Slow Rise Time Research Articles

Cyclin Dependent Kinase 2 (CDK2) Regulates Secretory Function in Pancreatic Beta-Cells

Cyclin dependent kinase 2 (CDK2) is a well-known regulator of the cell cycle in diverse types of cells. In pancreatic β-cells, CDKs have been the subject of much research devoted towards increasing β-cell mass by driving proliferation in diabetics, where there is a conspicuous loss of β-cell mass. However, mice lacking CDK2 in their pancreatic β-cells (Pdx1-Cre:CDK2–/– mice) exhibited glucose intolerance and reduced insulin release from isolated islets before losing β-cell mass. To examine the underlying defects, optical imaging of NAD(P)H/flavin autofluorescence was used to directly measure glycolytic and mitochondrial activity, which both trigger and control refilling of the readily-releasable granule pool in β-cells. By comparison to control islets, Pdx1-Cre:Cdk2–/– islets exhibited a reduced baseline, slower rise time, and a reduced maximal NAD(P)H/flavin response, which correlated with increased mitochondrial volume in EM images. Surprisingly, Ca2+ signaling - the direct trigger for β-cell insulin granule release - was not responsible for the observed secretory defect since the intracellular Ca2+ oscillations of Pdx1-Cre:Cdk2–/– islets were found to be more sensitive to glucose than control islets. Ca2+ channel activation is therefore uncoupled from both metabolism and secretion in this model. Uncoupling of secretion may further involve the exocytotic machinery, since mRNA levels of the SNARE protein SNAP25, and the glucose-dependent priming factor RAB27A, were reduced by CDK2 ablation. Taken together, our data indicates that CDK2 maintains insulin secretion by regulating metabolic output and plasma membrane electrical activity, and suggests further CDK2-dependent regulation of the exocytotic machinery, roles that are in distinct to regulation of β-cell proliferative capacity. Supported by F32DK085960 (MJM), the NIDDK Intramural Research Program (SGR), and R01DK46409 (LSS).

Characteristics of current pulses in rocket-triggered lightning

The waveform parameters of current pulses in negative rocket-triggered lightning are analyzed statistically using recorded channel-base current during Shandong Artificially Triggered Lightning Experiment (SHATLE) from 2005 to 2011. The current pulses are divided into 3 categories, including return strokes, ICC pulses in initial continuous current (ICC) stage and M-components superimposed on the continuing current following the return stroke. The geometric mean values (GM) of peak current, risetime from 10% to 90% peak, half peak width and charge transfer for return strokes are 12.1kA, 1.0μs, 14.8μs, and 0.86C, respectively. The corresponding parameters for ICC pulses are 0.09kA, 437μs, 712μs, and 0.10C, respectively, while they are 0.28kA, 251μs, 242μs and 0.10C for M-components, respectively. There is significant fraction of larger M-components (about 10% of 63 samples) demonstrating peak current of several kilo-amperes, comparable to weak return strokes. The ICC pulses show somewhat lower current peak and slower risetime than the M-components with a difference of roughly 2–3 times, resulting from the lower charge source of ICC pulses. In general, the ICC pulses and M-components show smaller peak current and charge transfer while longer risetime and half peak width than return strokes, differing with 1–2 orders of magnitude. The charge neutralized by an individual triggered lightning flash ranged from 6.3C to 68.1C, while the initial stage (IS), lasted about 245ms (GM) and lowered a GM charge of 21.2C to the ground. The average IS current in an individual lightning discharge varies from 45.8A to 140.6A with a GM value of 86.7A.

γ-Aminobutyric Acid Type A (GABAA) Receptor α Subunits Play a Direct Role in Synaptic Versus Extrasynaptic Targeting

GABA(A) receptors (GABA(A)-Rs) are localized at both synaptic and extrasynaptic sites, mediating phasic and tonic inhibition, respectively. Previous studies suggest an important role of γ2 and δ subunits in synaptic versus extrasynaptic targeting of GABA(A)-Rs. Here, we demonstrate differential function of α2 and α6 subunits in guiding the localization of GABA(A)-Rs. To study the targeting of specific subtypes of GABA(A)-Rs, we used a molecularly engineered GABAergic synapse model to precisely control the GABA(A)-R subunit composition. We found that in neuron-HEK cell heterosynapses, GABAergic events mediated by α2β3γ2 receptors were very fast (rise time ∼2 ms), whereas events mediated by α6β3δ receptors were very slow (rise time ∼20 ms). Such an order of magnitude difference in rise time could not be attributed to the minute differences in receptor kinetics. Interestingly, synaptic events mediated by α6β3 or α6β3γ2 receptors were significantly slower than those mediated by α2β3 or α2β3γ2 receptors, suggesting a differential role of α subunit in receptor targeting. This was confirmed by differential targeting of the same δ-γ2 chimeric subunits to synaptic or extrasynaptic sites, depending on whether it was co-assembled with the α2 or α6 subunit. In addition, insertion of a gephyrin-binding site into the intracellular domain of α6 and δ subunits brought α6β3δ receptors closer to synaptic sites. Therefore, the α subunits, together with the γ2 and δ subunits, play a critical role in governing synaptic versus extrasynaptic targeting of GABA(A)-Rs, possibly through differential interactions with gephyrin.

Functional Changes in Muscle Afferent Neurones in an Osteoarthritis Model: Implications for Impaired Proprioceptive Performance

BackgroundImpaired proprioceptive performance is a significant clinical issue for many who suffer osteoarthritis (OA) and is a risk factor for falls and other liabilities. This study was designed to evaluate weight-bearing distribution in a rat model of OA and to determine whether changes also occur in muscle afferent neurones.Methodology/Principal FindingsIntracellular recordings were made in functionally identified dorsal root ganglion neurones in acute electrophysiological experiments on the anaesthetized animal following measurements of hind limb weight bearing in the incapacitance test. OA rats but not naïve control rats stood with less weight on the ipsilateral hind leg (P = 0.02). In the acute electrophysiological experiments that followed weight bearing measurements, action potentials (AP) elicited by electrical stimulation of the dorsal roots differed in OA rats, including longer AP duration (P = 0.006), slower rise time (P = 0.001) and slower maximum rising rate (P = 0.03). Depolarizing intracellular current injection elicited more APs in models than in naïve muscle afferent neurones (P = 0.01) indicating greater excitability. Axonal conduction velocity in model animals was slower (P = 0.04).Conclusions/SignificanceThe present study demonstrates changes in hind limb stance accompanied by changes in the functional properties of muscle afferent neurones in this derangement model of OA. This may provide a possible avenue to explore mechanisms underlying the impaired proprioceptive performance and perhaps other sensory disorders in people with OA.

Effect of pulse profile and chirp on a laser wakefield generation

A laser wakefield driven by an asymmetric laser pulse with/without chirp is investigated analytically and through two-dimensional particle-in-cell simulations. For a laser pulse with an appropriate pulse length compared with the plasma wavelength, the wakefield amplitude can be enhanced by using an asymmetric un-chirped laser pulse with a fast rise time; however, the growth is small. On the other hand, the wakefield can be greatly enhanced for both positively chirped laser pulse having a fast rise time and negatively chirped laser pulse having a slow rise time. Simulations show that at the early laser-plasma interaction stage, due to the influence of the fast rise time the wakefield driven by the positively chirped laser pulse is more intense than that driven by the negatively chirped laser pulse, which is in good agreement with analytical results. At a later time, since the laser pulse with positive chirp exhibits opposite evolution to the one with negative chirp when propagating in plasma, the wakefield in the latter case grows more intensely. These effects should be useful in laser wakefield acceleration experiments operating at low plasma densities.

Decoupling the artificial special pair to slow down the rate of singlet energy transfer

Trimer 2, composed of a cofacial heterobismacrocycle, octamethyl-porphyrin zinc(II) and bisarylporphyrin zinc(II) held by an anthracenyl spacer, and a flanking acceptor, bisarylporphyrin free-base ( Ar = -3,5-(t Bu )2 C 6 H 3), has been studied by means of absorption spectroscopy, "steady state and time-resolved fluorescence" and fs transient absorption spectroscopy, and density functional theory (DFT) in order to assess the effect of decoupling the chromophores' low energy MOs on the rate of the singlet, S1, energy transfer, k ET , compared to a recently reported work on a heavily coupled trimeric system, Trimer 1, [biphenylenebis(n-nonyl)porphyrin zinc(II)]-bisarylporphyrin free-base ( Ar = -3,5-(t Bu )2 C 6 H 3). The position of the 0–0 peaks of the absorption and fluorescence spectra of Trimer 2 indicates that these porphyrin units are respectively energy donor 1, donor 2, and acceptor. The DFT computations confirm that the MOs of the cofacial donor 1-donor 2 dyad are decoupled, but significant MO coupling between donor 2 and acceptor 1 is still present despite the strong dihedral angle between their respective average planes (77.5°: geometry optimization by DFT). The fs transient absorption spectra exhibit a clear S1–Sn fingerprint of the bisarylporphyrin zinc(II) chromophore and the kinetic trace exhibits a slow rise time of 87 ps, due to a S1 donor 1 → donor 2 ET. The transient species donor 2 and acceptor decay respectively in the short (~1.5) ns and 6 ns time scale.

Altered profile of basket cell afferent synapses in hyper‐excitable dentate gyrus revealed by optogenetic and two‐pathway stimulations

Cholecystokinin (CCK-) positive basket cells form a distinct class of inhibitory GABAergic interneurons, proposed to act as fine-tuning devices of hippocampal gamma-frequency (30-90 Hz) oscillations, which can convert into higher frequency seizure activity. Therefore, CCK-basket cells may play an important role in regulation of hyper-excitability and seizures in the hippocampus. In normal conditions, the endogenous excitability regulator neuropeptide Y (NPY) has been shown to modulate afferent inputs onto dentate gyrus CCK-basket cells, providing a possible novel mechanism for excitability control in the hippocampus. Using GAD65-GFP mice for CCK-basket cell identification, and whole-cell patch-clamp recordings, we explored whether the effect of NPY on afferent synapses to CCK-basket cells is modified in the hyper-excitable dentate gyrus. To induce a hyper-excitable state, recurrent seizures were evoked by electrical stimulation of the hippocampus using the well-characterized rapid kindling protocol. The frequency of spontaneous and miniature excitatory and inhibitory post-synaptic currents recorded in CCK-basket cells was decreased by NPY. The excitatory post-synaptic currents evoked in CCK-basket cells by optogenetic activation of principal neurons were also decreased in amplitude. Interestingly, we observed an increased proportion of spontaneous inhibitory post-synaptic currents with slower rise times, indicating that NPY may inhibit gamma aminobutyric acid release preferentially in peri-somatic synapses. These findings indicate that increased levels and release of NPY observed after seizures can modulate afferent inputs to CCK-basket cells, and therefore alter their impact on the oscillatory network activity and excitability in the hippocampus.

Abstract 5387: Exploring effects of nanosecond and microsecond pulsed electric field ablation in a rat orthotopic hepatocellular carcinoma model

Abstract Hepatocellular carcinoma (HCC) is the third most common cancer in the world. A diagnosis of HCC carries a guarded prognosis, especially when metastasis is documented. Reliable, curative treatment for very early, early, and intermediate stages is still an inconsistent ideal. Nanosecond (ns), or microsecond (μs) pulsed electric fields (PEFs) that produce irreversible electroporation (IRE) could be used to ablate local tumor masses through minimal invasive or open surgery delivery. Both nsPEFs and μsPEFs induce permanent permeabilization of cells resulting in cell death without any thermal effect. NsPEFs can also impact intracellular organelles and induce apoptosis. Previous studies indicated under certain condition, there was a protective tumor response following nsPEFs treatment in mouse subcutaneous HCC model. In these studies, we specifically test the efficacy of nsPEFs and μsPEFs in an orthotopic HCC rat tumor model. N1S1 cells stably transfected with luciferase were used to generate tumors. Orthotropic insertion of transfected N1S1 HCC cells in Sprague Dawley rat livers was used as the model. Different pulses numbers, rise times and pulse durations were applied. The effect on treated tumor tissue suggests that a single treatment of 500 or 1000 100ns pulses with either slow (50ns) or fast (10ns) rise times and 50 kV/cm can eliminate HCC tumors. Continuous delivery of eighty 100μs microsecond pulses at 2.5 kV/cm dramatically reverses lesion enlargement. 1000 pulses of 100 ns PEFs with fast rise times provoke an innate immune response. Immune responses in this orthotopic HCC model are still under investigation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5387. doi:1538-7445.AM2012-5387

Digital pulse height correction in HgI2 γ-ray detectors

We report on the application of digital pulse processing algorithms to improve the spectroscopic performance of a 1.2 mm thick planar HgI2 γ-ray detector. We have used offline processing of pulses which were recorded using a high resolution waveform digitizer. The recovery processes include long duration shaping to avoid ballistic deficit in the case of slow pulses, and the application of biparametric correction techniques to compensate for charge loss. Pulses of duration as long as 100 μs were recorded to facilitate long duration shaping. Two different pulse processing algorithms, viz. semi-Gaussian and moving window deconvolution, were applied and their performance was compared. The application of long duration shaping and digital charge-loss correction improved the energy resolution at 662 keV by more than 20% and the peak to background ratio by a factor of two. The resolution and the peak to background ratio were further seen to improve drastically upon rejection of counts with very slow rise-time. A 2.6% energy resolution at 662 keV with 14:1 peak to background ratio was obtained.

Characteristics of Protein-Polymer Nanobiocomposite Films for Protein Devices

Here we report the characteristics of protein-polymer nanobiocomposite films and their solid state devices. The protein-polymer nanobiocomposite films (thickness = approximately 125 nm) were prepared by spin-coating the solution of cytochrome c (cyt c) and poly(vinyl alcohol) (PVA) (cyt c:PVA = 3:1 by weight). To understand the characteristics of the cyt c-PVA films (nanolayers), we employed the optical absorption and surface morphology measurement and then fabricated planar diode-type solid state devices. The optical absorption measurement showed that the heme part of cyt c in the cyt c-PVA nanolayer was well kept after the coating process, while the crack-like surface was found from the atomic force microscopy measurement. The planar device showed an ohmic type dark current, but the current gradually increased as the incident light intensity increased. In particular, the (photo) current was strongly dependent upon the voltage, which was assigned to the insulating role of cyt c surrounding groups and PVA. This trend was supported by the slow rise and decay time via photo-switching experiment.

The α1K276E Startle Disease Mutation Reveals Multiple Intermediate States in the Gating of Glycine Receptors

Loss-of-function mutations in human glycine receptors cause hyperekplexia, a rare inherited disease associated with an exaggerated startle response. We have studied a human disease mutation in the M2-M3 loop of the glycine receptor α1 subunit (K276E) using direct fitting of mechanisms to single-channel recordings with the program HJCFIT. Whole-cell recordings from HEK293 cells showed the mutation reduced the receptor glycine sensitivity. In single-channel recordings, rat homomeric α1 K276E receptors were barely active, even at 200 mM glycine. Coexpression of the β subunit partially rescued channel function. Heteromeric mutant channels opened in brief bursts at 300 μM glycine (a concentration that is near-maximal for wild type) and reached a maximum one-channel open probability of about 45% at 100 mm glycine (compared to 96% for wild type). Distributions of apparent open times contained more than one component in high glycine and, therefore, could not be described by mechanisms with only one fully liganded open state. Fits to the data were much better with mechanisms in which opening can also occur from more than one fully liganded intermediate (e.g., "primed" models). Brief pulses of glycine (∼3 ms, 30 mM) applied to mutant channels in outside-out patches activated currents with a slower rise time (1.5 ms) than those of wild-type channels (0.2 ms) and a much faster decay. These features were predicted reasonably well by the mechanisms obtained from fitting single-channel data. Our results show that, by slowing and impairing channel gating, the K276E mutation facilitates the detection of closed reaction intermediates in the activation pathway of glycine channels.

Point-to-plane pulsed discharge initiated flame structure modification in propane–air flame

The effect of a point-to-plane pulsed discharge on a propane/air flame has been investigated by phase-locked simultaneous measurements of the change in gas temperature and OH planar laser-induced-fluorescence (PLIF). Phase-locked simultaneous measurement of gas temperature through spontaneous Raman scattering and OH PLIF with the variation of pulsed plasma energy and plasma generation location with respect to the flame holder and flame reaction zone have been performed. A fast rise time (15 ns) and a slower rise time (150 ns) high voltage pulsers are used to produce OH radical densities 50% greater than the ambient flame produced OH radicals in both lean and rich premixed flames. The excess OH radical densities were found to decay to the 50% level with time constants greater than 100 µs in the burnt gas regions with gas temperatures greater than 1000 K. The flame perturbation was dependent on the pulse repetition rates as well as on the pulse rise time for similar energy deposition per pulse. A laser photo-deflection measurement of acoustic pressure pulse generation by the pulsed discharge suggests that flame perturbation by the downstream plasma is caused mostly by flow perturbation.

Sound-evoked network calcium transients in mouse auditory cortexin vivo

Population calcium signals generated by the action potential activity of local clusters of neurons have been recorded in the auditory cortex of mice using an optical fibre-based approach. These network calcium transients (NCaTs) occurred spontaneously as well as in response to sound stimulation. Two-photon calcium imaging experiments suggest that neurons and neuropil contribute about equally to the NCaT. Sound-evoked calcium signals had two components: an early, fast increase in calcium concentration, which corresponds to the short-latency spiking responses observed in electrophysiological experiments, and a late, slow calcium transient which lasted for at least 1 s. The slow calcium transients evoked by sound were essentially identical to spontaneous NCaTs. Their sizes were dependent on the spontaneous activity level at sound onset, suggesting that spontaneous and sensory-evoked NCaTs excluded each other. When using pure tones as stimulus, the early evoked calcium transients were more narrowly tuned than the slow NCaTs. The slow NCaTs were correlated with global ‘up states' recorded with epidural potentials, and sound presented during an epidural ‘down state' triggered a calcium transient that was associated with an epidural ‘up state'. Essentially indistinguishable calcium transients were evoked by optogenetic activation of local clusters of layer 5 pyramidal neurons in the auditory cortex, indicating that these neurons play an important role in the generation of the calcium signal. Taken together, our results identify sound-evoked slow NCaTs as an integral component of neuronal signalling in the mouse auditory cortex, reflecting the prolonged neuronal activity of local clusters of neurons that can be activated even by brief stimuli.

Age‐dependent remodelling of inhibitory synapses onto hippocampal CA1 oriens‐lacunosum moleculare interneurons

Stratum oriens-lacunosum moleculare interneurons (O-LM INs) represent the major element of the hippocampal feedback inhibitory circuit, which provides inhibition to the distal dendritic sites of CA1 pyramidal neurons. Although the intrinsic conductance profile and the properties of glutamatergic transmission to O-LM INs have become a subject of intense investigation, far less is known about the properties of the inhibitory synapses formed onto these cells. Here, we used whole-cell patch-clamp recordings in acute mouse hippocampal slices to study the properties and plasticity of GABAergic inhibitory synapses onto O-LM INs. Surprisingly, we found that the kinetics of inhibitory postsynaptic currents (IPSCs) were slower in mature synapses (P26-40) due to the synaptic incorporation of the α5 subunit of the GABA(A) receptor (a5-GABA(A)R). Moreover, this age-dependent synaptic expression of a5-GABA(A)Rs was directly associated with the emergence of long-term potentiation at IN inhibitory synapses. Finally, the slower time course of IPSCs observed in O-LM INs of mature animals had a profound effect on IN excitability by significantly delaying its spike firing. Our data suggest that GABAergic synapses onto O-LM INs undergo significant modifications during postnatal maturation. The developmental switch in IPSC properties and plasticity is controlled by the synaptic incorporation of the a5-GABA(A)R subunit and may represent a potential mechanism for the age-dependent modifications in the inhibitory control of the hippocampal feedback inhibitory circuit.

Reconstructing neuronal inputs from voltage recordings: application in the auditory system

There are neurons in the auditory midbrain of frog H. versicolor that respond selectively to slowly rising auditory stimuli (behaviorally relevant for this frog). We investigate possible mechanisms that underlie this rise-time selectivity. In particular, we want to find out whether the rise-time selectivity arises in midbrain or is inherited from lower level structures. To extract the relevant data from the existing recordings one needs to solve the inverse problem of computing the afferent inputs to the neuron. Reconstructing stimulus-evoked temporally-varying input to a neuron in vivo is a challenge. The existing model-based method allows us to resolve two synaptic conductances corresponding to two distinct reversal potentials. We present a new approach enabling the reconstruction of three input conductances. Our method is based on treating synaptic conductances and membrane voltage as random variables, generalizing the model to a stochastic differential equation, and deriving equations for both first and second moments. We apply reconstruction to simulated data and discuss applicability to experimental data. Based on conductance reconstructions, we present three computational models of possible slow rise-time selectivity mechanisms: local inactivation of inhibition, fast-rise-time sensitive inhibition and interval counting with adaptation. We also discuss the evidence from in vivo recordings in support of the models. Finally, we show model predictions, which allow to distinguish between the proposed mechanisms as more data become available.

Distinct functions of kainate receptors in the brain are determined by the auxiliary subunit Neto1

Ionotropic glutamate receptors principally mediate fast excitatory transmission in the brain. Among the three classes of ionotropic glutamate receptors, kainate receptors (KARs) display a categorical brain distribution, which has been historically defined by 3H-radiolabeled kainate binding. Compared with recombinant KARs expressed in heterologous cells, synaptic KARs exhibit characteristically slow rise-time and decay kinetics. However, the mechanisms responsible for these unique KAR properties remain unclear. Here we found that both the distinct high affinity biding pattern in the mouse brain and the channel properties of native KARs are determined by the KAR auxiliary subunit Neto1. Through modulation of agonist binding affinity and off-kinetics of KARs, but not trafficking of KARs, Neto1 determines both KAR high affinity binding pattern and the distinctively slow kinetics of postsynaptic KARs. By regulating KAR-EPSC kinetics, Neto1 can control synaptic temporal summation, spike generation and fidelity.

Effects of morpheme type on Deg Xinag ejectives.

Stem‐initially in Deg Xinag (Athabaskan), there is a three‐way contrast between voiceless unaspirated, voiceless aspirated and ejective stops, and affricates at a variety of places of articulation, including alveolar with lateral release. In prefixes, there is a single lateral ejective affricate, /tɫ’/, historically from *h‐s‐ɫ‐, [Leer (2000)]. In a previous acoustic study [Hargus (2008)], the prefixal lateral affricate had relatively few of the characteristics of ejectives in stems (VOT, f0 perturbation, jitter perturbation, rise time, and normalized fricative energy measures). However, it was not clear whether the reduced number of ejective characteristics of the prefix was a reflection of its evolution from non‐ejective *h‐s‐ɫ‐ or an expected effect of morpheme type (presumably via concomittant differences in stress). In a follow‐up study, the ejective characteristics of a prefixal retroflex ejective affricate (first person plural subject) are compared to those of retroflex ejective affricates in stems. Retroflex ejectives in stems have significantly longer VOT, slower rise time, lesser normalized fricative energy, and/or differences in normalized f0 relative to the prefix. This suggests that the variability of the prefixal lateral ejective affricate is an expected effect of morpheme type, and not a stage in the evolution to ejective. [(Work supported by NSF.)]

The K276E Startle Disease Mutation in the Glycine Receptor: Effects on Channel Gating

Glycine receptors mediate inhibitory synaptic currents that are essential for motor control. Loss-of-function mutations in human glycine receptors cause hyperekplexia, a rare inherited disease associated with an exaggerated startle response. We have studied a human disease mutation in the M2-M3 loop of the glycine receptor alpha1 subunit (K276E) using direct fitting of mechanisms to single-channel recordings with the program HJCFIT. Whole-cell recordings from HEK 293 cells suggested that mutant receptors, both homomeric alpha1 and heteromeric alpha1beta, were much less sensitive to glycine than their wild-type counterparts. Single-channel recordings showed that homomeric alpha1 K276E receptors are barely active, even at 200 mM glycine. In contrast, heteromeric channels show brief bursts of openings at 300 µM glycine (∼EC95 for wild-type) and reach a maximum one-channel open probability of about 45% at 100 mM glycine (cf. 96% for wild-type). Distributions of apparent open times contained more than one component at high concentrations, and so mechanisms that allowed only one open state in the fully liganded receptor were extremely poor at describing the observed single-channel behaviour. Instead, mechanisms in which opening can also occur from more than one fully liganded intermediate state (e.g. ‘primed’ models) gave a much better description of the data. Brief pulses of glycine (0.5-2 ms, 30 mM) applied to outside-out patches activated currents with a slower rise time (1.3 ms) than wild-type channels (0.2 ms), and a much faster decay. These features of the macroscopic currents were well predicted by the mechanisms obtained from fitting single-channel data. Our results show that the mutation K276E impairs the gating of the receptor, demonstrate an unforeseen role for the beta subunit in promoting channel opening, and give further evidence of reaction intermediates in the activation pathway.

Ablation of Glutamate Receptor GluRδ2 in Adult Purkinje Cells Causes Multiple Innervation of Climbing Fibers by Inducing Aberrant Invasion to Parallel Fiber Innervation Territory

Glutamate receptor GluRδ2 is exclusively expressed in Purkinje cells (PCs) from early development and plays key roles in parallel fiber (PF) synapse formation, elimination of surplus climbing fibers (CFs), long-term depression, motor coordination, and motor learning. To address its role in adulthood, we previously developed a mouse model of drug-induced GluRδ2 ablation in adult PCs (Takeuchi et al., 2005). In that study, we demonstrated an essential role to maintain the connectivity of PF-PC synapses, based on the observation that both mismatching of presynaptic and postsynaptic specializations and disconnection of PF-PC synapses are progressively increased after GluRδ2 ablation. Here, we pursued its role for CF wiring in adult cerebellum. In parallel with the disconnection of PF-PC synapses, ascending CF branches exhibited distal extension to innervate distal dendrites of the target and neighboring PCs. Furthermore, transverse CF branches, a short motile collateral rarely forming synapses in wild-type animals, displayed aberrant mediolateral extension to innervate distal dendrites of neighboring and remote PCs. Consequently, many PCs were wired by single main CF and other surplus CFs innervating a small part of distal dendrites. Electrophysiological recording further revealed that surplus CF-EPSCs characterized with slow rise time and small amplitude emerged after GluRδ2 ablation, and increased progressively both in number and amplitude. Therefore, GluRδ2 is essential for maintaining CF monoinnervation in adult cerebellum by suppressing aberrant invasion of CF branches to the territory of PF innervation. Thus, GluRδ2 fuels heterosynaptic competition and gives PFs the competitive advantages over CFs throughout the animal's life.

Ultrafast carrier dynamics in pristine and FeCl3-intercalated bilayer graphene

Ultrafast carrier dynamics of pristine bilayer graphene (BLG) and bilayer graphene intercalated with FeCl3 (FeCl3-G), were studied using time-resolved transient differential reflection (delta R/R). Compared to BLG, the FeCl3-G data showed an opposite sign of delta R/R, a slower rise time, and a single (instead of double) exponential relaxation. We attribute these differences in dynamics to the down-shifting of the Fermi level in FeCl3-G, as well as the formation of numerous horizontal bands arising from the d-orbitals of Fe. Our work shows that intercalation can dramatically change the electronic structure of graphene, and its associated carrier dynamics.