Dénes Pálfi

Dendritic spikes induce ripples in parvalbumin interneurons during hippocampal sharp waves.

Sharp-wave ripples are transient oscillatory events in the hippocampus that are associated with the reactivation of neuronal ensembles within specific circuits during memory formation. Fast-spiking, parvalbumin-expressing interneurons (FS-PV INs) are thought to provide fast integration in these oscillatory circuits by suppressing regenerative activity in their dendrites. Here, using fast 3D two-photon imaging and a caged glutamate, we challenge this classical view by demonstrating that FS-PV IN dendrites can generate propagating Ca(2+) spikes during sharp-wave ripples. The spikes originate from dendritic hot spots and are mediated dominantly by L-type Ca(2+) channels. Notably, Ca(2+) spikes were associated with intrinsically generated membrane potential oscillations. These oscillations required the activation of voltage-gated Na(+) channels, had the same frequency as the field potential oscillations associated with sharp-wave ripples, and controlled the phase of action potentials. Furthermore, our results demonstrate that the smallest functional unit that can generate ripple-frequency oscillations is a segment of a dendrite.

Local Postsynaptic Voltage-Gated Sodium Channel Activation in Dendritic Spines of Olfactory Bulb Granule Cells

Neuronal dendritic spines have been speculated to function as independent computational units, yet evidence for active electrical computation in spines is scarce. Here we show that strictly local voltage-gated sodium channel (Nav) activation can occur during excitatory postsynaptic potentials in the spines of olfactory bulb granule cells, which we mimic and detect via combined two-photon uncaging of glutamate and calcium imaging in conjunction with whole-cell recordings. We find that local Nav activation boosts calcium entry into spines through high-voltage-activated calcium channels and accelerates postsynaptic somatic depolarization, without affecting NMDA receptor-mediated signaling. Hence, Nav-mediated boosting promotes rapid output from the reciprocal granule cell spine onto the lateral mitral cell dendrite and thus can speed up recurrent inhibition. This striking example of electrical compartmentalization both adds to the understanding of olfactory network processing and broadens the general view of spine function.

Combined two-photon imaging, electrophysiological, and anatomical investigation of the human neocortex in vitro

Spontaneous synchronous population activity (SPA) can be detected by electrophysiological methods in cortical slices of epileptic patients, maintained in a physiological medium in vitro. In order to gain additional spatial information about the network mechanisms involved in the SPA generation, we combined electrophysiological studies with two-photon imaging. Neocortical slices prepared from postoperative tissue of epileptic and tumor patients were maintained in a dual perfusion chamber in a physiological incubation medium. SPA was recorded with a 24-channel extracellular linear microelectrode covering all neocortical layers. After identifying the electrophysiologically active regions of the slice, bolus loading of neuronal and glial markers was applied on the tissue. SPA-related [Formula: see text] transients were detected in a large population of neighboring neurons with two-photon microscopy, simultaneous with extracellular SPA and intracellular whole-cell patch-clamp recordings. The intracellularly recorded cells were filled for subsequent anatomy. The cells were reconstructed in three dimensions and examined with light- and transmission electron microscopy. Combining high spatial resolution two-photon [Formula: see text] imaging techniques and high temporal resolution extra- and intracellular electrophysiology with cellular anatomy may permit a deeper understanding of the structural and functional properties of the human neocortex.

Special Section on the BRAIN Initiative: Combined two-photon imaging, electrophysiological, and anatomical investigation of the human neocortex in vitro

Spontaneous synchronous population activity (SPA) can be detected by electrophysiological methods in cortical slices of epileptic patients, maintained in a physiological medium in vitro. In order to gain additional spatial information about the network mechanisms involved in the SPA generation, we combined electrophysiological studies with two-photon imaging. Neocortical slices prepared from postoperative tissue of epileptic and tumor patients were maintained in a dual perfusion chamber in a physiological incubation medium. SPA was recorded with a 24-channel extracellular linear microelectrode covering all neocortical layers. After identifying the electrophysiologically active regions of the slice, bolus loading of neuronal and glial markers was applied on the tissue. SPA-related [Formula: see text] transients were detected in a large population of neighboring neurons with two-photon microscopy, simultaneous with extracellular SPA and intracellular whole-cell patch-clamp recordings. The intracellularly recorded cells were filled for subsequent anatomy. The cells were reconstructed in three dimensions and examined with light- and transmission electron microscopy. Combining high spatial resolution two-photon [Formula: see text] imaging techniques and high temporal resolution extra- and intracellular electrophysiology with cellular anatomy may permit a deeper understanding of the structural and functional properties of the human neocortex.

[Fast three-dimensional two-photon scanning methods for studying neuronal physiology on cellular and network level].

INTRODUCTION Two-photon microscopy is the ideal tool to study how signals are processed in the functional brain tissue. However, early raster scanning strategies were inadequate to record fast 3D events like action potentials. AIM The aim of the authors was to record various neuronal activity patterns with high signal-to-noise ratio in an optical manner. METHOD Authors developed new data acquisition methods and microscope hardware. RESULTS Multiple Line Scanning enables the experimenter to select multiple regions of interests, doing this not just increases repetition speed, but also the signal-to-noise ratio of the fluorescence transients. On the same principle, an acousto-optical deflector based 3D scanning microscope has been developed with a sub-millisecond temporal resolution and a millimeter z-scanning range. Its usability is demonstrated by obtaining 3D optical recordings of action potential backpropagation in several hundred micrometers long neuronal processes of single neurons and by 3D random-access scanning of Ca(2+) transients in hundreds of neurons in the mouse visual cortex. CONCLUSIONS Region of interest scanning enables high signal-to-noise ratio and repetition speed, while keeping good depth penetration of the two-photon microscopes.

High efficiency two-photon uncaging coupled by the correction of spontaneous hydrolysis

Two-photon (TP) uncaging of neurotransmitter molecules is the method of choice to mimic and study the subtleties of neuronal communication either in the intact brain or in slice preparations. However, the currently available caged materials are just at the limit of their usability and have several drawbacks. The local and focal nature of their use may for example be jeopardized by a high spontaneous hydrolysis rate of the commercially available compounds with increased photochemical release rate. Here, using quantum chemical modelling we show the mechanisms of hydrolysis and two-photon activation, and synthesized more effective caged compounds. Furthermore, we have developed a new enzymatic elimination method removing neurotransmitters inadvertently escaping from their compound during experiment. This method, usable both in one and two-photon experiments, allows for the use of materials with an increased rate of photochemical release. The efficiency of the new compound and the enzymatic method and of the new compound are demonstrated in neurophysiological experiments.

Correction to Local Postsynaptic Voltage-Gated Sodium Channel Activation in Dendritic Spines of Olfactory Bulb Granule Cells [Neuron 85, (2015), 590-601]

(Neuron 85, 590–601; February 4, 2015) In the original publication, Figure 5 depicted the predicted time course of several electrical currents in the granule-cell spine based on computational modeling. In the original panel 5F, the timescale of the voltage trace in the spine head and the current traces were not properly matched; the resulting figure showed that the Ca current flows before the action potential, which is not correct. In addition, the parameter set in Table S4 did not correspond to these traces; the true parameter set was accidentally overwritten. We, therefore, chose a new combination of model parameters. The resulting traces are shown in panels 5B through 5F. Labels and scale bars are as in the original text. The values of the new parameters are listed in Table S4; the star in Figure S3 has been repositioned accordingly. The article and the Supplemental Information have been corrected online. The authors regret the error.