Voltage jump/capacitance relaxation studies of bilayer structure and dynamics. Studies on oxidized cholesterol membranes.

A voltage-jump technique for the study of the time course of small, voltage-induced changes in the structure of single bilayers is presented, and a method is introduced whereby electromechanical (electrostrictive) phenomena can be separated from dielectric relaxations. As no foreign molecules need be introduced into the bilayers, the question about probe artifacts is eliminated. The time constants and amplitudes of dielectric relaxations in oxidized cholesterol bilayers at 21 degrees C, along with their tentative identification are: (a) tau = 3.3 msec, deltaC/Co = 0.8% and tau = 0.7 msec, deltaC/Co = 0.6%: reorientation in the plane of the membrane of domains or clusters of dipoles. (b) tau = 155 musec, deltaC/Co = 1.5-3%: rotational reorientation of individual molecules. (c) tau = 18 musec, deltaC/Co - 1.4%: small amplitude reorientations individual dipoles about an axis lying in the plane of the membrane. Electrostictive effects with time constants between about 2 and 50 msec were also detected. A temperature study of both the dielectric and electrostrictive phenomena is reported. The application of the technique to other membrane compositions and to a variety of BLM problems is discussed.     

Effect of coherence time of the applied magnetic field on ornithine decarboxylase activity

Skepticism over the possibility of weak electromagnetic fields affecting cell function exists because endogenous thermal noise fields are larger than those reported to cause effects. Four-hour exposure to a 55- or 65-Hz field approximately doubles the specific activity of ornithine decarboxylase (ODC) in L929 cells. To test the idea that the cell discriminates against this thermal noise because it is incoherent, partial incoherence was introduced into the applied field by shifting the frequency between 55- to 65-Hz at intervals of tau coh--delta tau where tau coh is a predetermined time interval and delta tau much less than tau coh varies randomly from one frequency shift to the next. To obtain the full ODC enhancement, coherence of the impressed signal must be maintained for a minimum of about 10s. For tau coh = 5.0s a partial enhancement is elicited, and at 1.0s there is no response. Unfortunately coherence times of this duration are too short to solve the thermal noise puzzle.     

Two distinct modulatory effects on calcium channels in adult rat sensory neurons.

D-ala2-D-leu5-enkephalin (100 to 1000 nM) reduces HVA Ca2+ currents of approximately 60% in 92% of the adult rat sensory neurons tested. In 80% of the cells sensitive to enkephalin, the reduction in Ca2+ current amplitude was associated with a prolongation of the current activation that was relieved by means of conditioning pulses in a potential range only about 10 mV positive to the current activation range in control conditions. The time course of the current activation was fitted to a single exponential in control, (tau = 2.23 msec +/- 0.14 n = 38) and double exponential with enkephalin, (tau 1 = 2.18 msec +/- 0.25 and tau 2 = 9.6 msec +/- 1, test pulse to -10 mV, 22 degrees C). A strong conditioning depolarizing prepulse speeded up the activation time course, completely eliminating the slow, voltage-sensitive exponential component, but it was only partial effective in restoring the current amplitude to control values. The voltage-independent inhibitory component that was not relieved could be recovered only by washing out enkephalin. In the remaining 20% of the cells affected, enkephalin decreased Ca2+ current amplitude without prolongation of Ca2+ channel activation. In these cases the conditioning voltage pulse was not effective in relieving the inhibition that persisted also at strong positive test potentials, on the outward currents. The voltage-dependent inhibition occurred slowly after enkephalin superfusion (tau congruent to 12 sec), whereas the voltage-independent one developed about ten times more rapidly. Dopamine (100 microM) could also induce both voltage-dependent and independent modulations.(ABSTRACT TRUNCATED AT 250 WORDS)     

An extrapolation method for reducing equilibration times in sedimentation equilibrium experiments.

We present a detailed investigation of the use of an extrapolation technique to decrease running times of sedimentation equilibrium experiments. If concentration profiles are available at time delta tau, 2delta tau, 3delta tau,...., cn(r) = c(r, n delta tau), then the Aitken transformation replaces the cn(r) + ĉn(r) = [cn + 1(r) cn - 1(r) - c2n(r)]/[cn + 1(r) + cn - 1(r) - 2cn(r)]. We show that the ĉn(r) converge to the equilibrium values c infinity (r) much more quickly than the cn(r). Savings in time are shown to range from a factor of approximately 2 for meniscus depletion experiments to factors of between 4 and 8 for lower speeds or smaller molecular weights. It is also shown that the technique is quite sensitive to noise, so that an accurate optical system is required to allow its optimal use.     

Cholinergic anti-inflammatory pathway activity and High Mobility Group Box-1 (HMGB1) serum levels in patients with rheumatoid arthritis

High Mobility Group Box-1 (HMGB1) is a cytokine implicated in the pathogenesis of rheumatoid arthritis (RA) and other inflammatory diseases. The cholinergic anti-inflammatory pathway, a vagus nerve-dependent mechanism, inhibits HMGB1 release in experimental disease models. Here, we examine the relationship between vagus nerve activity and HMGB1 in patients with RA. We compared RR interval variability, an index of cardiac vagal modulation, HMGB1 and hsCRP serum levels, and disease activity scores in thirteen RA patients and eleven age- and sex-matched controls. In RA patients, serum levels of HMGB1 and hsCRP were elevated as compared with controls (HMGB1=71 ng/mL [45-99] vs. 18 ng/mL [0-40], P<0.0001; hsCRP=14.5 mg/L [0.7-59] vs. 1 mg/L [0.4-2.9], P<0.001). RR interval variability in RA patients was significantly decreased as compared with controls (HF=38 msec2 [14-80] vs. 288 msec2 [38-364], P<0.0001; rMSSD=20.9+/-9.79 msec, 52.6+/-35.3 msec, P<0.01). HMGB1 levels and RR interval variability were significantly related (rho=-0.49, P<0.01). HMGB1 serum levels significantly correlated with disease activity scores (DAS-28) in patients with RA (P=0.004). The study design does not enable a determination of causality, but the results are consistent with the hypothesis that decreased cholinergic anti-inflammatory pathway activity is associated with increased HMGB1 levels in patients with RA.     

Estimation of heritability and repeatability of resting metabolic rate in birds, with free-living pied flycatchers Ficedula hypoleuca (Aves: Passeriformes) as an example

Estimates of a trait heritability and repeatability can get at an idea of its usefulness for being an individual characteristic and its ability to change under selection pressure. Heritability and repeatability of energetic parameters still poorly studied in birds. The most important physiological characteristic of homoiotherms is resting metabolic rate (RMR), which, in the absence of productive processes, does not exceed basal metabolic rate (BMR). We estimated BMR repeatability in free-living pied flycatchers in Moscow Region (55 degrees 44' N, 36 degrees 51' E; 1992-2008) and Tomsk (56 degrees 20' N, 84 degrees 56' E; 2008-2009) populations over intervals from 40 days to 3 years. In Moscow Region population, BMR repeatability amounted to tau = 0.34 +/- 0.10 (n=80) if measured over 1 year interval, tau = 0.60 +/- 0.15 (n=19) if measured over 2 years interval, and tau = 0.85 +/- 0.13 (n=6) if measured over 3 years interval providing that consecutive BMR measurements were done in the same period of reproductive season. In Tomsk population, BMR repeatability, measured over 1 year interval, amounted to tau = 0.49 +/- 0.11 (n=50). Repeatability is a measure of a trait constancy and sets the upper limit of its heritability. To estimate RMR heritability, cross-fostering experiments have been conducted in 2003-2005 with flycatchers of Moscow Region population. RMR of chicks positively correlated with BMR of their biological fathers, whereas such correlation in metabolic rates between chicks and their foster fathers was absent. The RMR heritability estimate turned out to be h2 = 0.43 +/- 0.17 (n=210). The obtained estimates of heritability and repeatability of fundamental energetic traits are rather high for physiological features. This suggests the existence of a potential for direct selection on BMR and evolutionary stable diversity of avian populations with regard to basal metabolic rate.     

Efficiency of a visual system under conditions of uncertainty of the appearance of a moving object

The effect of uncertainty of a moving object appearance in the noise field upon the coefficient efficiency, we studied. At short durations of presentation (40-80 msec) and high level of external noise, this effect was maximal: magnified 100 times. The efficiency coefficient dependence on the duration of a moving object presentation was shown to be characterized by two maximums. The position of minimum situated between these two maximums was found to be independent of either presence or absence of uncertainty of a number of parameters: such as initial position of the object the image, time of its appearance, noise level, velocity and direction of the movement, and has a latency approximately 120 sec. A functional model of the observed phenomena, has been proposed.     

Membrane pores: a computer simulation of interacting pores analyzed by g1(tau) and g2(tau) correlation functions

Ion channels in a cell membrane were modeled by a computer simulation of fluctuating pores distributed in a spatial array, a cellular automata. The sum of the currents through such a set of pores models a noise analysis experiment. These currents were analyzed by using the optical correlation functions gn(tau) = (fn(t)fn(t + tau))/(f2(t))n, where f(t) are the current deviations around the mean current and () denotes the time average. These functions can be easily used to determine if the noise is Gaussian. If the noise is not Gaussian, they provide additional information not already contained in the power spectrum. When the pores do not interact with each other, the noise is Gaussian and the power spectrum a Lorentzian. When the pores interact in a strongly cooperative way the noise was still Gaussian and the power spectrum still a Lorentzian, but the usual analysis applied to such a case would over-estimate the single channel conductance. If the kinetics of the pore opening and closing vary on the time scale of the experiment then the relationship g2(tau) = 1 + 2[g1(tau)]2 is no longer satisfied.     

Lateral mobility in membranes as detected by fluorescence recovery after photobleaching.

The evaluation of lateral diffusion coefficients of membrane components by the technique of fluorescence recovery after photobleaching (FRAP) is often complicated by uncertainties in the values of the intensities F(O), immediately after bleaching, and F(infinity), after full recovery. These uncertainties arise from instrumental settling time immediately after bleaching and from cell, tissue, microscope, or laser beam movements at the long times required to measure F(infinity). We have developed a method for precise analysis of FRAP data that minimizes these problems. The method is based on the observation that a plot of the reciprocal function R(tau) = F(infinity)/[F(infinity)-F(tau)] is linear over a large time range when (a) the laser beam has a Gaussian profile, (b) recovery involves a single diffusion coefficient, and (c) there is no membrane flow. Moreover, the ratio of intercept to slope of the linear plot is equal to tau 1/2, the time required for the bleached fluorescence to rise to 50% of the full recovery value, F(infinity). The lateral diffusion coefficient D is related to tau 1/2 by tau 1/2 = beta w2/4D where beta is a defined parameter and w is the effective radius of the focused laser beam. These results are shown to indicate that the recovery of fluorescence F(tau) can be represented over a large range of percent bleach, and recovery time tau by the relatively simple expression F(tau) = [ F(o) + F(infinity) (tau/tau 1/2)]/[1 + tau/tau 1/2)]. FRAP data can therefore be easily evaluated by a nonlinear regression analysis with this equation or by a linear fit to the reciprocal function R(tau). It is shown that any error in F(infinity) can be easily detected in a plot of R(tau) vs. tau which deviates significantly from a straight line when F(infinity) is in error by as little as 5%. A scheme for evaluating D by linear analysis is presented. It is also shown that the linear reciprocal plot provides a simple method for detecting flow or multiple diffusion coefficients and for establishing conditions (data precision, differences in multiple diffusion coefficients, magnitude of flow rate compared to lateral diffusion) under which flow or multiple diffusion coefficients can be detected. These aspects are discussed in some detail.     

A TEA-insensitive flickering potassium channel active around the resting potential in myelinated nerve

A novel potassium-selective channel which is active at membrane potentials between -100 mV and +40 mV has been identified in peripheral myelinated axons of Xenopus laevis using the patch-clamp technique. At negative potentials with 105 mM-K on both sides of the membrane, the channel at 1 kHz resolution showed a series of brief openings and closings interrupted by longer closings, resulting in a flickery bursting activity. Measurements with resolution up to 10 kHz revealed a single-channel conductance of 49 pS with 105 mM-K and 17 pS with 2.5 mM-K on the outer side of the membrane. The channel was selective for K ions over Na ions (PNa/PK = 0.033). The probability of being within a burst in outside-out patches varied from patch to patch (> 0.2, but often > 0.9), and was independent of membrane potential. Open-time histograms were satisfactorily described with a single exponential (tau o = 0.09 msec), closed times with the sum of three exponentials (tau c = 0.13, 5.9, and 36.6 msec). Sensitivity to external tetraethylammonium was comparatively low (IC50 = 19.0 mM). External Cs ions reduced the apparent unitary conductance for inward currents at Em = -90 mV (IC50 = 1.1 mM). Ba and, more potently, Zn ions lowered not only the apparent single-channel conductance but also open probability. The local anesthetic bupivacaine with high potency reduced probability of being within a burst (IC50 = 165 nM). The flickering K channel is clearly different from the other five types of K channels identified so far in the same preparation. We suggest that this channel may form the molecular basis of the resting potential in vertebrate myelinated axons.     

Alterations in outward K(+) currents on removal of external Ca(2+) in human atrial myocytes

External divalent cations are known to play an important role in the function of voltage-gated ion channels. The purpose of this study was to examine the sensitivity of the voltage-gated K(+) currents of human atrial myocytes to external Ca(2+) ions. Myocytes were isolated by collagenase digestion of atrial appendages taken from patients undergoing coronary artery-bypass surgery. Currents were recorded from single isolated myocytes at 37 degrees C using the whole-cell patch-clamp technique. With 0.5 mM external Ca(2+), voltage pulses positive to -20 mV (holding potential = -60 mV) activated outward currents which very rapidly reached a peak (I(peak)) and subsequently inactivated (tau = 7.5 +/- 0.7 msec at +60 mV) to a sustained level, demonstrating the contribution of both rapidly inactivating transient (I(to1)) and non-inactivating sustained (I(so)) outward currents. The I(to1) component of I(peak), but not I(so), showed voltage-dependent inactivation using 100 msec prepulses (V(1/2) = -35.2 +/- 0.5 mV). The K(+) channel blocker, 4-aminopyridine (4-AP, 2 mM), inhibited I(to1) by approximately 76% and reduced I(so) by approximately 33%. Removal of external Ca(2+) had several effects: (i) I(peak) was reduced in a manner consistent with an approximately 13 mV shift to negative voltages in the voltage-dependent inactivation of I(to1). (ii) I(so) was increased over the entire voltage range and this was associated with an increase in a non-inactivating 4-AP-sensitive current. (iii) In 79% cells (11/14), a slowly inactivating component was revealed such that the time-dependent inactivation was described by a double exponential time course (tau(1) = 7.0 +/- 0.7, tau(2) = 90 +/- 21 msec at +60 mV) with no effect on the fast time constant. Removal of external Ca(2+) was associated with an additional component to the voltage-dependent inactivation of I(peak) and I(so) (V(1/2) = -20.5 +/- 1.5 mV). The slowly inactivating component was seen only in the absence of external Ca(2+) ions and was insensitive to 4-AP (2 mM). Experiments with Cs(+)-rich pipette solutions suggested that the Ca(2+)-sensitive currents were carried predominantly by K(+) ions. External Ca(2+) ions are important to voltage-gated K(+) channel function in human atrial myocytes and removal of external Ca(2+) ions affects I(to1) and 4-AP-sensitive I(so) in distinct ways.     

The pressure dependent dynamic elasticity of 35 thoracic and 16 abdominal human aortas in vitro described by a five component model

Segments of 35 thoracic and 16 abdominal human aortas, including nine pairs, aged 30-78 yr at autopsy, were perfused with 37 degrees C Tyrode's solution at in situ length. Diameter changes due to 20 mmHg pressure steps between 20 and 180 mmHg were measured to 1 micron accuracy at an equivalent noise level of 0.1 micron RMS, using balanced transducers. Aortic creep curves at each pressure level were described individually by a constant plus bi-exponential creep model characterized by two creep fractions (alpha 1 and alpha 2) and two time constants (tau 1 and tau 2). Creep fractions and time constants increased substantially with the pressure level, indicating a significant effect of pressure or distension on aortic viscoelasticity. At 110 mmHg the mean +/- 1 S.D. parameter values were: thoracic aorta: alpha 1 = 0.076 +/- 0.017, alpha 2 = 0.102 +/- 0.028, tau 1 = 0.73 +/- 0.29 s, tau 2 = 14.0 +/- 4.1 s; abdominal aorta: alpha 1 = 0.078 +/- 0.017, alpha 2 = 0.101 +/- 0.025, tau 1 = 0.61 +/- 0.12 s, tau 2 = 12.1 +/- 3.4 s. Nine paired comparisons at each pressure level showed that creep fractions and time constants of thoracic and abdominal segments were not significantly different (p = 0.05).     

The inhibitory chloride channel of the lobster Panulirus penicillatus neuromuscular junction.

1. Single channel activity was recorded from muscle membranes of the lobster Panulirus penicillatus using the patch-clamp technique. 2. Cell-attached, outside-out and inside-out patches were prepared from the deep abdominal extensor muscle. 3. Low amplitude single channel currents were observed in most patches, and were identified as being chloride-currents. 4. The chloride channel was active spontaneously, and tended to desensitize when outside-out patches were exposed to a small jet of glutamate. 5. Amplitude histograms of single channel currents presented a well defined peak of 8 pA at a membrane potential of -160 mV, while open and closed time histograms were fit to single exponential functions with tau open of 3.27 msec and tau closed of 31.58 msec.     

Evoked potentials in the visual and association cortex of alert cats during paired uniform stimulation of the lateral geniculate body and pulvinar

Recovery curves of evoked potentials in the association and visual cortex during paired stimulation of the pulvinar in chronic experiments on alert cats were shown to be similar in character. Depression of the test response was observed only if the interval between stimuli was of the order of 10 msec, but if it was 40 msec considerable (2–4 times) facilitation of the second response was observed, mainly on account of an increase in the negative component N₁. Facilitation was less marked if the intervals were from 60 to 100 msec, and they decreased gradually to an interval of 200 msec. The recovery curve of cortical evoked potentials during paired stimulation of the lateral geniculate body differed considerably from the recovery curve during paired stimulation of the pulvinar and was characterized by a gradual increase in amplitude of the second response — from its almost total suppression with an interval of 10 msec to slight facilitation with an interval of 200 msec. If intervals of 10 to 80 msec were used, the test response was restored more slowly in the association cortex than in the visual cortex. The results are discussed from the standpoint of differences in the character of intracortical spread of excitation as a result of activation of geniculo-cortical and pulvinar-cortical pathways of conduction of information.Institute of Higher Nervous Activity and Neurophysiology, Academy of Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 16, No. 4, pp. 497–505, July–August, 1984.     

QT interval and QT dispersion before and after diet therapy in patients with simple obesity

To evaluate whether a disordered QT interval and its dispersion in obese patients, if any, may be improved by therapeutic weight reduction, 36 obese patients admitted to our university hospital were examined over a 5-year period from April 1, 1992 to March 31, 1997. Participants included 18 males and 18 females whose mean age +/- SD was 28 +/- 9 and 33 +/- 14 years, respectively, and whose mean body mass index +/- SD was 35 +/- 5 and 38 +/- 6 kg/m2, respectively. Thirty-six control patients were matched in age and gender with the obese patients. All the obese patients were treated with behavioral therapy together with very-low-calorie conventional Japanese diet (VLCD: 370 kcal/day). A standard 12-lead electrocardiogram (ECG) revealed longer maximum (445 +/- 32 msec, mean +/- SD) and minimum (388 +/- 29 msec) heart rate corrected QT intervals (QTc intervals) in the obese group than in the control group (P < 0.0001 for each). QTc dispersion, defined as the difference between maximum and minimum QTc intervals derived from 12-lead ECG, was greater in the obese group (57 +/- 19 msec) than in the control group (32 +/- 13 msec) (P < 0.0001). Both the maximum and minimum QTc intervals in the obese patients were shortened, respectively, to 434 +/- 28 msec and 377 +/- 29 msec (P < 0.05 for each) with no significant change in either QTc dispersion, QRS voltage, or QRS duration following weight reduction. The coefficient value from the linear regression line between QT interval and RR interval in the obese group was less than in the control group. Together, the results show that obesity per se causes both a prolongation of QTc interval and an increase in QTc dispersion, and that weight reduction improves the prolonged QTc interval observed in obese patients.     

Ligand binding on ladder lattices

The ligand binding problems on two-dimensional ladders, which model many important binding phenomena in molecular biology, are studied in details. The model is represented by four parameters, the interactions between ligands when bound to adjacent sites on opposite legs of the ladder (tau), the interactions between bound ligands in the longitudinal direction of the ladder (sigma), the number of binding sites that are covered by a bound ligand (m), and the intrinsic binding constant (K). The partition functions of ring ladders are approached with the transfer matrix method. A general relation is derived which connects the partition function of a linear ladder with that of a ring ladder. The results obtained apply to the general situation of multivalent binding, in which m>1. Special attention is paid to the case where the ligand covers one site (m=1). In this case explicit formulas are given for the partition functions of ring and linear ladders. Closed-form expressions are obtained for various properties of the system, including the degree of binding (theta), the midpoint in the binding isotherm (1/square root(tau sigma)), the initial and end slopes of the Scatchard plots (2sigma + tau - 4 and -sigma2 tau, respectively). From these closed-form formulas, sigma and tau may be extracted from experimental data. The model reveals certain features which do not exist in one-dimensional models. Using the general method discussed in [1], the recurrence relation is found for the partition functions. The analytical solution found for this model provides test cases to verify the numerical results for more complex two-dimensional models.     

Non-proline-dependent protein kinases phosphorylate several sites found in tau from Alzheimer disease brain

Of 21 phosphorylation sites identified in PHF-tau 11 are on ser/thr-X motifs and are probably phosphorylated by non-proline-dependent protein kinases (non-PDPKs). The identities of the non-PDPKs and how they interact to hyperphosphorylate PHF-tau are still unclear. In a previous study we have shown that the rate of phosphorylation of human tau 39 by a PDPK (GSK-3) was increased several fold if tau were first prephosphorylated by non-PDPKs (Singh et al., FEBS Lett 358: 267-272, 1995). In this study we have examined how the specificity of a non-PDPK for different sites on human tau 39 is modulated when tau is prephosphorylated by other non-PDPKs (A-kinase, C-kinase, CK-1, CaM kinase II) as well as a PDPK (GSK-3). We found that the rate of phosphorylation of tau 39 by a non-PDPK can be stimulated if tau were first prephosphorylated by other non-PDPKs. Of the four non-PDPKs only CK-1 can phosphorylate sites (thr 231, ser 396, ser 404) known to be present in PHF-tau. Further, these sites were phosphorylated more rapidly and to a greater extent by CK-1 if tau 39 were first prephosphorylated by A-kinase, CaM kinase II or GSK-3. These results suggest that the site specificities of the non-PDPKs that participate in PHF-tau hyperphosphorylation can be modulated at the substrate level by the phosphorylation state of tau.Abbreviations PHF paired helical filaments - A-kinase cyclic AMP-dependent protein kinase - CaM kinase II calcium/calmodulin-dependent protein kinase II - C-kinase calcium/phospholipid-dependent protein kinase - CK-1 casein kinase-1 - CK-2 casein kinase-2 - GSK-3 glycogen synthase kinase-3 - MAP kinase mitogen-activated protein kinase - PDPK proline-dependent protein kinase     

Electrophysiological properties of isolated rat liver cells

The electrophysiological properties of isolated rat liver cells were studied using the patch clamp method in whole-cell configuration. The membrane potential in isolated hepatocytes was -42 +/- 7 mV (n = 20). The input resistance (Rin) and the time constant (tau m) were 51 +/- 17 M (the range of 34 to 180 M omega) (n = 20) and 4.2 +/- 1.0 msec (the range of 3 to 16.5 ms) (n = 20). Assuming that the specific membrane capacitance is 1 microF/cm2, the membrane resistance and membrane capacitance were 42. +/- 9.0 K omega cm2 and 87 +/- 27 pF. These values indicate that isolated rat hepatocytes are not abnormally permeable or leaky. The current-voltage relationship was linear with no rectification. The depolarizing pulse from the resting potential did not induce fast or slow inward currents even when norepinephrine or high Ca2 (3.6 mM) were applied. This indicates that there is no voltage-sensitive Ca2+ channel in the isolated hepatocytes.     

Relation of the Escherichia coli dnaX gene to its two products--the tau and gamma subunits of DNA polymerase III holoenzyme.

The Escherichia coli DNA polymerase III holoenzyme 71.1 kDa tau subunit is a 643 amino acid protein encoded by the dnaX gene. This gene also encodes the holoenzyme 56.5 kDa gamma subunit. The tau factor (as a tau'-LacZ' fusion protein) has been isolated and shown to be cleaved in vitro to form gamma and a 135 kda C-terminal cleavage product. The tau'-LacZ' fusion protein, gamma, and the C-terminal cleavage product have been isolated. N-terminal sequencing has demonstrated that tau and gamma share the same N-terminal sequences and that tau is proteolytically cleaved in vitro between residues 498 and 499 to form gamma. In addition, residues 420-440 were shown to be present in both tau and gamma by use of antibody specific for a synthetic peptide corresponding to that sequence. Some mechanism functions in vivo to ensure that tau and gamma are synthesized in a ratio of about one-to-one, as shown by radioimmune precipitation of tau and gamma from cellular extracts.     

Comparison of excitatory currents activated by different transmitters on crustacean muscle. I. Acetylcholine-activated channels

The properties of acetylcholine-activated excitatory currents on the gm1 muscle of three marine decapod crustaceans, the spiny lobsters Panulirus argus and interruptus, and the crab Cancer borealis, were examined using either noise analysis, analysis of synaptic current decays, or analysis of the voltage dependence of ionophoretically activated cholinergic conductance increases. The apparent mean channel open time (tau n) obtained from noise analysis at -80 mV and 12 degrees C was approximately 13 ms; tau n was prolonged e-fold for about every 100-mV hyperpolarization in membrane potential; tau n was prolonged e- fold for every 10 degrees C decrease in temperature. Gamma, the single- channel conductance, at 12 degrees C was approximately 18 pS and was not affected by voltage; gamma was increased approximately 2.5-fold for every 10 degrees C increase in temperature. Synaptic currents decayed with a single exponential time course, and at -80 mV and 12 degrees C, the time constant of decay of synaptic currents, tau ejc, was approximately 14-15 ms and was prolonged e-fold about every 140-mV hyperpolarization; tau ejc was prolonged about e-fold for every 10 degrees C decrease in temperature. The voltage dependence of the amplitude of steady-state cholinergic currents suggests that the total conductance increase produced by cholinergic agonists is increased with hyperpolarization. Compared with glutamate channels found on similar decapod muscles (see the following article), the acetylcholine channels stay open longer, conduct ions more slowly, and are more sensitive to changes in the membrane potential.