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PubMed2006

Developmental regulation of whole cell capacitance and membrane current in identified interneurons in C. elegans.

Postembryonic developmental changes in electrophysiological properties of the AIY interneuron class were investigated using whole cell voltage clamp. AIY interneurons displayed an increase in cell capacitance during larval development, whereas steady-state current amplitude did not increase. The time course of the outward membrane current, carried at least in part by K+ ions, matured, from a slowly activating, sustained current to a rapidly activating, decaying current. We also investigated how the development of capacitance and outward current was altered by loss-of-function mutations in genes expressed in AIY. One such gene, the LIM homeobox gene ttx-3, is known to be involved in the specification of the AIY neuronal subtype. In ttx-3 mutants, capacitance and outward current matured precociously. In mutants of the gene wrk-1, an immunoglobulin superfamily (IgSF) member whose expression is regulated by ttx-3, capacitance matured normally, whereas outward current matured precociously. We conclude that AIY interneurons contain distinct pathways for regulating capacitance and membrane current.

Aging↗

PubMed2006

Ultrasonic bath depth control and regulation in single cell recordings.

Control of the bath depth is critical in many applications of the patch-clamp technique, particularly when the capacitance of cells is determined to assess secretion or transmitter release or in studies of ion currents sensitive to small changes in the hydrostatic pressure. We describe an inexpensive technique for tight control of the bath depth with the aid of a commercially available ultrasound sensor. The sensor continuously determines changes in the distance to the bath surface with a resolution of about 10 mum. The signal from the sensor is digitized in a microcontroller card and used to send on or off signals at 100 Hz to a peristaltic pump that removes volume from the bath. The inflow into the bath can be realized in a versatile way. The capacitance of Sylgard-coated patch-clamp glass electrodes, demonstrated to be extremely sensitive to small changes in the area moistened by bath solution, is constant within the noise level of +/-3 fF when immersed into a depth-controlled bath, even during exchange of the bath medium. Thus, when small changes in the cell capacitance are measured in patch-clamp experiments, errors due to alterations in the pipette capacitance caused by bath depth fluctuations are eliminated.

Animals↗

PubMed1997

Structure-function relations of variant and fragment nisins studied with model membrane systems.

Nisin, a 34 residue lantibiotic produced by strains of Lactococcus lactis subsp. lactis, exerts antimicrobial activity against Gram-positive bacteria at the cytoplasmic membrane. The structural aspects of nisin which facilitate membrane interaction and permeabilization have been investigated in planar lipid bilayers and liposomes with proteolytic fragments and site-directed variants. N-Terminal nisin fragments N1-12 and N1-20 had little effect on phospholipid mobility, on macroscopic electrical conductance, or on calcein release from liposomes. By contrast, the I30W nisin A variant induced a time-dependent reduction in lipid mobility, indicative of nisin-membrane surface interactions, as well as a decline in membrane capacitance, rise in conductance, and calcein release from liposomes. In these respects I30W nisin A is similar to native nisin. Charge substitutions were also engineered to generate K12L and H27K nisin A variants, both of which were similar to I30W nisin A with respect to an overall reduction in phospholipid mobility. While the K12L nisin A variant elicited a higher increase in membrane capacitance and electrical conductance than I30W nisin A, the H27K nisin A variant elicited weaker effects. These results point to a substantial role for intramembrane charged residues in controlling ion flow through nisin-doped membranes. Native nisin and variants elicit an enhanced release of calcein from liposomes composed of the negatively-charged phospholipids cardiolipin and phosphatidylserine, compared with phospholipid bearing no net charge, suggesting that an electrostatic attraction encourages the initial nisin-membrane association. The results are discussed in the context of other recently proposed models for nisin action.

Amino Acid Sequence↗

PubMed2003

Membrane activity of biotechnological peptide drugs.

Charged Langmuir-Blodgett monolayers deposited at an immobilised liquid-liquid interface have been used as a simple model for a biological membrane to investigate the membrane activity of biotechnological oligopeptide drugs.

Biotechnology↗

PubMed2001

Membrane capacitance measurement using patch clamp with integrated self-balancing lock-in amplifier.

A lock-in amplifier was incorporated directly into the resistance and capacitance compensation circuitry of a patch clamp set-up, to allow exocytosis to be monitored in the whole-cell mode by measuring changes in cell membrane capacitance. The integration of these two systems enabled us to provide a novel operating mode, which we term the "track-in" mode, where the output signals from the lock-in amplifier are used to make compensating electronic adjustments of the resistance and capacitance control settings. The lock-in amplifier outputs remain near zero, and the control voltages generated by the feedback circuits provide linear and calibrated resistance and capacitance measurements. Results obtained from model cells and mouse inner hair cells show that this is achieved without loss of sensitivity or of significant time resolution compared with the conventional lock-in amplifier technique, whereas the sensitivity to switching phase errors is effectively eliminated. An automatic phase tracking system using a low-frequency dithering of the resistance control setting can nevertheless be used to preserve the correct switching phase if required. The track-in approach has considerable advantages over software implementations in terms of economy and convenience, since the resistance and capacitance signals can be recorded directly on any general-purpose data-acquisition system.

Animals↗

PubMedDate not supplied

A majority-logic nanodevice using a balanced pair of single-electron boxes.

This paper describes a majority-logic gate device that will be useful in developing single-electron integrated circuits. The gate device consists of two identical single-electron boxes combined to form a balanced pair. It accepts three inputs and produces a majority-logic output by using imbalances caused by the input signals; it produces a 1 output if two or three inputs are 1, and a 0 output if two or three inputs are 0. We combine these gate devices into two subsystems, a shift register and an adder, and demonstrate their operation by computer simulation. We also propose a method of fabricating the unit element of the gate device, a minute dot with four coupling arms. We demonstrate by experiments that it is possible to arrange these unit elements on a GaAs substrate, in a self-organizing manner, by means of a process technology that is based on selective-area metalorganic vapor-phase epitaxy.

Aluminum↗

PubMed2005

On the charge regulation of proteins.

It is known that the overall charge of a protein can change as the molecule approaches a charged object like another protein or a cell membrane. We have formalized this mechanism using a statistical mechanical framework and show how this rather overlooked interaction increases the attraction between protein molecules. From the theory, we can identify a unique property, the protein charge capacitance, that contains all information needed to describe the charge regulation mechanism. The capacitance can be obtained from experiment or theory and is a function of pH, salt concentration, and the number of titrating residues. For a range of different protein molecules, we calculate the capacitance and demonstrate how it can be used to quantify the charge regulation interaction. With minimal effort, the derived formulas can be used to improve existing models by including a charge regulation term. Good agreement is found between theory, simulations, and experimental data.

Electric Capacitance↗

PubMed2003

Monitoring DNA hybridization on alkyl modified silicon surface through capacitance measurement.

Single strand oligodeoxynucleotide is attached to the alkyl modified silicon surface through a peptide bond. The oligodeoxynucleotide-modified silicon substrate is used as a working electrode in an electrochemical cell system. After the electrode is treated by a solution containing strands of complementary oligodeoxynucleotide the Mott-Schottky measurements exhibit obvious negative shift in the flat band potential of the electrode, while in a control experiment treated with a solution of non-complementary oligodeoxynucleotide such a shift does not occur. The DNA hybridization is also manifested in a real time capacitance measurement. A DNA sensor based on the capacitance measurement could be more convenient than that based on a fluorescence detection.

Alkylation↗

PubMed2006

Up-regulation of matrix in bovine articular cartilage explants by electric fields.

Electric stimulation has long been used as a tool to promote connective tissue healing, but the mechanism(s) by which this is accomplished are not yet known. We have previously determined, using mass cultures of fetal bovine articular chondrocytes, a specific set of capacitively coupled electrical stimulation parameters (e.g., duration of stimulation, response time, amplitude, frequency, and duty cycle) that significantly elevated production of collagen and proteoglycan, and up-regulated type II collagen and aggrecan mRNA expression in vitro. In the present study, we applied our best signal parameters (30-min continuous stimulation (100% duty cycle) followed by a pulsed (1h on, 5h off, 4x/day) 50% (1 min on, 1 min off) duty cycle) to cultures of adult bovine articular cartilage explants and obtained similar results. Since the latter system more closely mimics the in vivo chondrocyte environment, these data argue for the utility of electric stimulation for the maintenance of adult cartilage matrix in situ.

Aggrecans↗

PubMed2001

The actions of barium and strontium on exocytosis and endocytosis in the synaptic terminal of goldfish bipolar cells.

1. We investigated the properties of Ca2+-sensitive steps in the cycling of synaptic vesicles by comparing the actions of Ca2+, Ba2+ and Sr2+ in the synaptic terminal of depolarizing bipolar cells isolated from the retina of goldfish. FM1-43 fluorescence and capacitance measurements demonstrated that exocytosis, endocytosis and vesicle mobilization were maintained when external Ca2+ was replaced by either Ba2+ or Sr2+. 2. The rapidly releasable pool of vesicles (RRP) was equivalent to 1.5 % of the membrane surface area when measured in the presence of 2.5 mM Ca2+, but only 0.4 % in 2.5 mM Sr2+. The relative sizes of the RRP in Ca2+, Sr2+ and Ba2+ were 1.0, 0.28 and 0.1, respectively. We conclude that a smaller proportion of docked vesicles are available for fast exocytosis triggered by the influx of Sr2+ or Ba2+ compared to Ca2+. 3. The slow phase of exocytosis was not altered when Ca2+ was replaced by Ba2+, but it was accelerated 1.6-fold in Sr2+. The peak concentrations of Ca2+, Sr2+ and Ba2+ (measured using Mag-fura-5) were approximately 4, approximately 14 and approximately 60 microM, respectively. The order of efficiency for the stimulation of slow exocytosis was Ca2+ approximately Sr2+ > Ba2+. 4. Exocytosis was prolonged after the influx of Sr2+ and Ba2+. Sr2+ was cleared from the synaptic terminal with the same time constant as Ca2+ (1.3 s), but Ba2+ was cleared 10-100 times more slowly. Although Ba(2+) stimulates the slow release of a large number of vesicles, it did so less efficiently than Ca2+ or Sr2+. 5. The recovery of the membrane capacitance was equally rapid in Sr2+ and Ca2+, demonstrating that the fast mode of endocytosis could be triggered by either cation.

Animals↗

PubMed2006

Voltage-dependent capacitance of human embryonic kidney cells.

We determine membrane capacitance, C as a function of dc voltage for the human embryonic kidney (HEK) cell. C was calculated from the admittance, Y, obtained during a voltage ramp when the HEK cell was held in whole-cell patch-clamp configuration. Y was determined at frequencies of 390.625 and from the measured current, i obtained with a dual-sinusoidal stimulus. We find that the fractional increase in the capacitance, C is small ( < 1%) and grows with the square of the voltage, Psi. C can be described by: C=C(0)(1+alpha(Psi+psi(s))2)[where C(0): Capacitance at 0 volts, psi(s): Difference in surface potential between cytoplasmic and extracellular leaflets and alpha: Proportionality constant]. We find that alpha and psi(s) are 0.120 (+/- 0.01) V(-2) and -0.073 (+/-0.017 V in solutions that contain ion channel blockers and 0.108 (+/- 0.29) V(-2) and -0.023 (+/- 0.009) V when 10 mM sodium salicylate was added to the extracellular solution. This suggests that salicylate does not affect the rate at which C grows with Psi, but reduces the charge asymmetry of the membrane. We also observe an additional linear differential capacitance of about (-46 fFV(-1)) in about 60% of the cells, this additional component acts simultaneously with the quadratic component and was not observed when salicylate was added to the solution. We suggest that the voltage dependent capacitance originates from electromechanical coupling either by electrostriction and/or Maxwell stress effects and estimate that a small electromechanical force (approximately equal to 1 pN) acts at physiological potentials. These results are relevant to understand the electromechanical coupling in outer hair cells (OHCs) of the mammalian cochlea, where an asymmetric bell-shaped C versus Psi relationship is observed upon application of a similar field. Prestin, a membrane protein expressed in OHCs is required to observe this function. When we compare the total charge contributions from HEK cell membrane (7 x 10(4) electrons, 10 pF cell) with that determined for prestin transfected cells (up to 5 x 10(6) electrons) we conclude that the charge contributions from the collective motion of membrane proteins and lipids in the field is dwarfed relative to that when prestin is present. We suggest that the capacitance-voltage relationships should be similar to that observed for HEK cells for OHCs that do not express prestin in their membranes.

Cell Line↗

PubMed2002

Regulation of transferrin-induced endocytosis by wild-type and C282Y-mutant HFE in transfected HeLa cells.

The hereditary hemochromatosis protein HFE is known to complex with the transferrin receptor; however, its function regarding endocytosis of transferrin is unclear. We performed patch-clamp capacitance measurements in transfected HeLa cells carrying wild-type or C282Y-mutant HFE cDNA under the control of a tetracycline-sensitive promoter. Whole cell experiments in cells with suppressed expression of wild-type HFE revealed a decrease in membrane capacitance, reflecting predominance of endocytosis in the presence of transferrin. Cells overexpressing C282Y-mutant HFE displayed less intense capacitance decreases, whereas no significant decrease was observed in cells overexpressing wild-type HFE. The formation of single endocytic vesicles in cells with suppressed expression of wild-type HFE was greatly increased in the presence of transferrin as revealed by cell-attached recordings. According to their calculated diameters, many of these vesicles corresponded to clathrin-coated vesicles. These results suggest that wild-type HFE negatively modulates the endocytic uptake of transferrin. This inhibitory effect is attenuated in cells expressing C282Y-mutant HFE. Time-resolved measurements of cell membrane capacitance provide a powerful tool to study transferrin-induced endocytosis in single cells.

Electric Capacitance↗

PubMed2006

Capacitance measurements in the mouse rod bipolar cell identify a pool of releasable synaptic vesicles.

The mouse is an important model system for understanding the molecular basis of neuronal signaling and diseases of synaptic communication. However, the best-characterized retinal ribbon-style synapses are those of nonmammalian vertebrates. To remedy this situation, we asked whether it would be feasible to track synaptic vesicle dynamics in the isolated mouse rod bipolar cell using time-resolved capacitance measurements. The results demonstrate that membrane depolarization triggered an increase in membrane capacitance that was Ca(2+) dependent and restricted to the synaptic compartment, consistent with exocytosis. The amplitude of the capacitance response recorded from the easily accessible soma of an intact mouse rod bipolar cell was identical to that recorded directly from the small synaptic terminal, suggesting that in the carefully selected cohort of cells presented here, axonal resistance was not a significant barrier to current flow. This supposition was supported by the analysis of passive membrane properties and a comparison of membrane capacitance measurements in cells with and without synaptic terminals and reinforced by the lack of an effect of sine-wave frequency (200-1,600 Hz) on the measured capacitance increase. The magnitude of the capacitance response increased with Ca(2+) entry until a plateau was reached at a spatially averaged intraterminal calcium of about 600 nM. We interpret this plateau, nominally 30 fF, as corresponding to a releasable pool of synaptic vesicles. The robustness of this measure suggests that capacitance measurements may be used in the mouse rod bipolar cell to compare pool size across treatment conditions.

Animals↗

PubMed2006

Skin capacitance imaging and corneosurfametry. A comparative assessment of the impact of surfactants on stratum corneum.

Silicon image sensor (SIS) technology was recently introduced as an innovative tool (SkinChip, L'Oréal) providing sensitive imaging of the skin capacitance. This method can detect discrete focal variations in skin surface hydration, and thus early discrete manifestations of skin irritation induced by surfactants. In the present in vivo study, 2 neat and diluted shampoos, and 5% and 10% sodium laurylsulfate solutions were tested on human skin. Each surfactant solution was gently rubbed on the skin using wet hair wicks mimicking the casual use of a shampoo on the scalp. Clinical and SIS evaluations were carried out. In addition, the same products were tested using the ex vivo corneosurfametry bioassay performed on human stratum corneum (SC) harvested by cyanoacrylate skin surface strippings. The colourimetric index of mildness (CIM) was measured on these samples. The product reactivity with the SC was recognized by darker skin capacitance images, and by both lowered SkinChip-generated values and lowered CIM values. The extent in changes varied according to the nature of the test products and their concentrations. The SkinChip image changes likely corresponded to the acute surfactant-induced water swelling of the corneocytes. Skin capacitance imaging and corneosurfametry allow to disclose discrete surfactant-induced alterations of corneocytes.

Dermatitis, Allergic Contact↗

PubMed2005

Transepidermal water loss and skin capacitance alterations among workers in an ultra-low humidity environment.

No studies have been performed evaluating skin barrier alterations in humans exposed to ultra-low humidity (ULH) in spite of several lines of evidence from animal experiments suggesting that the skin barrier is altered on exposure to ULH. The objectives of this study were to assess barrier function changes in workers occupationally exposed to ULH (relative humidity 1.5%), and to evaluate whether the exposure duration shows a dose-response relationship with transepidermal water loss (TEWL) and skin capacitance. A total of 49 male workers exposed to ULH for 12 h per working day were classified into five subgroups based on their ULH exposure duration (<0.5 months, 0.5-1.0 month, 1.1-10.0 months, 10.1-20.0 months, and >20.0 months). A group of 12 age-matched male laboratory workers from a normal humidity environment were recruited as a control group. TEWL and skin capacitance were measured to evaluate their skin barrier function. TEWL measurements showed a significant decline (8.3+/-0.4 vs 10.0+/-0.4 g m(-2) h(-1), P < 0.05) but no differences were found in skin capacitance (39.7+/-1.3 a.u. vs 45.0+/-2.4 a.u., P = 0.68) between the whole ULH exposure group and the control group. Maximum decreases in TEWL and skin capacitance were seen in the subgroups exposed for <0.5 months and 0.5-1.0 month, respectively. Almost completely natural recovery occurred in skin capacitance after 20 months ULH exposure, in contrast to less than 90% recovery in TEWL. Three stages were defined according to the pattern of alterations in TEWL and skin capacitance in relation to ULH exposure duration. A positive association between TEWL and skin capacitance occurred in the control group and stage I but a negative correlation in stage II. No correlation was found in stage III. Our study demonstrated that workers exposed to a ULH environment could exhibit skin barrier alterations. Both TEWL and skin capacitance decreased within 2 weeks of ULH exposure. The maximum alterations in TEWL and skin capacitance occurred during 0.5-1.0 months and 2 weeks, respectively. TEWL recovered partially and more slowly than skin capacitance which recovered earlier and almost completely.

Adult↗

PubMed2006

Application of power spectra patterns in Fourier transform square wave voltammetry to evaluate electrode kinetics of surface-confined proteins.

This paper describes an application of Fourier transform (FT) voltammetry that provides a quantitative evaluation of the electron-transfer kinetics of protein molecules attached to electrode surfaces. The potential waveform applied in these experiments consists of a large-amplitude square wave of frequency f superimposed onto the traditional triangular voltage used in dc cyclic voltammetry. The resultant current-time response, when Fourier transformed into the frequency domain, provides patterns of data at the even harmonic frequencies that arise from nonlinearity in the Faradaic response. These even harmonic contributions are ideally suited for kinetic evaluation of electron-transfer processes because they are highly selective to quasi-reversible behavior (insensitive to reversible or irreversible processes) and almost devoid of background charging current. Inverse FT methods can then be used to provide the wave shapes of the dc as well as the ac voltammetric components and other characteristics employed to detect the level of nonideality present relative to theoretical models based upon noninteracting surface-confined molecules. The new form of data evaluation has been applied to the electron-transfer properties of a typical biological electron carrier, the blue copper protein azurin, immobilized on polycrystalline gold electrodes modified with self-assembled monolayers of different length alkanethiols. Details of the electrode kinetics (rates of electron transfer, dispersion, and charge-transfer coefficients) as a function of alkanethiol, apparent surface coverage, and capacitance are all deduced from the square wave (FT-inverse FT) protocol, and the implications of these findings are considered.

Azurin↗

PubMed2001

Clinical evaluation of defibrillation efficacy with a new single-capacitor biphasic waveform in patients undergoing implantation of an implantable cardioverter defibrillator.

AIMS: Improvements in the size and shape of implantable cardioverter defibrillators (ICDs) might be obtained by using one capacitor instead of the series connection of two capacitors traditionally used in ICDs. The aim of this study was to determine whether a biphasic waveform delivered from a single 336 microF capacitor had the same defibrillation efficacy as a standard biphasic waveform. METHODS AND RESULTS: Randomized, paired defibrillation threshold testing was acutely performed in 54 patients undergoing ICD implantation. A standard 140 microF 80% tilt biphasic waveform (two 280 microF capacitors connected in series) was compared with an experimental biphasic waveform delivered from a single 336 microF capacitor at either 60% tilt (33 patients) or 80% tilt (21 patients). All waveforms had a 60/40 phase1/phase2 duration ratio. Compared with the standard waveform, the 60% tilt experimental waveform had a lower delivered energy (6.7 +/- 2.8 vs 7.9 +/- 3.3 joules, P<0.02), lower peak voltage (218 +/- 43 vs 333 +/- 68 V, P<0.01), and a slightly longer pulse duration (13.4 +/- 1.4 vs 10.7 +/- 1.1 ms, P<0.01). Conversely, the 80% tilt experimental waveform had a higher delivered energy (9.1 +/- 3.5 vs 6.3 +/- 2.4 joules, P<0.01), a lower peak voltage (234 +/- 44 vs 302 +/- 51 V, P<0.01) and a much longer pulse duration (25.7 +/- 2.5 vs 1.13 +/- 1 ms, P<0.01). CONCLUSION: Waveforms delivered from a large capacitance are feasible but require a lower tilt. This technique may allow smaller, thinner ICDs without jeopardizing defibrillation success.

Aged↗

PubMed1995

Tibial nonunion treated with direct current, capacitive coupling, or bone graft.

Two hundred seventy-one tibial nonunions of average duration of 23.5 months (range, 9-69 months) were treated with direct current (167 patients), capacitive coupled electrical stimulation (56 patients), or bone graft surgery (48 patients). Logistic regression analysis was used to compare heal rates among the 3 treatment methods, to identify risk factors adversely affecting the heal rate, and to predict the probability of successful healing of a nonunion of any given risk profile treated with each of the 3 forms of therapy. Seven risk factors were identified: duration of nonunion, prior bone graft surgery, prior electrical treatment, open fracture, osteomyelitis, comminuted or oblique fracture, and atrophic nonunion. When no risk factors were present, there were no significant differences among the 3 treatment methods. As progressively more risk factors were present, the predicted heal rates decreased significantly regardless of the treatment method. Some differences among the treatment groups did appear in the heal rates: bone graft surgery yielded a worse heal rate when there was a previous bone graft failure, and capacitive coupling had a worse heal rate in the presence of an atrophic nonunion.

Adult↗