Adenosine Signaling through A1 Receptors Inhibits Chemosensitive Neurons in the Retrotrapezoid Nucleus.

A subset of neurons in the retrotrapezoid nucleus (RTN) function as respiratory chemoreceptors by regulating depth and frequency of breathing in response to changes in tissue CO2/H+. The activity of chemosensitive RTN neurons is also subject to modulation by CO2/H+-dependent purinergic signaling. However, mechanisms contributing to purinergic regulation of RTN chemoreceptors are not entirely clear. Recent evidence suggests adenosine inhibits RTN chemoreception in vivo by activation of A1 receptors. The goal of this study was to characterize effects of adenosine on chemosensitive RTN neurons and identify intrinsic and synaptic mechanisms underlying this response. Cell-attached recordings from RTN chemoreceptors in slices from rat or wild-type mouse pups (mixed sex) show that exposure to adenosine (1 µM) inhibits chemoreceptor activity by an A1 receptor-dependent mechanism. However, exposure to a selective A1 receptor antagonist (8-cyclopentyl-1,3-dipropylxanthine, DPCPX; 30 nM) alone did not potentiate CO2/H+-stimulated activity, suggesting activation of A1 receptors does not limit chemoreceptor activity under these reduced conditions. Whole-cell voltage-clamp from chemosensitive RTN neurons shows that exposure to adenosine activated an inward rectifying K+ conductance, and at the network level, adenosine preferentially decreased frequency of EPSCs but not IPSCs. These results show that adenosine activation of A1 receptors inhibits chemosensitive RTN neurons by direct activation of a G-protein-regulated inward-rectifier K+ (GIRK)-like conductance, and presynaptically, by suppression of excitatory synaptic input to chemoreceptors.

Expression of Kir4.1 and Kir5.1 inwardly rectifying potassium channels in oligodendrocytes, the myelinating cells of the CNS.

The inwardly rectifying K+ channel subtype Kir5.1 is only functional as a heteromeric channel with Kir4.1. In the CNS, Kir4.1 is localised to astrocytes and is the molecular basis of their strongly negative membrane potential. Oligodendrocytes are the specialised myelinating glia of the CNS and their resting membrane potential provides the driving force for ion and water transport that is essential for myelination. However, little is known about the ion channel profile of mature myelinating oligodendrocytes. Here, we identify for the first time colocalization of Kir5.1 with Kir4.1 in oligodendrocytes in white matter. Immunolocalization with membrane-bound Na+/K+-ATPase and western blot of the plasma membrane fraction of the optic nerve, a typical CNS white matter tract containing axons and the oligodendrocytes that myelinate them, demonstrates that Kir4.1 and Kir5.1 are colocalized on oligodendrocyte cell membranes. Co-immunoprecipitation provides evidence that oligodendrocytes and astrocytes express a combination of homomeric Kir4.1 and heteromeric Kir4.1/Kir5.1 channels. Genetic knock-out and shRNA to ablate Kir4.1 indicates plasmalemmal expression of Kir5.1 in glia is largely dependent on Kir4.1 and the plasmalemmal anchoring protein PSD-95. The results demonstrate that, in addition to astrocytes, oligodendrocytes express both homomeric Kir4.1 and heteromeric Kir4.1/Kir5.1 channels. In astrocytes, these channels are essential to their key functions of K+ uptake and CO2/H+ chemosensation. We propose Kir4.1/Kir5.1 channels have equivalent functions in oligodendrocytes, maintaining myelin integrity in the face of large ionic shifts associated with action potential propagation along myelinated axons.

Detection of viral bioagents using a shear horizontal surface acoustic wave biosensor.

Viruses are of high medical and biodefense concern and their detection at concentrations well below the threshold necessary to cause health hazards continues to be a challenge with respect to sensitivity, specificity, and selectivity. Ideally, assays for accurate and real time detection of viral agents would not necessitate any pre-processing of the analyte, which would make them applicable for example to bodily fluids (blood, sputum) and man-made as well as naturally occurring bodies of water (pools, rivers). We describe herein a robust biosensor that combines the sensitivity of surface acoustic waves (SAW) generated at a frequency of 325MHz with the specificity provided by antibodies for the detection of viral agents. A lithium tantalate-based SAW transducer with silicon dioxide waveguide sensor platform featuring three test and one reference delay lines was used to adsorb antibodies directed against either Coxsackie virus B4 or the category A bioagent Sin Nombre virus (SNV), a member of the genus Hantavirus, family Bunyaviridae, negative-stranded RNA viruses. Rapid detection (within seconds) of increasing concentrations of viral particles was linear over a range of order of magnitude for both viruses, although the sensor was approximately 5 x 10(5)-fold more sensitive for the detection of SNV. For both pathogens, the sensor's selectivity for its target was not compromised by the presence of confounding Herpes Simplex virus type 1. The biosensor was able to detect SNV at doses lower than the load of virus typically found in a human patient suffering from hantavirus cardiopulmonary syndrome (HCPS). Further, in a proof-of-principle real world application, the SAW biosensor was capable to selectively detect SNV agents in complex solutions, such as naturally occurring bodies of water (river, sewage effluent) without analyte pre-processing. This is the first study that reports on the detection of viral agents using an antibody-based SAW biosensor that has the potential to be used as a hand-held and self-contained device for rapid viral detection in the field.

Colloid solutions for fluid resuscitation.

BACKGROUND: Colloids are widely used in the replacement of fluid volume. However doubts remain as to which colloid is best. Different colloids vary in their molecular weight and therefore in the length of time they remain in the circulatory system. Because of this and their other characteristics, they may differ in their safety and efficacy. OBJECTIVES: To compare the effects of different colloid solutions in patients thought to need volume replacement. SEARCH STRATEGY: We searched the Cochrane Injuries Group specialised register, the Cochrane Controlled Trials Register (2002 Issue 3), MEDLINE (1994-2002/07), EMBASE (1974-2002 August week 1), and the National Research Register (2002 issue 3). Bibliographies of trials retrieved were searched, and drug companies manufacturing colloids were contacted for information. The search was last updated in September 2002. SELECTION CRITERIA: Randomised and quasi-randomised trials comparing colloid solutions in critically ill and surgical patients thought to need volume replacement. The main outcomes measured were death, amount of whole blood transfused, and incidence of adverse reactions. DATA COLLECTION AND ANALYSIS: Two authors independently extracted the data and assessed the quality of the trials. MAIN RESULTS: Fifty-seven trials met the inclusion criteria, with a total of 3659 participants. Quality of allocation concealment was judged to be adequate in 20 trials and poor or uncertain in 37. Deaths were obtained from 36 trials. For albumin or PPF versus hydroxyethyl starch (HES) 20 trials (n=1029) reported mortality. The pooled relative risk (RR) was 1.17 (95% CI 0.91, 1.50). For albumin or PPF versus gelatin four trials (n=542) reported mortality. The RR was 0.99 (0.69, 1.42). For gelatin vs HES 11 trials (n=945) reported mortality, RR was 1.00 (0.78,1.28). RR was not estimable in the albumin vs dextran, gelatin vs dextran, and HES vs dextran groups. Thirty-six trials recorded the amount of blood transfused, however quantitative analysis was not possible due to skewness and variable reporting. Fifteen trials recorded adverse reactions, but none occurred. REVIEWER'S CONCLUSIONS: From this review, there is no evidence that one colloid solution is more effective or safe than any other, although the confidence intervals are wide and do not exclude clinically significant differences between colloids. Larger trials of fluid therapy are needed if clinically significant differences in mortality are to be detected or excluded.

Colloid solutions for fluid resuscitation.

BACKGROUND: Colloids are widely used in the replacement of fluid volume, however doubts remain as to their benefits. Different colloids vary in their molecular weight and therefore in the length of time they remain in the circulatory system. Because of this and their other characteristics, they may differ in their safety and efficacy. OBJECTIVES: To compare the effects of different colloid solutions in patients thought to need volume replacement. SEARCH STRATEGY: The Cochrane Injuries Group specialised register, The Cochrane Controlled trials register (all years), MEDLINE (1994-2000), EMBASE (1974-2000) were searched. Bibliographies of trials retrieved were searched, and drug companies manufacturing colloids were contacted for information. SELECTION CRITERIA: Randomised and quasi randomised trials comparing colloid solutions in critically ill and surgical patients thought to need volume replacement. The main outcomes measured were death, amount of whole blood transfused and incidence of adverse reactions. DATA COLLECTION AND ANALYSIS: Two authors independently extracted the data and assessed the quality of the trials. MAIN RESULTS: 52 trials met the inclusion criteria, with a total of 3311 participants. Quality of allocation concealment was judged to be adequate in 17 trials and poor or uncertain in 35. Deaths were obtained from 31 trials. For albumin or PPF versus hydroxyethyl starch (HES) 20 trials (n=1029) reported mortality. The pooled relative risk (RR) was 1.17 (95% CI 0.91, 1.50). For albumin or PPF versus gelatin four trials (n=542) reported mortality. The RR was 0.99 (0.69, 1.42). For gelatin vs HES six trials (n=597) reported mortality, RR was 0.96 (0.69, 1.33). RR was not estimable in the albumin vs dextran, gelatin vs dextran, and HES vs dextran groups. Thirty-one trials recorded the amount of blood transfused, however quantitative analysis was not possible due to skewness and variable reporting. Fifteen trials recorded adverse reactions, but none occurred. REVIEWER'S CONCLUSIONS: From this review, there is no evidence that one colloid solution is more effective or safe than any other, although the confidence intervals are wide and do not exclude clinically significant differences between colloids. Larger trials of fluid therapy are needed if clinically significant differences in mortality are to be detected or excluded.

Colloid solutions for fluid resuscitation.

BACKGROUND: Colloids are widely used in the replacement of fluid volume, however doubts remain as to their benefits. Different colloids vary in their molecular weight and therefore in the length of time they remain in the circulatory system. Because of this and their other characteristics, they may differ in their safety and efficacy. OBJECTIVES: To compare the effects of different colloid solutions in patients thought to need volume replacement. SEARCH STRATEGY: The Cochrane Injuries Group specialised register, The Cochrane Controlled trials register (all years), MEDLINE (1994-98), EMBASE (1974-98) were searched. Bibliographies of trials retrieved were searched and drug companies manufacturing colloids were contacted for information. SELECTION CRITERIA: Randomised and quasi randomised trials comparing colloid solutions in critically ill and surgical patients thought to need volume replacement. The main outcomes measured were death, amount of whole blood transfused and incidence of adverse reactions. DATA COLLECTION AND ANALYSIS: Two authors independently extracted the data and assessed the quality of the trials. MAIN RESULTS: 46 trials met the inclusion criteria, with a total of 2884 participants. Many of the trials were small. In the majority of trials quality was poor or uncertain. Deaths were obtained from 27 trials. Twenty three trials recorded the amount of blood transfused, however quantitative analysis was not possible due to skewness and variable reporting. Thirteen trials recorded adverse reactions, but none occurred. For albumin or PPF versus hydroxyethyl starch (HES) 20 trials reported mortality. The pooled relative risk (RR) was 1.17 (95% CI 0.91, 1.50). For albumin or PPF versus gelatin 3 trials reported mortality. The RR was 0.99 (0.69, 1.42). For gelatin vs HES 3 trials reported mortality, RR was 0.97 (0.65, 1.44). RR was not estimable in the albumin vs dextran, gelatin vs dextran, and HES vs dextran groups. REVIEWER'S CONCLUSIONS: From this review, there is no evidence that one colloid solution is more effective or safe than any other, although the confidence intervals are wide and do not exclude clinically significant differences between colloids. Larger trials of fluid therapy are needed if clinically significant differences in mortality are to be detected or excluded.

The prostate expression database (PEDB): status and enhancements in 2000.

The Prostate Expression Database (PEDB) is an online resource designed to access and analyze gene expression information derived from the human prostate. PEDB archives >55 000 expressed sequence tags (ESTs) from 43 cDNA libraries in a curated relational database that provides detailed library information including tissue source, library construction methods, sequence diversity and sequence abundance. The differential expression of each EST species can be viewed across all libraries using a Virtual Expression Analysis Tool (VEAT), a graphical user interface written in Java for intra- and inter-library species comparisons. Recent enhancements to PEDB include: (i) the functional categorization of annotated EST assemblies using a classification scheme developed at The Institute for Genome Research; (ii) catalogs of expressed genes in specific prostate tissue sources designated as transcriptomes; and (iii) the addition of prostate proteome information derived from two-dimensional electrophoreses and mass spectrometry of prostate cancer cell lines. PEDB may be accessed via the WWW at http://www.mbt.washington.edu/PEDB/

Negative selection: a method for obtaining low-abundance cDNAs using high-density cDNA clone arrays.

The identification of the entire complement of genes expressed in a cell, tissue, or organism provides a framework for understanding biological properties and establishes a tool set for subsequent functional studies. The large-scale sequencing of randomly selected clones from cDNA libraries has been successfully employed as a method for identifying a large fraction of these expressed genes. However, this approach is limited by the inherent redundancy of cellular transcripts reflecting widely variant levels of gene transcription. As a result, a high percentage of transcript duplications are encountered as the number of sequenced clones accrues. To address this problem, we have developed a negative hybridization selection method that employs the hybridization of complex cDNA probes to high-density arrays of cDNA clones and the subsequent selection of clones with a null or low hybridization signal. This approach was applied to a cDNA library constructed from normal human prostate tissue and resulted in the reduction of highly expressed prostate cDNAs from 6.8 to 0.57% with an overall decline in clone redundancy from 33 to 11%. The selected clones also reflected a more diverse cDNA population, with 89% of the clones representing distinctly different cDNAs compared with 67% of the randomly selected clones. This method compares favorably with cDNA library re-association normalization approaches and offers several distinct advantages, including the flexibility to use previously prepared libraries, and the ability to employ an iterative screening approach for continued accrual of cDNAs representing rare transcripts.

Kynostatin and 17beta-estradiol prevent the apoptotic death of human neuroblastoma cells exposed to HIV-1 protease.

A significant number of adult male patients with acquired immunodeficiency syndrome develop cerebral atrophy and progressive brain disorders such as dementia complex and neuropsychiatric problems. Upon entering the brain via activated macrophages or microglias, the human immunodeficiency type 1 virus (HIV-1) may produce cytotoxic factors such as HIV-1 envelope protein (gp120) and protease. Owing to significant proteolysis of nonviral proteins, the protease derived from HIV-1 may be detrimental to brain cells and neurons. Our results revealed that HIV-1 protease, at nanomolar concentrations, was as potent as gp120 in causing neurotoxicity in human neuroblastoma neurotypic SH-SY5Y cells. As shown by the Oncor ApopTag staining procedure, HIV-1 protease significantly increased the number of apoptotic cells over the serum-free controls. Moreover, HIV-1 protease-induced neurotoxicity was blocked by a selective protease inhibitor, kynostatin (KNI-272). Antioxidants such as 17beta-estradiol, melatonin, and S-nitrosoglutathione also prevented protease-induced neurotoxicity. These findings indicate that oxidative proteolysis may mediate HIV-1 protease-induced apoptosis and the degeneration of neurons and other brain cells. Centrally active protease inhibitors and antioxidants may play an important role in preventing cerebral atrophy and associated dementia complex caused by HIV-1.

The Pyramid Model: an integrated approach for evaluating continuing education programs and outcomes.

Recent mergers and downsizing of health care agencies have made resources for continuing education (CE) increasingly scarce. Nurse educators must demonstrate the effectiveness and sustainability of CE programs and establish the link between nursing professionalism and positive patient outcomes. The Pyramid Evaluation Model expands and enhances previous evaluation frameworks. Simple steps are outlined to evaluate CE programs and outcomes systematically and comprehensively through an impact model that examines goals, reviews program design, monitors program implementation, assesses outcomes and impact, and analyzes efficiency.

PEDB: the Prostate Expression Database.

The Prostate Expression Database (PEDB) is a curated relational database and suite of analysis tools designed for the study of prostate gene expression in normal and disease states. Expressed Sequence Tags (ESTs) and full-length cDNA sequences derived from more than 40 human prostate cDNA libraries are maintained and represent a wide spectrum of normal and pathological conditions. Detailed library information including tissue source, library construction methods, sequence diversity and abundance are available in a library archive. Prostate ESTs are assembled into distinct species groups using the multiple alignment program CAP2 and are annotated with information from the GenBank, dbEST and Unigene public sequence databases. Annotated sequences in PEDB are searched using the BLAST algorithm. The differential expression of each EST species can be viewed across all libraries using a Virtual Expression Analysis Tool (VEAT), a graphical user interface written in Java for intra- and inter-library species comparisons. PEDB may be accessed via the World Wide Web at http://www.mbt.washington.edu/PEDB/

Justification for a pediatric satellite pharmacy at a tertiary-care institution.

The justification for a pediatric satellite pharmacy is described. Of the 742 inpatient beds in a tertiary-care medical center, 150 are dedicated to pediatric patients. The pharmacy department provides services to both adult and pediatric patients from a central pharmacy. However, unit dose drug distribution services are not provided to the nurseries. Furthermore, the department lacks a core of pharmacists specializing in pediatric practice. As a result, the ability of the pharmacy to support pediatric drug use has been compromised, and medication errors and infection control problems have occurred that can be attributed to this situation. A pediatric satellite pharmacy was proposed to improve the quality of pediatric care. Service objectives were established, and benefits were identified. A financial analysis was included in the proposal that documented a total gain attributable to a pediatric satellite pharmacy of approximately $285,000 over five years. The satellite pharmacy was approved by hospital administration. The pharmacy department's quality assurance committee will monitor the satellite's impact. The justification for a proposed pediatric satellite pharmacy included indications that the satellite could correct medication-related problems and improve pediatric care and that implementation would have a positive effect on finances.