Biology subjects

Papagiakoumou, E.

Publications and source records attributed to Papagiakoumou, E..

6 recordsLinked to original sources

bioRxiv2025

The Rod Bipolar Cell Pathway Contributes To Surround Responses In OFFRetinal Ganglion Cells

Sensory neurons can be influenced by stimuli beyond their receptive field center, yet the mechanisms underlying this surround modulation remain poorly understood. In the retina, many OFF ganglion cells exhibit responses to ON stimulation outside their receptive field center. However, disentangling the pathways and cell types contributing to these responses has been challenging with traditional experimental approaches. Here, we combined optogenetics, two-photon holographic stimulation, and multi-electrode array recordings to identify the intermediate retinal cell types involved in this circuit. We found that the pathway formed by rod bipolar cells and AII amacrine cells--one of the primary relays of rod-driven signals under low-light conditions--plays a key role in mediating this surround modulation. Specifically, crossover inhibition exploits the same amacrine cells responsible for surround suppression to disinhibit distant ganglion cells. This suggests that the retina repurposes existing circuits for surround modulation, optimizing resources through multifunctional inhibitory pathways.

neuroscience↗

bioRxiv2024

Two-photon voltage imaging with rhodopsin-based sensors

The recent advances in sophisticated optical techniques, coupled with two-photon sensitive genetic voltage indicators (GEVIs), have enabled in-depth voltage imaging in vivo at single spike and single-cell resolution. To date, these results have been only achieved using ASAP-type sensors, as the complex photocycle of rhodopsin-based voltage indicators posed challenges for their two-photon use, restricting their application to one-photon approaches. In this work, we demonstrate that rhodopsin-based GEVIs (FRET-opsin) can be used under two-photon illumination when their peculiar light intensity dependence of kinetics and sensitivity are considered. We rationally engineer a fully genetically-encoded, rhodopsin-based voltage indicator with the brightest known fluorophore AaFP1, Jarvis, and demonstrate its utility under both one- and two-photon illumination. We also showed two-photon usability of the similar FRET-opsin sensor pAce. Our comparison of 2P scanless with fast 2P scanning illumination revealed that the latter approach is less suitable for this class of indicators and, on the contrary, both sensors responded well when scanless approaches were used. Furthermore, utilising Jarvis, we demonstrated high-fidelity, high-SNR action potential detection at kilohertz-imaging rates both in mouse hippocampal slices and in zebrafish larvae. To the best of our knowledge, this study represents the first report of a fully genetically-encoded rhodopsin-based voltage indicator for high contrast action potential detection under two-photon illumination in vitro and in vivo.

neuroscience↗

bioRxiv2019

Multiplexed temporally focused light shaping through a GRIN lens for precise in-depth optogenetic photostimulation

In the past 10 years, the use of light has become irreplaceable for the optogenetic study and control of neurons and neural circuits. Optical techniques are however limited by scattering and can only see through a depth of few hundreds {micro}m in living tissues. GRIN lens based micro-endoscopes represent a powerful solution to reach deeper regions. In this work we demonstrate that cutting edge optical methods for the precise photostimulation of multiple neurons in three dimensions can be performed through a GRIN lens. By spatio-temporally shaping a laser beam in the two-photon regime we project several tens of targets, spatially confined to the size of a single cell, in a volume of 150x150x400 m3. We then apply such concept to the optogenetic stimulation of multiple neurons simultaneously in vivo in mice. Our work paves the way for an all-optical investigation of neural circuits at previously unattainable depths.

neuroscience↗

bioRxiv2019

All-optical interrogation of a direction selective retinal circuit by holographic wave front shaping

Direction selective (DS) ganglion cells (GC) in the retina maintain their tuning across a broad range of light levels. Yet very different circuits can shape their responses from bright to dim light, and their respective contributions are difficult to tease apart. In particular, the contribution of the rod bipolar cell (RBC) primary pathway, a key player in dim light, is unclear. To understand its contribution to DSGC response, we designed an all-optical approach allowing precise manipulation of single retinal neurons. Our system activates single cells in the bipolar cell (BC) layer by two-photon (2P) temporally focused holographic illumination, while recording the activity in the ganglion cell layer by 2P Ca2 imaging. By doing so, we demonstrate that RBCs provide an asymmetric input to DSGCs, suggesting they contribute to their direction selectivity. Our results suggest that every circuit providing an input to direction selective cells can generate direction selectivity by itself. This hints at a general principle to achieve robust selectivity in sensory areas.

neuroscience↗

bioRxiv2018

Parallel holographic illumination enables sub-millisecond two-photon optogenetic activation in mouse visual cortex in vivo

Selective control of action potential generation in individual cells from a neuronal ensemble is desirable for dissecting circuit mechanisms underlying perception and behavior. Here, by using two-photon (2P) temporally focused computer-generated holography (TF-CGH), we demonstrate optical manipulation of neuronal excitability at the supragranular layers of anesthetized mouse visual cortex. Utilizing amplified laser-pulses delivered via a localized holographic spot, our optical system achieves suprathreshold activation by exciting either of the three optogenetic actuators, ReaChR, CoChR or ChrimsonR, with brief illumination ([≤] 10 ms) at moderate excitation power ((in average [≤] 0.2 mW/m2 corresponding to [≤] 25 mW/cell). Using 2P-guided whole-cell or cell-attached recordings in positive neurons expressing respective opsin in vivo, we find that parallel illumination induces spikes of millisecond temporal resolution and sub-millisecond precision, which are preserved upon repetitive illuminations up to tens of Hz. Holographic stimulation thus enables temporally precise optogenetic activation independently of opsins channel kinetics. Furthermore, we demonstrate that parallel optogenetic activation can be combined with functional imaging for all-optical control of a neuronal sub-population that co-expresses the photosensitive opsin ReaChR and the calcium indicator GCaMP6s. Parallel optical control of neuronal activity with cellular resolution and millisecond temporal precision should be advantageous for investigating neuronal connections and further yielding causal links between connectivity, microcircuit dynamics, and brain functions.

neuroscience↗

bioRxiv2017

Multiplexed temporally focused light shaping for high-resolutionmulti-cell targeting

Patterning light at the single-cell level over multiple neurons in the brain is crucial for optogenetic photostimulation that can recapitulate natural activity patterns and, thereby, determine the role of specific components of brain activity in behavior. To this end we have developed a method for projecting three-dimensional, 2-photon excitation patterns that are confined to many individual neurons. The new versatile optical scheme generates multiple extended excitation spots in a large volume with micrometric lateral and axial resolution. Two-dimensional temporally focused shapes are multiplexed several times over selected positions, thanks to the precise spatial phase modulation of the pulsed beam. This permits, under multiple configurations, the generation of tens of axially confined spots in an extended volume, spanning a range in depth of up to 500 m. We demonstrate the potential of the approach by performing multi-cell volumetric excitation of photoactivatable GCaMP in the central nervous system of Drosophila larvae, a challenging structure with densely arrayed and small diameter neurons, and by photoconverting the fluorescent protein Kaede in zebrafish larvae. Our technique paves the way for the optogenetic manipulation of a large number of neurons in intact circuits.

neuroscience↗