Biology subjects

Mathieu, B.

Publications and source records attributed to Mathieu, B..

12 recordsLinked to original sources

bioRxiv2026

Streamlined Production of Recombinant Adeno-Associated Viruses Using Gateway Cloning Technology for Neural Circuit All-Optical Interrogation

The advent of optogenetics and the rapid development of genetically encoded actuators and reporters for calcium, voltage, neurotransmitters, and other molecules have revolutionized neuroscience research by enabling precise, non-invasive optical interrogation of neural circuits. Successful implementation, however, critically depends on efficient reporter expression in defined neuronal populations. We present a Gateway(R)-based cloning platform providing a robust pipeline for rapid, efficient construction of recombinant adeno-associated virus (rAAV) vectors optimized for neuroscience applications. This platform addresses promoter selection, indicator engineering, and capsid-type optimization, and enables systematic optimization of vector components across a range of optical tools. The benefit of this modular approach is illustrated in the case of ULoVE, a two-photon microscopy method based on acousto-optic deflectors (AODs). This method provides serial light-targeting with kHz sampling rates and high signal-to-noise ratio in vivo, which imposes stringent requirements on indicator expression -- including cell-type specificity, sparse labelling, and precise control of expression levels-- efficiently met through the combinatorial flexibility of the Gateway pipeline. As specific examples, Gateway-constructed Cre-driver viruses combined with Cre-dependent reporters enabled cell-type-specific labelling for two complementary applications. In cerebellar Purkinje cells, an L7::Cre driver virus paired with a Gateway-constructed voltage indicator (JEDI2P-Kv) and ULoVE two-photon excitation at ~5 kHz enables resolution of sub-millisecond dendritic voltage dynamics in awake, behaving mice, including optical detection of dendritic spikelets previously accessible only via intracellular electrophysiology. The same driver virus paired with calcium indicators (GCaMP6f, jRGECO1a) resolves climbing fiber-evoked calcium kinetics under the same conditions. In acute cerebellar slices, a kit::Cre driver virus enabled cell-type-specific expression of the optogenetic actuator ChR2(H134R) in molecular layer interneurons, where ULoVE doughnut-pattern photostimulation achieved single-cell, sub-millisecond optogenetic activation with micron-scale spatial resolution. Beyond ULoVE applications, the same sparse, strong labelling also proved suitable for anatomical tracing of axonal projections in cleared cerebellar tissue, resolving individual DCN axon terminals at the granule cell layer --molecular layer boundary. Taken together, our dual-front approach --combining a modular AAV expression pipeline with AOD-based optical acquisition --provides an integrated platform for developing sophisticated optical tools, including but not limited to AOD-based microscopy, for neural circuit interrogation.

neuroscience↗

bioRxiv2026

A versatile, positive-going voltage indicator that enables accessible two-photon recordings in vivo

Genetically encoded voltage indicators (GEVIs) enable cell-type-specific optical readout of membrane potential, but two-photon (2P) spike detection has been hampered by low signal-to-noise and ultrafast off-kinetics, restricting use to specialized microscopes. We introduce FORCE1s, a green, positive-going GEVI engineered to make robust 2P voltage imaging broadly accessible. FORCE1s brightens from a dark baseline during depolarization, reports spikes with [~]100% {Delta}F/F in awake mice, and displays repolarization kinetics that are tuned for reliable spike detection at sub-kilohertz frame rates. As a result, FORCE1s supports spike-resolved multi-cell recordings on standard resonant-scanning microscopes, and further scales to larger fields of view and neuron counts on advanced modalities. FORCE1s also enables multiplexed voltage-neurotransmitter imaging and extended recordings in freely moving mice using a compact, affordable MEMS-based 2P miniscope. Together, these advances establish FORCE1s as a community-ready tool that democratizes deep-tissue voltage imaging across platforms and experimental contexts.

neuroscience↗

bioRxiv2026

Temporal Focusing for Enhanced Background Rejection in AOD-Based Two-Photon Serial Holography

The ability to record 3D neuronal activity with cellular resolution, high signal-to-noise ratio (SNR) and millisecond temporal resolution is a major challenge in neuroscience. One powerful method is random-access two-photon microscopy based on acousto-optic deflectors (AODs), which uses a holographically-shaped point spread function (PSF) scanned in 3D to maximize the sampling rate and SNR. However, this approach suffers from greater background contamination due to the holographically shaped PSF than standard two-photon microscopy with diffraction-limited PSF. To overcome this limitation, we implemented a new version of an AOD scanning system, which integrates temporal focusing. The complex spatiotemporal distortions encountered in this configuration, including a significant group delay dispersion associated with the pulse front tilt generated by the AOD, were compensated for by introducing an acousto-optic modulator before the AOD. We designed extended patterns by combining temporal focusing on one direction and holographic wavefront shaping in the perpendicular axis. Taking advantage of the AODs ability to shape the wavefront at the same speed as the scan, we were able to accurately superimpose the spatial and temporal foci over the entire field of view. Finally, we generated complex, extended two-photon excitation patterns by combining temporal focusing in one direction and holographic multiplexing in the perpendicular direction. These patterns provide significantly improved background rejection compared to 2D holographic patterns, thus offering promising prospects for in vivo recordings of neuronal activity in dense samples with improved SNR.

biophysics↗

bioRxiv2025

Human mitochondrial DNA variants influence telomere length: evidence from a transmitochondrial cybrid model

Telomere shortening is a hallmark of aging, yet telomere length (TL) varies considerably among individuals and is strongly influenced by inheritance. In mice, efficient mitochondrial function-characterized by low reactive oxygen species (ROS) production-is critical for telomere elongation during early embryogenesis. Since mitochondrial DNA (mtDNA) encodes several subunits of the electron transport chain, it may influence TL at birth by regulating mitochondrial function in utero. To explore the relationship between mtDNA and TL in human, we used a transmitochondrial cybrid approach, introducing mitochondria from donor platelets with varying telomere lengths into mtDNA-depleted cells. This revealed an inverse correlation between donor blood TL and mitochondrial ROS levels measured in the resulting cybrids, suggesting that specific mtDNA variants may contribute to the maintenance of long telomeres in humans by enhancing mitochondrial fitness. During in vitro cybrid formation, a transient phase of oxidative stress precedes cellular adaptation. In this specific window, mtDNA variants associated with reduced complex I (CI) activity induced rapid telomere shortening--an effect rescued by antioxidant and NAD precursor supplementation. While these variants occur naturally in certain individuals with long telomeres, our data suggest that, at least under in vitro conditions of acute oxidative stress, CI is critical to support PARP1 activity by maintaining the NAD/NADH balance, thereby preserving telomere integrity. Collectively, these findings solidify the link between mtDNA variants and human TL regulation, highlighting potential therapeutic opportunities for mitochondrial replacement strategies. Significance StatementTelomere length at birth influences aging trajectories and disease risk later in life, yet the mechanisms governing this trait remain incompletely understood. Using a transmitochondrial cybrid approach, we show that single-nucleotide variants in the mitochondrial genome of healthy donors directly affect mitochondrial metabolism and reactive oxygen species production. In addition, mitochondrial ROS levels measured in cybrids inversely correlate with blood cell telomere length in donors. During cybrid formation, mitochondrial DNA variants associated with reduced CI activity promote telomere shortening. Attrition was reversed by antioxidant and NAD precursor supplementation, pointing to an essential role for robust CI function in sustaining telomere length during acute oxidative stress, at least under in vitro conditions. Together, these findings establish a direct link between mitochondrial genetics, redox homeostasis, and telomere maintenance in human cells.

cell biology↗

bioRxiv2024

A novel rhodopsin-based voltage indicator for simultaneous two-photon optical recording with GCaMP in vivo

Genetically encoded voltage indicators (GEVIs) allow optical recording of membrane potential from targeted cells in vivo. However, red GEVIs that are compatible with two-photon microscopy and that can be multiplexed in vivo with green reporters like GCaMP, are currently lacking. To address this gap, we explored diverse rhodopsin proteins as GEVIs and engineered a novel GEVI, 2Photron, based on a rhodopsin from the green algae Klebsormidium nitens. 2Photron, combined with two photon ultrafast local volume excitation (ULoVE), enabled multiplexed readout of spiking and subthreshold voltage simultaneously with GCaMP calcium signals in visual cortical neurons of awake, behaving mice. These recordings revealed the cell-specific relationship of spiking and subthreshold voltage dynamics with GCaMP responses, highlighting the challenges of extracting underlying spike trains from calcium imaging.

neuroscience↗

bioRxiv2024

Large-Scale COI Gene Data Analysis for the Development of a Gel-Based Multiplex PCR Test Capable of Identifying Biting Midge Vector Species and Haplotypes (Diptera: Ceratopogonidae) of the Culicoides subgenus Avaritia Fox, 1955

The emergence of culicoid-transmitted bluetongue and Schmallenberg viruses in several European countries demonstrated the ability of indigenous biting midge species to transmit pathogens. Entomologic research programmes identified members of the Obsoletus Group (Culicoides subgenus Avaritia) as keyplayers in disease epidemiology in Europe. However, morphological identification of potential vectors to species level is challenging due to the existence of isomorphic species. PCR tests developed to facilitate genetic species determination have been dismantled by the discovery of new genetic variants (haplotypes) of C. obsoletus sensu stricto (s.s.), forming distinct clades. In this study, 4,422 GenBank entries of the mitochondrial cytochrome c oxidase subunit I (COI) gene of subgenus Avaritia members of the genus Culicoides were analyzed to develop a conventional multiplex PCR, capable of detecting all vector species and recently described clades of the western Palaearctic in this subgenus. Numerous GenBank entries incorrectly assigned to a species were identified, analyzed and reassigned. The results suggest that the three clades of C. obsoletus represent independent species, whereas C. montanus should rather be regarded as a genetic variant of C. obsoletus s.s.. Based on these findings, specific primers were designed and validated with DNA material from field-caught biting midges which achieved very high diagnostic sensitivity (100%) when compared to an established reference PCR (82.6%). Hence, the newly developed multiplex PCR represents the first molecular tool which enables both the identification of accepted species and of the three clades of C. obsoletus s.s. and could provide new insights into the ecology of the latter.

genetics↗

bioRxiv2023

Identification and organization of a postural anti-gravity module in the cerebellar vermis

The cerebellum is known to control the proper balance of isometric muscular contractions that maintain body posture. Current optogenetic manipulations of the cerebellar cortex output, however, have focused on ballistic body movements, examining movement initiation or perturbations. Here, by optogenetic stimulations of cerebellar Purkinje cells, the output of the cerebellar cortex, we evaluate body posture maintenance. By sequential analysis of body movement, we dissect the effect of optogenetic stimulation into a directly induced movement that is then followed by a compensatory reflex to regain body posture. We identify a module in the medial part of the anterior vermis which, through multiple muscle tone regulation, is involved in postural anti-gravity maintenance of the body. Moreover, we report an antero-posterior and medio-lateral functional segregation over the vermal lobules IV/V/VI. Taken together our results open new avenues for better understanding of the modular functional organization of the cerebellar cortex and its role in postural anti-gravity maintenance. HighlightsO_LIVermal Purkinje cell activation elicits a graded postural collapse in the standing mouse C_LIO_LIThe collapse triggers a secondary composite postural reflex C_LIO_LIAn identified cerebellar module is involved in postural anti-gravity tone maintenance C_LIO_LIThe anti-gravity function is somatotopically organized within this module C_LI

neuroscience↗

bioRxiv2023

Assessing vector competence of mosquitoes from northeastern France to West Nile virus and Usutu virus

West Nile virus (WNV) and Usutu virus (USUV) are two arthropod-borne viruses that circulate in mainland France. Assessing vector competence has only been conducted so far with mosquitoes from southern France while an increasingly active circulation of WNV and USUV has been reported in the last years. The main vectors are mosquitoes of the Culex genus and the common mosquito Culex pipiens. Here, we measure the vector competence of five mosquito species (Aedes rusticus, Aedes albopictus, Anopheles plumbeus, Culex pipiens and Culiseta longiareolata) present in northeastern France. Field-collected populations were exposed to artificial infectious blood meal containing WNV or USUV and examined at different days post-infection. We show that (i) Cx. pipiens transmitted WNV and USUV, (ii) Ae. rusticus only WNV, and (iii) unexpectedly, Ae. albopictus transmitted both WNV and USUV. Less surprising, An. plumbeus was not competent for both viruses. Combined with data on distribution and population dynamics, these assessments of vector competence will help in developing a risk map and implementing appropriate prevention and control measures. Author summaryWest Nile virus (WNV) and Usutu virus (USUV) are on the rise in Europe and in France. WNV is reported in France as early as the 1960s in the Camargue and USUV more recently, in 2015 in eastern France. The re-emergence of WNV infections in the Camargue is associated with an expansion towards the North which is also favorable to maintain a viral transmission cycle. USUV frequently co-circulates with WNV sharing the same mosquito vectors. Culex pipiens, able to feed on birds and humans, is considered to be the main vector in France. Our study is the first to investigate the vector competence to WNV and USUV of five different mosquito species collected in northeastern France. We ascertain that French Cx. pipiens mosquitoes are competent to both WNV and USUV. More surprisingly, the mosquito Aedes albopictus from northeastern France was able to transmit WNV and USUV. Based on our result, we propose that surveillance of mosquitoes combined with viral detections must be implemented in northeastern France to allow early viral detection and timely intervention to prevent outbreaks of these two neurological diseases.

microbiology↗

bioRxiv2022

Seeing our hand or a tool during visually-guided actions: different effects on the somatosensory and visual cortices

The processing of proprioceptive information in the context of a conflict between visual and somatosensory feedbacks deteriorates motor performance. Previous studies have shown that seeing ones hand increases the weighting assigned to arm somatosensory inputs. In this light, we hypothesized that the sensory conflict, when tracing the contour of a shape with mirror-reversed vision, will be greater for participants who trace with a stylus seen in their hand (Hand group, n=17) than for participants who trace with the tip of rod without seen their hand (Tool group, n=15). Based on this hypothesis, we predicted that the tracing performance with mirror vision will be more deteriorated for the Hand group than for the Tool group, and we predicted a greater gating of somatosensory information for the Hand group to reduce the sensory conflict. The participants of both groups followed the outline of a shape in two visual conditions. Direct vision: the participants saw the hand or portion of a light 40 cm rod directly. Mirror Vision: the hand or the rod was seen through a mirror. We measured tracing performance using a digitizing tablet and the cortical activity with electroencephalography. Behavioral analyses revealed that the tracing performance of both groups was similarly impaired by mirror vision. However, contrasting the spectral content of the cortical oscillatory activity between the Mirror and Direct conditions, we observed that tracing with mirror vision resulted in significantly larger alpha (8-12 Hz) and beta (15-25 Hz) powers in the somatosensory cortex for participants of the Hand group. The somatosensory alpha and beta powers did not significantly differ between Mirror and Direct vision conditions for the Tool group. For both groups, tracing with mirror vision altered the activity of the visual cortex: decreased alpha power for the Hand group, decreased alpha and beta power for the Tool group. Overall, these results suggest that seeing the hand enhanced the sensory conflict when tracing with mirror vision and that the increase of alpha and beta powers in the somatosensory cortex served to reduce the weight assigned to somatosensory information. The increased activity of the visual cortex observed for both groups in the mirror vision condition suggests greater visual processing with increased task difficulty. Finally, the fact that the participants of the Tool group did not show better tracing performance than those of the Hand group suggests that tracing deterioration resulted from a sensorimotor conflict (as opposed to a visuo-proprioceptive conflict).

neuroscience↗

bioRxiv2021

KEEPING IN TOUCH WITH OUR HIDDEN SIDE

Previous studies have shown that the sensory modality used to identify the position of proprioceptive targets hidden from sight, but frequently viewed, influences the type of the body representation employed for reaching them with the finger. The question then arises as to whether this observation also applies to proprioceptive targets which are hidden from sight, and rarely, if ever, viewed. We used an established technique for pinpointing the type of body representation used for the spatial encoding of targets which consisted of assessing the effect of peripheral gaze fixation on the pointing accuracy. More precisely, an exteroceptive, visually dependent, body representation is thought to be used if gaze deviation induces a deviation of the pointing movement. Three light-emitting diodes (LEDs) were positioned at the participants eye level at -25 deg, 0 deg and +25 deg with respect to the cyclopean eye. Without moving the head, the participant fixated the lit LED before the experimenter indicated one of the three target head positions: topmost point of the head (vertex) and two other points located at the front and back of the head. These targets were either verbal-cued or tactile-cued. The goal of the subjects (n=27) was to reach the target with their index finger. We analysed the accuracy of the movements directed to the topmost point of the head, which is a well-defined, yet out of view anatomical point. Based on the possibility of the brain to create visual representations of the body areas that remain out of view, we hypothesized that the position of the vertex is encoded using an exteroceptive body representation, both when verbally or tactile-cued. Results revealed that the pointing errors were biased in the opposite direction of gaze fixation for both verbal-cued and tactile-cued targets, suggesting the use of a vision-dependent exteroceptive body representation. The enhancement of the visual body representations by sensorimotor processes was suggested by the greater pointing accuracy when the vertex was identified by tactile stimulation compared to verbal instruction. Moreover, we found in a control condition that participants were more accurate in indicating the position of their own vertex than the vertex of other people. This result supports the idea that sensorimotor experiences increase the spatial resolution of the exteroceptive body representation. Together, our results suggest that the position of rarely viewed body parts are spatially encoded by an exteroceptive body representation and that non-visual sensorimotor processes are involved in the constructing of this representation.

neuroscience↗

bioRxiv2018

Fast two-photon volumetric imaging of an improved voltage indicator reveals electrical activity in deeply located neurons in the awake brain

Imaging of transmembrane voltage deep in brain tissue with cellular resolution has the potential to reveal information processing by neuronal circuits in living animals with minimal perturbation. Multi-photon voltage imaging in vivo, however, is currently limited by speed and sensitivity of both indicators and imaging methods. Here, we report the engineering of an improved genetically encoded voltage indicator, ASAP3, which exhibits up to 51% fluorescence responses in the physiological voltage range, sub-millisecond activation kinetics, and full responsivity under two-photon illumination. We also introduce an ultrafast local volume excitation (ULOVE) two-photon scanning method to sample ASAP3 signals in awake mice at kilohertz rates with increased stability and sensitivity. ASAP3 and ULOVE allowed continuous single-trial tracking of spikes and subthreshold events for minutes in deep locations, with subcellular resolution, and with repeated sampling over multiple days. By imaging voltage in visual cortex neurons, we found evidence for cell type-dependent subthreshold modulation by locomotion. Thus, ASAP3 and ULOVE enable continuous high-speed high-resolution imaging of electrical activity in deeply located genetically defined neurons during awake behavior.

neuroscience↗

bioRxiv2018

Active intermixing of indirect and direct neurons builds the striatal mosaic

The striatum controls behaviors via the activity of direct and indirect pathway projection neurons (dSPN and iSPN) that are intermingled in all compartments. While such mosaic ensures the balanced activity of the two pathways, how it emerges remains largely unknown. Here, we show that both SPN populations are specified embryonically and progressively intermix through multidirectional iSPN migration. Using conditional mutants of the dSPN-specific transcription factor Ebf1, we found that inactivating this gene impaired selective dSPN properties, including axon pathfinding, whereas molecular and functional features of iSPN were preserved. Remarkably, Ebf1 mutation disrupted iSPN/dSPN intermixing, resulting in an uneven distribution. Such architectural defect was selective of the matrix compartment, revealing that intermixing is a parallel process to compartment formation. Our study reveals that, while iSPN/dSPN specification is largely independent, their intermingling emerges from an active migration of iSPN, thereby providing a novel framework for the building of striatal architecture.

neuroscience↗