Single Exponential Time Constant Research Articles

Acid-base equilibria of axial ligand and peripheral pyridyl group with stepwise formation of nine species of aluminum (III) tera(4-pyridyl) porphyrin

As one of the promising molecular catalysts for the two-electron water oxidation initiated through one-electron oxidation of the catalyst, trans-diaqua-coordinated 5, 10, 15, 20-tetra(4-pyridyl)porphyrinatealuminum (III) (AlTPyP(OH2)2) was studied on the protolytic behavior of the axial ligating water molecules and the redox properties in aqueous methanol solution. The reversible stepwise protolytic reactions of the two axially ligated water molecules, as well as four peripheral pyridyl groups at the meso-position of AlTPyP, were characterized using UV–vis absorption spectroscopy, 1H NMR, fluorescence lifetime measurements, and cyclic voltammetry. The result showed that four-step protolytic equilibrium reactions of the axial ligands proceeded under the basic pH condition and those of meso-substituted pyridyl groups did also under the acidic pH condition. All the nine species are in equilibria in the ground states as characterized with each pKa value. On the other hand, they exist as individual species in the excited states having each single exponential decay time constant. Here we will report the first detailed study on the formation of nine species from a single molecule of AlTPyP through protolytic reactions in the pH range between 0.54 and 12.3.

New Tl2LaBr5: Ce3+ crystal scintillator for γ-rays detection

In this study we present our preliminary report on the scintillation properties of new Ce-doped Tl2LaBr5 single crystal. Two zones vertical Bridgman technique is used for the growth of this compound. Pure and Ce-doped samples showed maximum emission peaks at 435nm and 415nm, respectively. Best light yield of 43,000±4300ph/MeV with 6.3% (FWHM) energy resolution is obtained for 5% Ce-doped sample under γ-ray excitation. Single exponential decay time constant of 25ns is observed for 5% Ce doped sample. Effective Z-number is found to be 67, therefore efficient detection of X- and γ-ray will be possible. Preliminary results revealed that this compound will be an ideal candidate for the medical imaging techniques. Further investigations are under way for the determination of optimized conditions of this compound.

Saccadic and Postsaccadic Disconjugacy in Zebrafish Larvae Suggests Independent Eye Movement Control.

Spontaneous eye movements of zebrafish larvae in the dark consist of centrifugal saccades that move the eyes from a central to an eccentric position and postsaccadic centripetal drifts. In a previous study, we showed that the fitted single-exponential time constants of the postsaccadic drifts are longer in the temporal-to-nasal (T->N) direction than in the nasal-to-temporal (N->T) direction. In the present study, we further report that saccadic peak velocities are higher and saccadic amplitudes are larger in the N->T direction than in the T->N direction. We investigated the underlying mechanism of this ocular disconjugacy in the dark with a top-down approach. A mathematic ocular motor model, including an eye plant, a set of burst neurons and a velocity-to-position neural integrator (VPNI), was built to simulate the typical larval eye movements in the dark. The modeling parameters, such as VPNI time constants, neural impulse signals generated by the burst neurons and time constants of the eye plant, were iteratively adjusted to fit the average saccadic eye movement. These simulations suggest that four pools of burst neurons and four pools of VPNIs are needed to explain the disconjugate eye movements in our results. A premotor mechanism controls the synchronous timing of binocular saccades, but the pools of burst and integrator neurons in zebrafish larvae seem to be different (and maybe separate) for both eyes and horizontal directions, which leads to the observed ocular disconjugacies during saccades and postsaccadic drifts in the dark.

Ultrafast Interfacial Electron and Hole Transfer from CsPbBr3 Perovskite Quantum Dots.

Recently reported colloidal lead halide perovskite quantum dots (QDs) with tunable photoluminescence (PL) wavelengths covering the whole visible spectrum and exceptionally high PL quantum yields (QYs, 50-90%) constitute a new family of functional materials with potential applications in light-harvesting and -emitting devices. By transient absorption spectroscopy, we show that the high PL QYs (∼79%) can be attributed to negligible electron or hole trapping pathways in CsPbBr3 QDs: ∼94% of lowest excitonic states decayed with a single-exponential time constant of 4.5 ± 0.2 ns. Furthermore, excitons in CsPbBr3 QDs can be efficiently dissociated in the presence of electron or hole acceptors. The half-lives of electron transfer (ET) to benzoquinone and subsequent charge recombination are 65 ± 5 ps and 2.6 ± 0.4 ns, respectively. The half-lives for hole transfer (HT) to phenothiazine and the subsequent charge recombination are 49 ± 6 ps and 1.0 ± 0.2 ns, respectively. The lack of electron and hole traps and fast interfacial ET and HT rates are key properties that may enable the development of efficient lead halide perovskite QDs-based light-harvesting and -emitting devices.

Heteromeric Kv7.2/7.3 channels differentially regulate action potential initiation and conduction in neocortical myelinated axons.

Rapid energy-efficient signaling along vertebrate axons is achieved through intricate subcellular arrangements of voltage-gated ion channels and myelination. One recently appreciated example is the tight colocalization of K(v)7 potassium channels and voltage-gated sodium (Na(v)) channels in the axonal initial segment and nodes of Ranvier. The local biophysical properties of these K(v)7 channels and the functional impact of colocalization with Na(v) channels remain poorly understood. Here, we quantitatively examined K(v)7 channels in myelinated axons of rat neocortical pyramidal neurons using high-resolution confocal imaging and patch-clamp recording. K(v)7.2 and 7.3 immunoreactivity steeply increased within the distal two-thirds of the axon initial segment and was mirrored by the conductance density estimates, which increased from ~12 (proximal) to 150 pS μm(-2) (distal). The axonal initial segment and nodal M-currents were similar in voltage dependence and kinetics, carried by K(v)7.2/7.3 heterotetramers, 4% activated at the resting membrane potential and rapidly activated with single-exponential time constants (~15 ms at 28 mV). Experiments and computational modeling showed that while somatodendritic K(v)7 channels are strongly activated by the backpropagating action potential to attenuate the afterdepolarization and repetitive firing, axonal K(v)7 channels are minimally recruited by the forward-propagating action potential. Instead, in nodal domains K(v)7.2/7.3 channels were found to increase Na(v) channel availability and action potential amplitude by stabilizing the resting membrane potential. Thus, K(v)7 clustering near axonal Na(v) channels serves specific and context-dependent roles, both restraining initiation and enhancing conduction of the action potential.

Neurogliaform cells of amygdala: a source of slow phasic inhibition in the basolateral complex

Synaptic inhibition in the amygdala actively participates in processing emotional information. To improve the understanding of interneurons in amygdala networks it is necessary to characterize the GABAergic cell types, their connectivity and physiological roles. We used a mouse line expressing a green fluorescent protein (GFP) under the neuropeptide Y (NPY) promoter. Paired recordings between presynaptic NPY-GFP-expressing (+) cells and postsynaptic principal neurons (PNs) of the basolateral amygdala (BLA) were performed. The NPY-GFP+ neurons displayed small somata and short dendrites embedded in a cloud of highly arborized axon, suggesting a neurogliaform cell (NGFC) type. We discovered that a NPY-GFP+ cell evoked a GABA(A) receptor-mediated slow inhibitory postsynaptic current (IPSC) in a PN and an autaptic IPSC. The slow kinetics of these IPSCs was likely caused by the low concentration and spillover of extracellular GABA. We also report that NGFCs of the BLA fired action potentials phase-locked to hippocampal theta oscillations in anaesthetized rats. When this firing was re-played in NPY+-NGFCs in vitro, it evoked a transient depression of the IPSCs. Presynaptic GABA(B) receptors and functional depletion of synaptic vesicles determined this short-term plasticity. Synaptic contacts made by recorded NGFCs showed close appositions, and rarely identifiable classical synaptic structures. Thus, we report here a novel interneuron type of the amygdala that generates volume transmission of GABA. The peculiar functional mode of NGFCs makes them unique amongst all GABAergic cell types of the amygdala identified so far.

Molecular Beacon Probes of Oligonucleotides Photodamaged by Psoralen

Ultraviolet A (UVA)-irradiated 4'-hydroxymethyl-4,5',8-trimethyl psoralen (HMT) in the presence of a poly-dT(17) and dA(7) TTA(8) oligonucleotides produces HMT-dT(17) and HMT-dA(7) TTA(8) adducts in aqueous solution. In this article, we determine whether these HMT-dT(17) and HMT-dA(7) TTA(8) adducts can be detected with a molecular beacon (MB) probe. We measure the degree of damage in dT(17) and dA(7) TTA(8) solutions containing UVA-activated HMT via monitoring the decrease in MB fluorescence. Photoproduct formation is confirmed by MALDI-TOF MS (matrix-assisted laser desorption/ionization time-of-fight mass spectrometry measurements) and absorption spectroscopy. The MB fluorescence decreases upon UVA irradiation in the presence of HMT with a single-exponential time constants of 114.2 ± 6.5 min for HMT-dT(17) adducts and 677.8 ± 181.8 min for HMT-dA(7) TTA(8) adducts. Our results show that fluorescent MB probes are a selective, robust and accurate tool for detecting UVA-activated HMT-induced DNA damage.

New Developments in Superconducting Tunnel Junction X-Ray Spectrometers for Synchrotron Science

We are developing Ta-based superconducting tunnel junction (STJ) X-ray detectors for high-resolution soft X-ray spectroscopy at the synchrotron. For scaling to large detector arrays, we have also built a compact, low-cost, remote-controllable preamplifier with <3 eV electronic noise. Current Ta-STJs attain an energy resolution between 6.5 and 9 eV FWHM for energies up to ∼2 keV, and can be operated at rates up to ∼5,000 counts/s as long as the signals decay with a single exponential time constant.

Modification of carrier localization in basal‐plane stacking faults: The effect of Si‐doping in a‐plane GaN

AbstractThe optical properties of Si‐doped a‐plane GaN epilayers grown on r‐plane sapphire are studied. The low temperature emission is dominated by basal‐plane stacking fault (BSF) recombination throughout the investigated doping range (1 × 1017 to 5 × 1019 cm−3). From temperature dependent photoluminescence (PL) measurements in conjunction with PL excitation studies, the carrier localization energy within the BSF is inferred to decrease from 17 meV to a negligible level as the doping density increases from 1 × 1017 to 5 × 1018 cm−3. It is proposed that electrons, ionized from the Si‐donor atoms at the growth temperature, are able to transfer to the BSFs, where they progressively fill the available density of localized states. For doping levels in excess of 1 × 1018 cm−3, the luminescence linewidth broadens significantly and the luminescence transients decay with a single exponential time constant. This behaviour is attributed to the onset of band‐filling which causes a marked increase in the free electron density within the BSFs.

High-field Overhauser dynamic nuclear polarization in silicon below the metal–insulator transition

Single crystal silicon is an excellent system to explore dynamic nuclear polarization (DNP), as it exhibits a continuum of properties from metallic to insulating as a function of doping concentration and temperature. At low doping concentrations DNP has been observed to occur via the solid effect, while at very high-doping concentrations an Overhauser mechanism is responsible. Here we report the hyperpolarization of (29)Si in n-doped silicon crystals, with doping concentrations in the range of (1-3) × 10(17) cm(-3). In this regime exchange interactions between donors become extremely important. The sign of the enhancement in our experiments and its frequency dependence suggest that the (29)Si spins are directly polarized by donor electrons via an Overhauser mechanism within exchange-coupled donor clusters. The exchange interaction between donors only needs to be larger than the silicon hyperfine interaction (typically much smaller than the donor hyperfine coupling) to enable this Overhauser mechanism. Nuclear polarization enhancement is observed for a range of donor clusters in which the exchange energy is comparable to the donor hyperfine interaction. The DNP dynamics are characterized by a single exponential time constant that depends on the microwave power, indicating that the Overhauser mechanism is a rate-limiting step. Since only about 2% of the silicon nuclei are located within 1 Bohr radius of the donor electron, nuclear spin diffusion is important in transferring the polarization to all the spins. However, the spin-diffusion time is much shorter than the Overhauser time due to the relatively weak silicon hyperfine coupling strength. In a 2.35 T magnetic field at 1.1 K, we observed a DNP enhancement of 244 ± 84 resulting in a silicon polarization of 10.4 ± 3.4% following 2 h of microwave irradiation.

An Improved Cerulean Fluorescent Protein with Enhanced Brightness and Reduced Reversible Photoswitching

Cyan fluorescent proteins (CFPs), such as Cerulean, are widely used as donor fluorophores in Förster resonance energy transfer (FRET) experiments. Nonetheless, the most widely used variants suffer from drawbacks that include low quantum yields and unstable flurorescence. To improve the fluorescence properties of Cerulean, we used the X-ray structure to rationally target specific amino acids for optimization by site-directed mutagenesis. Optimization of residues in strands 7 and 8 of the β-barrel improved the quantum yield of Cerulean from 0.48 to 0.60. Further optimization by incorporating the wild-type T65S mutation in the chromophore improved the quantum yield to 0.87. This variant, mCerulean3, is 20% brighter and shows greatly reduced fluorescence photoswitching behavior compared to the recently described mTurquoise fluorescent protein in vitro and in living cells. The fluorescence lifetime of mCerulean3 also fits to a single exponential time constant, making mCerulean3 a suitable choice for fluorescence lifetime microscopy experiments. Furthermore, inclusion of mCerulean3 in a fusion protein with mVenus produced FRET ratios with less variance than mTurquoise-containing fusions in living cells. Thus, mCerulean3 is a bright, photostable cyan fluorescent protein which possesses several characteristics that are highly desirable for FRET experiments.

A Single Channel Description of an Inactivation Deficient Sodium Channel Mutant

A characteristic of many voltage-gated ion channels including sodium and potassium channels is that macroscopic currents elicited with a voltage step subsequently decline, or inactivate. Because entry into inactivated states tends to limit the activity of single sodium channels to a small number of openings, identifying the statistical properties of other less stable gating processes can be difficult. Here, we examined the single channel characteristics of a mutant skeletal muscle sodium channel (rat Nav1.4 L435W/L437C/A438W) that exhibits very little macroscopic inactivation (Wang et al., 2003) in response to voltage steps to −60 and −40 mV. Although open dwell time distributions could be more or less described by a single exponential component, both deviations from the single exponential fit and time constants from bi-exponential fits were consistent across patches, suggesting the existence of two open states with time constants of approximately 0.5-1 ms and 2-4 ms. Bursts of openings separated by closures of less than 2 ms were observed frequently, with the number of bursts in a given sweep increasing at more depolarized voltages. The number of opening within each burst was also voltage dependent, increasing from about 3 to 5 openings at −60 and −40 mV, respectively. These data will be presented with a preliminary kinetic description of sodium channel activity in the absence of inactivation. This work was funded by NIH grant GM084140.

Versatility of Trigger Factor Interactions with Ribosome-Nascent Chain Complexes

Trigger factor (TF) is the first molecular chaperone that interacts with nascent chains emerging from bacterial ribosomes. TF is a modular protein, consisting of an N-terminal ribosome binding domain, a PPIase domain, and a C-terminal domain, all of which participate in polypeptide binding. To directly monitor the interactions of TF with nascent polypeptide chains, TF variants were site-specifically labeled with an environmentally sensitive NBD fluorophore. We found a marked increase in TF-NBD fluorescence during translation of firefly luciferase (Luc) chains, which expose substantial regions of hydrophobicity, but not with nascent chains lacking extensive hydrophobic segments. TF remained associated with Luc nascent chains for 111 +/- 7 s, much longer than it remained bound to the ribosomes (t((1/2)) approximately 10-14 s). Thus, multiple TF molecules can bind per nascent chain during translation. The Escherichia coli cytosolic proteome was classified into predicted weak and strong interactors for TF, based on the occurrence of continuous hydrophobic segments in the primary sequence. The residence time of TF on the nascent chain generally correlated with the presence of hydrophobic regions and the capacity of nascent chains to bury hydrophobicity. Interestingly, TF bound the signal sequence of a secretory protein, pOmpA, but not the hydrophobic signal anchor sequence of the inner membrane protein FtsQ. On the other hand, proteins lacking linear hydrophobic segments also recruited TF, suggesting that TF can recognize hydrophobic surface features discontinuous in sequence. Moreover, TF retained significant affinity for the folded domain of the positively charged, ribosomal protein S7, indicative of an alternative mode of TF action. Thus, unlike other chaperones, TF appears to employ multiple mechanisms to interact with a wide range of substrate proteins.

Ortho-para transition of interstitialH2andD2in Si

The ortho-para transition of interstitial ${\text{H}}_{2}$ in Si has been observed in a recent Raman study [M. Hiller et al., Phys. Rev. Lett. 98, 055504 (2007); M. Hiller et al., Phys. Rev. Lett. 99, 209901 (2007)]. In order to address issues that are difficult to study by Raman spectroscopy, we have performed IR-absorption experiments to further investigate this ortho-para transition. We find that when a Si sample containing ${\text{H}}_{2}$ that has been equilibrated at room temperature is subsequently held at 77 K, the $3618.4\text{ }{\text{cm}}^{\ensuremath{-}1}$ IR line assigned to ortho-${\text{H}}_{2}$ is reduced in intensity with a single exponential time constant of 229 h as the system relaxes to the para-${\text{H}}_{2}$ state. The previous Raman data had been analyzed by a model in which the ortho-para transition is caused by an interaction with the nuclear magnetic moment of $^{29}\text{S}\text{i}$ (4.7% abundant) that is present in natural Si. However, in order for every ${\text{H}}_{2}$ molecule to interact with a $^{29}\text{S}\text{i}$, at least six shells of Si neighbors must be considered. Because of the strong dependence of the transition rate on the $^{29}\text{S}{\text{i-H}}_{2}$ distance, the kinetics of such a transition should be highly nonexponential. Thus, our IR data are not consistent with this mechanism. We have also investigated the para-ortho transition of interstitial ${\text{D}}_{2}$ and have found decay rates similar to those for the ortho-para transition of ${\text{H}}_{2}$. We have considered two possible mechanisms for this transition: ${\text{H}}_{2}$ or ${\text{D}}_{2}$ tunneling, which would allow the molecule to sample tetrahedral sites adjacent to $^{29}\text{S}\text{i}$, and spin-rotation coupling that can act while the molecule is dynamically off-center. Consideration of a possible tunneling rate by theory and the similarity of the ${\text{H}}_{2}$ and ${\text{D}}_{2}$ results seem to rule out the tunneling mechanism but not the spin-rotation mechanism. These results clearly show that the mechanism of the ortho-para transition for ${\text{H}}_{2}$ and ${\text{D}}_{2}$ in Si remains an experimental and a theoretical challenge.

Skeletal Myotubes from Adult Mice in a Cardiac Environment

The present study was designed to evaluate the functional impact of cardiac environment on the differentiation of skeletal myoblasts and Ca2+ signaling in electrically stimulated myotubes. Adult mice FDB muscle myoblasts were cultured alone or in co-culture with cardiomyocytes contractile monolayers. After 5 days of differentiation in a cardiac environment, the number of skeletal myogenic cells and myotubes were fourfold higher than control. Cardiac environment changed parameters of myotube calcium transients (Fluo-4AM) associated with Ca2+ removal mechanisms without affecting parameters related to Ca2+ release. The values (mean ± sem) of half width (HW) and single exponential decay time constant (τ), obtained from mono-cultured myotubes (MMy) were 209.42±34.72 ms and 415.16±55.12 ms (n=9), while for co-cultured myotubes (CMy) they were 554.94±67.38 ms and 1340.17±330.73 ms (n=9), respectively. HW and τ from neighboring cardiomyocytes (CM) were 274.78±50.17 ms and 387.72±47.98 ms (n=5). The transient rise time (RT) values for MMy and CMy were 33.46±8.79 (n=9) and 29.66±8.97 ms (n=9) while the amplitude values (ΔF/F) for the two cases were 0.94±0.09 (n=9) and 1.00±0.07 (n=9), respectively. In the absence of external Ca2+ (0.5mM EGTA) the parameters associated with Ca2+ removal were not affected, while those related to release mechanisms were as follows: RT = 105.38±24.93 ms (n=5) and 80.87±18.28 ms (n=5) for MMy and CMy respectively, and ΔF/F=0.56±0.11 (n=5) and 0.71±0.12 (n=5) for MMy and CMy, respectively. Thus, transient sensitivity to extracellular Ca2+ was not affected by coculture since both CMy and MMy were similarly modified by exposure to 0Ca2+. We conclude that at an early coculture stage, a cardiac environment facilitates skeletal muscle differentiation without affecting functional attributes characteristic of skeletal muscle, with the exception of a selective effect on Ca2+ removal parameters.

Short-Term Depression at the Reciprocal Synapses between a Retinal Bipolar Cell Terminal and Amacrine Cells

Visual adaptation is thought to occur partly at retinal synapses that are subject to plastic changes. However, the locus and properties of this plasticity are not well known. Here, we studied short-term plasticity at the reciprocal synapse between bipolar cell terminals and amacrine cells in goldfish retinal slices. Depolarization of a single bipolar cell terminal for 100 ms triggers the release of glutamate onto amacrine cell processes, which in turn leads to GABAergic feedback from amacrine cells onto the same terminal. We find that this release of GABA undergoes paired-pulse depression (PPD) that recovers in <1 min (single exponential time constant, tau approximately = 12 s). This disynaptic PPD is independent of mGluR-mediated plasticity and depletion of glutamatergic synaptic vesicle pools, because exocytosis assayed via capacitance jumps (deltaC(m)) recovered completely after 10 s (tau approximately = 2 s). Fast application of GABA (10 mM) onto outside-out patches excised from bipolar cell terminals showed that the recovery of GABA(A) receptors from desensitization depends on the duration of the application [fast recovery (<2 s) for short applications; slow (tau approximately = 12 s) for prolonged applications]. We thus blocked GABA(A) receptors and retested the GABAergic response mediated by nondesensitizing GABA(C) receptors to two rapid glutamate puffs onto the bipolar cell terminal. These responses consistently displayed PPD. Furthermore, blocking AMPA-receptor desensitization with cyclothiazide, or evoking GABA release with NMDA receptors, did not reduce PPD. We thus suggest that depletion of synaptic vesicle pools in GABAergic amacrine cells is a major contributor to PPD.

Sprint performance-duration relationships are set by the fractional duration of external force application

We hypothesized that the maximum mechanical power outputs that can be maintained during all-out sprint cycling efforts lasting from a few seconds to several minutes can be accurately estimated from a single exponential time constant (k(cycle)) and two measurements on individual cyclists: the peak 3-s power output (P(mech max)) and the maximum mechanical power output that can be supported aerobically (P(aer)). Tests were conducted on seven subjects, four males and three females, on a stationary cycle ergometer at a pedal frequency of 100 rpm. Peak mechanical power output (P(mech max)) was the highest mean power output attained during a 3-s burst; the maximum power output supported aerobically (P(aer)) was determined from rates of oxygen uptake measured during a progressive, discontinuous cycling test to failure. Individual power output-duration relationships were determined from 13 to 16 all-out constant load sprints lasting from 5 to 350 s. In accordance with the above hypothesis, the power outputs measured during all-out sprinting efforts were estimated to within an average of 34 W or 6.6% from P(mech max), P(aer), and a single exponential constant (k(cycle) = 0.026 s(-1)) across a sixfold range of power outputs and a 70-fold range of sprint trial durations (R2 = 0.96 vs. identity, n = 105; range: 180 to 1,136 W). Duration-dependent decrements in sprint cycling power outputs were two times greater than those previously identified for sprint running speed (k(run) = 0.013 s(-1)). When related to the respective times of pedal and ground force application rather than total sprint time, decrements in sprint cycling and running performance followed the same time course (k = 0.054 s(-1)). We conclude that the duration-dependent decrements in sprinting performance are set by the fractional duration of the relevant muscular contractions.

Presynaptic Na+Channels: Locus, Development, and Recovery from Inactivation at a High-Fidelity Synapse

Na+ channel recovery from inactivation limits the maximal rate of neuronal firing. However, the properties of presynaptic Na+ channels are not well established because of the small size of most CNS boutons. Here we study the Na+ currents of the rat calyx of Held terminal and compare them with those of postsynaptic cells. We find that presynaptic Na+ currents recover from inactivation with a fast, single-exponential time constant (24 degrees C, tau of 1.4-1.8 ms; 35 degrees C, tau of 0.5 ms), and their inactivation rate accelerates twofold during development, which may contribute to the shortening of the action potential as the terminal matures. In contrast, recordings from postsynaptic cells in brainstem slices, and acutely dissociated, reveal that their Na+ currents recover from inactivation with a double-exponential time course (tau(fast) of 1.2-1.6 ms; tau(slow) of 80-125 ms; 24 degrees C). Surprisingly, confocal immunofluorescence revealed that Na+ channels are mostly absent from the calyx terminal but are instead highly concentrated in an unusually long (approximately 20-40 microm) unmyelinated axonal heminode. Outside-out patch recordings confirmed this segregation. Expression of Na(v)1.6 alpha-subunit increased during development, whereas the Na(v)1.2alpha-subunit was not present. Serial EM reconstructions also revealed a long pre-calyx heminode, and biophysical modeling showed that exclusion of Na+ channels from the calyx terminal produces an action potential waveform with a shorter half-width. We propose that the high density and polarized locus of Na+ channels on a long heminode are critical design features that allow the mature calyx of Held terminal to fire reliably at frequencies near 1 kHz.

Molecular Beacon Probes of Photodamage in Thymine and Uracil Oligonucleotides¶

Abstract Molecular beacons (MB) are becoming more common as sequence-selective detectors of nucleic acids. Although they can easily detect single-base mismatches, they have never been used to directly detect DNA or RNA damage. To measure the degree of ultraviolet (UV) light damage in oligonucleotides, we report a novel MB approach for general detection of photoproducts in UV-irradiated rU17 and dT17 oligonucleotides. With monochromatic UV light irradiation at ca 280 nm under anoxic conditions, the oligonucleotide absorption decays with a single-exponential time constant of 123 ± 1 min for rU17 and with double-exponential time constants of 78 ± 0.5 min (99%) and 180 ± 5 min (0.05%) for dT17 oligonucleotides. Under the same conditions, the MB fluorescence decays more quickly, with single-exponential time constants of 19 ± 2 and 26 ± 3 min for rU17 and dT17, respectively. Similar kinetics were observed with broadband UV light irradiation of oligonucleotides. The differences in the UV damage kinetics of dT17 ...

Molecular Beacon Probes of Photodamage in Thymine and Uracil Oligonucleotides

Molecular beacons (MB) are becoming more common as sequence-selective detectors of nucleic acids. Although they can easily detect single-base mismatches, they have never been used to directly detect DNA or RNA damage. To measure the degree of ultraviolet (UV) light damage in oligonucleotides, we report a novel MB approach for general detection of photoproducts in UV-irradiated rU17 and dT17 oligonucleotides. With monochromatic UV light irradiation at ca 280 nm under anoxic conditions, the oligonucleotide absorption decays with a single-exponential time constant of 123+/-1 min for rU17 and with double-exponential time constants of 78+/-0.5 min (99%) and 180+/-5 min (0.05%) for dT17 oligonucleotides. Under the same conditions, the MB fluorescence decays more quickly, with single-exponential time constants of 19+/-2 and 26+/-3 min for rU17 and dT17, respectively. Similar kinetics were observed with broadband UV light irradiation of oligonucleotides. The differences in the UV damage kinetics of dT17 and rU17 and their detection by absorption and fluorescence techniques will be discussed in the context of differential instabilities introduced in the nucleic acid-MB duplex by the different photoproducts formed.