Indicadores de cálcio e de voltagem codificados geneticamente na detecção de potenciais de ação e inputs sinápticos em cultura de neurônios hipocampais

Recently, genetically encoded optical indicators have emerged as noninvasive tools of high spatial and temporal resolution utilized to monitor the activity of individual neurons and specific neuronal populations. The increasing number of new optogenetic indicators, together with the absence of comparisons under identical conditions, has generated difficulty in choosing the most appropriate protein, depending on the experimental design. Therefore, the purpose of our study was to compare three recently developed reporter proteins: the calcium indicators GCaMP3 and R-GECO1, and the voltage indicator VSFP butterfly1.2. These probes were expressed in hippocampal neurons in culture, which were subjected to patchclamp recordings and optical imaging. The three groups (each one expressing a protein) exhibited similar values of membrane potential (in mV, GCaMP3: -56 ±8.0, R-GECO1: -57 ±2.5; VSFP: -60 ±3.9, p = 0.86); however, the group of neurons expressing VSFP showed a lower average of input resistance than the other groups (in Mohms, GCaMP3: 161 ±18.3; GECO1-R: 128 ±15.3; VSFP: 94 ±14.0, p = 0.02). Each neuron was submitted to current injections at different frequencies (10 Hz, 5 Hz, 3 Hz, 1.5 Hz, and 0.7 Hz) and their fluorescence responses were recorded in time. In our study, only 26.7% (4/15) of the neurons expressing VSFP showed detectable fluorescence signal in response to action potentials (APs). The average signal-to-noise ratio (SNR) obtained in response to five spikes (at 10 Hz) was small (1.3 ± 0.21), however the rapid kinetics of the VSFP allowed discrimination of APs as individual peaks, with detection of 53% of the evoked APs. Frequencies below 5 Hz and subthreshold signals were undetectable due to high noise. On the other hand, calcium indicators showed the greatest change in fluorescence following the same protocol (five APs at 10 Hz). Among the GCaMP3 expressing neurons, 80% (8/10) exhibited signal, with an average SNR value of 21 ±6.69 (soma), while for the R-GECO1 neurons, 50% (2/4) of the neurons had signal, with a mean SNR value of 52 ±19.7 (soma). For protocols at 10 Hz, 54% of the evoked APs were detected with GCaMP3 and 85% with R-GECO1. APs were detectable in all the analyzed frequencies and fluorescence signals were detected from subthreshold depolarizations as well. Because GCaMP3 is the most likely to yield fluorescence signal and with high SNR, some experiments were performed only with this probe. We demonstrate that GCaMP3 is effective in detecting synaptic inputs (involving Ca2+ influx), with high spatial and temporal resolution. Differences were also observed between the SNR values resulting from evoked APs, compared to spontaneous APs. In recordings of groups of cells, GCaMP3 showed clear discrimination between activated and silent cells, and reveals itself as a potential tool in studies of neuronal synchronization. Thus, our results indicate that the presently available calcium indicators allow detailed studies on neuronal communication, ranging from individual dendritic spines to the investigation of events of synchrony in neuronal networks genetically defined. In contrast, studies employing VSFPs represent a promising technology for monitoring neural activity and, although still to be improved, they may become more appropriate than calcium indicators, since neurons work on a time scale faster than events of calcium may foresee

Efeitos da nicotina no complexo do septo medial na via septo hipocampal em camundongos anestesiados por Ketamina

Nicotine administration in humans and rodents enhances memory and attention, and also has a positive effect in Alzheimer's Disease. The Medial Septum / Diagonal Band of Broca complex (MS/DBB) – a main cholinergic system – massively projects to the hippocampus through the fimbria-fornix, and this pathway is called the septohippocampal pathway. It has been demonstrated that the MS/DBB acts directly on the local field potential (LFP) rhythmic organization of the hippocampus, especially in the rhythmogenesis of Theta (4-8Hz) – an oscillation intrinsically linked to hippocampus mnemonic function. In vitro experiments gave evidence that nicotine applied to the MS/DBB generates a local network Theta rhythm within the MS/DBB. Thus, the present study proposes to elucidate the function of nicotine in the MS/DBB on the septo-hippocampal pathway. In vivo experiments compared the effect of MS/DBB microinfusion of saline (n=5) and nicotine (n=8) on Ketamine/Xylazine anaesthetized mice. We observed power spectrum density in the Gamma range (35 to 55 Hz) increasing in both structures (Wilcoxon Rank-Sum test, p=0.038) but with no change in coherence between these structures in the same range (Wilcoxon Rank-Sum test, p=0.60). There was also a decrease in power of the ketamineinduced Delta oscillation (1 to 3 Hz). We also performed in vitro experiments on the effect of nicotine on membrane voltage and action potential. We patch-clamped 22 neurons in current-clamp mode; 12 neurons were responsive to nicotine, half of them increased firing rate and other 6 decreased, and they significantly differed in action potential threshold (-47.3±0.9 mV vs. -41±1.9 mV, respectively, p=0.007) and halfwidth time (1.6±0.08 ms vs. 2±0.12 ms, respectively, p=0.01). Furthermore, we performed another set of in vitro experiments concerning the connectivity of the three major neuronal populations of MS/DBB that use acetylcholine, GABA or glutamate as neurotransmitter. Paired patch-clamp recordings found that glutamatergic and GABAergic neurons realize intra-septal connections that produce sizable currents in MS/DBB postsynaptic neurons. The probability of connectivity between different neuronal populations gave rise to a MS/DBB topology that was implemented in a realistic model, which corroborates that the network is highly sensitive to the generation of Gamma rhythm. Together, the data available in the full set of experiments suggests that nicotine may act as a cognitive enhancer, by inducing gamma oscillation in the local circuitry of the MS/DBB.

Desenvolvimento de antenas de microfita com Patch em anel utilizando materiais ferrimagnéticos e metamateriais

In general, the materials used as substrates in the project of microstrip antennas are: isotropic, anisotropic dielectrics and ferrimagnetic materials (magnetic anisotropy). The use of ferrimagnetic materials as substrates in microstrip patch antennas has been concentrated on the analysis of antennas with circular and rectangular patches. However, a new class of materials, called metamaterials, has been currently the focus of a great deal of interest. These materials exhibit bianisotropic characteristics, with permittivity and permeability tensors. The main objective of this work is to develop a theoretical and numerical analysis for the radiation characteristics of annular ring microstrip antennas, using ferrites and metamaterials as substrates. The full wave analysis is performed in the Hankel transform domain through the application of the Hertz vector potentials. Considering the definition of the Hertz potentials and imposing the boundary conditions, the dyadic Green s function components are obtained relating the surface current density components at the plane of the patch to the electric field tangential components. Then, Galerkin s method is used to obtain a system of matrix equations, whose solution gives the antenna resonant frequency. From this modeling, it is possible to obtain numerical results for the resonant frequency, radiation pattern, return loss, and antenna bandwidth as a function of the annular ring physical parameters, for different configurations and substrates. The theoretical analysis was developed for annular ring microstrip antennas on a double ferrimagnetic/isotropic dielectric substrate or metamaterial/isotropic dielectric substrate. Also, the analysis for annular ring microstrip antennas on a single ferrimagnetic or metamaterial layer and for suspended antennas can be performed as particular cases

Antenas de microfita com patch quase-fractal para aplicações em Redes WPAN/WLAN

The microstrip antennas are in constant evidence in current researches due to several advantages that it presents. Fractal geometry coupled with good performance and convenience of the planar structures are an excellent combination for design and analysis of structures with ever smaller features and multi-resonant and broadband. This geometry has been applied in such patch microstrip antennas to reduce its size and highlight its multi-band behavior. Compared with the conventional microstrip antennas, the quasifractal patch antennas have lower frequencies of resonance, enabling the manufacture of more compact antennas. The aim of this work is the design of quasi-fractal patch antennas through the use of Koch and Minkowski fractal curves applied to radiating and nonradiating antenna s edges of conventional rectangular patch fed by microstrip inset-fed line, initially designed for the frequency of 2.45 GHz. The inset-fed technique is investigated for the impedance matching of fractal antennas, which are fed through lines of microstrip. The efficiency of this technique is investigated experimentally and compared with simulations carried out by commercial software Ansoft Designer used for precise analysis of the electromagnetic behavior of antennas by the method of moments and the neural model proposed. In this dissertation a study of literature on theory of microstrip antennas is done, the same study is performed on the fractal geometry, giving more emphasis to its various forms, techniques for generation of fractals and its applicability. This work also presents a study on artificial neural networks, showing the types/architecture of networks used and their characteristics as well as the training algorithms that were used for their implementation. The equations of settings of the parameters for networks used in this study were derived from the gradient method. It will also be carried out research with emphasis on miniaturization of the proposed new structures, showing how an antenna designed with contours fractals is capable of a miniaturized antenna conventional rectangular patch. The study also consists of a modeling through artificial neural networks of the various parameters of the electromagnetic near-fractal antennas. The presented results demonstrate the excellent capacity of modeling techniques for neural microstrip antennas and all algorithms used in this work in achieving the proposed models were implemented in commercial software simulation of Matlab 7. In order to validate the results, several prototypes of antennas were built, measured on a vector network analyzer and simulated in software for comparison

Arranjo de antenas de microfita com substrato anisotrópico com patch supercondutor e aplicações em nanotecnologia

The main objective in this work is the analysis of resonance frequency microstrip structures with glass fiber and electromagnetic band gap (EBG/PBG) substrate and analysis of microstrip antennas with rectangular patch of superconductor of high critical temperature (HTS). In this work was used the superconductors YBCO (critical temperature of 90K), SnBaCaCuOy (critical temperature of 160K), and Sn5InCa2Ba4Cu10Oy (critical temperature of 212K) with results in Gigahertz and Terahertz. Was used microstrip antennas arrays planar and linear phase and linear phase planar with patch with superconductor. It presents a study of the major theories that explain superconductivity. In phase arrays were obtained the factors arrays for such configurations, and the criteria of phase and spacing between the elements compound in the array, which were examined in order to get a main lobe with high directivity and high gain. In the analysis we used the method of Transverse Transmission Line (TTL) used in domain of the Fourier Transform (FTD). The LTT is a full wave method, which obtains the electromagnetic field in terms of the components transverse of the structure. The addition of superconductive patch is made using the boundary condition resistive complex. Results are obtained resonance frequency as a function of the parameters of the antenna, radiation patterns of the E and H Planes, for the phase antenna arrays in linear and planar configurations, for different values of the phase and the spacing between elements

Aplicações de arranjos de antenas de microfita com patch supercondutor

This work has as main objective the study of arrays of microstrip antennas with superconductor rectangular patch. The phases and the radiation patterns are analyzed. A study of the main theories is presented that explain the microscopic and macroscopic phenomena of superconductivity. The BCS, London equations and the Two Fluid Model, are theories used in the applications of superconductors, at the microstrip antennas and antennas arrays. Phase Arrangements will be analyzed in linear and planar configurations. The arrangement factors of these configurations are obtained, and the phase criteria and the spacing between the elements, are examined in order to minimize losses in the superconductor, compared with normal conductors. The new rectangular patch antenna, consist of a superconducting material, with the critical temperature of 233 K, whose formula is Tl5Ba4Ca2Cu9Oy, is analyzed by the method of the Transverse nTransmission Line (TTL), developed by H. C. C. Fernandes, applied in the Fourier Transform Domain (FTD). The TTL is a full-wave method, which has committed to obtaining the electromagnetic fields in terms of the transverse components of the structure. The inclusion of superconducting patch is made using the complex resistive boundary condition, using the impedance of the superconductor in the Dyadic Green function, in the structure. Results are obtained from the resonance frequency depending on the parameters of the antenna using superconducting material, radiation patterns in E-Plane and H -Plane, the phased antennas array in linear and planar configurations, for different values of phase angles and different spacing between the elements

Antenas monopolo planar com patch em anel circular para sistemas UWB

The microstrip antennas are largely used in wireless communication systems due to their low cost, weight, less complex construction and manufacturing, in addition to its versatility. UWB systems have emerged as an alternative to wireless communications over short distances because they offer of higher capacity and lower multipath distortion than other systems with the same purpose. Combining the advantages of microstrip antennas to the characteristics of UWB, it is possible to develop more and more smaller devices, with diverse geometries to operate satisfactorily in these systems. This paper aims to propose alternatives to microstrip antennas for UWB systems operate in the range between 3.1 and 10.6 GHz, with a patch on circular ring. Some techniques are analyzed and employed to increase the bandwidth of proposed antenna: the insertion of a parasitic elements and a rectangular slit in the displaced ground plane. For this, key issues are presented as the basic principles of UWB systems, the fundamental theory of antennas and microstrip antennas. The simulations and experimental characterization of constructed antennas are presented, as well as analysis of parameters such as bandwidth and radiation pattern

Antenas de microfita com patch supercondutor a 212 K

This work has as main objective to study the application of microstrip antennas with patch and use of superconducting arrays of planar and linear phase. Was presented a study of the main theories that explain clearly the superconductivity. The BCS theory, Equations of London and the Two Fluid Model are theories that supported the implementation of the superconducting microstrip antennas. Arrangements phase was analyzed in linear and planar configuration of its antennas are reported factors such arrays to settings and criteria of phase and the spacing between the elements that make the arrayst was reviewed in order to minimize losses due to secondary lobes. The antenna used has a rectangular patch Sn5InCa2Ba4Cu10Oy the superconducting material was analyzed by the method of Transverse Transmission Line (TTL) applied in the field of Fourier transform (FTD). The TTL is a full-wave method, which has committed to obtaining the electromagnetic fields in terms of cross-cutting components of the structure. The inclusion of superconducting patch is made using the boundary condition, complex resistive. Are obtained when the resonant frequency depending on the parameters of the antenna, radiation pattern of E-Plan and H-Plan for the M-phase arrangements of antennas in the linear and planar configurations for different values of phase and spacing between the elements.

Antenas de microfita com patch em anel e múltiplas camadas dielétricas anisotrópicas uniaxiais

This work presents an analysis of the annular ring microstrip antennas printed on uniaxial anisotropic substrates and with superstrate.The analysis uses the full-wave formulation by means of the Hertz vector potentials method, in the Hankel transform domain. The definition of the Hertz vector potentials and the application of the appropriate boundary conditions to the structure allow determining the dyadic Green functions, relating the current densities in the conducting patch to the transforms of the tangential electric field components. Galerkin s method is then used to obtain the matrix equation whose nontrivial solution gives the complex resonant frequency of the antenna. From the modeling, it is possible to obtain results for the resonant frequency, bandwidth and quality factor, as a function of several parameters of the antenna, for different configurations. We have considered annular ring microstrip antennas on a single dielectric layer, antennas with two anisotropic dielectric layers, and annular ring microstrip antennas on suspended substrates. Numerical results for the resonant frequency of the these structures printed on isotropic substrates are also presented and compared with those published by other authors, showing a good agreement

Estudo de arranjos de antenas de microfita com Patch quase-fractal para comunicações sem fio

In this dissertation, are presented two microstrip antennas and two arrays for applications in wireless communication systems multiband. Initially, we studied an antenna and a linear array consisting of two elements identical to the patch antenna isolated. The shape of the patch used in both structures is based on fractal geometry and has multiband behavior. Next a new antenna is analyzed and a new array such as initial structure, but with the truncated ground plane, in order to obtain better bandwidths and return loss. For feeding the structures, we used microstrip transmission line. In the design of planar structures, was used HFSS software for the simulation. Next were built and measures electromagnetic parameters such as input impedance and return loss, using vector network analyzer in the telecommunications laboratory of Federal University of Rio Grande do Norte. The experimental results were compared with the simulated and showed improved return loss for the first array and also appeared a fourth band and increased directivity compared with the isolated antenna. The first two benefits are not commonly found in the literature. For structures with a truncated ground planes, the technique improved impedance matching, bandwidth and return loss when compared to the initial structure with filled ground planes. Moreover, these structures exhibited a better distribution of frequency, facilitating the adjustment of frequencies. Thus, it is expected that the planar structures presented in this study, particularly arrays may be suitable for specific applications in wireless communication systems when frequency multiband and wideband transmission signals are required.