Heterozygous expression of a Kcnt1 gain-of-function variant has differential effects on somatostatin- and parvalbumin-expressing cortical GABAergic neurons.

More than 20 recurrent missense gain-of-function (GOF) mutations have been identified in the sodium-activated potassium (KNa) channel gene KCNT1 in patients with severe developmental and epileptic encephalopathies (DEEs), most of which are resistant to current therapies. Defining the neuron types most vulnerable to KCNT1 GOF will advance our understanding of disease mechanisms and provide refined targets for precision therapy efforts. Here, we assessed the effects of heterozygous expression of a Kcnt1 GOF variant (Kcnt1Y777H) on KNa currents and neuronal physiology among cortical glutamatergic and GABAergic neurons in mice, including those expressing vasoactive intestinal polypeptide (VIP), somatostatin (SST), and parvalbumin (PV), to identify and model the pathogenic mechanisms of autosomal dominant KCNT1 GOF variants in DEEs. Although the Kcnt1Y777H variant had no effects on glutamatergic or VIP neuron function, it increased subthreshold KNa currents in both SST and PV neurons but with opposite effects on neuronal output; SST neurons became hypoexcitable with a higher rheobase current and lower action potential (AP) firing frequency, whereas PV neurons became hyperexcitable with a lower rheobase current and higher AP firing frequency. Further neurophysiological and computational modeling experiments showed that the differential effects of the Kcnt1Y777H variant on SST and PV neurons are not likely due to inherent differences in these neuron types, but to an increased persistent sodium current in PV, but not SST, neurons. The Kcnt1Y777H variant also increased excitatory input onto, and chemical and electrical synaptic connectivity between, SST neurons. Together, these data suggest differential pathogenic mechanisms, both direct and compensatory, contribute to disease phenotypes, and provide a salient example of how a pathogenic ion channel variant can cause opposite functional effects in closely related neuron subtypes due to interactions with other ionic conductances.

Intrinsic Molecular Proton Sensitivity Underlies GPR4 Effects on Retrotrapezoid Nucleus Neuronal Activation and CO2-Stimulated Breathing.

An interoceptive homeostatic reflex monitors levels of CO2/H+ to maintain blood gas homeostasis and rapidly regulate tissue acid-base balance by driving lung ventilation and CO2 excretion-this CO2-evoked increase in respiration is the hypercapnic ventilatory reflex (HCVR). Retrotrapezoid nucleus (RTN) neurons provide crucial excitatory drive to downstream respiratory rhythm/pattern-generating circuits, and their activity is directly modulated by changes in CO2/H+ RTN neurons express GPR4 and TASK-2, global deletion of which abrogates CO2/H+ activation of RTN neurons and the HCVR. It has not been determined if the intrinsic pH sensitivity of these proton detectors is required for these effects. We used CRISPR/Cas9 genome editing to generate mice with mutations in either of two pH-sensing histidine residues in GPR4 to determine effects on RTN neuronal CO2/H+ sensitivity and the HCVR. In global GPR4(H81F) and GPR4(H167F) mice, CO2-stimulated breathing and CO2-induced RTN neuronal activation were strongly blunted, with no effect on hypoxia-stimulated breathing. In brainstem slices from GPR4(H81F) mice, peak firing of RTN neurons during bath acidification was significantly reduced compared with GPR4 wild-type mice, and a subpopulation of RTN neurons was rendered pH-insensitive, phenocopying previous results from GPR4-deleted mice. These effects were independent of changes in RTN number/distribution, neuronal excitability or transcript levels for GPR4 and TASK-2. CO2-stimulated breathing was reduced to a similar extent in GPR4(H81F) and TASK-2-deleted mice, with combined mutation yielding no additional deficit in the HCVR. Together, these data demonstrate that the intrinsic pH sensitivity of GPR4 is necessary for full elaboration of the HCVR.

Liver-Selective Imidazolopyrazine Mitochondrial Uncoupler SHD865 Reverses Adiposity and Glucose Intolerance in Mice.

Excess body fat is a risk factor for metabolic diseases and is a leading preventable cause of morbidity and mortality worldwide. There is a strong need to find new treatments that decrease the burden of obesity and lower the risk of obesity-related comorbidities, including cardiovascular disease and type 2 diabetes. Pharmacologic mitochondrial uncouplers represent a potential treatment for obesity through their ability to increase nutrient oxidation. Herein, we report the in vitro and in vivo characterization of compound SHD865, the first compound to be studied in vivo in a newly discovered class of imidazolopyrazine mitochondrial uncouplers. SHD865 is a derivative of the furazanopyrazine uncoupler BAM15. SHD865 is a milder mitochondrial uncoupler than BAM15 that results in a lower maximal respiration rate. In a mouse model of diet-induced adiposity, 6-week treatment with SHD865 completely restored normal body composition and glucose tolerance to levels like those of chow-fed controls, without altering food intake. SHD865 treatment also corrected liver steatosis and plasma hyperlipidemia to normal levels comparable with chow-fed controls. SHD865 has maximal oral bioavailability in rats and slow clearance in human microsomes and hepatocytes. Collectively, these data identify the potential of imidazolopyrazine mitochondrial uncouplers as drug candidates for the treatment of obesity-related disorders.

The astrocytic Na+ -HCO3 - cotransporter, NBCe1, is dispensable for respiratory chemosensitivity.

The interoceptive homeostatic mechanism that controls breathing, blood gases and acid-base balance in response to changes in CO2 /H+ is exquisitely sensitive, with convergent roles proposed for chemosensory brainstem neurons in the retrotrapezoid nucleus (RTN) and their supporting glial cells. For astrocytes, a central role for NBCe1, a Na+ -HCO3 - cotransporter encoded by Slc4a4, has been envisaged in multiple mechanistic models (i.e. underlying enhanced CO2 -induced local extracellular acidification or purinergic signalling). We tested these NBCe1-centric models by using conditional knockout mice in which Slc4a4 was deleted from astrocytes. In GFAP-Cre;Slc4a4fl/fl mice we found diminished expression of Slc4a4 in RTN astrocytes by comparison to control littermates, and a concomitant reduction in NBCe1-mediated current. Despite disrupted NBCe1 function in RTN-adjacent astrocytes from these conditional knockout mice, CO2 -induced activation of RTN neurons or astrocytes in vitro and in vivo, and CO2 -stimulated breathing, were indistinguishable from NBCe1-intact littermates; hypoxia-stimulated breathing and sighs were likewise unaffected. We obtained a more widespread deletion of NBCe1 in brainstem astrocytes by using tamoxifen-treated Aldh1l1-Cre/ERT2;Slc4a4fl/fl mice. Again, there was no difference in effects of CO2 or hypoxia on breathing or on neuron/astrocyte activation in NBCe1-deleted mice. These data indicate that astrocytic NBCe1 is not required for the respiratory responses to these chemoreceptor stimuli in mice, and that any physiologically relevant astrocytic contributions must involve NBCe1-independent mechanisms. KEY POINTS: The electrogenic NBCe1 transporter is proposed to mediate local astrocytic CO2 /H+ sensing that enables excitatory modulation of nearby retrotrapezoid nucleus (RTN) neurons to support chemosensory control of breathing. We used two different Cre mouse lines for cell-specific and/or temporally regulated deletion of the NBCe1 gene (Slc4a4) in astrocytes to test this hypothesis. In both mouse lines, Slc4a4 was depleted from RTN-associated astrocytes but CO2 -induced Fos expression (i.e. cell activation) in RTN neurons and local astrocytes was intact. Likewise, respiratory chemoreflexes evoked by changes in CO2 or O2 were unaffected by loss of astrocytic Slc4a4. These data do not support the previously proposed role for NBCe1 in respiratory chemosensitivity mediated by astrocytes.

[Development of Nasal Continuum Minimally Invasive Surgical Robot System].

In order to improve the operation difficulties in the narrow space of the nasal maxillary sinus, the nasal continuum minimally invasive surgical robot system is designed. The ball-and-socket joints and NiTiNol tubes are used as the main body of the continuum structure to improve the degree of freedom. The hardware systems and software systems are designed. The security control policies are planned. Finally, the robot confirmed prototype experiments are conducted and the feasibility of continuum robot confirmed through master-slave control experiment and animal experiment.

Development and experiments of a continuum robotic system for transoral laryngeal surgery.

PURPOSE: Currently, self-retaining laryngoscopic surgery is not suitable for some patients, and there are dead zones relating to surgical field exposure and operation. The quality of the surgery can also be affected by the long periods of time required to complete it. Teleoperated continuum robots with flexible joints are expected to solve these issues. However, at the current stage of developing transoral robotic surgery systems, their large size affects the precision of surgical operative actions and there are high development and treatment costs. METHODS: We fabricated a flexible joint based on selective laser melting technology and designed a shallow neural network-based kinematic modeling approach for a continuum surgical robot. Then, human model and animal experiments were completed by master-slave teleoperation to verify the force capability and dexterity of the robot, respectively. RESULTS: As verified by human model and animal experiments, the designed continuum robot was demonstrated to achieve transoral laryngeal surgical field exposure without laryngoscope assistance, with sufficient load capability to finish the biopsy of vocal fold tissue in living animals. CONCLUSION: The designed continuum robotic system allows the biopsy of vocal fold tissue without laryngoscope assistance. Its stiffness and dexterity indicate the system's potential for applications in the diagnosis and treatment of vocal fold nodules and polyps. The limitations of this robotic system as shown in the experiments were also analyzed.

Development and validation of a two-segment continuum robot for maxillary sinus surgery.

BACKGROUND: Existing rigid instruments have difficulties in backward inspection and operation. Moreover, the pathway to the maxillary sinus is curved and narrow, resulting in complex and repetitive manual operations. There is a necessity to develop a retro-flexing robot and achieve path-following motion. METHODS: A continuum robotic system is developed for maxillary sinus surgery (MSS). And the system adopts an anatomical constraint-based optimization of the follow-the-leader strategy to generate a safe control scheme along a given path. RESULTS: The accuracy of the system is evaluated, and the task of reaching deep-seated targets is performed in a constrained anatomical space. The simulations and experiments of the path-following motion have validated the feasibility of the proposed method. Furthermore, a preliminary porcine study is performed to assess the capability of instruments. CONCLUSIONS: The developed continuum robotic system can meet the requirements of MSS.

TRPM4 Contributes to Subthreshold Membrane Potential Oscillations in Multiple Mouse Pacemaker Neurons.

Select neuronal populations display steady rhythmic neuronal firing that provides tonic excitation to drive downstream networks and behaviors. In noradrenergic neurons of the locus coeruleus (LC), circadian neurons of the suprachiasmatic nucleus (SCN), and CO2/H+-activated neurons of the brainstem retrotrapezoid nucleus (RTN), large subthreshold membrane potential oscillations contribute to the pacemaker-like action potential discharge. The oscillations and firing in LC and SCN involve contributions from leak sodium (NALCN) and L-type calcium channels while recent work from RTN suggested an additional pivotal role for a secondary calcium-activated and voltage-gated cationic current sensitive to TRPM4 channel blockers. Here, we tested whether TRPM4 contributes to subthreshold oscillations in mouse LC and SCN. By RNAscope in situ hybridization, Trpm4 transcripts were detected in both cell groups. In whole-cell recordings from acute slice preparations, prominent voltage-dependent membrane potential oscillations were revealed in LC and SCN after blocking action potentials. These oscillations were inhibited by two chemically-distinct blockers of TRPM4, 9-phenanthrol (9-pt) and 4-chloro-2-[[2-(2-chlorophenoxy)acetyl]amino]benzoic acid (CBA). Under whole-cell voltage clamp, inward currents evoked by oscillation voltage waveforms were inhibited in LC by blocking L-type calcium channels and TRPM4. These data implicate TRPM4 in the large subthreshold membrane potential oscillations that underlie tonic action potential discharge in LC and SCN, providing a voltage-dependent and calcium-dependent cationic current to augment the depolarizing inward Na+ and Ca2+ currents previously associated with this distinctive electroresponsive property.

TRPM4 mediates a subthreshold membrane potential oscillation in respiratory chemoreceptor neurons that drives pacemaker firing and breathing.

Brainstem networks that control regular tidal breathing depend on excitatory drive, including from tonically active, CO2/H+-sensitive neurons of the retrotrapezoid nucleus (RTN). Here, we examine intrinsic ionic mechanisms underlying the metronomic firing activity characteristic of RTN neurons. In mouse brainstem slices, large-amplitude membrane potential oscillations are evident in synaptically isolated RTN neurons after blocking action potentials. The voltage-dependent oscillations are abolished by sodium replacement; blocking calcium channels (primarily L-type); chelating intracellular Ca2+; and inhibiting TRPM4, a Ca2+-dependent cationic channel. Likewise, oscillation voltage waveform currents are sensitive to calcium and TRPM4 channel blockers. Extracellular acidification and serotonin (5-HT) evoke membrane depolarization that augments TRPM4-dependent oscillatory activity and action potential discharge. Finally, inhibition of TRPM4 channels in the RTN of anesthetized mice reduces central respiratory output. These data implicate TRPM4 in a subthreshold oscillation that supports the pacemaker-like firing of RTN neurons required for basal, CO2-stimulated, and state-dependent breathing.

A brainstem peptide system activated at birth protects postnatal breathing.

Among numerous challenges encountered at the beginning of extrauterine life, the most celebrated is the first breath that initiates a life-sustaining motor activity1. The neural systems that regulate breathing are fragile early in development, and it is not clear how they adjust to support breathing at birth. Here we identify a neuropeptide system that becomes activated immediately after birth and supports breathing. Mice that lack PACAP selectively in neurons of the retrotrapezoid nucleus (RTN) displayed increased apnoeas and blunted CO2-stimulated breathing; re-expression of PACAP in RTN neurons corrected these breathing deficits. Deletion of the PACAP receptor PAC1 from the pre-Bötzinger complex-an RTN target region responsible for generating the respiratory rhythm-phenocopied the breathing deficits observed after RTN deletion of PACAP, and suppressed PACAP-evoked respiratory stimulation in the pre-Bötzinger complex. Notably, a postnatal burst of PACAP expression occurred in RTN neurons precisely at the time of birth, coinciding with exposure to the external environment. Neonatal mice with deletion of PACAP in RTN neurons displayed increased apnoeas that were further exacerbated by changes in ambient temperature. Our findings demonstrate that well-timed PACAP expression by RTN neurons provides an important supplementary respiratory drive immediately after birth and reveal key molecular components of a peptidergic neural circuit that supports breathing at a particularly vulnerable period in life.

Clinical characteristics and prognosis of community-acquired pneumonia in autoimmune disease-induced immunocompromised host: A retrospective observational study.

BACKGROUND: Community-acquired pneumonia (CAP) in autoimmune diseases (AID)-induced immunocompromised host (ICH) had a high incidence and poor prognosis. However, only a few studies had determined the clinical characteristics of these patients. Our study was to explore the characteristics and predictors of mortality in CAP patients accompanied with AID-induced ICH. METHODS: From 2013 to 2018, a total of 94 CAP patients accompanied with AID-induced ICH, admitted to Emergency Department of Zhongshan Hospital, Fudan University, were enrolled in this study. Clinical data and the risk regression estimates of repeated predictors were evaluated by generalized estimating equations (GEEs) analysis. An open-cohort approach was used to classify patient's outcomes into the survival or non-survival group. RESULTS: The hospital mortality of patients with CAP occurring in AID-induced ICH was 60.64%. No significant differences were found with respect to clinical symptoms and lung images between survival and non-survival groups, while renal insufficiency and dysfunction of coagulation had higher proportions in non-survival patients (P<0.05). Both noninvasive ventilation (NIV) and invasive mechanical ventilation (IMV) were performed more frequently in non-survival group (P< 0.05). By the multivariate GEEs analysis, the repeated measured longitudinal indices of neutrophil-to-lymphocyte ratio (NLR) (odds ratio [OR]=1.055, 95% confidence interval [95%CI] 1.025-1.086), lactate dehydrogenase (LDH) (OR=1.004, 95%CI 1.002-1.006) and serum creatinine (sCr) (OR=1.018, 95%CI 1.008-1.028), were associated with a higher risk of mortality. CONCLUSION: The CAP patients in AID-induced ICH had a high mortality. A significant relationship was demonstrated between the factors of NLR, LDH, sCr and mortality risk in these patients.

Effects of Micronucleus Frequencies and Mitochondrial DNA Copy Numbers among Benzene-Exposed Workers in China.

To provide a more comprehensive understanding of genotoxic effects from benzene exposure, its effects on induction of mitochondrial DNA copy number (MtDNAcn) and of micronucleus (MN) were investigated using peripheral blood from workers in China. Changes in mtDNAcn and MN were determined using quantitative real-time polymerase chain reaction (PCR) and cytokinesis-block micronucleus assays (CBMN), respectively, in 58 control and 174 benzene-exposed workers in Shanghai, China. Among the exposed workers, relative mtDNAcn increased and then decreased with increasing doses of benzene exposure. Significant and dose-dependent increase in MN frequencies were observed among the different exposure groups. In addition, the relative mtDNAcn were significantly associated with the MN frequencies in the low-level exposure group (P = 0.046), but not in the high dose groups. Therefore, the mechanisms for induction of MtDNAcn and MN by benzene may be similar from exposure to low doses but different from high doses. Similar increase of MN frequencies and MtDNAcn may be due to oxidative stress induced by benzene at low concentrations, while higher concentrations may start to initiate the cell death pathway. The pathway may be associated with excessive MtDNAcn which can initiate apoptosis while MN can continue to be induced. However, the differential mechanisms need to be investigated because they may represent different levels of risk for different health consequences. On the other hand, our data indicate that induction of MtDNAcn may be a sensitive genotoxic biomarker for workers with exposure to low dose of benzene. Environ. Mol. Mutagen. 61:355-360, 2020. © 2020 Wiley Periodicals, Inc.

Clinical characteristics, risk factors, immune status and prognosis of secondary infection of sepsis: a retrospective observational study.

BACKGROUND: Secondary infection has a higher incidence in septic patients and affects clinical outcomes. This study aims to investigate the clinical characteristics, risk factors, immune status and prognosis of secondary infection of sepsis. METHODS: A four-year retrospective study was carried out in Zhongshan Hospital, Fudan University, enrolling septic patients admitted between January, 2014 and January, 2018. Clinical data were acquired from medical records. CD14+ monocyte human leukocyte antigen-D related (HLA-DR) expression and serum cytokines levels were measured by flow cytometry and enzyme-linked immunosorbent assay (ELISA) respectively. RESULTS: A total of 297 septic patients were enrolled, 92 of whom developed 150 cases of secondary infections. Respiratory tract was the most common site of secondary infection (n = 84, 56%) and Acinetobacter baumanii the most commonly isolated pathogen (n = 40, 31%). Urinary and deep venous catheterization increased the risk of secondary infection. Lower HLA-DR expression and elevated IL-10 level were found in secondary infection group. The expected prolonged in-hospital stay owing to secondary infection was 4.63 ± 1.87 days. Secondary infection was also associated with higher in-hospital, 30-day and 90-day mortality. Kaplan-Meier survival analysis and Log-rank test revealed that secondary infection group had worse survival between day 15 and day 90. CONCLUSIONS: Urinary and deep venous catheterization increased the risk of secondary infection, in which underlying immunosuppression might also play a role. Secondary infection affected the prognosis of septic patients and prolonged in-hospital length of stay.

Gpr174-deficient regulatory T cells decrease cytokine storm in septic mice.

G protein-coupled receptor 174 (GPR174) is mainly expressed in thymus, spleen, lymph nodes, and leukocytes, and genetic variation in GPR174 is associated with susceptibility to autoimmune diseases, indicating that GPR174 is involved in the immune response. However, the function of GPR174 in regulating inflammatory responses against bacterial infection in sepsis remains unclear. In this study, we investigated the role of GPR174 in regulating suppressive function of regulatory T cells (Treg cells) and the underlying mechanism of Gpr174-deficient Treg cells in controlling cytokine storm of sepsis. We showed that Gpr174-dedicient mice were resistant to inflammatory shock induced by lipopolysaccharide (LPS) and cecal ligation and puncture (CLP). Moreover, Gpr174 was highly expressed in Treg cells, and its deficiency in mice promoted the expression of cytotoxic T lymphocyte associated antigen 4 (CTLA-4) and interleukin (IL)-10 in Treg cells. By using the LPS-induced sepsis model, we demonstrated that anti-inflammatory macrophages (M2 macrophages) induction was Treg cell-dependent and Gpr174-deficient Treg cells protected mice against sepsis-induced lung damage through prompting M2 macrophages polarization. In vitro, Gpr174-deficient Treg cells also promoted the polarization of macrophages toward M2 cells and dampened the secretions of pro-inflammatory cytokines (IL-6 and tumor necrosis factor-α (TNF-α)) in macrophages. In conclusion, these findings suggested that GPR174 plays an important role in the initial period of sepsis through the regulation of macrophage polarization and pro- and anti-inflammatory cytokine secretions. Therefore, GPR174 may be a promising target for therapeutic agents to regulate inflammatory disorders.

Clinical outcomes of otitis media with effusion following palatoplasty in patients with incomplete cleft palate.

BACKGROUND: Diagnosis and treatment strategies for CP with OME and the timing of ear tube insertion remain controversial. OBJECTIVES: To analyze the clinical outcomes of otitis media with effusion in children with incomplete cleft palate after palatoplasty prospectively. METHODS: A total of 30 children (10 months-2 years old) with incomplete CP were enrolled in this study and received at least 6 months of follow-up evaluations after palatoplasty. RESULTS: The overall improvement rate of OME was as high as 26% in this group. Average air conduction hearing threshold was significantly better than that before surgery in the 8 patients with improved OME (p < .05). Among the 8 children with improved OME, 7 (87.5%) were found to have middle ear effusion that improved within 6 months after CP repair. There was no significant difference in the improvement rate between the severe degree II group and the mild group. CONCLUSION: A 6-month follow-up period is recommended. The severity of incomplete CP is not fully related to the function of the eustachian tube. SIGNIFICANCE: The overall improvement rate was as high as 26%, and effusion in the tympanic cavity subsided in 7 out of 8 cases within 6 months after the CP repair.

Clinical characteristics and prognosis of serous body cavity effusions in patients with sepsis: a retrospective observational study.

BACKGROUND: Cavity effusion is common in patients with infectious diseases. However, the incidence rate and characteristics of serous cavity effusions (SCE) in septic patients are not clear to date. The objective of this study was to investigate the incidence and characteristics of SCE in septic patients and to explore the correlations between the bloody effusions and the illness severity/prognosis in septic patients. METHODS: From January 2010 to January 2015, a total of 214 patients with severe sepsis and septic shock were enrolled in this retrospective observational study. Thoracentesis or abdominal paracentesis was performed in 45 septic patients because of massive pleural effusions or ascites. The serum concentrations of VEGF, VEGFR, Ang, sICAM-1, sVCAM-1, E-selectin, Serpine1 and VE-cadherin in 45 septic patients underwent paracentesis were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Of the 214 septic patients, 155 (72.4%) had SCE according to imaging or ultrasound manifestations. 45 subjects with SCE underwent therapeutic thoracentesis or abdominal paracentesis. Effusion laboratory analysis showed that exudates were predominant when compared with transudates (95.6% vs. 4.4%), and 16 (35.6%) patients suffered bloody effusions. Compared with patients with non-bloody effusions, those with bloody effusions showed higher critical illness scores (13 vs. 17 for APACHE II; 7 vs. 9 for SOFA), and higher mortality (6.9% vs. 62.5%). Moreover, patients with bloody effusions had delayed TT and APTT, increased D-dimer concentration, and higher serum levels of CRP and PCT (P < 0.05). In addition, the serum levels of Ang2, sVCAM-1 and E-selectin were significantly higher in patients with bloody effusions than in those with non-bloody effusions (P < 0.05). However, the serum level of VEGFR2 was lower in patients with bloody fluids (P = 0.025). CONCLUSIONS: The incidence of serous cavity effusion is high in patients with sepsis. The septic patients with bloody effusions suffer a more inflammatory burden and a worse prognosis compared to septic patients with non-bloody effusions.

Protein docking refinement by convex underestimation in the low-dimensional subspace of encounter complexes.

We propose a novel stochastic global optimization algorithm with applications to the refinement stage of protein docking prediction methods. Our approach can process conformations sampled from multiple clusters, each roughly corresponding to a different binding energy funnel. These clusters are obtained using a density-based clustering method. In each cluster, we identify a smooth "permissive" subspace which avoids high-energy barriers and then underestimate the binding energy function using general convex polynomials in this subspace. We use the underestimator to bias sampling towards its global minimum. Sampling and subspace underestimation are repeated several times and the conformations sampled at the last iteration form a refined ensemble. We report computational results on a comprehensive benchmark of 224 protein complexes, establishing that our refined ensemble significantly improves the quality of the conformations of the original set given to the algorithm. We also devise a method to enhance the ensemble from which near-native models are selected.

The Role of Ca2+ and BK Channels of Locus Coeruleus (LC) Neurons as a Brake to the CO2 Chemosensitivity Response of Rats.

The cellular mechanisms by which LC neurons respond to hypercapnia are usually attributed to an "accelerator" whereby hypercapnic acidosis causes an inhibition of K+ channels or activation of Na+ and Ca+2 channels to depolarize CO2-sensitive neurons. Nevertheless, it is still unknown if this "accelerator" mechanism could be controlled by a brake phenomenon. Whole-cell patch clamping, fluorescence imaging microscopy and plethysmography were used to study the chemosensitive response of the LC neurons. Hypercapnic acidosis activates L-type Ca2+ channels and large conductance Ca-activated K+ (BK) channels, which function as a "brake" on the chemosensitive response of LC neurons. Our findings indicate that both Ca2+ and BK currents develop over the first 2 weeks of postnatal life in rat LC slices and that this brake pathway may cause the developmental decrease in the chemosensitive firing rate response of LC neurons to hypercapnic acidosis. Inhibition of this brake by paxilline (BK channel inhibitor) returns the magnitude of the chemosensitive firing rate response from LC neurons in rats older than P10 to high values similar to those in LC neurons from younger rats. Inhibition of BK channels in LC neurons by bilateral injections of paxilline into the LC results in a significant increase in the hypercapnic ventilatory response of adult rats. Our findings indicate that a BK channel-based braking system helps to determine the chemosensitive respiratory drive of LC neurons and contributes to the hypercapnic ventilatory response. Perhaps, abnormalities of this braking system could result in hypercapnia-induced respiratory disorders and panic responses.

Distortion product otoacoustic emission together with tympanometry for assessing otitis media with effusion in children.

OBJECTIVE: To investigate the diagnostic value of distortion product otoacoustic emission (DPOAE) together with tympanometry in assessing otitis media with effusion in children. METHODS: Three hundred and thirty-nine patients, who were diagnosed with obstructive sleep apnea-hypopnea syndrome (OSAHS) and prepared to undergo adenotonsillectomy and had a unilateral or bilateral type 'B' or 'C' tympanogram were enrolled in this study. Patients were divided into the following four groups: Group 1, type 'B' tympanogram with positive DPOAE; Group 2, type 'B' tympanogram with negative DPOAE; Group 3, type 'C' tympanogram with positive DPOAE and Group 4, type 'C' tympanogram with negative DPOAE. RESULTS: Tympanometry showed a type 'B' pattern in 467 ears and type 'C' pattern in 163 ears. Among 163 ears with type 'C' tympanogram, negative DPOAE results were seen in 96 ears. Group 4 (56/96) had a significant high rate of middle ear effusion than Group 3 (14/67) (p < .05). In patients with a type 'C' tympanogram, those with a low compliance value (∼0.25 ml), concurrent chronic tonsillitis and large tonsils are likely to have middle ear effusion. CONCLUSIONS: DPOAE could be a great help in screening for middle ear effusion in patients with a type 'C' tympanogram.