosis by yeasts are those wearing open shoes (2.59%); technicians and professionals (10.49%) and alcohol drinkers (3.72%). CONCLUSION: The fact that Candida albicans is not present in this study as causal agent of onychomycosis, and emerging and non-common yeasts were indeed isolated, creates new challenges. It is remarked the clinical criterion that when onychomycosis is suspected in diabetics, the diagnosis for culturing dermatophytes and yeasts should be included.

The neuroergonomic evaluation of human machine interface design in air traffic control using behavioral and EGG/ERP measures.

The Air Traffic Control (ATC) environment is complex and safety-critical. Whilst exchanging information with pilots, controllers must also be alert to visual notifications displayed on the radar screen (e.g., warning which indicates a loss of minimum separation between aircraft). Under the assumption that attentional resources are shared between vision and hearing, the visual interface design may also impact the ability to process these auditory stimuli. Using a simulated ATC task, we compared the behavioral and neural responses to two different visual notification designs--the operational alarm that involves blinking colored "ALRT" displayed around the label of the notified plane ("Color-Blink"), and the more salient alarm involving the same blinking text plus four moving yellow chevrons ("Box-Animation"). Participants performed a concurrent auditory task with the requirement to react to rare pitch tones. P300 from the occurrence of the tones was taken as an indicator of remaining attentional resources. Participants who were presented with the more salient visual design showed better accuracy than the group with the suboptimal operational design. On a physiological level, auditory P300 amplitude in the former group was greater than that observed in the latter group. One potential explanation is that the enhanced visual design freed up attentional resources which, in turn, improved the cerebral processing of the auditory stimuli. These results suggest that P300 amplitude can be used as a valid estimation of the efficiency of interface designs, and of cognitive load more generally.

A neurophysiological training evaluation metric for air traffic management.

The aim of this work was to analyze the possibility to apply a neuroelectrical cognitive metrics for the evaluation of the training level of subjects during the learning of a task employed by Air Traffic Controllers (ATCos). In particular, the Electroencephalogram (EEG), the Electrocardiogram (ECG) and the Electrooculogram (EOG) signals were gathered from a group of students during the execution of an Air Traffic Management (ATM) task, proposed at three different levels of difficulty. The neuroelectrical results were compared with the subjective perception of the task difficulty obtained by the NASA-TLX questionnaires. From these analyses, we suggest that the integration of information derived from the power spectral density (PSD) of the EEG signals, the heart rate (HR) and the eye-blink rate (EBR) return important quantitative information about the training level of the subjects. In particular, by focusing the analysis on the direct and inverse correlation of the frontal PSD theta (4-7 (Hz)) and HR, and of the parietal PSD alpha (10-12 (Hz)) and EBR, respectively, with the degree of mental and emotive engagement, it is possible to obtain useful information about the training improvement across the training sessions.

Natural killer cells and malignant haemopathies: a model for the interaction of cancer with innate immunity.

Despite recent progress in the therapeutic approach of malignant haemopathies, their prognoses remain frequently poor. Immunotherapy offers an alternative of great interest in this context but defect or abnormal expression of human leukocyte antigens (HLA), frequently observed in cancer cells, limits its efficiency. Natural killer (NK) cells, which are able to kill target cells in a HLA-independent way, represent a novel tool in the treatment of haematological malignancies. Abnormal NK cytolytic function is observed in all the haematological malignancies studied, such as acute leukaemia, myelodysplastic syndromes or chronic myeloid/lymphoid leukaemia. Several mechanisms are involved in the alterations of NK cytotoxicity: decreased expression of activating receptors, increased expression of inhibitory receptors or defective expression of NK ligands on target cells. Further studies are needed to identify how each type of haematological malignancy escapes from the innate immune response. Attempts to increase the expression of activating receptors, to counteract inhibitory receptors expression, or to increase NK cell cytotoxic capacities could overcome tumour escape from innate immunity. These therapies are based on monoclonal antibodies or culture of NK cells in presence of cytokines or dendritic cells. Moreover, many novel drugs used in haematological malignancies [tyrosine kinase inhibitors, IMIDs(®), proteasome inhibitors, demethylating agents, histone deacetylase inhibitors (HDACis), histamine dihydrochloride] display interesting immunomodulatory properties that affect NK cells. These data suggest that combined modalities associating cytotoxic drugs with innate immunity modulators may represent a major breakthrough in tumour eradication.

Thinning affects nutrient resorption and nutrient-use efficiency in two Pinus sylvestris stands in the Pyrenees.

Needle chemical composition was measured, and nutrient resorption, nutrient-use efficiency (NUE), and other indexes were estimated for 24 months in two contrasting natural Pinus sylvestris L. forests in the western Pyrenees in Spain. For each location (Aspurz, 650 m elevation, 7% slope; Garde, 1335 m elevation, 40% slope), there were three reference plots (P0), three plots with 20% of the basal area removed (P20), and three with 30% of the basal area removed (P30). Needle P, Ca, and Mg concentrations were higher in Garde, but N concentration was higher for Aspurz, without differences for K. Nutrient-resorption efficiency of P was higher in Aspurz, of Mg higher in Garde, and there were no differences between sites in N and K. Nutrient-resorption proficiency was significantly higher in the site with lower soil nutrient availability, i.e., for P, Ca, and Mg in Aspurz, but N in Garde (no differences in K); this may be an indicator of nutrient conservation strategy. Annual nutrient productivity (A) was higher for all nutrients in Aspurz, whereas the mean residence time (MRT) was higher in Garde in all nutrients but P. NUE was significantly higher in Garde for all nutrients but P, which was more efficiently used in Aspurz. In both sites, N, P, and K concentrations were higher in the 2002 cohort, Ca in the 2000 cohort, and maximum Mg was found in the 2001 cohort. Thinning caused a reduction of Mg concentration in the 2001 cohort in Aspurz, an increase of Ca resorption proficiency in Aspurz and Mg resorption at both sites, and reduction of P, K, and Mg nutrient response efficiency (NRE) in Garde. Thinning may have caused an increase of the C:Mg ratio through facilitating the development of more biosynthesis apparatus in a more illuminated canopy, but it seemed not to affect resorption in a significant way. Changes in NRE in Garde after thinning show that forest management can affect how trees use nutrients. Our results indicate that the strategy to optimize NUE is different in each stand. In Aspurz (a Mediterranean ecosystem), pine trees carried out resorption more efficiently, while in Garde (a continental forest), trees used nutrients for longer periods of time and reduced their efficiency in using the available soil nutrients after reduced competition by thinning.

[Imaging of the postoperative knee in athletes: articular cartilage, menisci, and ligaments]. / Imagerie du genou du sportif opéré: cartilage, ménisques et ligaments.

This article focuses on the common surgical procedures and reviews the clinical and imaging features of the postoperative knee following prior surgical treatment of meniscal, articular cartilage, and ligamentous injuries.

New separation method for organic and inorganic selenium compounds based on anion exchange chromatography followed by inductively coupled plasma mass spectrometry.

We describe a new method for separating the organic and inorganic selenocompounds methaneseleninic acid, selenite, selenate, methylselenocysteine, selenocystine as well as both selenomethionine and its oxidized form. The separation is performed on a Hamilton PRP-X100 column. According to the literature, the oxidized form of selenomethionine-which is easily formed-is eluted close to the dead volume when this column is used. The choice of parahydroxybenzoic acid as mobile phase enabled us to elute all of these species after this oxidized form, resulting in better identification and quantification. The factors determining separation (eluent concentration, pH, gradient) were optimized via an experimental design. Application of the method to yeast and commercial tablets showed that the principal Se compound present was selenomethionine, which was also present in its oxidized form.

Vanin-1-/- mice exhibit a glutathione-mediated tissue resistance to oxidative stress.

Vanin-1 is an epithelial ectoenzyme with pantetheinase activity and generating the amino-thiol cysteamine through the metabolism of pantothenic acid (vitamin B(5)). Here we show that Vanin-1(-/-) mice, which lack cysteamine in tissues, exhibit resistance to oxidative injury induced by whole-body gamma-irradiation or paraquat. This protection is correlated with reduced apoptosis and inflammation and is reversed by treating mutant animals with cystamine. The better tolerance of the Vanin-1(-/-) mice is associated with an enhanced gamma-glutamylcysteine synthetase activity in liver, probably due to the absence of cysteamine and leading to elevated stores of glutathione (GSH), the most potent cellular antioxidant. Consequently, Vanin-1(-/-) mice maintain a more reducing environment in tissue after exposure to irradiation. In normal mice, we found a stress-induced biphasic expression of Vanin-1 regulated via antioxidant response elements in its promoter region. This process should finely tune the redox environment and thus change an early inflammatory process into a late tissue repair process. We propose Vanin-1 as a key molecule to regulate the GSH-dependent response to oxidative injury in tissue at the epithelial level. Therefore, Vanin/pantetheinase inhibitors could be useful for treatment of damage due to irradiation and pro-oxidant inducers.

Novel CD28-responsive enhancer activated by CREB/ATF and AP-1 families in the human interleukin-2 receptor alpha-chain locus.

The interaction of interleukin-2 (IL-2) with its receptor (IL-2R) critically regulates the T-cell immune response, and the alpha chain CD25/IL-2Ralpha is required for the formation of the high-affinity receptor. Tissue-specific, inducible expression of the IL-2Ralpha gene is regulated by at least three positive regulatory regions (PRRI, PRRII, and PRRIII), but none responded to CD28 engagement in gene reporter assays although CD28 costimulation strongly amplifies IL-2Ralpha gene transcription. By DNase I hypersensitivity analysis, we have identified a novel TCR-CD3- and CD28-responsive enhancer (CD28rE) located 8.5 kb 5' of the IL-2Ralpha gene. PRRIV/CD28rE contains a functional CRE/TRE element required for CD28 signaling. The T-cell-specific, CD28-responsive expression of the IL-2Ralpha gene appears controlled through PRRIV/CD28rE by cooperation of CREB/ATF and AP-1 family transcription factors.

Structure-activity relationship studies of melanin-concentrating hormone (MCH)-related peptide ligands at SLC-1, the human MCH receptor.

Melanin-concentrating hormone (MCH) is a cyclic nonadecapeptide involved in the regulation of feeding behavior, which acts through a G protein-coupled receptor (SLC-1) inhibiting adenylcyclase activity. In this study, 57 analogues of MCH were investigated on the recently cloned human MCH receptor stably expressed in HEK293 cells, on both the inhibition of forskolin-stimulated cAMP production and guanosine-5'-O-(3-[(35)S]thiotriphosphate ([(35)S]- GTPgammaS) binding. The dodecapeptide MCH-(6-17) (MCH ring between Cys(7) and Cys(16), with a single extra amino acid at the N terminus (Arg(6)) and at the C terminus (Trp(17))) was found to be the minimal sequence required for a full and potent agonistic response on cAMP formation and [(35)S]- GTPgammaS binding. We Ala-scanned this dodecapeptide and found that only 3 of 8 amino acids of the ring, namely Met(8), Arg(11), and Tyr(13), were essential to elicit full and potent responses in both tests. Deletions inside the ring led either to inactivity or to poor antagonists with potencies in the micromolar range. Cys(7) and Cys(16) were substituted by Asp and Lys or one of their analogues, in an attempt to replace the disulfide bridge by an amide bond. However, those modifications were deleterious for agonistic activity. In [(35)S]- GTPgammaS binding, these compounds behaved as weak antagonists (K(B) 1-4 microm). Finally, substitution in MCH-(6-17) of 6 out of 12 amino acids by non-natural residues and concomitant replacement of the disulfide bond by an amide bond led to three compounds with potent antagonistic properties (K(B) = 0.1-0.2 microm). Exploitation of these structure-activity relationships should open the way to the design of short and stable MCH peptide antagonists.

Specific deficiency of p56lck expression in T lymphocytes from type 1 diabetic patients.

Peripheral T lymphocyte activation in response to TCR/CD3 stimulation is reduced in type 1 diabetic patients. To explore the basis of this deficiency, a comprehensive analysis of the signal transduction pathway downstream of the TCR/CD3 complex was performed for a cohort of patients (n = 38). The main result of the study shows that T cell hyporesponsiveness is positively correlated with a reduced amount of p56(lck) in resting T lymphocytes. Upon CD3-mediated activation, this defect leads to a hypophosphorylation of the CD3zeta-chain and few other polypeptides without affecting the recruitment of ZAP70. Other downstream effectors of the TCR/CD3 transduction machinery, such as phosphatidylinositol 3-kinase p85alpha, p59(fyn), linker for activation of T cells (LAT), and phospholipase C-gamma1, are not affected. In some patients, the severity of this phenotypic deficit could be linked to low levels of p56(lck) mRNA and resulted in the failure to efficiently induce the expression of the CD69 early activation marker. We propose that a primary deficiency in human type 1 diabetes is a defect in TCR/CD3-mediated T cell activation due to the abnormal expression of the p56(lck) tyrosine kinase.

TCRalpha enhancer activation occurs via a conformational change of a pre-assembled nucleo-protein complex.

The TCR alpha enhancer (Ealpha) has served as a paradigm for studying how enhancers organize trans-activators into nucleo-protein complexes thought to recruit and synergistically stimulate the transcriptional machinery. Little is known, however, of either the extent or dynamics of Ealpha occupancy by nuclear factors during T cell development. Using dimethyl sulfate (DMS) in vivo footprinting, we demonstrate extensive Ealpha occupancy, encompassing both previously identified and novel sites, not only in T cells representing a developmental stage where Ealpha is known to be active (CD4(+)CD8(+)-DP cells), but surprisingly, also in cells at an earlier developmental stage where Ealpha is not active (CD4(-)CD8(-)-DN cells). Partial occupancy was also established in B-lymphoid but not non-lymphoid cells. In vivo DNase I footprinting, however, implied developmentally induced changes in nucleo-protein complex topography. Stage-specific differences in factor composition at Ealpha sequences were also suggested by EMSA analysis. These results, which indicate that alterations in the structure of a pre-assembled nucleo-protein complex correlate with the onset of Ealpha activity, may exemplify one mechanism by which enhancers can rapidly respond to incoming stimuli.

IL-2 and long-term T cell activation induce physical and functional interaction between STAT5 and ETS transcription factors in human T cells.

Activation of Stat5 by many cytokines implies that it cannot alone insure the specificity of the regulation of its target genes. We have evidenced a physical and functional interaction between members of two unrelated transcription factor families, Ets-1, Ets-2 and Stat5, which could contribute to the proliferative response to interleukin 2. Competition with GAS- and EBS-specific oligonucleotides and immunoassays with a set of anti-Stat and anti-Ets families revealed that the IL-2-induced Stat5-Ets complex recognizes several GAS motifs identified as target sites for activated Stat5 dimers. Coimmunoprecipitation experiments evidenced that a Stat5/Ets-1/2 complex is formed in vivo in absence of DNA. GST-pull down experiments demonstrated that the C-terminal domain of Ets-1 is sufficient for this interaction in vitro. Cotransfection experiments in Kit225 T cells resulted in cooperative transcriptional activity between both transcription factors in response to a combination of IL-2, PMA and ionomycin. A Stat5-Ets protein complex was the major inducible DNA-binding complex bound to the human IL-2rE GASd/EBSd motif in long-term proliferating normal human T cells activated by CD2 and CD28. These results suggest that the inducible Stat5-Ets protein interaction plays a role in the regulation of gene expression in response to IL-2 in human T lymphocytes.

[Treatment of chronic postinfectious fatigue: randomized double-blind study of two doses of sulbutiamine (400-600 mg/day) versus placebo]. / Traitement de l'asthénie en période postinfectieuse: étude randomisée en double aveugle de deux doses de sulbutiamine (400-600 mg/j) versus placebo.

PURPOSE: Chronic fatigue remains a medical mystery and a therapeutic failure. The subgroup of chronic fatigue postinfectious fatigue (CPIF) is an interesting one since it is quite frequent in general practice. METHODS: We studied sulbutiamine (Su), isobutyryl-thiamine disulfide in this context. We included 326 general-practice patients suffering from CPIF: they received randomly either Su, 400 mg daily (n = 106), or Su, 600 mg daily (n = 111), or placebo (n = 109) for 28 days in a double-blind, parallel-group study. 315 patients completed the study. RESULTS: The evaluation of fatigue, by multiple means including mainly MFI, a validated multidimensional fatigue scale, showed overall no significant difference between the groups. On the 7th day, however, women receiving Su, 600 mg had less fatigue (P < 0.01), but the figures were quite diverse and no persistent effect was noted at the 28th day. CONCLUSION: Thus, we showed for the first time that a high level general-practice study of fatigue is feasible using specific tools. Whether the effect observed after 1 week in women represents a true finding needs additional research. Further studies are in progress in order to characterize better the potential usefulness of Su in chronic fatigue.

[A paraneoplastic acrosyndrome]. / Un acrosyndrome paranéoplasique.

BACKGROUND: Paraneoplastic digital ischemia is an uncommon complication of metastatic adenocarcinomas. CASE REPORT: Two years after remission of an uterine adenocarcinoma, the patient developed an acrosyndrome involving all four limbs with digital ischemia. Recurrent carcinoma was evidenced by a very high antinuclear antibody titer. Chemotherapy improved the acrosyndrome. DISCUSSION: Vasomotor disorders which developed in older subjects with no other signs of autoimmune disorders should suggest a neoplastic origin. Icshemia of the fingers would be caused by vasculitis. An elevated antinuclear antibody titer may be a supplementary argument suggesting a neoplastic etiology.

Physical interaction of the bHLH LYL1 protein and NF-kappaB1 p105.

The LYL1 gene was first identified upon the molecular characterization of the t(7;9)(q35;p13) translocation associated with some human T-cell acute leukemias (T-ALLs). In adult tissues, LYL1 expression is restricted to hematopoietic cells with the notable exclusion of the T cell lineage. LYL1 encodes a basic helix-loop-helix (bHLH) protein highly related to TAL-1, whose activation is also associated with a high proportion of human T-ALLs. A yeast two-hybrid system was used to identify proteins that specifically interact with LYL1 and might mediate its activities. We found that p105, the precursor of NF-kappaB1 p50, was the major LYL1-interacting protein in this system. The association between LYL1 and p105 was confirmed both in vitro and in vivo in mammalian cells. Biochemical studies indicated that the interaction was mediated by the bHLH motif of LYL1 and the ankyrin-like motifs of p105. Ectopic expression of LYL1 in a human T cell line caused a significant decrease in NF-kappaB-dependent transcription, associated with a reduced level of NF-kappaB1 proteins.

[Access to social coverage of uninsured patients attending a hospital clinic: a historical cohort study at the Baudelaire outpatient clinic in Paris]. / Accès à une couverture sociale des patients non assurés consultant en milieu hospitalier: une étude de cohorte historique au sein de la consultation "Baudelaire" à Paris.

BACKGROUND: In France health insurance coverage is universal (see note at the end of the text), nevertheless some people remain uninsured. In this high-risk population, the lack of insurance coverage contributes to the aggravation of health, by reducing access to medical care. In 1992, the Baudelaire consultation was incorporated into the outpatient clinic of Saint-Antoine hospital (Paris, France), to provide the uninsured with the same access as any other patient--but free of charge--to medical care. Social care was also provided in particular by assisting the uninsured in applying for insurance coverage. Our objectives were to quantify the delay in obtaining insurance coverage and to study whether the sociodemographic characteristics of these patients were associated with inequalities in terms of delays. METHODS: All patients attending the consultation for the first time in 1994 were included (n = 623). Because of differences linked to the French social security system, analysis was performed into two groups according to the existence of a prior insurance coverage. Delay in obtaining or recovering insurance coverage was considered as the key variable. The socio-demographic factors linked to the rates of access to insurance coverage were determined using Cox proportional hazards regression models. We also examined the factors linked with the existence of a prior insurance coverage by logistic regression modeling. RESULTS: Within one year 96% of the patients who had had insurance coverage in the past, and 63% of the patients who had not, were insured. No factor, whether nationality, educational level, socio-professional category, family situation, type of housing, made of income was found to be linked with obtaining or recovering insurance coverage. However, nearly all these factors were related with the existence of prior insurance coverage. CONCLUSIONS: Our approach of systematically providing social care allows 70% of uninsured patients to obtain insurance coverage within one year. This approach probably contributes to an improvement by facilitating access to mainstream health care. Moreover, no difference in delay in obtaining insurance coverage was found associated with sociodemographic characteristics.

NPY receptor subtypes involved in the contraction of the proximal colon of the rat.

Experiments were designed to determine the receptor subtype(s) involved in the contraction of the rat proximal colon to NPY. In this tissue, mRNA of Y2 and Y4 NPY receptor subtypes were highly expressed, whereas Y5 mRNA levels were very low and Y1 mRNA levels were intermediate. NPY analogues induced contractions with the following order of potency: rPP > hPP = PYY = NPY = [Leu31,Pro34]NPY > NPY(2-36) = [D-Trp32]NPY > NPY(33-36). Responses to NPY, PYY and NPY(13-36) were not or partially affected by tetrodotoxin, in contrast to the responses to [Leu31,Pro34]NPY, rPP, hPP and [D-Trp32]NPY which were fully blocked. Atropine did not inhibit the contractions to NPY, PYY and [Leu31,Pro34]NPY but significantly affected those to NPY(13-36), [D-Trp32]NPY, rPP and hPP. The specific Y1 receptor antagonist BIBP 3226 was ineffective but JCF 104 and JCF 105 (two compounds with preferential affinity toward the hY5 receptor versus the hY1 or hY2 receptor) abolished the contractions provoked by the NPY analogues. These results suggest that NPY activates three receptor subtypes, a Y2 subtype possibly by a direct action on the smooth muscle cells, as well as a Y4 and a Y5 (or 'Y5-like') subtype which, respectively, release acetylcholine and an unknown neurotransmitter.