A novel role for MLC1 in regulating astrocyte-synapse interactions.

Loss of function of the astrocyte membrane protein MLC1 is the primary genetic cause of the rare white matter disease Megalencephalic Leukoencephalopathy with subcortical Cysts (MLC), which is characterized by disrupted brain ion and water homeostasis. MLC1 is prominently present around fluid barriers in the brain, such as in astrocyte endfeet contacting blood vessels and in processes contacting the meninges. Whether the protein plays a role in other astrocyte domains is unknown. Here, we show that MLC1 is present in distal astrocyte processes, also known as perisynaptic astrocyte processes (PAPs) or astrocyte leaflets, which closely interact with excitatory synapses in the CA1 region of the hippocampus. We find that the PAP tip extending toward excitatory synapses is shortened in Mlc1-null mice. This affects glutamatergic synaptic transmission, resulting in a reduced rate of spontaneous release events and slower glutamate re-uptake under challenging conditions. Moreover, while PAPs in wildtype mice retract from the synapse upon fear conditioning, we reveal that this structural plasticity is disturbed in Mlc1-null mice, where PAPs are already shorter. Finally, Mlc1-null mice show reduced contextual fear memory. In conclusion, our study uncovers an unexpected role for the astrocyte protein MLC1 in regulating the structure of PAPs. Loss of MLC1 alters excitatory synaptic transmission, prevents normal PAP remodeling induced by fear conditioning and disrupts contextual fear memory expression. Thus, MLC1 is a new player in the regulation of astrocyte-synapse interactions.

Faecal carriage, risk factors, acquisition and persistence of ESBL-producing Enterobacteriaceae in dogs and cats and co-carriage with humans belonging to the same household.

BACKGROUND: ESBL-producing Enterobacteriaceae (ESBL-E) are observed in many reservoirs. Pets might play an important role in the dissemination of ESBL-E to humans since they live closely together. OBJECTIVES: To identify prevalence, risk factors, molecular characteristics, persistence and acquisition of ESBL-E in dogs and cats, and co-carriage in human-pet pairs belonging to the same household. METHODS: In a nationwide study, one person per household was randomly invited to complete a questionnaire and to submit a faecal sample. Dog and cat owners were invited to also submit a faecal sample from their pet. Repeated sampling after 1 and 6 months was performed in a subset. ESBL-E were obtained through selective culture and characterized by WGS. Logistic regression analyses and random forest models were performed to identify risk factors. RESULTS: The prevalence of ESBL-E carriage in these cohorts was 3.8% (95% CI: 2.7%-5.4%) for human participants (n=550), 10.7% (95% CI: 8.3%-13.7%) for dogs (n=555) and 1.4% (95% CI: 0.5%-3.8%) for cats (n=285). Among animals, blaCTX-M-1 was most abundant, followed by blaCTX-M-15. In dogs, persistence of carriage was 57.1% at 1 month and 42.9% at 6 months. Eating raw meat [OR: 8.8, 95% CI: 4.7-16.4; population attributable risk (PAR): 46.5%, 95% CI: 41.3%-49.3%] and dry food (OR: 0.2, 95% CI: 0.1-0.5; PAR: 56.5%, 95% CI: 33.2%-66.6%) were predictors for ESBL-E carriage in dogs. Human-dog co-carriage was demonstrated in five households. Human-cat co-carriage was not observed. CONCLUSIONS: ESBL-E prevalence was higher in dogs than in humans and lowest in cats. The main risk factor for ESBL-E carriage was eating raw meat. Co-carriage in dogs and household members was uncommon.

An automated home-cage-based 5-choice serial reaction time task for rapid assessment of attention and impulsivity in rats.

RATIONALE: The 5-choice serial reaction time task (5-CSRTT) is a widely used operant task for measuring attention and motor impulsivity in rodents. Training animals in this task requires an extensive period of daily operant sessions. Recently, a self-paced, automated version of this task has been developed for mice, which substantially reduces training time. Whether a similar approach is effective for rats is currently unknown. OBJECTIVE: Here, we tested whether attention and impulsivity can be assessed in rats with a self-paced version of the 5-CSRTT. METHODS: Operant boxes were connected to home-cages with tunnels. Two groups of rats self-paced their training by means of an automated script. The first group of animals was allowed unlimited access (UA) to start trials in the task; for the second group, trial availability was restricted to the first 2.5 h of the dark cycle (TR). Task parameter manipulations, such as variable inter-trial intervals and stimulus durations as well as pharmacological challenges with scopolamine, were tested to validate the task. RESULTS: Self-paced training took less than 1 week. Animals in the UA group showed higher levels of omissions compared with the TR group. In both protocols, variable inter-trial intervals increased impulsivity, and variable stimulus durations decreased attentional performance. Scopolamine affected cognitive performance in the TR group only. CONCLUSIONS: Home-cage-based training of the 5-CSRTT in rats, especially the TR protocol, presents a valid and fast alternative for measuring attention and impulsivity.

Adenosine A2A Receptors Control Glutamatergic Synaptic Plasticity in Fast Spiking Interneurons of the Prefrontal Cortex.

Adenosine A2A receptors (A2AR) are activated upon increased synaptic activity to assist in the implementation of long-term plastic changes at synapses. While it is reported that A2AR are involved in the control of prefrontal cortex (PFC)-dependent behavior such as working memory, reversal learning and effort-based decision making, it is not known whether A2AR control glutamatergic synapse plasticity within the medial PFC (mPFC). To elucidate that, we tested whether A2AR blockade affects long-term plasticity (LTP) of excitatory post-synaptic potentials in pyramidal neurons and fast spiking (FS) interneurons in layer 5 of the mPFC and of population spikes. Our results show that A2AR are enriched at mPFC synapses, where their blockade reversed the direction of plasticity at excitatory synapses onto layer 5 FS interneurons from LTP to long-term depression, while their blockade had no effect on the induction of LTP at excitatory synapses onto layer 5 pyramidal neurons. At the network level, extracellularly induced LTP of population spikes was reduced by A2AR blockade. The interneuron-specificity of A2AR in controlling glutamatergic synapse LTP may ensure that during periods of high synaptic activity, a proper excitation/inhibition balance is maintained within the mPFC.

Hippocampal extracellular matrix alterations contribute to cognitive impairment associated with a chronic depressive-like state in rats.

Patients with depression often suffer from cognitive impairments that contribute to disease burden. We used social defeat-induced persistent stress (SDPS) to induce a depressive-like state in rats and then studied long-lasting memory deficits in the absence of acute stressors in these animals. The SDPS rat model showed reduced short-term object location memory and maintenance of long-term potentiation (LTP) in CA1 pyramidal neurons of the dorsal hippocampus. SDPS animals displayed increased expression of synaptic chondroitin sulfate proteoglycans in the dorsal hippocampus. These effects were abrogated by a 3-week treatment with the antidepressant imipramine starting 8 weeks after the last defeat encounter. Next, we observed an increase in the number of perineuronal nets (PNNs) surrounding parvalbumin-expressing interneurons and a decrease in the frequency of inhibitory postsynaptic currents (IPSCs) in the hippocampal CA1 region in SDPS animals. In vivo breakdown of the hippocampus CA1 extracellular matrix by the enzyme chondroitinase ABC administered intracranially restored the number of PNNs, LTP maintenance, hippocampal inhibitory tone, and memory performance on the object place recognition test. Our data reveal a causal link between increased hippocampal extracellular matrix and the cognitive deficits associated with a chronic depressive-like state in rats exposed to SDPS.

A cross sectional and longitudinal study of endodontic and periapical status in an Australian population.

BACKGROUND: This study describes the cross sectional and longitudinal data of endodontic and periapical status of new patients presenting to a major dental hospital, and assesses the relationships between tooth-related variables with apical periodontitis. METHODS: The records of 695 patients were randomly selected and the orthopantomograms of these patients up to 31 October 2014 were reviewed by two endodontists. The periapical status of teeth was recorded using the periapical index. The presence and quality of root fillings and coronal restorations were recorded. Statistical analysis included Fleiss' kappa, Cohen's kappa and logistic regression set at P < 0.05. RESULTS: Of 695 patient records and 16 936 teeth examined, 138 (19.9%) patients or 284 (1.7%) teeth had root fillings and 179 (25.8%) patients or 325 (1.9%) teeth had apical periodontitis. Root fillings and coronal restorations were adequate in 34.6% and 69.4% teeth, respectively. A large proportion (47%) of teeth with apical periodontitis remained unchanged in subsequent orthopantomograms. CONCLUSIONS: There was lower prevalence of root filled teeth or apical periodontitis in the present study compared with international studies. The frequency of adequate root fillings must be considered unacceptably low. Teeth with apical periodontitis may remain quiescent in the absence of caries or restorative breakdown.

A practical approach to programmatic assessment design.

Assessment of complex tasks integrating several competencies calls for a programmatic design approach. As single instruments do not provide the information required to reach a robust judgment of integral performance, 73 guidelines for programmatic assessment design were developed. When simultaneously applying these interrelated guidelines, it is challenging to keep a clear overview of all assessment activities. The goal of this study was to provide practical support for applying a programmatic approach to assessment design, not bound to any specific educational paradigm. The guidelines were first applied in a postgraduate medical training setting, and a process analysis was conducted. This resulted in the identification of four steps for programmatic assessment design: evaluation, contextualisation, prioritisation and justification. Firstly, the (re)design process starts with sufficiently detailing the assessment environment and formulating the principal purpose. Key stakeholders with sufficient (assessment) expertise need to be involved in the analysis of strengths and weaknesses and identification of developmental needs. Central governance is essential to balance efforts and stakes with the principal purpose and decide on prioritisation of design decisions and selection of relevant guidelines. Finally, justification of assessment design decisions, quality assurance and external accountability close the loop, to ensure sound underpinning and continuous improvement of the assessment programme.

First Report of Goss's Bacterial Leaf Blight and Wilt of Corn Caused by Clavibacter michiganensis subsp. nebraskensis in North Dakota.

In August of 2011, the North Dakota State University Plant Diagnostic Lab received a hybrid corn (Zea mays) leaf sample from Burleigh County in south-central North Dakota (ND). The leaf had long, irregular, water-soaked lesions consistent with Goss's leaf blight of corn. Using a light microscope at 10× magnification, bacterial streaming was observed from the excised edge of leaf tissue. A bacterial suspension was created, streaked onto a semi-selective CNS medium (1), and incubated at 22°C. Dark yellow-orange colonies appeared on the medium after 5 days. Single colonies were subcultured onto additional CNS media. To verify the identity of the bacterial isolate, PCR amplification of the 16S ribosomal DNA from this isolate along with a known Clavibacter michiganensis spp. nebraskensis (Cmn) isolate collected in Indiana (4) was performed using the eubacterial universal primers 27f and 1525r (3). The 1,431-bp 16S rDNA region was obtained for each isolate and they were compared with each other and with those deposited in NCBI GenBank. Sequence alignment identified only one nucleotide difference between the ND isolate and the Indiana isolate. BLASTn search against the NCBI database showed the first 100 hits were described as C. michiganensis or unidentified Clavibacter sp. The ND isolate had a two-nucleotide difference with Cmn isolate NCPPB2581 (HE614873), and a three nucleotide difference was found with the C. michiganensis spp. michiganensis isolate NCPPB 382 (AM711867). To satisfy Koch's postulates, eight corn plants (Golden Cross Bantam) were grown in the greenhouse at 22 to 24°C. Four corn plants were inoculated at growth stage V4-V5 using a suspension of approximately 1 × 109 CFU/ml from cultures grown on CNS for 6 days. Wounds were created on the fifth leaf approximately 7 cm from the leaf tip using a tongue-seizing forceps outfitted with a rubber stopper composed of pins (2). Simultaneously, 1 ml of the bacterial suspension was delivered into the wounds through a hole on top of the rubber stopper. Four control plants were inoculated with sterile water in a similar fashion. No symptoms were observed on the control plants. After 6 days, long water-soaked symptoms were observed on leaves inoculated with the bacterial suspension. Using leaves with water-soaked lesions, the pathogen was re-isolated onto CNS media and subjected to PCR amplification, and the resulting amplicons were sequenced as before. The sequence of the amplicon from the re-isolation matched that of the original ND isolate. To our knowledge, this is the first account of Goss's leaf blight and wilt identified in ND. As the corn acreage and no-till production systems in the state have increased, the economic implications of this disease may become more significant. Recognition of symptoms and proper identification of this bacterial disease in the field should help reduce unnecessary foliar fungicide sprays. References: (1) D. C. Gross and A. K. Vidaver. Phytopathology 69:82, 1979. (2) W. A. Hagborg. Can. J. Bot. 48:1135, 1970. (3) X. Li and S. H. DeBoer. Can. J. Microbiol. 41:925, 1995. (4) G. Ruhl et al. Plant Dis. 93:841, 2009.

Occurrence and Distribution of Triticum mosaic virus in the Central Great Plains.

Wheat curl mite (WCM)-transmitted viruses-namely, Wheat streak mosaic virus (WSMV), Triticum mosaic virus (TriMV), and the High Plains virus (HPV)-are three of the wheat-infecting viruses in the central Great Plains of the United States. TriMV is newly discovered and its prevalence and incidence are largely unknown. Field surveys were carried out in Colorado, Kansas, Nebraska, and South Dakota in spring and fall 2010 and 2011 to determine TriMV prevalence and incidence and the frequency of TriMV co-infection with WSMV or HPV in winter wheat. WSMV was the most prevalent and was detected in 83% of 185 season-counties (= s-counties), 73% of 420 season-fields (= s-fields), and 35% of 12,973 samples. TriMV was detected in 32, 6, and 6% of s-counties, s-fields, and samples, respectively. HPV was detected in 34, 15, and 4% of s-counties, s-fields, and samples, respectively. TriMV was detected in all four states. In all, 91% of TriMV-positive samples were co-infected with WSMV, whereas WSMV and HPV were mainly detected as single infections. The results from this study indicate that TriMV occurs in winter wheat predominantly as a double infection with WSMV, which will complicate breeding for resistance to WCM-transmitted viruses.

α2-containing GABAA receptors expressed in hippocampal region CA3 control fast network oscillations.

GABA(A) receptors are critically involved in hippocampal oscillations. GABA(A) receptor α1 and α2 subunits are differentially expressed throughout the hippocampal circuitry and thereby may have distinct contributions to oscillations. It is unknown which GABA(A) receptor α subunit controls hippocampal oscillations and where these receptors are expressed. To address these questions we used transgenic mice expressing GABA(A) receptor α1 and/or α2 subunits with point mutations (H101R) that render these receptors insensitive to allosteric modulation at the benzodiazepine binding site, and tested how increased or decreased function of α subunits affects hippocampal oscillations. Positive allosteric modulation by zolpidem prolonged decay kinetics of hippocampal GABAergic synaptic transmission and reduced the frequency of cholinergically induced oscillations. Allosteric modulation of GABAergic receptors in CA3 altered oscillation frequency in CA1, while modulation of GABA receptors in CA1 did not affect oscillations. In mice having a point mutation (H101R) at the GABA(A) receptor α2 subunit, zolpidem effects on cholinergically induced oscillations were strongly reduced compared to wild-type animals, while zolpidem modulation was still present in mice with the H101R mutation at the α1 subunit. Furthermore, genetic knockout of α2 subunits strongly reduced oscillations, whereas knockout of α1 subunits had no effect. Allosteric modulation of GABAergic receptors was strongly reduced in unitary connections between fast spiking interneurons and pyramidal neurons in CA3 of α2H101R mice, but not of α1H101R mice, suggesting that fast spiking interneuron to pyramidal neuron synapses in CA3 contain α2 subunits. These findings suggest that α2-containing GABA(A) receptors expressed in the CA3 region provide the inhibition that controls hippocampal rhythm during cholinergically induced oscillations.

First Report of Goss's Bacterial Wilt and Leaf Blight (Clavibacter michiganensis subsp. nebraskensis) of Corn in Texas.

In September 2009, the University of Nebraska-Lincoln Plant and Pest Diagnostic Clinic received leaf samples of hybrid corn (Zea mays L.) displaying long, necrotic lesions with wavy margins. The lesions had discontinuous water-soaked spots that are indicative of Goss's bacterial wilt and leaf blight. The symptomatic leaves were submitted from Dallam County, located in the Texas Panhandle (northwest Texas). According to the USDA Farm Service Agency and the National Agricultural Statistics Service, in 2009 Dallam County had 54,025 ha planted to corn. This is approximately 19% of the total corn planted in the 26 counties in the Texas Panhandle and 6% of the total corn planted in the state of Texas. Extracts from the infected leaf tissue tested positive for Clavibacter michiganensis subsp. nebraskensis with a commercially available ELISA test (Neogen Inc., Scotland, UK). Isolation from the infected tissue onto CNS selective media (1) resulted in round, dark orange, mucoid colonies that tested gram positive with the Gram-stain test. BLAST nucleotide sequence alignments of the amplified 500-bp 16S rRNA region of the suspect culture's genome (2) revealed a 96% similarity for C. michiganensis subsp. nebraskensis (NCBI BLAST Accession No. U09381.1). To fulfill Koch's postulates, three sweet corn plants (Golden Cross Bantam) at growth stage V3 to V4 were inoculated in the greenhouse with a suspension of approximately 1 × 109 CFU/ml from suspect cultures grown on CNS for 5 days. Wounds approximately 6.5 cm long were created with sterile scissors on the fifth leaf from the bottom running parallel to the veins on either side of the midrib at the leaf apex. The leaf apex was dipped into 150 ml of the inoculum suspension for 5 s. Approximately 6 days after inoculation, discontinuous, water-soaked spots consistent with the symptoms on the original symptomatic leaves appeared on all the inoculated leaves near the site of infection. Colonies consistent with C. michiganensis subsp. nebraskensis (dark orange, mucoid) were reisolated onto CNS, completing Koch's postulates. To our knowledge, this is the first report of Goss's bacterial wilt and leaf blight on corn in Texas and because it is a residue-borne pathogen, the probability of it becoming a resident disease is relatively high. References: (1) D. C. Gross and A. K. Vidaver. Phytopathology 69:82, 1979. (2) X. Li and S. H. De Boer. 1995. Phytopathology 85:837, 1995.

GABAergic synapse properties may explain genetic variation in hippocampal network oscillations in mice.

Cognitive ability and the properties of brain oscillation are highly heritable in humans. Genetic variation underlying oscillatory activity might give rise to differences in cognition and behavior. How genetic diversity translates into altered properties of oscillations and synchronization of neuronal activity is unknown. To address this issue, we investigated cellular and synaptic mechanisms of hippocampal fast network oscillations in eight genetically distinct inbred mouse strains. The frequency of carbachol-induced oscillations differed substantially between mouse strains. Since GABAergic inhibition sets oscillation frequency, we studied the properties of inhibitory synaptic inputs (IPSCs) received by CA3 and CA1 pyramidal cells of three mouse strains that showed the highest, lowest and intermediate frequencies of oscillations. In CA3 pyramidal cells, the frequency of rhythmic IPSC input showed the same strain differences as the frequency of field oscillations. Furthermore, IPSC decay times in both CA1 and CA3 pyramidal cells were faster in mouse strains with higher oscillation frequencies than in mouse strains with lower oscillation frequency, suggesting that differences in GABA(A)-receptor subunit composition exist between these strains. Indeed, gene expression of GABA(A)-receptor ß2 (Gabrb2) and ß3 (Gabrb2) subunits was higher in mouse strains with faster decay kinetics compared with mouse strains with slower decay kinetics. Hippocampal pyramidal neurons in mouse strains with higher oscillation frequencies and faster decay kinetics fired action potential at higher frequencies. These data indicate that differences in genetic background may result in different GABA(A)-receptor subunit expression, which affects the rhythm of pyramidal neuron firing and fast network activity through GABA synapse kinetics.

Psychometric qualities of questionnaires for the assessment of otitis media impact.

BACKGROUND: The assessment of impact and evaluation of treatment effects in chronic otitis media (OM) calls for a much broader approach than just examining the presence of middle ear effusion or hearing loss. It is increasingly recognised that this condition may result in a comprised quality of life. Several studies have used proxy completed questionnaires to objectify the illness experience associated with chronic OM. OBJECTIVE OF REVIEW: To review questionnaires which have been developed to describe the effects of chronic OM on the daily functioning of children. Psychometric properties have been evaluated, in addition to discriminative and evaluative qualities. TYPE OF REVIEW: A systematic review of publications pertaining to developed questionnaires related with chronic OM. SEARCH STRATEGY: Systematic literature searches of PubMed (1966-January 2007) and EMBASE (1989-January 2007) were conducted, supplemented by using free text words to identify publications after January 2005. RESULTS: The included 15 questionnaires were developed for children with recurrent or persistent OM, describing functional health status (FHS), while two questionnaires also evaluate the effect of tympanostomy tubes insertion. The questionnaires generally cover six impact areas (physical symptoms, child development, educational performance, emotional/practical burden and general health status) with physical symptoms being the most prominant. CONCLUSIONS: The OM8-30, OMO-22 and OM-6 adequately reflect the multidimensional aspects of FHS in chronic OM. The OMO-22 and OM8-30 show the best psychometric properties for the discrimination of impact severity between children, while the OM-6 was found to have the best qualities for the evaluation of clinical change. Clinical applicability is crucial for the assessment of FHS in chronic OM, but requires a trade-off with necessary psychometric properties.

Reduced 5-HT1A- and GABAB receptor function in dorsal raphé neurons upon chronic fluoxetine treatment of socially stressed rats.

Autoinhibitory serotonin 1A receptors (5-HT(1A)) in dorsal raphé nucleus (DRN) have been implicated in chronic depression and in actions of selective serotonin reuptake inhibitors (SSRI). Due to experimental limitations, it was never studied at single-cell level whether changes in 5-HT(1A) receptor functionality occur in depression and during SSRI treatment. Here we address this question in a social stress paradigm in rats that mimics anhedonia, a core symptom of depression. We used whole cell patch-clamp recordings of 5-HT- and baclophen-induced G-protein-coupled inwardly rectifying potassium (GIRK) currents as a measure of 5-HT(1A)- and GABA(B) receptor functionality. 5-HT(1A)- and GABA(B) receptor-mediated GIRK-currents were not affected in socially stressed rats, suggesting that there was no abnormal (auto)inhibition in the DRN on social stress. However, chronic fluoxetine treatment of socially stressed rats restored anticipatory behavior and reduced the responsiveness of 5-HT(1A) receptor-mediated GIRK currents. Because GABA(B) receptor-induced GIRK responses were also suppressed, fluoxetine does not appear to desensitize 5-HT(1A) receptors but rather one of the downstream components shared with GABA(B) receptors. This fluoxetine effect on GIRK currents was also present in healthy animals and was independent of the animal's "depressed" state. Thus our data show that symptoms of depression after social stress are not paralleled by changes in 5-HT(1A) receptor signaling in DRN neurons, but SSRI treatment can alleviate these behavioral symptoms while acting strongly on the 5-HT(1A) receptor signaling pathway.

NMDA receptors trigger neurosecretion of 5-HT within dorsal raphe nucleus of the rat in the absence of action potential firing.

Activity and calcium-dependent release of neurotransmitters from the somatodendritic compartment is an important signalling mechanism between neurones throughout the brain. NMDA receptors and vesicles filled with neurotransmitters occur in close proximity in many brain areas. It is unknown whether calcium influx through these receptors can trigger the release of somatodendritic vesicles directly, or whether postsynaptic action potential firing is necessary for release of these vesicles. Here we addressed this question by studying local release of serotonin (5-HT) from dorsal raphé nucleus (DRN) neurones. We performed capacitance measurements to monitor the secretion of vesicles in giant soma patches, in response to short depolarizations and action potential waveforms. Amperometric measurements confirmed that secreted vesicles contained 5-HT. Surprisingly, two-photon imaging of DRN neurones in slices revealed that dendritic calcium concentration changes in response to somatic firing were restricted to proximal dendritic areas. This implied that alternative calcium entry pathways may dominate the induction of vesicle secretion from distal dendrites. In line with this, transient NMDA receptor activation, in the absence of action potential firing, was sufficient to induce capacitance changes. By monitoring GABAergic transmission onto DRN 5-HT neurones in slices, we show that endogenous NMDA receptor activation, in the absence of postsynaptic firing, induced release of 5-HT, which in turn increased the frequency of GABAergic inputs through activation of 5-HT(2) receptors. We propose here that calcium influx through NMDA receptors can directly induce postsynaptic 5-HT release from DRN neurones, which in turn may facilitate GABAergic input onto these cells.

[A case of very serious Sarcoptes mange in alpacas (Lama pacos)]. / Een geval van ernstige Sarcoptes-schurft bij alpaca's (Lama pacos).

After the diagnosis sarcoptic mange in four alpaca's (Lama pacos) we have tried to control this infection. Despite three treatments with doramectin, three with ivermectin, four with amitraz and two with diazinon we were unable to get the animals free of Sarcoptes mites and their condition deteriorated. One animal died six month after the first treatment. The three remaining animals were euthanized one month thereafter.

Response shift in the measurement of quality of life in hearing impaired adults after hearing aid fitting.

UNLABELLED: REASON FOR THE STUDY: Response shift is the change in the meaning of one's self-evaluation of a target construct, like quality of life (QOL). The objective of this study was to investigate whether response shift in the measurement of generic and specific QOL occurred in persons with a relatively mild health condition. For this purpose hearing impairment was used as a research model. MAJOR FINDINGS: Response shift effects were observed in the scores on the dimensions of hearing related QOL. In the scores on overall hearing related QOL, and in the scores on the generic control items, no response shift occurred. CONCLUSIONS: This study showed that response shift effects can take place in a relatively mild condition as well. The occurrence of response shift in QOL ratings over time could have large implications for the estimation of the effectiveness of medical interventions and for the use of these estimations in cost-effectiveness analyses. After a successful treatment the conventional change could be an underestimation of the effectiveness of the treatment, although it has also been argued that psychological adaptation is a welcome capacity of human beings, and that then-test changes do no justice to this capacity.

Long-term cannulation of the vena cava of rats for blood sampling: local and systemic effects observed by histopathology after six weeks of cannulation.

A cannulation system with fixation by a metal cuff around the tail was used for blood sampling. The cannula was guided subcutaneously and positioned in the vena cava after entering the body via the femoral vein. Histopathology was performed after long-term cannulation of up to 35 and 45 days. The presence of the cannula in the vena cava induced endothelial hypertrophy and hyperplasia accompanied by stromal hypertrophy. The endothelial activation was not limited to the vena cava but was also observed in both the cannulated vena iliaca and the contralateral control vena iliaca, the latter showing only minor alterations. In the lung, thrombi were noted in the larger lung arteries; and foreign body emboli, probably situated in the alveolar septi, could be detected occasionally. Inflammatory reactions in the tail at the site of cuff fixation consisted of a mixture of acute and chronic inflammatory responses. The chronic inflammation extended into the tail muscles, as shown by the presence of fibrous tissue associated with muscle degeneration. In conclusion, prolonged venous cannulation in rats resulted in local alterations in the veins, small emboli in the lungs and a moderate to marked inflammation in the tail. However, the procedure itself was well tolerated by the animals.

Subchronic mild noise stress increases HRP permeability in rat small intestine in vitro.

Recently we reported an increased trans- and paracellular protein permeability in rat small intestine after acute cold restraint stress. In the present study, we applied randomized 95- or 105-dB white noise pulses during 45 min/h, 12 h/day, duration 8 days, as a milder, but more chronic stressor to male rats. At 8 days before the noise experiments, 50% of the animals were cannulated in the vena cava for blood sampling during the experimental period. The other 50% of the animals were sacrificed at Day 9, segments of ileum were mounted in Ussing chambers and perfused at 37 degrees C. Horseradish peroxidase (HRP) was added mucosally, serosal appearance was detected enzymatically and tissues were fixed for electron microscopy. In the animals exposed to 95-dB noise, plasma corticosterone levels were enhanced twofold compared to controls, and ileal HRP flux was enhanced twofold. Electron micrographs of tissue from stressed or control animals showed no detectable paracellular staining of HRP. Quantification of HRP-containing endosomes in enterocytes revealed a twofold increase in endosome number in the animals exposed to 95-db noise indicating that the increased HRP permeability was primarily due to increased endocytosis. In contrast to the animals exposed to 95-dB noise, rats exposed to 105-dB noise showed no increase in corticosterone levels and ileal HRP fluxes were not significantly different from controls. We conclude that mild subchronic noise stress may cause a decrease in intestinal barrier function by increased transcytosis of luminal antigens.

Protease-dependent activation of epithelial cells by fungal allergens leads to morphologic changes and cytokine production.

BACKGROUND: Proteases in extracts of Aspergillus fumigatus cause epithelial cell desquamation and release of proinflammatory cytokines. OBJECTIVE: We sought to assess protease activity in Alternaria alternata, Cladosporium herbarum, and Aspergillus fumigatus extracts and study the ability of these extracts to cause desquamation and release of proinflammatory cytokines from epithelial cells. METHODS: Protease activities of the fungal extracts were quantified. Changes with respect to cell morphology, cell desquamation, and cytokine production (IL-6 and IL-8) were measured in the absence and presence of the fungal extracts in an airway-derived epithelial cell line (A549) and primary epithelial nasal cells. RESULTS: Fungal proteases differentially induced morphologic changes, cell desquamation, and production of IL-6 and IL-8 in a dose- and time-dependent fashion. Alternaria alternata extracts induced cell shrinking and cell desquamation and strongly enhanced the production of IL-6 and IL-8 at higher concentrations. Aspergillus fumigatus extracts caused cell shrinking, cell desquamation, and production of IL-6 and IL-8, even at low concentrations. The Aspergillus fumigatus-derived extract grown on collagen medium induced a strong dose-dependent decline in cytokine production at higher concentrations. Cladosporium herbarum extracts did not induce morphologic changes or cell desquamation but enhanced IL-6 and IL-8 productions at higher concentrations. The dependence of these effects on intact protease activity was shown by their abrogation by protease inhibitors. CONCLUSION: Proteases present in fungal extracts interact with epithelial cells, leading to morphologic changes, cell desquamation, and induction of proinflammatory cytokines. It is proposed that these fungal proteases may activate epithelial cells through a protease-activated receptor type 2-driven mechanism.