bluuebunny/crossref_metadata_2025 huggingface dataset:
Two forms of electrical resonance at theta frequencies, generated by M‐current, h‐current and persistent Na+current in rat hippocampal pyramidal cells
Authors: Hu Hua , Vervaeke Koen , Storm Johan F.
Published: December 2002

Coherent network oscillations in the brain are correlated with different behavioural states. Intrinsic resonance properties of neurons provide a basis for such oscillations. In the hippocampus, CA1 pyramidal neurons show resonance at theta (θ) frequencies (2‐7 Hz). To study the mechanisms underlying θ‐resonance, we performed whole‐cell recordings from CA1 pyramidal cells (n= 73) in rat hippocampal slices. Oscillating current injections at different frequencies (ZAP protocol), revealed clear resonance with peak impedance at 2‐5 Hz at ≈33 °C (increasing to ≈7 Hz at ≈38 °C). The θ‐resonance showed a U‐shaped voltage dependence, being strong at subthreshold, depolarized (≈‐60 mV) and hyperpolarized (≈‐80 mV) potentials, but weaker near the resting potential (‐72 mV). Voltage clamp experiments revealed three non‐inactivating currents operating in the subthresold voltage range: (1) M‐current (IM), which activated positive to ‐65 mV and was blocked by the M/KCNQ channel blocker XE991 (10 μm); (2) h‐current (Ih), which activated negative to ‐65 mV and was blocked by the h/HCN channel blocker ZD7288 (10 μm); and (3) a persistent Na+current (INaP), which activated positive to ‐65 mV and was blocked by tetrodotoxin (TTX, 1 μm). In current clamp, XE991 or TTX suppressed the resonance at depolarized, but not hyperpolarized membrane potentials, whereas ZD7288 abolished the resonance only at hyperpolarized potentials. We conclude that these cells show two forms of θ‐resonance: ‘M‐resonance’ generated by the M‐current and persistent Na+current in depolarized cells, and ‘H‐resonance’ generated by the h‐current in hyperpolarized cells. Computer simulations supported this interpretation. These results suggest a novel function for M/KCNQ channels in the brain: to facilitate neuronal resonance and network oscillations in cortical neurons, thus providing a basis for an oscillation‐based neural code.
Leptin inhibits epileptiform‐like activity in rat hippocampal neurones via PI 3‐kinase‐driven activation of BK channels
Authors: Shanley L. J. , O'Malley D. , Irving A. J. , Ashford M. L. , Harvey J.
Published: December 2002

The obese gene product, leptin is an important circulating satiety factor that regulates energy balance via its actions in the hypothalamus. However, leptin receptors are also expressed in brain regions not directly associated with energy homeostasis, such as the hippocampus. Here, leptin inhibits hippocampal neurones via activation of large conductance Ca2+‐activated K+ (BK) channels, a process that may be important in regulating neuronal excitability. We now show that leptin receptor labelling is expressed on somata, dendrites and axons, and is also concentrated at synapses in hippocampal cultures. In functional studies, leptin potently and reversibly reduces epileptiform‐like activity evoked in lean, but not leptin‐resistant Zucker fa/fa rats. Furthermore, leptin also depresses enhanced Ca2+ levels evoked following Mg2+ removal in hippocampal cultures. The ability of leptin to modulate this activity requires activation of BK, but not KATP, channels as the effects of leptin were mimicked by the BK channel activator NS‐1619, and inhibited by the BK channel inhibitors, iberiotoxin and charybdotoxin. The signalling mechanisms underlying this process involve stimulation of phosphoinositide 3‐kinase (PI 3‐kinase), but not mitogen‐activated protein kinase (MAPK), as two structurally unrelated inhibitors of PI 3‐kinase, LY294002 and wortmannin, blocked the actions of leptin. These data indicate that leptin, via PI 3‐kinase‐driven activation of BK channels, elicits a novel mechanism for controlling neuronal excitability. As uncontrolled excitability in the hippocampus is one underlying cause of temporal lobe epilepsy, this novel action of leptin could provide an alternative therapeutic target in the management of epilepsy.
Cellular mechanisms of orexin actions on paraventricular nucleus neurones in rat hypothalamus
Authors: Follwell Matthew J. , Ferguson Alastair V.
Published: December 2002

Using whole‐cell patch clamp techniques we have examined the cellular mechanisms underlying the effects of orexin A (OX‐A) on electrophysiologically identified magnocellular and parvocellular neurones in the rat hypothalamic paraventricular nucleus (PVN). The majority of magnocellular neurones (67 %) showed concentration‐dependent, reversible depolarizations in response to OX‐A. These effects were abolished in tetrodotoxin (TTX), suggesting them to be indirect effects on this population of neurones. OX‐A also caused increases in excitatory postsynaptic current (EPSC) frequency and amplitude in magnocellular neurones. The former effects were again blocked in TTX while increases in mini‐EPSC amplitude remained. Depolarizing effects of OX‐A on magnocellular neurones were also found to be abolished by kynurenic acid, supporting the conclusion that these effects were the result of activation of a glutamate interneurone. Parvocellular neurones (73 % of those tested) also showed concentration‐dependent, reversible depolarizations in response to OX‐A. In contrast to magnocellular neurones, these effects were maintained in TTX, indicating direct effects of OX‐A on this population of neurones. Voltage clamp analysis using slow voltage ramps demonstrated that OX‐A enhanced a non‐selective cationic conductance with a reversal potential of ‐40 mV in parvocellular neurones, effects which probably explain the depolarizing effects of this peptide in this subpopulation of PVN neurones. These studies have identified separate cellular mechanisms through which OX‐A influences the excitability of magnocellular and parvocellular PVN neurones.
Determinants of relaxation rate in rabbit skinned skeletal muscle fibres
Authors: Luo Ye , Davis Jonathan P. , Smillie Lawrence B. , Rall Jack A.
Published: December 2002

The influence of Ca2+‐activated force, the rate of dissociation of Ca2+ from troponin C (TnC) and decreased crossbridge detachment rate on the time course of relaxation induced by flash photolysis of diazo‐2 in rabbit skinned psoas fibres was investigated at 15 °C. The rate of relaxation increased as the diazo‐2 chelating capacity (i.e. free [diazo‐2]/free [Ca2+]) increased. At a constant diazo‐2 chelating capacity, the rate of relaxation was independent of the pre‐photolysis Ca2+‐activated force in the range 0.3‐0.8 of maximum isometric force. A TnC mutant that exhibited increased Ca2+ sensitivity caused by a decreased Ca2+ dissociation rate in solution (M82Q TnC) also increased the Ca2+ sensitivity of steady‐state force and decreased the rate of relaxation in fibres by approximately twofold. In contrast, a TnC mutant with decreased Ca2+ sensitivity caused by an increased Ca2+ dissociation rate in solution (NHdel TnC) decreased the Ca2+ sensitivity of steady‐state force but did not accelerate relaxation. Decreasing the rate of crossbridge kinetics by reducing intracellular inorganic phosphate concentration ([Pi]) slowed relaxation by approximately twofold and led to two phases of relaxation, a slow linear phase followed by a fast exponential phase. In fibres, M82Q TnC further slowed relaxation in low [Pi] conditions by approximately twofold, whereas NHdel TnC had no significant effect on relaxation. These results are consistent with the interpretation that the Ca2+‐dissociation rate and crossbridge detachment rate are similar in fast‐twitch skeletal muscle, such that decreasing either rate slows relaxation, but accelerating Ca2+ dissociation has little effect on relaxation.
Smooth muscle membrane potential modulates endothelium‐dependent relaxation of rat basilar artery via myo‐endothelial gap junctions
Authors: Allen Tracy , Iftinca Mircea , Cole William C. , Plane Frances
Published: December 2002

The release of endothelium‐derived relaxing factors, such as nitric oxide (NO), is dependent on an increase in intracellular calcium levels ([Ca2+]i) within endothelial cells. Endothelial cell membrane potential plays a critical role in the regulation of [Ca2+]i in that calcium influx from the extracellular space is dependent on membrane hyperpolarization. In this study, the effect of inhibition of vascular smooth muscle delayed rectifier K+ (KDR) channels by 4‐aminopyridine (4‐AP) on endothelium‐dependent relaxation of rat basilar artery to acetylcholine (ACh) was assessed. ACh‐evoked endothelium‐dependent relaxations were inhibited by N‐(Ω)‐nitro‐l‐arginine (l‐NNA) or 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ), confirming a role for NO and guanylyl cyclase. 4‐AP (300 μm) also suppressed ACh‐induced relaxation, with the maximal response reduced from ≈92 to ≈33 % (n= 11; P < 0.01). However, relaxations in response to exogenous NO, applied in the form of authentic NO, sodium nitroprusside or diethylamineNONOate (DEANONOate), were not affected by 4‐AP treatment (n= 3‐11). These data are not consistent with the view that 4‐AP‐sensitive KDR channels are mediators of vascular hyperpolarization and relaxation in response to endothelium‐derived NO. Inhibition of ACh‐evoked relaxation by 4‐AP was reversed by pinacidil (0.5‐1 μm; n= 5) or 18β‐glycyrrhetinic acid (18βGA; 5 μm; n= 5), indicating that depolarization and electrical coupling of the smooth muscle to the endothelium were involved. 4‐AP caused depolarization of both endothelial and vascular smooth muscle cells of isolated segments of basilar artery (mean change 11 ± 1 and 9 ± 2 mV, respectively; n= 15). Significantly, 18βGA almost completely prevented the depolarization of endothelial cells (n= 6), but not smooth muscle cells (n= 6) by 4‐AP. ACh‐induced hyperpolarization of endothelium and smooth muscle cells was also reduced by 4‐AP, but this inhibition was not observed in the combined presence of 4‐AP and 18βGA. These data indicate that 4‐AP can induce an indirect inhibition of endothelium‐dependent relaxation in the rat basilar artery by electrical coupling of smooth muscle membrane depolarization to the endothelium via myo‐endothelial gap junctions.
The history of contraction of the wrist flexors can change cortical excitability
Authors: Stuart Meg , Butler Jane E. , Collins David F. , Taylor Janet L. , Gandevia Simon C.
Published: December 2002

Voluntary contractions induce thixotropic changes in intrafusal muscle fibres and hence, by induction or removal of ‘slack’, the background discharge and sensitivity of spindle endings to stretch is altered. This study assessed whether such changes also altered the ‘excitability’ of the motor cortex. Eleven subjects performed a series of voluntary conditioning contractions of the wrist flexors designed to remove slack in the intrafusal fibres (contract and test at intermediate length, termed ‘contract‐test’) or to introduce slack (contract at long length and test at intermediate length, termed ‘contract‐long’). Surface electromyographic recordings were made from one wrist flexor, flexor carpi radialis. Subjects relaxed after each contraction, and 10 s later a test stimulus was applied to elicit a tendon tap response, H‐reflex, or motor‐evoked potential (MEP) to transcranial magnetic stimulation in the flexor carpi radialis. Each of the three test stimuli was applied during 15 consecutive pairs of contractions (‘contract‐long’ and ‘contract‐test’). Three subjects repeated the protocol using transmastoid electrical stimulation as the test stimulus to evoke a cervicomedullary motor‐evoked potential (CMEP). For the group of subjects, after conditioning contractions designed to induce slack there was a significant reduction in the amplitude of the tendon reflex, no significant change in the H‐reflex, and a small but significant reduction in the amplitude of the MEP. In one subject the CMEP was significantly reduced, while it was unchanged in two others. In the absence of corresponding changes in the H‐reflex (or CMEP), changes in the size of the response to motor cortical stimulation suggest that the level of motor cortical ‘excitability’ changes according to naturally induced variations in the discharge of muscle spindle afferents.
Multiple variations of abdominal vessels in a single cadaver
Authors: Bilge Okan , Aktan A. Asli
Published: December 2002

Variations of abdominal vessels are common and there are many reports about this subject. We report numerous variations of the superior part of the abdominal aorta and renal vessels which were determined in a single cadaver. Clin. Anat. 16:37–39, 2003. © 2002 Wiley‐Liss, Inc.
Index forecasting and model selection
Authors: Haefke Christian , Helmenstein Christian
Published: December 2002

In this paper we derive a trading strategy that exploits the informational difference implied by different stock market index construction principles. In order to gain a competitive advantage over other market participants we forecast the indexes one day ahead and subsequently generate buy and sell signals through the trading rule. To illustrate how the system works we apply it to select from those stocks that are included in the Austrian Traded Index (ATX). The forecasting of the indexes is performed on the basis of standard financial econometric techniques and feedforward neural networks. We discuss the importance of parsimonious modeling and the applicability of information criteria for architecture selection in artificial neural networks. Copyright © 2002 John Wiley & Sons, Ltd.
Genetic basis of male sexual behavior
Authors: Emmons Scott W. , Lipton Jonathan
Published: December 2002

Male sexual behavior is increasingly the focus of genetic study in a variety of animals. Genetic analysis in the soil roundworm Caenorhabditis elegans and the fruit fly Drosophila melanogaster has lead to identification of genes and circuits that govern behaviors ranging from motivation and mate‐searching to courtship and copulation. Some worm and fly genes have counterparts with related functions in higher animals and many more such correspondences can be expected. Analysis of mutations in mammals can potentially lead to insights into such issues as monogamous versus promiscuous sexual behavior and sexual orientation. Genetic analysis of sexual behavior has implications for understanding how the nervous system generates and controls a complex behavior. It can also help us to gain an appreciation of how behavior is encoded by genes and their regulatory sequences. © 2003 Wiley Periodicals, Inc. J Neurobiol 54: 93–110, 2003
Differential scanning calorimetric, circular dichroism, and fourier transform infrared spectroscopic characterization of the thermal unfolding of xylanase A from Streptomyces lividans
Authors: Roberge Martin , Lewis Ruthven N.A.H. , Shareck François , Morosoli Rolf , Kluepfel Dieter , Dupont Claude , McElhaney Ronald N.
Published: December 2002

The thermal unfolding of xylanase A from Streptomyces lividans, and of its isolated substrate binding and catalytic domains, was studied by differential scanning calorimetry and Fourier transform infrared and circular dichroism spectroscopy. Our calorimetric studies show that the thermal denaturation of the intact enzyme is a complex process consisting of two endothermic events centered near 57 and 64°C and an exothermic event centered near 75°C, all of which overlap slightly on the temperature scale. A comparison of the data obtained with the intact enzyme and isolated substrate binding and catalytic domains indicate that the lower‐ and higher‐temperature endothermic events are attributable to the thermal unfolding of the xylan binding and catalytic domains, respectively, whereas the higher‐temperature exothermic event arises from the aggregation and precipitation of the denatured catalytic domain. Moreover, the thermal unfolding of the two domains of the native enzyme are thermodynamically independent and differentially sensitive to pH. The unfolding of the substrate binding domain is a reversible two‐state process and, under appropriate conditions, the refolding of this domain to its native conformation can occur. In contrast, the unfolding of the catalytic domain is a more complex process in which two subdomains unfold independently over a similar temperature range. Also, the unfolding of the catalytic domain leads to aggregation and precipitation, which effectively precludes the refolding of the protein to its native conformation. These observations are compatible with the results of our spectroscopic studies, which show that the catalytic and substrate binding domains of the enzyme are structurally dissimilar and that their native conformations are unaffected by their association in the intact enzyme. Thus, the calorimetric and spectroscopic data demonstrate that the S. lividans xylanase A consists of structurally dissimilar catalytic and substrate binding domains that, although covalently linked, undergo essentially independent thermal denaturation. These observations provide valuable new insights into the structure and thermal stability of this enzyme and should assist our efforts at engineering xylanases that are more thermally robust and otherwise better suited for industrial applications. Proteins 2003;50:341–354. © 2002 Wiley‐Liss, Inc.
Protein side‐chain rearrangement in regions of point mutations
Authors: Eyal Eran , Najmanovich Rafael , Edelman Marvin , Sobolev Vladimir
Published: December 2002

A major problem in predicting amino acid side‐chain rearrangements following point mutations is the potentially large search space. We analyzed a nonredundant data set of 393 Protein Data Bank protein pairs, each consisting of structures differing in one amino acid, to determine the number of residues changing conformation in the region of mutation. In 91–95% of cases, two or fewer residues underwent side‐chain conformational change. If mutation sites with backbone displacements were excluded, the number increased to 97%. The majority of rearrangements (over 60%) were due to the inherent flexibility of side‐chains, as derived from analysis of a control set of protein subunits whose crystal structures were determined more than once. Different amino acids demonstrated different degrees of flexibility near mutation sites. Large polar or charged residues, and serine, are more flexible, while the aromatic amino acids, and cysteine, are less so. This pattern is common to the inherent side‐chain flexibility, as well as the increased flexibility at ligand binding sites and mutation sites. The probability for conformational change was correlated with B‐factor, frequency of the side‐chain conformation in proteins and solvent accessibility. The last trend was stronger for aromatic and hydrophilic residues than for hydrophobic ones. We conclude that the search space for predicting side‐chain conformations in the region of mutation can be effectively restricted. However, the overall ability to predict a particular side‐chain conformation, or to check predictions according to individual existing structures, is limited. These findings may be useful in deriving empirical rules for modeling side‐chain conformations. Proteins 2003;50:272–282. © 2002 Wiley‐Liss, Inc.
Late detection of breast and colorectal cancer in Minnesota counties: an application of spatial smoothing and clustering
Authors: J. Thomas Avis , P. Carlin Bradley
Published: December 2002

Cancers detected at a later disease stage are associated with significantly higher mortality risk. Lack of uniformity in county‐level cancer detection rates is thus of substantial interest to state health departments and other public health professionals. In this paper, we perform several spatial analyses of breast and colorectal cancer detection data for Minnesota counties for 1995–1997. We look for outliers and clusters in the late detection rates using a number of techniques: (i) applying various mapping schemes, (ii) smoothing the data using Bayesian methods implemented via Markov chain Monte Carlo, and (iii) applying maximum likelihood techniques to test for the presence of clusters and to identify the most likely clusters

Our results suggest a fairly uniform spatial pattern in both sets of detection rates. Spatially smoothed rates did not reveal clusters of counties with significantly higher late detection risk, nor were county‐level covariates (such as income, education, and race) particularly helpful in explaining the rates. However, our spatial clustering approach (using the scan statistic) did produce statistically significant clusters of counties which may indicate differences of practical importance for public health. Copyright © 2003 John Wiley & Sons, Ltd.
ICH E9 guideline ‘Statistical principles for clinical trials’: a case study
Authors: Phillips Alan , Haudiquet Vincent
Published: December 2002

The International Conference on Harmonization (ICH) E9 guideline ‘Statistical principles for clinical trials’ was adopted by the Committee for Proprietary Medicinal Products in March 1998, and consequently is operational in Europe. It has also been adopted in the U.S.A. and Japan. The aim of this paper is to relate the problems encountered during a recent regulatory submission to those discussed in the ICH E9 guideline. Statistical principles discussed in the guideline, but not comprehensively addressed when the clinical development programme was initiated in the mid‐1990s, will be reviewed. The impact of each issue on the approvability of the dossier will be discussed, together with recommendations on how to avoid such problems with other ongoing clinical development programmes. Copyright © 2003 John Wiley & Sons, Ltd.
Probability density of displacement and overturning length scales under diverse stratification
Authors: Lorke Andreas , Wüest Alfred
Published: December 2002

Vertical overturns, produced by turbulence in density‐stratified lakes and oceans, are often quantified by the Thorpe scale, LT. The correlation between LT and the Ozmidov (energy‐containing) scale can be used to estimate rates of turbulent dissipation and vertical diffusivities. Based on temperature microstructure measurements from several stratified lakes, the probability density functions (pdf's) of Thorpe displacements and overturning length scales are distinguished and discussed. The stratification was varying over five orders of magnitude between the different data sets, resulting in Thorpe scales between 1 cm and 100 m. It is shown that the analyzed pdf's follow a universal form that can be empirically described by an exponential function and parameterized by a single length scale. The functional form of the pdf of overturning length scales can be related to the inertial subrange, which is found in temperature, as well as in length‐scale fluctuation spectra. As a result, the ratio between the maximum displacement length scale, Lmax, and the Thorpe scale depends on the rate of turbulent kinetic energy dissipation. Based on the universal pdf of overturning length scales, we show with numerical simulations that Lmax is a more appropriate macroscopic length scale than LT for the estimation of turbulence based on displacements in temperature profiles.
Radiative and microphysical properties of Arctic stratus clouds from multiangle downwelling infrared radiances
Authors: Rathke Carsten , Neshyba Steven , Shupe Matthew D. , Rowe Penny , Rivers Aaron
Published: December 2002

The information content of multiangle downwelling infrared radiance spectra of stratus clouds is investigated. As an example, 76 sets of spectra were measured at angles of 0, 15, 30 and 45° from zenith, using an interferometer based at the Surface Heat Budget of the Arctic Ocean (SHEBA) drifting ice camp. Exploiting the angular variation of radiance in infrared microwindows, a “geometric” algorithm is used to determine cloud temperature and optical depth without auxiliary information. For comparison, a spectral method allows us to infer cloud microphysical properties for each angle; each multiangle set therefore constitutes a microphysical characterization of horizontal inhomogeneity of the cloudy scene. We show that cloud temperatures determined with both approaches agree with temperatures obtained from lidar/radiosonde data. The multiangle radiance observations can also be used to calculate the longwave flux reaching the surface. We find that up to 14 W m−2 of the overcast fluxes can be attributed to horizontal variations in cloud microphysical properties.
Carbon isotope geochemistry and nanomorphology of soil black carbon: Black chernozemic soils in central Europe originate from ancient biomass burning
Authors: Schmidt Michael W. I. , Skjemstad Jan O. , Jäger Cornelia
Published: December 2002

A common paradigm is that chernozem soils developed in the Holocene under grassland steppes, with their formation largely determined by three factors, parent material, climate and faunal mixing. For European chernozems, however, pollen records show that steppes were rare. Here, using high‐resolution transmission electron microscopy, electron energy loss spectroscopy, micro Raman spectroscopy and radiocarbon dating, we characterized the nanomorphology and chemical structure of soil organic carbon (SOC) from central European chernozems. We identified submicron remnants of burned biomass (15–45 percent of SOC), coexisting as amorphous char‐black carbon (BC) derived from pyrolized cellulose or soot‐BC. The BC was several millenia in age (1160–5040 carbon‐14 years) and up to 3990 radiocarbon years older than bulk SOC, indicating significant residence times for BC in soils. These results challenge common paradigms on chernozem formation and add fire as an important novel factor. It is also clear that the role of fire in soil formation has been underestimated outside classical fire prone biomes. Furthermore, our results demonstrate the importance of quantifying BC in soils because of its large contribution, longevity and potential role in the global biogeochemical carbon cycle.
Nicotinic ACH receptor subtypes on gastrointestinally projecting neurones in the dorsal motor vagal nucleus of the rat
Authors: Sahibzada Niaz , Ferreira Manuel , Williams Bernice , Wasserman Adam , Vicini Stefano , Gillis Richard A.
Published: December 2002

To determine the predominant nicotinic ACh receptor (nAChR) located on neurones in the dorsal motor nucleus of the vagus (DMV) that project to the gastrointestinal tract, we used the rat brainstem slice preparation and whole‐cell recordings of DMV neurones identified by retrograde DiI tracing to pharmacologically characterize nAChRs. Pressure ejection of acetylcholine (ACh, 250 μm for 200 ms) from a patch pipette placed ≈10‐20 μm from the surface of the recorded cell produced an inward current in most DMV neurones sampled. The average currents for neurones projecting to the fundus, antrum and caecum were 149 ± 38 (n= 25), 115 ± 18 (n= 29) and 117 ± 23 pA (n= 6), respectively. Blockade of the α7 subtype of nAChR with either α‐bungarotoxin (α‐BGT) or methyllycaconitine (MLA) counteracted 60‐75 % of the ACh‐evoked current in DMV neurones projecting to the fundus, antrum and caecum. In neurones projecting to the fundus and the antrum, currents resistant to α‐BGT were significantly blocked by dihydro‐β‐erythroidine (10‐20 nm), an antagonist of the α4β2 subtype of nAChR. In neurones projecting to the caecum, currents resistant to α‐BGT were significantly depressed by a low concentration of mecamylamine (1 μm). Cytisine (100 μm), an agonist of nAChRs that contain the α7 or the β4 subunit, evoked significant currents in caecum‐projecting neurones that were previously exposed to α‐BGT. In contrast, cytisine had no effect on DMV neurones previously exposed to α‐BGT that project to the fundus or antrum. Our data indicate that the prevailing nAChR subtype in DMV neurones projecting to the GI tract is the α7 subtype. In addition, we obtained evidence for the co‐expression of the α4β2 nAChR subtype on DMV neurones projecting to the fundus and antrum, and the α3β4 nAChR subtype on DMV neurones projecting to the caecum.
Opioid‐resistant respiratory pathway from the preinspiratory neurones to abdominal muscles: in vivo and in vitro study in the newborn rat
Authors: Janczewski Wiktor A. , Onimaru Hiroshi , Homma Ikuo , Feldman Jack L.
Published: December 2002

We report that after spontaneous breathing movements are stopped by administration of opioids (opioid‐induced apnoea) in neonatal rats, abdominal muscles continue to contract at a rate similar to that observed during periods of ventilation. Correspondingly, in vitro bath application of a μ opioid receptor agonist suppresses the activity of the fourth cervical root (C4) supplying the diaphragm, but not the rhythmic activity of the first lumbar root (L1) innervating the abdominal muscles. This indicates the existence of opioid‐resistant rhythmogenic neurones and a neuronal pathway transmitting their activity to the abdominal motoneurones. We have investigated this pathway by using a brainstem‐spinal cord preparation of the neonatal rat. We identified bulbospinal neurones with a firing pattern identical to that of the L1 root. These neurones were located caudal to the obex in the vicinity of the nucleus retroambiguus. Resting potentials ranged from ‐49 to ‐40 mV (mean ±s.d. ‐44.0 ± 4.3 mV). The mean input resistance was 315.5 ± 54.8 MΩ. The mean antidromic latency from the L1 level was 42.8 ± 4.4 ms. Axons crossed the midline at the level of the cell body. The activity pattern of the bulbospinal neurones and the L1 root consisted of two bursts per respiratory cycle with a silent period during inspiration. This pattern is characteristic of preinspiratory neurones. We found that 11 % of the preinspiratory neurones projected to the area where the bulbospinal neurones were located. These preinspiratory neurones were found in the rostral ventrolateral medulla close (200‐350 μm) to the ventral surface at the level of the rostral half of the nucleus retrofacialis. Our data suggest the operation of a disynaptic pathway from the preinspiratory neurones to the L1 motoneurones in the in vitro preparation. We propose that the same pathway is responsible for rhythmic activation of the abdominal muscles during opioid‐induced apnoea in the newborn rat.
Role of the flocculus in mediating vestibular nucleus neuron plasticity during vestibular compensation in the rat
Authors: Johnston Alex R. , Seckl Jonathan R. , Dutia Mayank B.
Published: December 2002

We investigated the role of the cerebellar flocculus in mediating the adaptive changes that occur in the intrinsic properties of brainstem medial vestibular nucleus (MVN) neurons during vestibular compensation. Ipsi‐lesional, but not contra‐lesional, flocculectomy prevented the compensatory increase in intrinsic excitability (CIE) that normally occurs in the de‐afferented MVN neurons within 4 h after unilateral labyrinthectomy (UL). Flocculectomy did not, however, prevent the down‐regulation of efficacy of GABA receptors that also occurs in these neurons after UL, indicating that these responses of the MVN neurons to deafferentation are discrete, parallel processes. CIE was also abolished by intra‐floccular microinjection of the metabotropic glutamate receptor (mGluR) antagonist AIDA, and the protein kinase C inhibitor bisindolymaleimide I (BIS‐I). The serene‐threonine kinase inhibitor H‐7 had no effect when microinjected at the time of de‐afferentation, but abolished CIE if microinjected 2 h later. These cellular effects are in line with the recently reported retardatory effects of BIS‐I and H‐7 on behavioural recovery after UL. They demonstrate that the increase in intrinsic excitability in MVN neurons during vestibular compensation is cerebellum dependent, and requires mGluR activation and protein phosphorylation in cerebellar cortex. Furthermore, microinjection of the glucocorticoid receptor (GR) antagonist RU38486 into the ipsi‐lesional flocculus also abolished CIE in MVN neurons. Thus an important site for glucocorticoids in facilitating vestibular compensation is within the cerebellar cortex. These observations ascribe functional significance to the high levels of GR and 11‐β‐HSD Type 1 expression in cerebellum.
Expression of a calpastatin transgene slows muscle wasting and obviates changes in myosin isoform expression during murine muscle disuse
Authors: Tidball James G. , Spencer Melissa J.
Published: December 2002

Muscle wasting is a prominent feature of several systemic diseases, neurological damage and muscle disuse. The contribution of calpain proteases to muscle wasting in any instance of muscle injury or disease has remained unknown because of the inability to specifically perturb calpain activity in vivo. We have generated a transgenic mouse with muscle‐specific overexpression of calpastatin, which is the endogenous inhibitor of calpains, and induced muscle atrophy by unloading hindlimb musculature for 10 days. Expression of the transgene resulted in increases in calpastatin concentration in muscle by 30‐ to 50‐fold, and eliminated all calpain activity that was detectable on zymograms. Muscle fibres in ambulatory, transgenic mice were smaller in diameter, but more numerous, so that muscle mass did not differ between transgenic and non‐transgenic mice. This is consistent with the role of the calpain‐calpastatin system in muscle cell fusion that has been observed in vitro. Overexpression of calpastatin reduced muscle atrophy by 30 % during the 10 day unloading period. In addition, calpastatin overexpression completely prevented the shift in myofibrillar myosin content from slow to fast isoforms, which normally occurs in muscle unloading. These findings indicate that therapeutics directed toward regulating the calpain‐calpastatin system may be beneficial in preventing muscle mass loss in muscle injury and disease.
Activation of a ptx‐insensitive g protein is involved in histamine‐induced recombinant m‐channel modulation
Authors: Guo Juan , Schofield Geoffery G.
Published: December 2002

The M‐type potassium current (IM) plays a dominant role in regulating membrane excitability and is modulated by many neurotransmitters. However, except in the case of bradykinin, the signal transduction pathways involved in M‐channel modulation have not been fully elucidated. The channels underlying IM are produced by the coassembly of KCNQ2 and KCNQ3 channel subunits and can be expressed in heterologous systems where they can be modulated by several neurotransmitter receptors including histamine H1 receptors. In HEK293T cells, histamine acting via transiently expressed H1R produced a strong inhibition of recombinant M‐channels but had no overt effects on the voltage dependence or voltage range of IM activation. In addition, the modulation of IM by histamine was not voltage sensitive, whereas channel gating, particularly deactivation, was accelerated by histamine. Non‐hydrolysable guanine nucleotide analogues (GDP‐β‐S and GTP‐γ‐S) and pertussis toxin (PTX) treatment demonstrated the involvement of a PTX‐insensitive G protein in the signal transduction pathway mediating histamine‐induced IM modulation. Abrogation of the histamine‐induced modulation of IM by expression of a C‐terminal construct of phospholipase C (PLC‐β1‐ct), which buffers activated Gαq/11 subunits, implicates this G protein α subunit in the modulatory pathway. On the other hand, abrogation of the histamine‐induced modulation of IM by expression of two constructs which buffer free βγ subunits, transducin (Gαt) and a C‐terminal construct of a G protein receptor kinase (MAS‐GRK2‐ct), implicates βγ dimers in the modulatory pathway. These findings demonstrate that histamine modulates recombinant M‐channels in HEK293T cells via a PTX‐insensitive G protein, probably Gαq/11, in a similar manner to a number of other G protein‐coupled receptors. However, histamine‐induced IM modulation in HEK293T cells is novel in that βγ subunits in addition to Gαq/11 subunits appear to be involved in the modulation of KCNQ2/3 channel currents.
The relation between charge movement and transport‐associated currents in the rat GABA cotransporter rGAT1
Authors: Fesce Riccardo , Giovannardi Stefano , Binda Francesca , Bossi Elena , Peres Antonio
Published: December 2002

Most cotransporters characteristically display two main kinds of electrical activity: in the absence of organic substrate, transient presteady‐state currents (Ipre) are generated by charge relocation during voltage steps; in the presence of substrate, sustained, transport‐associated currents (Itr) are recorded. Quantitative comparison of these two currents, in Xenopus oocytes expressing the neural GABA cotransporter rGAT1, revealed several unforeseen consistencies between Ipre and Itr, in terms of magnitude and kinetic parameters. The decay rate constant (r) of Ipre and the quantity of charge displaced to an inner position in the transporter (Qin(0)) depended on voltage and ionic conditions. Saturating GABA concentrations, applied under the same conditions, suppressed Ipre (i.e. Qin(∞) = 0) and produced a transport‐associated current with amplitude Itr=Qin(0)r. At non‐saturating levels of GABA, changes of Itr were compensated by corresponding variations in Qin, such that Ipre and Itr complemented each other, according to the relation: Itr= (Qin(0) ‐ Qin) r. Complementarity of magnitude, superimposable kinetic properties and equal dependence on voltage and [Na+]o point to the uniqueness of the charge carrier for both processes, suggesting that transport and charge migration arise from the same molecular mechanism. The observed experimental relations were correctly predicted by a simple three‐state kinetic model, in which GABA binding takes place after charge binding and inward migration have occurred. The model also predicts the observed voltage dependence of the apparent affinity of the transporter for GABA, and suggests a voltage‐independent GABA binding rate with a value around 0.64 μm−1 s−1.
Distinct effect of actin cytoskeleton disassembly on exo‐ and endocytic events in a membrane patch of rat melanotrophs
Authors: Chowdhury Helena H. , Kreft Marko , Zorec Robert
Published: December 2002

We used the cell‐attached mode of patch‐clamp technique to measure discrete attofarad steps in membrane capacitance (Cm), reporting area changes in the plasma membrane due to unitary exocytic and endocytic events. To investigate the role of the actin cytoskeleton in elementary exocytic and endocytic events, neuroendocrine rat melanotrophs were treated with Clostridium spiroforme toxin (CST), which specifically depolymerises F‐actin. The average amplitude of exocytic events was not significantly different in control and in CST‐treated cells. However, the amplitude of endocytic events was significantly smaller in CST‐treated cells as compared to controls. The frequency of exocytic events increased by 2‐fold in CST‐treated cells relative to controls. In control cells the average frequency of exocytic events (νexo) was lower than the frequency of endocytic events (νendo) with a ratio νexo/νendo < 1. In the toxin treated cells, the predominant process was exocytosis with a ratio (νexo/νendo > 1). To study the coupling between the two processes, the slopes of regression lines relating νexo and νendo in a given patch of membrane were studied. The slopes of regression lines were similar, whereas the line intercepts with the y‐axis were significantly different. The increased frequency of unitary exocytic events in CST‐treated cells is consistent with the view, that the actin cytoskeleton acts as a barrier for exocytosis. While the disassembly of the actin cytoskeleton diminishes the size of unitary endocytic events, suggesting an important role of the actin cytoskeleton in determining the size of endocytic vesicles, the coupling between exocytosis and endocytosis in a given patch of membrane was independent of the state of the actin cytoskeleton.
Rapid non‐genomic inhibition of ATP‐induced Cl− secretion by dexamethasone in human bronchial epithelium
Authors: Urbach V. , Walsh D. E. , Mainprice B. , Bousquet J. , Harvey B. J.
Published: December 2002

A non‐genomic antisecretory role for dexamethasone at low concentrations (0.1 nm to1 μm) is described in monolayers of human bronchial epithelial cells in primary culture and in a continuous cell line (16HBE14o‐ cells). Dexamethasone produced a rapid decrease of [Ca2+]i (measured with fura‐2 spectrofluorescence) to a new steady‐state concentration. After 15 min exposure to dexamethasone (1 nm), [Ca2+]i was reduced by 32 ± 11 nm (n= 7, P < 0.0001) from a basal value of 213 ± 36 nm (n= 7). We have shown previously that aldosterone (1 nm) also produces a rapid fall in [Ca2+]i; however, after the decrease in [Ca2+]i induced by dexamethasone, subsequent addition of aldosterone did not produced any further lowering of [Ca2+]i. The rapid response to dexamethasone was insensitive to pretreatment with cycloheximide and unaffected by the glucocorticoid type II and mineralocorticoid receptor antagonists RU486 and spironolactone, respectively. The rapid [Ca2+]i decrease induced by dexamethasone was inhibited by the Ca2+‐ATPase pump inhibitor thapsigargin (1 μm), the adenylate cyclase inhibitor MDL hydrochloride (500 μm) and the protein kinase A inhibitor Rp‐adenosine 3′,5′‐cyclic monophosphorothioate (200 μm), but was not affected by the protein kinase C inhibitor, chelerythrine chloride (0.1 μm). Treatment of 16HBE14o‐ cell monolayers with dexamethasone (1 nm) inhibited the large and transient [Ca2+]i increase induced by apical exposure to ATP (10−4m). Dexamethasone (1 nm) also reduced by 30 % the Ca2+‐dependant Cl− secretion induced by apical exposure to ATP (measured as the Cl−‐sensitive short‐circuit current across monolayers mounted in Ussing chambers). Our results demonstrate, for the first time, that dexamethasone at low concentrations inhibits Cl− secretion in human bronchial epithelial cells. The rapid inhibition of Cl− secretion induced by the synthetic glucocorticoid is associated with a rapid decrease in [Ca2+]i via a non‐genomic mechanism that does not involve the classical glucocorticoid or mineralocorticoid receptor. Rather, it is a result of rapid non‐genomic stimulation of thapsigargin‐sensitive Ca2+‐ATPase, via adenylate cyclase and protein kinase A signalling.
Freshly isolated bovine coronary endothelial cells do not express the BKca channel gene
Authors: Gauthier Kathryn M. , Liu Caiqiong , Popovic Aleksandra , Albarwani Sulayma , Rusch Nancy J.
Published: December 2002

Recent reports have suggested that different types of Ca2+‐activated K+ channels may be selectively expressed either in the vascular endothelial cells (ECs) or smooth muscle cells (SMCs) of a single artery. In this study, we directly compared mRNA, protein and functional expression of the high‐conductance Ca2+‐activated K+ (BKCa) channel between freshly isolated ECs and SMCs from bovine coronary arteries. Fresh ECs and SMCs were enzymatically isolated, and their separation verified by immunofluorescent detection of α‐actin and platelet/endothelium cell adhesion molecule (PECAM) proteins, respectively. Subsequently, studies using a sequence‐specific antibody directed against the pore‐forming α‐subunit of the BKCa channel only detected its expression in the SMCs, whereas PECAM‐positive ECs were devoid of the α‐subunit protein. Additionally, multicell RT‐PCR performed using cDNA derived from either SMCs or ECs only detected mRNA encoding the BKCaα‐subunit in the SMCs. Finally, whole‐cell recordings of outward K+ current detected a prominent iberiotoxin‐sensitive BKCa current in SMCs that was absent in ECs, and the BKCa channel opener NS 1619 only enhanced K+ current in the SMCs. Thus, bovine coronary SMCs densely express BKCa channels whereas adjacent ECs in the same artery appear to lack the expression of the BKCa channel gene. These findings indicate a cell‐specific distribution of Ca2+‐activated K+ channels in SMCs and ECs from a single arterial site.
The effects of ischaemic preconditioning, diazoxide and 5‐hydroxydecanoate on rat heart mitochondrial volume and respiration
Authors: Lim Kelvin H. H. , Javadov Sabzali A. , Das Manika , Clarke Samantha J. , Suleiman M.‐Saadeh , Halestrap Andrew P.
Published: December 2002

Studies with different ATP‐sensitive potassium (KATP) channel openers and blockers have implicated opening of mitochondrial KATP (mitoKATP) channels in ischaemic preconditioning (IPC). It would be predicted that this should increase mitochondrial matrix volume and hence respiratory chain activity. Here we confirm this directly using mitochondria rapidly isolated from Langendorff‐perfused hearts. Pre‐ischaemic matrix volumes for control and IPC hearts (expressed in μl per mg protein ±s.e.m., n= 6), determined with 3H2O and [14C]sucrose, were 0.67 ± 0.02 and 0.83 ± 0.04 (P < 0.01), respectively, increasing to 1.01 ± 0.05 and 1.18 ± 0.02 following 30 min ischaemia (P < 0.01) and to 1.21 ± 0.13 and 1.26 ± 0.25 after 30 min reperfusion. Rates of ADP‐stimulated (State 3) and uncoupled 2‐oxoglutarate and succinate oxidation increased in parallel with matrix volume until maximum rates were reached at volumes of 1.1 μl ml−1 or greater. The mitoKATP channel opener, diazoxide (50 μm), caused a similar increase in matrix volume, but with inhibition rather than activation of succinate and 2‐oxoglutarate oxidation. Direct addition of diazoxide (50 μm) to isolated mitochondria also inhibited State 3 succinate and 2‐oxoglutarate oxidation by 30 %, but not that of palmitoyl carnitine. Unexpectedly, treatment of hearts with the mitoKATP channel blocker 5‐hydroxydecanoate (5HD) at 100 or 300 μm, also increased mitochondrial volume and inhibited respiration. In isolated mitochondria, 5HD was rapidly converted to 5HD‐CoA by mitochondrial fatty acyl CoA synthetase and acted as a weak substrate or inhibitor of respiration depending on the conditions employed. These data highlight the dangers of using 5HD and diazoxide as specific modulators of mitoKATP channels in the heart.
SIGNATURES OF THE NEUTRAL TOP-PION IN eγ COLLISIONS
Authors: YUE CHONGXING , LI HONG , WANG XUELEI
Published: December 2002

We calculate the contributions of the neutral top-pion [Formula: see text] to the process [Formula: see text] in the framework of topcolor-assisted technicolor (TC2) models. Our results show that [Formula: see text] can give significant contributions to this process. With reasonable values of the parameters in TC2 models, the cross-section σ can reach 1.3 pb which may be detected at the eγ collisions based on the future high energy linear e+e- collider experiments.
QFT, STRINGS AND LOW-DIMENSIONAL TOPOLOGY
Authors: SEN HU
Published: December 2002

In between the 80's and 90's we witnessed deep interactions between mathematics and theoretical physics, especially in the understanding of low-dimensional topology in terms of quantum field theory. For example, Jones polynomials (Chern–Simons–Witten theory), Donaldson and Seiberg–Witten invariants (SUSY Yang–Mills theory) and mirror symmetry (T duality in strings) are all naturally understood in terms of QFT and strings. Recent developments indicate a close relationship between gauge theory and gravity theory both in physics and in low-dimensional topology. We shall survey these developments and report some of our work. We shall also find that the keys to connect geometric and physical objects are through symmetry and quantization.
DOPING AND GROWTH OF THIN Si EPILAYER AND SiGe BY UHV/CVD
Authors: HUANG JINGYUN , WANG LEI , ZHAO BINGHUI , YE ZHIZHEN
Published: December 2002

The real time B-doped thin Si and SiGe epilayers on diameter 3-inch Si substrates were grown by an ultrahigh vacuum chemical vapor deposition (UHV/CVD) system. The substrate temperature during growth was from 550 ~ 780°C. The properties of the epilayers were characterized by secondary ion mass spectrum (SIMS), Fourier transform infrared spectroscopy (FTIR), spreading resistance profile (SRP) and double crystal x-ray diffraction (DCXRD). The carrier concentration in the intrinsic Si epilayer was about 1012 cm -3 and the resistivity was as high as about 10000 Ωcm. In addition, the abrupt transition and B-doped concentration of 1015 ~ 1019 cm -3 were achieved for Si and SiGe epilayers.