Quantitative detection of Streptococcus pneumoniae in nasopharyngeal secretions by real-time PCR

Streptococcus pneumoniae is an important cause of community-acquired pneumonia. However, in this setting the diagnostic sensitivity of blood cultures is below 30%. Since during such infections changes in the amounts of S. pneumoniae may also occur in the upper respiratory tract, quantification of these bacteria in nasopharnygeal secretions (NPSs) may offer a suitable diagnostic approach. Real-time PCR offers a sensitive, efficient, and routinely reproducible approach to quantification. Using primers and a fluorescent probe specific for the pneumolysin gene, we were able to detect DNA from serial dilutions of S. pneumoniae cells in which the quantities of DNA ranged from the amounts extracted from 1 to 10(6) cells. No difference was noted when the same DNA was mixed with DNA extracted from NPSs shown to be deficient of S. pneumoniae following culture, suggesting that this bacterium can be detected and accurately quantitated in clinical samples. DNAs from Haemophilus influenzae, Moraxella catarrhalis, or alpha-hemolytic streptococci other than S. pneumoniae were not amplified or were only weakly amplified when there were > or =10(6) cells per reaction mixture. When the assay was applied to NPSs from patients with respiratory tract infections, the assay performed with a sensitivity of 100% and a specificity of up to 96% compared to the culture results. The numbers of S. pneumoniae organisms detected by real-time PCR correlated with the numbers detected by semiquantitative cultures. A real-time PCR that targeted the pneumolysin gene provided a sensitive and reliable means for routine rapid detection and quantification of S. pneumoniae present in NPSs. This assay may serve as a tool to study changes in the amounts of S. pneumoniae during lower respiratory tract infections.     

Sleep as a behavioral model of neuro-immune interactions

The central nervous system, by a variety of mechanisms engages in constant surveillance of the peripheral immune system. Alterations in the status of the peripheral immune system induced by an invading pathogen for example, are quickly detected by the central nervous system, which then responds by altering physiological processes and behavior in an attempt to support the immune system in its efforts to eliminate the pathogen. Sleep is one of several behaviors that are dramatically altered in response to infection. Immune-active substances such as the pro-inflammatory cytokines interleukin-1 and tumor necrosis factor, either directly or indirectly via interactions with neurotransmitters or neurohormones are involved in the regulation of sleep. Because these cytokines increase during infection, they are likely candidates for mediating the profound alterations in sleep that occur during infection. Since regulation of behavior is the function of the central nervous system, infection-induced alterations in behavior provide a unique model for the study of neuro-immune interactions.     

Hypothalamic serotonergic activity correlates better with brain temperature than with sleep-wake cycle and muscle tone in rats.

The activity of the serotonergic system varies in phase with the sleep-wake cycle, which is associated with changes in several physiological functions, including electroencephalographic activity, brain temperature, and locomotion. The aim of the present study was to clarify which of these parameters correlates better with serotonergic activity in spontaneous conditions. Voltammetric recordings by telemetry of serotonergic metabolism in the medial preoptic area and polygraphic recordings of sleep-wake activity (by means of electroencephalographic delta band, brain cortical temperature and neck electromyographic activity recordings) were simultaneously performed in freely moving rats. Univariate analyses of variance revealed that each variable under investigation was statistically correlated with serotonergic metabolism. When the variables were entered into the model simultaneously, both partial correlation and step-wise multiple regression analyses indicated that the highest correlation exists between serotonergic metabolism and brain cortical temperature. The present data show that serotonergic activity in the medial preoptic area is closely linked to physiological changes in brain temperature.     

Intercellular adhesion molecule-1 expression induced by interleukin (IL)-1β or an IL-1β fragment is blocked by an IL-1 receptor antagonist and a soluble IL-1 receptor

The effects of a recombinant human interleukin-1 (IL-1) receptor antagonist (IL-1ra) and a recombinant human soluble IL-1 receptor (sIL-1R) on cytokine-induced intercellular adhesion molecule-1 (ICAM-1) expression in a human glioblastoma cell line and a neuroblastoma cell line were determined. Cells were incubated with IL-1β, tumor necrosis factor (TNF)α and interferon (IFN)γ. Cells were also tested under identical conditions with an IL-1β synthetic peptide fragment (IL-1β_208–240) previously shown to possess biological activity. IL-1β, TNFα and IFNγ potentiated ICAM-1 expression in both cell lines in a dose-related manner. The IL-1β_208–240 fragments, corresponding to the rabbit, rat and human sequences, enhanced ICAM-1 expression in glioblastoma cells at high doses. ICAM-1 expression induced by IL-1β, rabbit IL-1β_208–240 and human IL-1β_208–240 was blocked by the IL-1ra, while TNFα- and IFNγ-induced ICAM-1 expression were not. ICAM-1 expression induced by IL-1β and human IL-1β_208–240 was also blocked by the sIL-1R. Our findings suggest that IL1β_208–240 acts as an IL-1β agonist in enhancing ICAM-1 expression in vitro and that this effect is receptor-mediated.     

Changes in spontaneous activity of medialis dorsalis thalamic neurones during sleep and wakefulness

In chronic unanaesthetized cats unitary activity of medialis dorsalis (MD) thalamic neurones was recorded during wakefulness (W), slow wave (SWS) and desynchronized (DS) sleep. The discharge pattern of these neurones changes during SWS compared to W. Comparison between desynchronization of W and DS shows a change in the mean frequency, being higher in W than in DS. The results suggest that MD neurones participate in the organization of the sleep-wakefulness cycle.     

Interleukin-1 Inhibits Putative Cholinergic Neurons in Vitro and REM Sleep when Microinjected into the Rat Laterodorsal Tegmental Nucleus

Study Objectives:

REM sleep is suppressed during infection, an effect mimicked by the administration of cytokines such as interleukin-1 (IL-1). In spite of this observation, brain sites and neurochemical systems mediating IL-1-induced suppression of REM sleep have not been identified. Cholinergic neurons in the brainstem laterodorsal tegmental nucleus (LDT) are part of the neuronal circuitry responsible for REM sleep generation. Since IL-1 inhibits acetylcholine synthesis and release, the aim of this study was to test the two different, but related hypotheses. We hypothesized that IL-1 inhibits LDT cholinergic neurons, and that, as a result of this inhibition, IL-1 suppresses REM sleep.

Design, Measurement, and Results:

To test these hypotheses, the electrophysiological activity of putative cholinergic LDT neurons was recorded in a rat brainstem slice preparation. Interleukin-1 significantly inhibited the firing rate of 76% of recorded putative cholinergic LDT neurons and reduced the amplitude of glutamatergic evoked potentials in 60% of recorded neurons. When IL-1 (1 ng) was microinjected into the LDT of freely behaving rats, REM sleep was reduced by about 50% (from 12.7% ± 1.5% of recording time [after vehicle] to 6.1% ± 1.4% following IL-1 administration) during post-injection hours 3-4.

Conclusions:

Results of this study support the hypothesis that IL-1 can suppress REM sleep by acting at the level of the LDT nucleus. Furthermore this effect may result from the inhibition of evoked glutamatergic responses and of spontaneous firing of putative cholinergic LDT neurons.

Citation:

Brambilla D; Barajon I; Bianchi S; Opp MR; Imeri L. Interleukin-1 inhibits putative cholinergic neurons in vitro and REM sleep when microinjected into the rat laterodorsal tegmental nucleus. SLEEP 2010;33(7):919-929.     

Effects of serotonergic activation by 5-hydroxytryptophan on sleep and body temperature of C57BL/6J and interleukin-6-deficient mice are dose and time related

STUDY OBJECTIVES: Extensive data implicate serotonin (5-hydroxytryptamine [5-HT]) in the regulation of sleep. Jouvet has hypothesized that 5-HT promotes wakefulness, yet is necessary for subsequent non-rapid eye movement (NREM) sleep, actions he proposes to be mediated by sleep factors. Studies in rat support this dual role for 5-HT. The objectives of this study were to (1) determine effects of serotonergic activation on sleep of mice and (2) elucidate a potential role for the cytokine interleukin-6 as a sleep factor mediating serotonergic effects on sleep. DESIGN: C57BL/6J and B6.129S6-II6(tm1Kopf)(interleukin-6 knockout [IL-6 KO]) mice were purchased from the Jackson Laboratory and instrumented for recording the electroencephalogram and body temperature. After recovery, separate groups of mice were injected intraperitoneally at either light or dark onset with vehicle or with the 5-HT precursor 5-hydroxytryptophan (5-HTP). Sleep-wake behavior was determined and body temperature recorded for 24 hours after injections. RESULTS: 5-HTP induced hypothermia in both mouse strains. When injected at dark onset, the highest dose of 5-HTP (200 mg/kg) increased NREM sleep. Light onset administration initially increased wakefulness, with increases in NREM sleep apparent only during the subsequent dark period. For most parameters, there were no differences in responses between strains. However IL-6 KO mice at some doses exhibited a greater increase in NREM sleep. CONCLUSIONS: 5-HTP alters sleep-wake behavior and body temperature of mice in a manner similar to that of rats. Increases in NREM sleep after 5-HTP are apparent only during the dark period, which may represent a fundamental property of the serotonergic system. These results suggest that 5-HT should not be considered either wake promoting or NREM sleep promoting. Rather, the role of 5-HT in the regulation of sleep-wake behavior must be considered within the context of the degree to which the system is activated and the time at which the activation occurs.     

Examining subjective well-being among older adults using pain medications

Purpose

To examine the relationship between social participation and subjective well-being (SWB) among older adults using pain medications and evaluate the impact of sex on this relationship.

Methods

A cross-sectional analysis was conducted using the 2019 National Health and Aging Trends Study data, a nationally-representative sample of Medicare beneficiaries 65 years and older. Individuals were included if they reported taking pain medications for five days or more per week over the last month. Social participation was operationalized using the sum score of four items: visiting family and friends, going out for enjoyment, attending religious services, and participation in other organized activities. SWB was operationalized as a latent variable using four items reflecting positive and negative emotions, and three items reflecting self-evaluation. Structural equation modeling was used to test the relationship between key study constructs, as well as the moderating effect of sex on the relationship between social participation and SWB.

Results

A total of 964 (weighted N?=?7,660,599) participants were included in the study. Most participants were female (61.3%), White (81.0%), community-dwelling (94.9%) older adults. Confirmatory factor analysis showed appropriate fit for SWB. Social participation had a statistically significant association with SWB (unstandardized regression coefficient?=?0.133; 95% CI 0.071, 0.196; p?<?0.001) after adjusting for covariates. However, this relationship was not moderated by sex (p?=?0.836).

Conclusion

Social participation is positively and significantly associated with SWB among older adults using pain medications. Interventions aimed at improving SWB should consider incorporating a social activities component.

     

Generation of renal Epo-producing cell lines by conditional gene tagging reveals rapid HIF-2 driven Epo kinetics,cell autonomous feedback regulation,and a telocyte phenotype

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5-Hydroxytryptophan, but not L-tryptophan, alters sleep and brain temperature in rats

The precise role of serotonin (5-hydroxytryptamine) in the regulation of sleep is not fully understood. To further clarify this role for 5-hydroxytryptamine, the 5-hydroxytryptamine precursors L-tryptophan (40 and 80 mg/kg) and L-5-hydroxytryptophan (25-, 50-, 75-, 100 mg/kg) were injected intraperitoneally into freely behaving rats 15 min prior to dark onset, and subsequent effects on sleep-wake activity and cortical brain temperature were determined. L-5-hydroxytryptophan, but not L-tryptophan, induced dose-dependent changes in sleep-wake activity. During the 12-h dark period, non-rapid eye movement sleep was inhibited in post-injection hours 1-2 by the two lowest L-5-hydroxytryptophan doses tested, while the two highest doses induced a delayed increase in non-rapid eye movement sleep in post-injection hours 3-12. These highest doses inhibited non-rapid eye movement sleep during the subsequent 12-h light period. The finding that L-5-hydroxytryptophan, but not L-tryptophan, induced a dose-dependent and long-lasting decrease in cortical brain temperature regardless of whether or not non-rapid eye movement sleep was suppressed or enhanced contributes to a growing list of conditions showing that sleep-wake activity and thermoregulation, although normally tightly coupled, may be dissociated. The initial non-rapid eye movement sleep inhibition observed following low doses of L-5-hydroxytryptophan may be attributable to increased serotonergic activity since 5-hydroxytryptamine may promote wakefulness per se, whereas the delayed non-rapid eye movement sleep enhancement after higher doses may be due to the induction by 5-hydroxytryptamine of sleep-inducing factor(s), as previously hypothesized. The period of non-rapid eye movement sleep inhibition beginning 12 h after administration of L-5-hydroxytryptophan doses that increase non-rapid eye movement sleep is characteristic of physiological manipulations in which non-rapid eye movement sleep is enhanced. The results of the present study suggest that the complex effects of 5-HT on sleep depend on the degree and time course of activation of the serotonergic system such that 5-HT may directly inhibit sleep, yet induce a cascade of physiological processes that enhance subsequent sleep.