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- Ultrasensitive gold micro-structured electrodes enabling the detection of extra-cellular long-lasting potentials in astrocytes populationsPublication . Mestre, Ana L. G.; Cerquido, Monica; INÁCIO, PEDRO; Asgarifar, Sanaz; Lourenco, Ana S.; Lurdes S. Cristiano, M.; Aguiar, Paulo; Medeiros, Maria C. R.; Araújo, Inês; Ventura, Joao; Gomes, Henrique L.Ultra-sensitive electrodes for extracellular recordings were fabricated and electrically characterized. A signal detection limit defined by a noise level of 0.3-0.4 mu V for a bandwidth of 12.5 Hz was achieved. To obtain this high sensitivity, large area (4 mm(2)) electrodes were used. The electrode surface is also micro-structured with an array of gold mushroom-like shapes to further enhance the active area. In comparison with a flat gold surface, the micro-structured surface increases the capacitance of the electrode/electrolyte interface by 54%. The electrode low impedance and low noise enable the detection of weak and low frequency quasi-periodic signals produced by astrocytes populations that thus far had remained inaccessible using conventional extracellular electrodes. Signals with 5 mu V in amplitude and lasting for 5-10 s were measured, with a peak-to-peak signal-to-noise ratio of 16. The electrodes and the methodology developed here can be used as an ultrasensitive electrophysiological tool to reveal the synchronization dynamics of ultra-slow ionic signalling between non-electrogenic cells.
- Extracellular electrophysiological based sensor to monitor cancer cells cooperative migration and cell-cell connectionsPublication . Asgarifar, Sanaz; Mestre, Ana L. G.; Félix, Rute; Inacio, Pedro M. C.; Lurdes S. Cristiano, M.; Medeiros, Maria C. R.; Araújo, Inês; Power, Deborah; Gomes, Henrique L.Herein, we describe an electrophysiological based sensor that reproducibly monitors and quantifies in real-time collective migration and the formation of cell-cell junctions by C6 glioma cells seeded on top of electrodes. The signal amplitude and frequency generated by the migrating cells changed over time and these parameters were used to accurately calculate the migration speed. Electrophysiological measurements could also distinguish individual from collective cell migration. The migration of densely packed cells generated strong signals, while dispersed cells showed weak bioelectrical activity. We propose this electrophysiological technique as a cell-based biosensor to gain insight into the mechanisms of cooperative migration of cancer cells. Possible applications include screening for anti-migratory compounds, which may lead to the development of novel strategies for antineoplastic chemotherapy.
- Extracellular electrical recording of pH-triggered bursts in C6 glioma cell populationsPublication . Rocha, Paulo R. F.; Medeiros, Maria C. R.; Kintzel, Ulrike; Vogt, Johannes; Araújo, Inês; Mestre, Ana L. G.; Mailaender, Volker; Schlett, Paul; Droege, Melanie; Schneider, Leonid; Biscarini, Fabio; de Leeuw, Dago M.; Gomes, Henrique L.Glioma patients often suffer from epileptic seizures because of the tumor's impact on the brain physiology. Using the rat glioma cell line C6 as a model system, we performed long-term live recordings of the electrical activity of glioma populations in an ultrasensitive detection method. The transducer exploits large-area electrodes that maximize double-layer capacitance, thus increasing the sensitivity. This strategy allowed us to record glioma electrical activity. We show that although glioma cells are nonelectrogenic, they display a remarkable electrical burst activity in time. The low-frequency current noise after cell adhesion is dominated by the flow of Na+ ions through voltage-gated ion channels. However, after an incubation period of many hours, the current noise markedly increased. This electric bursting phenomenon was not associated with apoptosis because the cells were viable and proliferative during the period of increased electric activity. We detected a rapid cell culture medium acidification accompanying this event. By using specific inhibitors, we showed that the electrical bursting activity was prompted by extracellular pH changes, which enhanced Na+ ion flux through the psalmotoxin 1-sensitive acid-sensing ion channels. Our model of pH-triggered bursting was unambiguously supported by deliberate, external acidification of the cell culture medium. This unexpected, acidosis-driven electrical activity is likely to directly perturb, in vivo, the functionality of the healthy neuronal network in the vicinity of the tumor bulk and may contribute to seizures in glioma patients.
- Performance assessment of polymer based electrodes for in vitro electrophysiological sensing: the role of the electrode impedancePublication . Medeiros, Maria C. R.; Mestre, Ana L. G.; INÁCIO, PEDRO; Santos, João M. L.; Araújo, Inês; Bragança, José; Biscarini, Fabio; Gomes, Henrique L.Conducting polymer electrodes based on poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) are used to record extracellular signals from autonomous cardiac contractile cells and glioma cell cultures. The performance of these conducting polymer electrodes is compared with Au electrodes. A small-signal impedance analysis shows that in the presence of an electrolyte, both Au and polymer electrodes establish high capacitive double-layers. However, the polymer/electrolyte interfacial resistance is 3 orders of magnitude lower than the resistance of the metal/electrolyte interface. The polymer low interfacial resistance minimizes the intrinsic thermal noise and increases the system sensitivity. However, when measurements are carried out in current mode a low interfacial resistance partially acts as a short circuit of the interfacial capacitance, this affects the signal shape.
- Extracellular electrophysiological measurements of cooperative signals in astrocytes populationsPublication . Mestre, Ana L. G.; INÁCIO, PEDRO; ELAMINE, Youssef; Asgarifar, Sanaz; Lourenco, Ana S.; Cristiano, Maria L. S.; Aguiar, Paulo; Medeiros, Maria C. R.; Araújo, Inês; Ventura, Joao; Gomes, Henrique L.Astrocytes are neuroglial cells that exhibit functional electrical properties sensitive to neuronal activity and capable of modulating neurotransmission. Thus, electrophysiological recordings of astroglial activity are very attractive to study the dynamics of glial signaling. This contribution reports on the use of ultra-sensitive planar electrodes combined with low noise and low frequency amplifiers that enable the detection of extracellular signals produced by primary cultures of astrocytes isolated from mouse cerebral cortex. Recorded activity is characterized by spontaneous bursts comprised of discrete signals with pronounced changes on the signal rate and amplitude. Weak and sporadic signals become synchronized and evolve with time to higher amplitude signals with a quasi-periodic behavior, revealing a cooperative signaling process. The methodology presented herewith enables the study of ionic fluctuations of population of cells, complementing the single cells observation by calcium imaging as well as by patch-clamp techniques.