Long-term memory in Staphylococcus aureus α-hemolysin ion channel kinetics.

The kinetics of an ion channel are classically understood as a random process. However, studies have shown that in complex ion channels, formed by multiple subunits, this process can be deterministic, presenting long-term memory. Staphylococcus aureus α-hemolysin (α-HL) is a toxin that acts as the major factor in Staphylococcus aureus virulence. α-HL is a water-soluble protein capable of forming ion channels into lipid bilayers, by insertion of an amphipathic  ß-barrel. Here, the α-HL was used as an experimental model to study memory in ion channel kinetics. We applied the approximate entropy (ApEn) approach to analyze randomness and the Detrended Fluctuation Analysis (DFA) to investigate the existence of long memory in α-HL channel kinetics. Single-channel currents were measured through experiments with α-HL channels incorporated in planar lipid bilayers. All experiments were carried out under the following conditions: 1 M NaCl solution, pH 4.5; transmembrane potential of + 40 mV and temperature 25 ± 1 °C. Single-channel currents were recorded in real-time in the memory of a microcomputer coupled to an A/D converter and a patch-clamp amplifier. The conductance value of the α-HL channels was 0.82 ± 0.0025 nS (n = 128). The DFA analysis showed that the kinetics of α-HL channels presents long-term memory ([Formula: see text] = 0.63 ± 0.04). The ApEn outcomes showed low complexity to dwell times when open (ApEno = 0.5514 ± 0.28) and closed (ApEnc = 0.1145 ± 0.08), corroborating the results of the DFA method.

Memory in Ion Channel Kinetics.

Ion channels are transport proteins present in the lipid bilayers of biological membranes. They are involved in many physiological processes, such as the generation of nerve impulses, hormonal secretion, and heartbeat. Conformational changes in the ion channel-forming protein allow the opening or closing of pores to control the ionic flux through the cell membranes. The opening and closing of the ion channel have been classically treated as a random kinetic process, known as a Markov process. Here the time the channel remains in a given state is assumed to be independent of the condition it had in the previous state. More recently, however, several studies have shown that this process is not random but a deterministic one, where both the open and closed dwell-times and the ionic current flowing through the channel are history-dependent. This property is called long memory or long-range correlation. However, there is still much controversy regarding how this memory originates, which region of the channel is responsible for this property, and which models could best reproduce the memory effect. In this article, we provide a review of what is, where it is, its possible origin, and the mathematical methods used to analyze the long-term memory present in the kinetic process of ion channels.

Omega-3 effects on electrocorticographic patterns of adult Wistar rats exposed to ionizing radiation.

This study aimed to assess the effect of supplementation with omega-3 in Wistar rats exposed to ionizing radiation in a dose of 18 Gy on the cortical electrical activity, using mathematical methods such as the power spectrum (PS) and the detrended fluctuation analysis (DFA) in the evaluation of the electrocorticogram (ECoG) record. The PS analysis showed that in non-irradiated animals but supplemented with omega-3 there was a decrease in the power of the beta rhythm, while the DFA applied to different frequency ranges of the ECoG showed a significant increase in the long-range correlation only for the theta wave when compared with non-supplemented animals. In the evaluation of the radiation effect through the PS, an increase in the power of the theta rhythm was observed in both groups (non-supplemented and supplemented animals) only when they were evaluated one week after irradiation. The DFA method also showed difference in this wave. The PS and DFA methods applied to the ECoG record allowed a quantitative analysis of the cortical electrical activity in rats in response to the omega-3 effects, ionizing radiation, or both.

Biomechanical and biochemical investigation of erythrocytes in late stage human leptospirosis.

Leptospirosis is a zoonotic disease caused by bacteria of the genus Leptospira, which can cause lipid changes in the erythrocyte membrane. Optical tweezers were used to characterize rheological changes in erythrocytes from patients with leptospirosis in the late stage. Biochemical methods were also used for quantification of plasma lipid, erythrocyte membrane lipid, and evaluation of liver function. Our data showed that the mean elastic constant of erythrocytes from patients with leptospirosis was around 67% higher than the control (healthy individuals), indicating that patient's erythrocytes were less elastic. In individuals with leptospirosis, several alterations in relation to control were observed in the plasma lipids, however, in the erythrocyte membrane, only phosphatidylcholine showed a significant difference compared to control, increasing around 41%. With respect to the evaluation of liver function of individuals with leptospirosis, there was a significant increase in levels of alanine transaminase (154%) and aspartate transaminase (150%), whereas albumin was 43.8% lower than control (P<0.01). The lecithin-cholesterol acyltransferase fractional activity was 3.6 times lower in individuals with leptospirosis than in the healthy individuals (P<0.01). The decrease of the erythrocyte elasticity may be related to the changes of erythrocyte membrane phospholipids composition caused by disturbances that occur during human leptospirosis, with phosphatidylcholine being a strong candidate in the erythrocyte rheological changes.

Biomechanical and biochemical investigation of erythrocytes in late stage human leptospirosis

Leptospirosis is a zoonotic disease caused by bacteria of the genus Leptospira, which can cause lipid changes in the erythrocyte membrane. Optical tweezers were used to characterize rheological changes in erythrocytes from patients with leptospirosis in the late stage. Biochemical methods were also used for quantification of plasma lipid, erythrocyte membrane lipid, and evaluation of liver function. Our data showed that the mean elastic constant of erythrocytes from patients with leptospirosis was around 67% higher than the control (healthy individuals), indicating that patient's erythrocytes were less elastic. In individuals with leptospirosis, several alterations in relation to control were observed in the plasma lipids, however, in the erythrocyte membrane, only phosphatidylcholine showed a significant difference compared to control, increasing around 41%. With respect to the evaluation of liver function of individuals with leptospirosis, there was a significant increase in levels of alanine transaminase (154%) and aspartate transaminase (150%), whereas albumin was 43.8% lower than control (P<0.01). The lecithin-cholesterol acyltransferase fractional activity was 3.6 times lower in individuals with leptospirosis than in the healthy individuals (P<0.01). The decrease of the erythrocyte elasticity may be related to the changes of erythrocyte membrane phospholipids composition caused by disturbances that occur during human leptospirosis, with phosphatidylcholine being a strong candidate in the erythrocyte rheological changes.

Long-term correlation of the electrocorticogram as a bioindicator of brain exposure to ionizing radiation.

Understanding the effects of radiation and its possible influence on the nervous system are of great clinical interest. However, there have been few electrophysiological studies on brain activity after exposure to ionizing radiation (IR). A new methodological approach regarding the assessment of the possible effects of IR on brain activity is the use of linear and nonlinear mathematical methods in the analysis of complex time series, such as brain oscillations measured using the electrocorticogram (ECoG). The objective of this study was to use linear and nonlinear mathematical methods as biomarkers of gamma radiation regarding cortical electrical activity. Adult Wistar rats were divided into 3 groups: 1 control and 2 irradiated groups, evaluated at 24 h (IR24) and 90 days (IR90) after exposure to 18 Gy of gamma radiation from a cobalt-60 radiotherapy source. The ECoG was analyzed using power spectrum methods for the calculation of the power of delta, theta, alpha and beta rhythms and by means of the α-exponent of the detrended fluctuation analysis (DFA). Using both mathematical methods it was possible to identify changes in the ECoG, and to identify significant changes in the pattern of the recording at 24 h after irradiation. Some of these changes were persistent at 90 days after exposure to IR. In particular, the theta wave using the two methods showed higher sensitivity than other waves, suggesting that it is a possible biomarker of exposure to IR.

Long-term correlation of the electrocorticogram as a bioindicator of brain exposure to ionizing radiation

Understanding the effects of radiation and its possible influence on the nervous system are of great clinical interest. However, there have been few electrophysiological studies on brain activity after exposure to ionizing radiation (IR). A new methodological approach regarding the assessment of the possible effects of IR on brain activity is the use of linear and nonlinear mathematical methods in the analysis of complex time series, such as brain oscillations measured using the electrocorticogram (ECoG). The objective of this study was to use linear and nonlinear mathematical methods as biomarkers of gamma radiation regarding cortical electrical activity. Adult Wistar rats were divided into 3 groups: 1 control and 2 irradiated groups, evaluated at 24 h (IR24) and 90 days (IR90) after exposure to 18 Gy of gamma radiation from a cobalt-60 radiotherapy source. The ECoG was analyzed using power spectrum methods for the calculation of the power of delta, theta, alpha and beta rhythms and by means of the α-exponent of the detrended fluctuation analysis (DFA). Using both mathematical methods it was possible to identify changes in the ECoG, and to identify significant changes in the pattern of the recording at 24 h after irradiation. Some of these changes were persistent at 90 days after exposure to IR. In particular, the theta wave using the two methods showed higher sensitivity than other waves, suggesting that it is a possible biomarker of exposure to IR.

Do physical fitness measures influence internal training load responses in high-level futsal players?

BACKGROUND: The aim of the present study was to verify whether aerobic fitness and ability to perform repeated high-intensity efforts influence the internal training load (ITL), which consists of the actual stress imposed in the athletes' organisms, in professional futsal players. METHODS: Twelve high-level futsal players (age: 26.3±4.9 years, body mass: 73.5±7.5 kg) participated in the study. The investigated athletes took part in a 5-week pre-season period. The ITL was quantified by means of the session-Rating of Perceived Exertion method. The athletes performed the Yo-yo Intermittent Recovery Test level 2 (YYIR2) in order to assess the ability to perform repeated high-intensity actions, and the multistage shuttle-run test (MSRT) in order to evaluate aerobic fitness, before (T0) and after (T1) the pre-season period. Maximal oxygen uptake (VO2max.), oxygen uptake at Respiratory compensation point (VO2-RCP) and maximal aerobic speed (MAS) obtained in MSRT were retained for analyses. RESULTS: The results from Pearson's correlation test showed significant and a very large correlation between ITL and YYIR2 performance (r=-0.75). Moreover, a significant and large correlation between ITL and VO2max. (r=-0.62), ITL and MAS (r=-0.67), and ITL and VO2-RCP (r=-0.58) were also observed. CONCLUSION: It can be concluded that aerobic fitness and ability to perform repeated high-intensity actions may influence ITL responses in professional futsal players.

Crescimento vascular em membrana do saco vitelínico e desenvolvimento embrionário de codornas japonesas (Coturnix japonica) expostas a campo magnético de baixa frequência / Vascular growth in yolk sac membrane and embryonic development of Japanese quail (Coturnix japonica) exposed to low frequency magnetic field

O objetivo deste trabalho foi observar a influência do campo magnético (CM) de baixa frequência na membrana do saco vitelínico (MSV) e no desenvolvimento do embrião de codornas japonesas (Coturnix japonica) em 72 horas de incubação. Ovos fertilizados foram expostos a nove horas consecutivas de CM, sendo um grupo a partir das 48 horas e o outro a partir das 63 horas de incubação. A quantificação da vascularização da MSV foi determinada pela obtenção da dimensão fractal por meio dos métodos de box-counting e de dimensão de informação, enquanto o peso corporal e o percentual de comprimento cefálico dos embriões foram utilizados como parâmetros de desenvolvimento embrionário. O CM não causou diferenças significativas na densidade vascular da MSV nem no desenvolvimento embrionário, quando comparados ao grupo controle...

The aim of this study was to observe the influence of the low frequency magnetic field (MF) on the yolk sac membrane (YSM) and embryonic development of Japanese quail (Coturnix japonica) in 72 hours of incubation. Fertilized eggs were exposed to 9 consecutive hours of MF, with a group from 48 hours and the second group from 63 hours of incubation. The evaluation of YSM vascularization was determined by the fractal dimensions obtained through box-counting method and information dimension, while body weight of the embryo and percentage of cephalic length were used as parameters for embryo development. The MF caused no significant differences in vessel density in the YSM, nor in the embryonic development considering the body weight and percentage cephalic length, when were compared to the control group...

Efeito dos ácidos graxos essenciais sobre lipidemia e vascularização da membrana vitelina de codornas japonesas / Effect of essential fatty acids on lipidemia and vitelline membrane vascularization in Japanese quails

Verificou-se o efeito de diferentes fontes de ácidos graxos essenciais ômega-3 e ômega-6 sobre o perfil lipídico de codornas japonesas e sobre o crescimento vascular na membrana vitelina dos embriões de aves suplementadas com óleo de peixe, 2% e 4%, e óleo de soja, 2% e 4%, em relação à dieta-controle, sem suplementação lipídica. Foi usado o método enzimático com reação colorimétrica para estimar o perfil lipídico sérico de colesterol total, triglicerídeos e HDL das aves. A vascularização na membrana vitelina foi quantificada por meio da dimensão fractal, utilizando-se o método de box-counting. A concentração de colesterol dos grupos controle e tratados não diferiu entre si. Para o HDL, o grupo que recebeu maior proporção de óleo de peixe, 4%, diferiu dos outros grupos. Os níveis de triglicerídeos das codornas suplementadas com óleo de peixe foram superiores aos dos demais grupos. A dimensão fractal da vascularização da membrana vitelina dos embriões de codornas dos grupos que receberam 4% de óleo de peixe e 4% de óleo de soja foi significativamente menor que a dos demais grupos, indicando efeitos antiangiogênicos no processo de formação vascular.

The effect of different sources of omega-3 and omega-6 essential fatty acids on serum lipid profile in Japanese quails and on vascular growth in the embryos vitelline membrane was verified. The birds were supplemented with fish oil, 2% and 4%, and soybean oil, 2% and 4%, and one control diet, without lipid supply. The enzymatic method with colorimetric reaction was used to estimate the profile of serum total cholesterol, triglycerides and HDL of the birds. Vitelline membrane vascularization was measured using the box-counting fractal dimension. The cholesterol concentration of the treated and control groups showed no difference. For HDL, the group treated with a higher proportion of fish oil, 4%, differed from other groups. The triglycerides levels of quails supplemented with fish oil were higher than the other groups. The fractal dimension of quail embryos vitelline membrane from the groups treated with 4% fish oil and 4% soybean oil was lower than the other groups, indicating antiangiogenic effects on the vascular formation process.

Can the fractal dimension be applied for the early diagnosis of non-proliferative diabetic retinopathy?

The fractal dimension has been employed as a useful parameter in the diagnosis of retinal disease. Avakian et al. (Curr Eye Res 2002; 24: 274-280), comparing the vascular pattern of normal patients with mild to moderate non-proliferative diabetic retinopathy (NPDR), found a significant difference between them only in the macular region. This significant difference in the box-counting fractal dimension of the macular region between normal and mild NPDR patients has been proposed as a method of precocious diagnosis of NPDR. The aim of the present study was to determine if fractal dimensions can really be used as a parameter for the early diagnosis of NPDR. Box-counting and information fractal dimensions were used to parameterize the vascular pattern of the human retina. The two methods were applied to the whole retina and to nine anatomical regions of the retina in 5 individuals with mild NPDR and in 28 diabetic but opthalmically normal individuals (controls), with age between 31 and 86 years. All images of retina were obtained from the Digital Retinal Images for Vessel Extraction (DRIVE) database. The results showed that the fractal dimension parameter was not sensitive enough to be of use for an early diagnosis of NPDR.

Can the fractal dimension be applied for the early diagnosis of non-proliferative diabetic retinopathy?

The fractal dimension has been employed as a useful parameter in the diagnosis of retinal disease. Avakian et al. (Curr Eye Res 2002; 24: 274-280), comparing the vascular pattern of normal patients with mild to moderate non-proliferative diabetic retinopathy (NPDR), found a significant difference between them only in the macular region. This significant difference in the box-counting fractal dimension of the macular region between normal and mild NPDR patients has been proposed as a method of precocious diagnosis of NPDR. The aim of the present study was to determine if fractal dimensions can really be used as a parameter for the early diagnosis of NPDR. Box-counting and information fractal dimensions were used to parameterize the vascular pattern of the human retina. The two methods were applied to the whole retina and to nine anatomical regions of the retina in 5 individuals with mild NPDR and in 28 diabetic but opthalmically normal individuals (controls), with age between 31 and 86 years. All images of retina were obtained from the Digital Retinal Images for Vessel Extraction (DRIVE) database. The results showed that the fractal dimension parameter was not sensitive enough to be of use for an early diagnosis of NPDR.

Fractal analysis of retinal vessel patterns in ophthalmically normal dogs.

OBJECTIVE: To study the applicability of the fractal dimension as a parameter for describing retinal vessel patterns in ophthalmically normal dogs. PROCEDURES: The following strategy was adopted: (i) development of an experimental procedure to obtain digitalized photographs of the fundus; (ii) development of software to segment retinal vessel images and calculate the box-counting and radius of gyration fractal dimensions of the retinal vessels and diffusion-limited aggregation (DLA), a process with similar characteristics to retinal vessel morphology, and (iii) establishment of a standard curve for the fractal dimensions of segmented vessels. RESULTS: Digitalized photographs of the fundus showed an adequate contrast between the vessels and the rest of the fundus for numerical analysis. The software developed produced a binary image of the retinal vessels permitting calculation of the fractal dimension. The mean values of the fractal dimensions calculated by the methods of box-counting and radius of gyration for the DLA were significantly different (t = -40.33, P approximately 0). The radius of gyration method was found to be more suitable for documenting the dimension of the DLA and, consequently, of the dog's retinal vessels. CONCLUSION: This methodology may be useful to differentiate between normal and pathologic states of canine retinal vascularization.

Theoretical study of dilute GaN-4d transition metal alloys.

Electronic calculations were carried out for the dilute ordered alloys Ga(0.94) (TM)(0.06)N (TM = Y, Zr, Nb, Mo, Tc, Ru, Rh, Pd, and Ag) in the zinc-blende structure. The theoretical framework used was the density functional theory, using the local spin density approximation, as implemented in the full-potential linearized augmented plane wave method. We examine energy band structures, densities of states, charge distributions, and local magnetic moments and anticipate the properties of these promising systems for applications in spin electronic devices.

Hurst analysis applied to the study of single calcium-activated potassium channel kinetics.

The gating of ion channels has been modeled by assuming that the transitions between open and closed states is a memoryless process. Nevertheless, analysis of records of unitary current events suggests that the kinetic process presents short-term memory, i.e. the open- and closed-dwell times are short-term correlated. Here the rescaled range analysis (R/S Hurst analysis) is used as a method to test long-term correlation, in single calcium-activated potassium channels present in Leydig cells. The Hurst coefficients, calculated for four different voltages (V) are: 0.634+/-0.022 (n=3) for V=+20 mV; 0.635+/-0.012 (n=4) for V=+40 mV; 0.606+/-0.020 (n=4) for V=+60 mV and 0.608+/-0.026 (n=4) for V=+80 mV. This indicates that open- and closed-dwell times are long-term correlated and do not change with the voltage applied to the patch at a 5% significance level (F=2.2402;p=0.140715). Randomly shuffling the experimental data removes the correlation in all voltages. When the Hurst method was applied to the results from a simulated three-state Markovian model, it could not account for the long-term correlation found in the experimental data. In this case, H has the following values: 0. 5498+/-0.018 (n=100) for V=+20 mV; 0.5557+/-0.0202 (n=100) for V=+40 mV; 0.5565+/-0.0246 (n=100) for V=60 mV and 0.5595+/-0.0247 (n=100) for V=+80 mV. Even a four-state Markovian model was not adequate to correctly simulate the long-term memory found experimentally, with H values significantly different from those found for the experimental data, in the same voltage range (F=15.0355;p=0.00001). In conclusion, this paper shows that: (1) the open- and closed-dwell times of the single calcium-activated potassium channel of Leydig cells are long-term correlated; (2) three- and four-state Markovian models, which describe very well the dwell time distributions, are not adequate to describe the long-term correlation found between the open and closed states of this ion channel.

Heparin influence on alpha-staphylotoxin formed channel.

The effects of heparin on ion channels formed by Staphylococcus aureus alpha-toxin (ST channel) in lipid bilayers were studied under voltage clamp conditions. Heparin concentrations as small as 100 pM induced a sharp dose-dependent increase in channel voltage sensitivity. This was only observed when heparin was added to the negative-potential side of lipid bilayers in the presence of divalent cations. Divalent cations differ in their efficiency: Zn2+>Ca2+>Mg2+. The apparent positive gating charge increased 2-3-fold with heparin addition as well as with acidification of the bathing solution. 'Free' carboxyl groups and carboxyl groups in ion pairs of the protein moiety are hypothesized to interact with sulfated groups of heparin through divalent cation bridges. The cis mouth of the channel (that protrudes beyond the membrane plane on the side of ST addition and to which voltage was applied) is less sensitive to heparin than the trans-mouth. It is suggested that charged residues which interact with heparin at the cis mouth of ST channels and which contribute to the effective gating charge at negative voltage may be physically different from those at the trans mouth and at positive voltage.

[How can we measure a diameter of each entrance of a ion channel recording only channel conductance?]. / Kak izmerit' diametr kazhdogo vkhoda u ionnogo kanala, regestriruia tol'ko ego provodimost'?

The channel formed by colicin Ia in planar lipid bilayer was used to develop a method determining a diameter of each channel entrance. The method is based on measuring of a channel filling with nonelectrolytes through cis- and trans-entrance separately. To do this, one need to establish the two things only: the conductance of an ion channel and the conductivity of used solutions. Using the method to predict the internal geometry of a channel lumen is discussed.

A novel approach to study the geometry of the water lumen of ion channels: colicin Ia channels in planar lipid bilayers.

This paper describes a new approach to evaluate the inner structure (including a main constriction and its localization) of the water lumen of an ion channel. The method is based on the determination of channel filling by different nonelectrolyte molecules through each side of an ion channel. The method has two characteristic features that make its use attractive: (i) the possibility to ascertain the existence, localization and size of a narrow part inside an ion channel water lumen and (ii) the chances to determine the maximal size of both entrances of an ion channel and to obtain additional information about the geometry of its water lumen at the same time. Determinations were made on colicin Ia ion channels inserted into planar lipid bilayers. This channel was chosen because there is an apparent contradiction between its low single channel conductance and the large diameter of its water lumen. Our results show that the water lumen of the colicin Ia channel has a funnel-like structure with a small trans-entrance, with a diameter of about 1.0 nm, and a large cis-entrance, with a diameter of approximately 1.8 nm. A constriction with a diameter of approximately 0.7 nm is shown to be located close to the trans-entrance of the channel. The method can also be applied to patch clamp studies of single ion channels.

Channel-sizing experiments in multichannel bilayers.

The possibility of obtaining information about the radius of high and low conductance states of channels in multichannel membranes was tested experimentally. In spite of the interference of non-electrolytes on the numbers of channels that appeared in the membrane, the non-electrolyte-exclusion method was successfully adapted to multichannel bilayers to estimate the radius of the larger opening of the low conductance state of the channel induced by Staphylococcus aureus alpha-toxin. At the pH used, the channel transition to a low conductance state was accompanied by a decrease of the opening radius from 1.3 +/- 0.2 nm to 0.9 +/- 0.1 nm. The determination criteria for maximum size of a channel opening when using the non-electrolyte exclusion method is discussed.

The hinge portion of the S. aureus alpha-toxin crosses the lipid bilayer and is part of the trans-mouth of the channel.

This paper compares the functional properties of ion channels formed in planar lipid membranes by the wild and mutant Staphylococcus aureus alpha-toxin. It was shown that replacement of the amino acid Gly at position 130 by Cys in the primary structure of the toxin decreases the single-channel conductance with a concomitant decrease in the pH at which the channel becomes unable to discriminate between Cl- and K+ ions. The mutation also induced an increase in the asymmetry in the current-voltage relationship of the channel. The results of our experiments suggest that the trans-mouth of the channel is responsible for all the observed changes in channel properties. It was assumed that this entrance is built by the glycine-rich hinge portion of the toxin and is situated close to the surface of monolayer facing the trans-compartment.