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Acute Slices (acute + slice)
Selected AbstractsSpontaneous recurrent network activity in organotypic rat hippocampal slicesEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2005Majid H. Mohajerani Abstract Organotypic hippocampal slices were prepared from postnatal day 4 rats and maintained in culture for >6 weeks. Cultured slices exhibited from 12 days in vitro spontaneous events which closely resembled giant depolarizing potentials (GDPs) recorded in neonatal hippocampal slices. GDP-like events occurred over the entire hippocampus with a delay of 30,60 ms between two adjacent regions as demonstrated by pair recordings from CA3,CA3, CA3,CA1 and interneurone,CA3 pyramidal cells. As in acute slices, spontaneous recurrent events were generated by the interplay of GABA and glutamate acting on AMPA receptors as they were reversibly blocked by bicuculline and 6,7-dinitroquinoxaline-2,3-dione but not by dl -2-amino-5-phosphonopentaoic acid. The equilibrium potentials for GABA measured in whole cell and gramicidin-perforated patch from interconnected interneurones,CA3 pyramidal cells were ,70 and ,56 mV, respectively. The resting membrane potential estimated from the reversal of N -methyl- d -aspartate-induced single-channel currents in cell-attach experiments was ,75 mV. In spite of its depolarizing action, in the majority of cases GABA was still inhibitory as it blocked the firing of principal cells. The increased level of glutamatergic connectivity certainly contributed to network synchronization and to the development of interictal discharges after prolonged exposure to bicuculline. In spite of its inhibitory action, in a minority of cells GABA was still depolarizing and excitatory as it was able to bring principal cells to fire, suggesting that a certain degree of immaturity is still present in cultured slices. This was in line with the transient bicuculline-induced block of GDPs and with the isoguvacine-induced increase of GDP frequency. [source] Involvement of post-synaptic kainate receptors during synaptic transmission between unitary connections in rat neocortexEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 11 2003Afia B. Ali Abstract The properties of functional kainate receptor-mediated EPSCs were studied in acute slices from 19,35-day-old rats. EPSCs elicited in pyramidal and fast-spiking cells in layers 2/3 and 5 of the rat motor cortex by extracellular single shock stimulus in the presence of GYKI 53655 and D-2-amino-5-phosphopentanoic resulted in a residual current. This current was not enhanced by cyclothiazide but was blocked by 6-cyano-7-nitroquinoxalin-2,3-dione and is thought to be mediated by kainate receptors. These kainate receptor-mediated currents displayed a wide range of time courses depending on which pre-synaptic fibres were activated. With paired recordings, unitary EPSCs elicited in pyramidal cells were almost totally blocked by GYKI 53655 and D-2-amino-5-phosphopentanoic. However, when L-transpyrrolidine-2,4-dicarboxylate (PDC), a glutamate uptake blocker, was introduced in the bath, the amplitude of kainate receptor-mediated currents, which is resistant to GYKI 53655 and D-2-amino-5-phosphopentanoic, was revealed. The rise and decay time constants of the kainate receptor-mediated currents were identical to control EPSCs. PDC was not required to reveal the kainate receptor-mediated currents elicited in fast-spiking cells which also displayed similar rise and decay time constants to the control EPSCs. Excitatory input onto pyramidal and fast-spiking cells in the neocortex mediated by kainate receptors contributed between 14 and 40% of the total control unitary EPSCs which displayed identical time courses to the AMPA receptor-mediated component of the EPSCs. Post-synaptic kainate receptors at connected pyramidal cell synapses may be located extra-synaptically. [source] Innervation of interneurons immunoreactive for VIP by intrinsically bursting pyramidal cells and fast-spiking interneurons in infragranular layers of juvenile rat neocortexEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2002Jochen F. Staiger Abstract Cortical columns contain specific neuronal populations with characteristic sets of connections. This wiring forms the structural basis of dynamic information processing. However, at the single-cell level little is known about specific connectivity patterns. We performed experiments in infragranular layers (V and VI) of rat somatosensory cortex, to clarify further the input patterns of inhibitory interneurons immunoreactive (ir) for vasoactive intestinal polypeptide (VIP). Neurons in acute slices were electrophysiologically characterized using whole-cell recordings and filled with biocytin. This allowed us to determine their firing pattern as regular-spiking, intrinsically bursting and fast-spiking, respectively. Biocytin was revealed histochemically and VIP immunohistochemically. Sections were examined for contacts between the axons of the filled neurons and the VIP-ir targets. Twenty pyramidal cells and five nonpyramidal (inter)neurons were recovered and sufficiently stained for further analysis. Regular-spiking pyramidal cells displayed no axonal boutons in contact with VIP-ir targets. In contrast, intrinsically bursting layer V pyramidal cells showed four putative single contacts with a proximal dendrite of VIP neurons. Fast-spiking interneurons formed contacts with two to six VIP neurons, preferentially at their somata. Single as well as multiple contacts on individual target cells were found. Electron microscopic examinations showed that light-microscopically determined contacts represent sites of synaptic interactions. Our results suggest that, within infragranular local cortical circuits, (i) fast-spiking interneurons are more likely to influence VIP cells than are pyramidal cells and (ii) pyramidal cell input probably needs to be highly convergent to fire VIP target cells. [source] Dual effects of NMDA receptor activation on polysialylated neural cell adhesion molecule expression during brainstem postnatal developmentEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 8 2001Farima Bouzioukh Abstract Here we show a dual role of N -methyl- d -aspartate receptor (NMDAR) activation in controlling polysialylated neural cell adhesion molecule (PSA-NCAM) dynamic expression in the dorsal vagal complex (DVC), a gateway for many primary afferent fibres. In this structure the overall expression of PSA-NCAM decreases during the first 2 weeks after birth to persist only at synapses in the adult. Electrical stimulation of the vagal afferents causes a rapid increase of PSA-NCAM expression both in vivo and in acute slices before postnatal day (P) 14 whereas a similar stimulation induces a decrease after P15. Inhibition of NMDAR activity in vitro completely prevented these changes. These regulations depend on calmodulin activation and cGMP production at all stages. By contrast, blockade of neuronal nitric oxide synthase (nNOS) prevented these changes only after P10 in agreement with its late expression in the DVC. The pivotal role of NMDAR is also supported by the observation that chronic blockade induces a dramatic decrease in PSA-NCAM expression. [source] GABAA -receptor expression in glioma cells is triggered by contact with neuronal cellsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 8 2001Michael Synowitz Abstract The expression of functional GABAA -receptors in glioma cells correlates with low malignancy of tumours and cell lines from glioma lack these receptors. Here we show that contact with neurons induces the expression of functional GABAA -receptors. C6 and F98 glioma cell lines were labelled by recombinant expression of enhanced green fluorescent protein injected into rat brain and studied in acute slices after two to three weeks of tumour growth. The cells responded to GABA or the specific agonist, muscimol with a current typical for GABAA -receptors, as studied with the patch-clamp technique. To get insight into the mechanism of GABAA receptor induction, the C6 or F98 cells were co-cultured with neurons, astrocytes, oligodendrocytes and microglia. Glioma cells expressed functional GABAA receptors within 24 h only in cultures where physical contact to neurons occurred. Activation of GABAA -receptors in the co-cultures attenuated glioma cell metabolism while blockade of the receptors increased metabolism. We conclude that with this form of interaction, neurons can influence tumour behaviour in the brain. [source] Dedifferentiation of intrinsic response properties of motoneurons in organotypic cultures of the spinal cord of the adult turtleEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 7 2000Jean-François Perrier Abstract Explant cultures from the spinal cord of adult turtles were established and used to study the sensitivity of the intrinsic response properties of motoneurons to the changes in connectivity and milieu imposed by isolation in culture. Transverse sections 700 ,m thick were explanted on cover slips and maintained in roller-tube cultures in medium containing serum and the growth factors brain-derived neurotrophin factor (BDNF), neurotrophin-3 (NT3), glial cell line-derived neurotrophic factor (GDNF) and ciliary neurotrophic factor (CNTF). The gross morphology of acute sections was maintained after 4 weeks in culture. Cell bodies of motoneurons remained stainable in fixed cultures with an antibody against choline acetyltransferase (ChAT) throughout the culture period. During culture, motoneurons maintained stable resting membrane potentials and were contacted by functional synapses. The ability to generate action potentials was also preserved as was delayed inward rectification and generation of calcium spikes in the presence of tetra-ethyl ammonium (TEA). In response to depolarization, however, motoneurons presented strong outward rectification, and only 41% of the cells recorded from maintained the ability to fire repetitively. By the second week in culture, a fraction of motoneurons displayed fast and slow transient outward rectification and low-threshold calcium spikes, features not seen in turtle motoneurons in acute slices. On the other hand, properties mediated by L-type Ca2+ channels disappeared during the first few days in culture. Our observations show that the phenotypical intrinsic response properties of mature spinal motoneurons are modified in explant cultures. The properties acquired resemble the properties in juvenile motoneurons in several species of terrestrial vertebrates. [source] Electrical and chemical synapses between relay neurons in developing thalamusTHE JOURNAL OF PHYSIOLOGY, Issue 13 2010Seung-Chan Lee Gap junction-mediated electrical synapses interconnect diverse types of neurons in the mammalian brain, and they may play important roles in the synchronization and development of neural circuits. Thalamic relay neurons are the major source of input to neocortex. Electrical synapses have not been directly observed between relay neurons in either developing or adult animals. We tested for electrical synapses by recording from pairs of relay neurons in acute slices of developing ventrobasal nucleus (VBN) of the thalamus from rats and mice. Electrical synapses were common between VBN relay neurons during the first postnatal week, and then declined sharply during the second week. Electrical coupling was reduced among cells of connexin36 (Cx36) knockout mice; however, some neuron pairs remained coupled. This implies that electrical synapses between the majority of coupled VBN neurons require Cx36 but that other gap junction proteins also contribute. The anatomical distribution of a ,-galactosidase reporter indicated that Cx36 was expressed in some VBN neurons during the first postnatal week and sharply declined over the second week, consistent with our physiological results. VBN relay neurons also communicated via chemical synapses. Rare pairs of relay neurons excited one another monosynaptically. Much more commonly, spikes in one relay neuron evoked disynaptic inhibition (via the thalamic reticular nucleus) in the same or a neighbouring relay neuron. Disynaptic inhibition between VBN cells emerged as electrical coupling was decreasing, during the second postnatal week. Our results demonstrate that thalamic relay neurons communicate primarily via electrical synapses during early postnatal development, and then lose their electrical coupling as a chemical synapse-mediated inhibitory circuit matures. [source] Efficient Ca2+ buffering in fast-spiking basket cells of rat hippocampusTHE JOURNAL OF PHYSIOLOGY, Issue 8 2008Yexica Aponte Fast-spiking parvalbumin-expressing basket cells (BCs) represent a major type of inhibitory interneuron in the hippocampus. These cells inhibit principal cells in a temporally precise manner and are involved in the generation of network oscillations. Although BCs show a unique expression profile of Ca2+ -permeable receptors, Ca2+ -binding proteins and Ca2+ -dependent signalling molecules, physiological Ca2+ signalling in these interneurons has not been investigated. To study action potential (AP)-induced dendritic Ca2+ influx and buffering, we combined whole-cell patch-clamp recordings with ratiometric Ca2+ imaging from the proximal apical dendrites of rigorously identified BCs in acute slices, using the high-affinity Ca2+ indicator fura-2 or the low-affinity dye fura-FF. Single APs evoked dendritic Ca2+ transients with small amplitude. Bursts of APs evoked Ca2+ transients with amplitudes that increased linearly with AP number. Analysis of Ca2+ transients under steady-state conditions with different fura-2 concentrations and during loading with 200 ,m fura-2 indicated that the endogenous Ca2+ -binding ratio was ,200 (,S= 202 ± 26 for the loading experiments). The peak amplitude of the Ca2+ transients measured directly with 100 ,m fura-FF was 39 nm AP,1. At ,23°C, the decay time constant of the Ca2+ transients was 390 ms, corresponding to an extrusion rate of ,600 s,1. At 34°C, the decay time constant was 203 ms and the corresponding extrusion rate was ,1100 s,1. At both temperatures, continuous theta-burst activity with three to five APs per theta cycle, as occurs in vivo during exploration, led to a moderate increase in the global Ca2+ concentration that was proportional to AP number, whereas more intense stimulation was required to reach micromolar Ca2+ concentrations and to shift Ca2+ signalling into a non-linear regime. In conclusion, dentate gyrus BCs show a high endogenous Ca2+ -binding ratio, a small AP-induced dendritic Ca2+ influx, and a relatively slow Ca2+ extrusion. These specific buffering properties of BCs will sharpen the time course of local Ca2+ signals, while prolonging the decay of global Ca2+ signals. [source] Neural Signal Manager: a collection of classical and innovative tools for multi-channel spike train analysisINTERNATIONAL JOURNAL OF ADAPTIVE CONTROL AND SIGNAL PROCESSING, Issue 11 2009Antonio Novellino Abstract Recent developments in the neuroengineering field and the widespread use of the micro electrode arrays (MEAs) for electrophysiological investigations made available new approaches for studying the dynamics of dissociated neuronal networks as well as acute/organotypic slices maintained ex vivo. Importantly, the extraction of relevant parameters from these neural populations is likely to involve long-term measurements, lasting from a few hours to entire days. The processing of huge amounts of electrophysiological data, in terms of computational time and automation of the procedures, is actually one of the major bottlenecks for both in vivo and in vitro recordings. In this paper we present a collection of algorithms implemented within a new software package, named the Neural Signal Manager (NSM), aimed at analyzing a huge quantity of data recorded by means of MEAs in a fast and efficient way. The NSM offers different approaches for both spike and burst analysis, and integrates state-of-the-art statistical algorithms, such as the inter-spike interval histogram or the post stimulus time histogram, with some recent ones, such as the burst detection and its related statistics. In order to show the potentialities of the software, the application of the developed algorithms to a set of spontaneous activity recordings from dissociated cultures at different ages is presented in the Results section. Copyright © 2008 John Wiley & Sons, Ltd. [source] | ||||||||||