Biology subjects

Gambino, F.

Publications and source records attributed to Gambino, F..

9 recordsLinked to original sources

bioRxiv2025

Adeno-associated virus delivered Cre recombinase as a versatile strategy for modeling focal neuronal lesions

Lesion studies remain central to neuroscience, offering key insights into brain-behavior relationships. Excitotoxic agents, though widely used, often produce inconsistent, non-specific, and ethically complex outcomes. In contrast, optogenetics and chemogenetics enable precise, reversible manipulations but pose technical and interpretational challenges, including cost and non-physiological activation. There remains a need for robust, controlled, and ethical lesion methods. Here, we present Cre recombinase expression as a reproducible alternative to excitotoxic lesions in mice. Using high titration adeno-associated viral (AAV) vectors, we show that Cre expression induces focal neurodegeneration across brain regions, affecting both excitatory and inhibitory neurons. This effect is independent of viral serotype or fluorescent tags and is not visible with standard staining but is revealed by neuronal markers. The resulting neuronal loss is followed by glial remodeling and significant behavioral alterations. AAV-mediated Cre expression thus provides a powerful, genetically targeted lesion model that bridges classical and modern approaches to circuit manipulation.

neuroscience↗

bioRxiv2024

Secondary motor cortex tracks decision value and supports behavioral flexibility during non-instructed choice

Optimal decision-making relies on interconnected frontal brain regions, which permit animals to adapt their decisions based on their internal state, experience, and environmental context. Among them, the secondary motor cortex (M2) shows earlier decision-related activity required for sensory-guided action selection. However, the role of M2 in adaptive decision-making in the absence of instructive sensory cues remains unclear. Under such conditions, action-selection relies on abstract representations of actions and their values. Using in vivo microscopy and modeling, we showed that M2 neurons in mice exhibited persistent activity encoding decision values (DV) predicting the probability of action-selection during a non-cue-guided lever task. This was confirmed by the reduced reversal performance upon M2 optogenetic inhibition prior to action-selection. Furthermore, updates in DV determined the rate at which learning is reversed. Together, these results provide strong evidence of the use of DV by M2 to adapt choice in the absence of instructive sensory cues. Declaration of interestsThe authors declare no competing financial interests.

neuroscience↗

bioRxiv2024

Link between respiratory pauses and vigilance states in freely moving mice

Brain activity and breathing rate influence each other but it remains unclear how fine respiratory features vary across vigilance states. Using simultaneous nasal pressure and hippocampal local field potential recordings in freely-moving mice, we show that the position of respiratory pauses within breathing cycles distinguish Wake, Rapid Eye Movement (REM) and non-REM (NREM) sleep states. Model-based predictions of vigilance states based on respiratory features perform well even on animals outside of the training set, suggesting the rules are generalized. Respiratory features underwent specific changes at state transitions, such as progressive elimination of pauses after inhalation foreshadowing REM. During NREM, respiratory changes predicted moment-to moment sigma power variations beyond movement-defined packets delineated by micro-arousals, as pauses after inhalation fragmented NREM sleep into [~]30s windows of high sigma power. Overall, our findings reveal that respiratory features structure the macro- and micro-architecture of sleep, opening new windows into brain states through respiration.

neuroscience↗

bioRxiv2023

Quantitative target engagement of RIPK1 in human whole blood via the cellular thermal shift assay for potential pre-clinical and clinical applications

The cellular thermal shift assay (CETSA(R)) is a target engagement method widely used for preclinical characterization of small molecule compounds. CETSA(R) has been used for semi-quantitative readouts in whole blood with PBMC isolation, and quantitative, plate-based readouts using cell lines. However, there has been no quantitative evaluation of CETSA(R) in unprocessed human whole blood, which is preferred for clinical applications. Here we report two separate assay formats - Alpha CETSA(R) and MSD CETSA(R) - that require less than 100 L of whole blood per sample without PBMC isolation. We chose RIPK1 as a proof-of-concept target and, by measuring engagement of seven different inhibitors, demonstrate high assay sensitivity and robustness. These quantitative CETSA(R) platforms enable possible applications in preclinical pharmacokinetic-pharmacodynamic studies, and direct target engagement with small molecules in clinical trials.

molecular biology↗

bioRxiv2022

NNeurite: artificial neuronal networks for the unsupervised extraction of axonal and dendritic time-lapse signals

Fluorescence microscopy of Ca2+ transients in small neurites of the behaving mouse provides an unprecedented view of the micrometer-scale mechanisms supporting neuronal communication and computation, and therefore opens the way to understanding their role in cognition. However, the exploitation of this growing and precious experimental data is impeded by the scarcity of methods dedicated to the analysis of images of neurites activity in vivo. We present NNeurite, a set of mathematical and computational techniques specialized for the analysis of time-lapse microscopy images of neurite activity in small behaving animals. Starting from noisy and unstable microscopy images containing an unknown number of small neurites, NNeurite simultaneously aligns images, denoises signals and extracts the location and the temporal activity of the sources of Ca2+ transients. At the core of NNeurite is a novel artificial neuronal network (NN) which we have specifically designed to solve the non-negative matrix factorization (NMF) problem modeling source separation in fluorescence microscopy images. For the first time, we have embedded non-rigid image alignment in the NMF optimization procedure, hence allowing to stabilize images based on the transient and weak neurite signals. NNeurite processing is free of any human intervention as NN training is unsupervised and the unknown number of Ca2+ sources is automatically obtained by the NN-based computation of a low-dimensional representation of time-lapse images. Importantly, the spatial shapes of the sources of Ca2+ fluorescence are not constrained in NNeurite, which allowed to automatically extract the micrometer-scale details of dendritic and axonal branches, such dendritic spines and synaptic boutons, in the cortex of behaving mice. We provide NNeurite as a free and open-source library to support the efforts of the community in advancing in vivo microscopy of neurite activity.

neuroscience↗

bioRxiv2020

Astroglial calcium transfer from endoplasmic reticulum to mitochondria determines synaptic integration

Intracellular calcium signaling underlies the astroglial control of synaptic transmission and plasticity. Mitochondria-endoplasmic reticulum contacts (MERCs) are key determinants of calcium dynamics, but their functional impact on astroglial regulation of brain information processing is currently unexplored. We found that the activation of astrocyte mitochondrial-associated CB1 receptors (mtCB1) determines MERCs-dependent intracellular calcium signaling and synaptic integration. The stimulation of mtCB1 receptors promotes calcium transfer from the endoplasmic reticulum to mitochondria through specific mechanisms regulating the activity of the mitochondrial calcium uniporter (MCU) channel. Physiologically, mtCB1-dependent mitochondrial calcium uptake determines the precise dynamics of cytosolic calcium events in astrocytes upon endocannabinoid mobilization. Accordingly, electrophysiological recordings in hippocampal slices showed that genetic exclusion of mtCB1 receptors or specific astroglial MCU inhibition blocks lateral synaptic potentiation, a key example of astrocyte-dependent integration of distant synapses activity. Altogether, these data reveal an unforeseen link between astroglial MERCs and the regulation of brain network functions.

neuroscience↗

bioRxiv2020

An increase in dendritic plateau potentials is associated with experience-dependent cortical map reorganization

The organization of sensory maps in the cerebral cortex depends on experience, which drives homeostatic and long-term synaptic plasticity of cortico-cortical circuits. In the mouse primary somatosensory cortex (S1) afferents from the higher-order, posterior medial thalamic nucleus (POm) gate synaptic plasticity in layer (L) 2/3 pyramidal neurons via disinhibition and the production of dendritic plateau potentials. Here we address whether these thalamocortically mediated responses play a role in whisker map plasticity in S1. We find that trimming all but two whiskers causes a partial fusion of the representations of the two spared whiskers, concomitantly with an increase in the occurrence of POm-driven, N-methyl-D-aspartate receptor (NMDAR)-dependent plateau potentials. Blocking the plateau potentials restores the archetypical organization of the sensory map. Our results reveal a novel mechanism for experience-dependent cortical map plasticity in which higher-order thalamocortically mediated plateau potentials facilitate the fusion of normally segregated cortical representations.

neuroscience↗

bioRxiv2020

AMPAR trafficking dependent LTP initiates cortical remapping and adaptive behaviors during sensory experience

Cortical plasticity improves behaviors and helps recover lost functions after injury by adapting neuronal computations. However, the underlying synaptic and circuit mechanisms remain unclear. In mice, we found that trimming all but one whisker enhances sensory responses from the spared whisker in the somatosensory barrel cortex and occludes whisker-mediated long-term potentiation (w-LTP) in vivo. In addition, whisking-dependent behaviors that are initially impaired by single whisker experience (SWE) rapidly recover when associated cortical regions remap. Blocking the surface diffusion of AMPA receptors (AMPARs) suppresses the expression of w-LTP in naive mice with all whiskers intact, demonstrating that physiologically induced LTP in vivo requires AMPARs trafficking. We used this approach to demonstrate that w-LTP is required for SWE-mediated strengthening of synaptic inputs and initiates the recovery of previously learned skills during the early phases of SWE. Taken together, our data reveal that w-LTP mediates cortical remapping and behavioral improvement upon partial sensory deafferentation and demonstrates that restoration of sensory function after peripheral injury can be manipulated.

neuroscience↗

bioRxiv2019

Non-linear dendritic integration in frontal circuits shapes sensory discrimination in rodents

Survival depends on the ability of animals to select the appropriate behavior in response to threat and safety sensory cues. However, the synaptic and circuit mechanisms by which the brain learns to encode accurate predictors of threat and safety remain largely unexplored. Here, we show that frontal association cortex (FrA) pyramidal neurons integrate auditory cues and basolateral amygdala (BLA) inputs non-linearly in a NMDAR-dependent manner. We found that the response of FrA pyramidal neurons was more pronounced to Gaussian noise than to pure 8 kHz tone, and that the activation of BLA-to-FrA axons was the strongest during safe periods in between conditioning pairings. Blocking BLA-to-FrA signaling specifically at the time of presentation of Gaussian noise (but not 8 kHz tone) as a safety signal impaired the formation of auditory fear memories. Taken together, our data reveal a circuit mechanism that facilitates the formation of fear traces in the FrA, thus providing a new framework for probing discriminative learning and related disorders.

neuroscience↗