Current and future applications of light-sheet imaging for identifying molecular and developmental processes in autism spectrum disorders.

Autism spectrum disorder (ASD) is identified by a set of neurodevelopmental divergences that typically affect the social communication domain. ASD is also characterized by heterogeneous cognitive impairments and is associated with cooccurring physical and medical conditions. As behaviors emerge as the brain matures, it is particularly essential to identify any gaps in neurodevelopmental trajectories during early perinatal life. Here, we introduce the potential of light-sheet imaging for studying developmental biology and cross-scale interactions among genetic, cellular, molecular and macroscale levels of circuitry and connectivity. We first report the core principles of light-sheet imaging and the recent progress in studying brain development in preclinical animal models and human organoids. We also present studies using light-sheet imaging to understand the development and function of other organs, such as the skin and gastrointestinal tract. We also provide information on the potential of light-sheet imaging in preclinical drug development. Finally, we speculate on the translational benefits of light-sheet imaging for studying individual brain-body interactions in advancing ASD research and creating personalized interventions.

Clonidine modulates the activity of the subthalamic-supplementary motor loop: evidence from a pharmacological study combining deep brain stimulation and electroencephalography recordings in Parkinsonian patients.

Clonidine is an anti-hypertensive medication which acts as an alpha-adrenergic receptor agonist. As the noradrenergic system is likely to support cognitive functions including attention and executive control, other clinical uses of clonidine have recently gained popularity for the treatment of neuropsychiatric disorders like attention-deficit hyperactivity disorder or Tourette syndrome, but the mechanism of action is still unclear. Here, we test the hypothesis that the noradrenergic system regulates the activity of subthalamo-motor cortical loops, and that this influence can be modulated by clonidine. We used pharmacological manipulation of clonidine in a placebo-controlled study in combination with subthalamic nucleus-deep brain stimulation (STN-DBS) in 16 Parkinson's disease patients performing a reaction time task requiring to refrain from reacting (proactive inhibition). We recorded electroencephalographical activity of the whole cortex, and applied spectral analyses directly at the source level after advanced blind source separation. We found only one cortical source localized to the supplementary motor area (SMA) that supported an interaction of pharmacological and subthalamic stimulation. Under placebo, STN-DBS reduced proactive alpha power in the SMA, a marker of local inhibitory activity. This effect was associated with the speeding-up of movement initiation. Clonidine substantially increased proactive alpha power from the SMA source, and canceled out the benefits of STN-DBS on movement initiation. These results provide the first direct neural evidence in humans that the tonic inhibitory activity of the subthalamocortical loops underlying the control of movement initiation is coupled to the noradrenergic system, and that this activity can be targeted by pharmacological agents acting on alpha-adrenergic receptors.

How Does Sensor-Space Group Blind Source Separation Face Inter-individual Neuroanatomical Variability? Insights from a Simulation Study Based on the PALS-B12 Atlas.

Because of volume conduction and inter-individual neuroanatomical variability, similar sources in different brains may lead to variable topographies. This represents a major limitation for sensor-space group level decomposition of electroencephalographic data, a technique which introduces potential biases when aggregating individual data. To which extent this impedes subsequent source separation and localization was quantified in the present study. To this end, several simulations using an atlas of human cerebral cortex that takes into account the variability of cortical morphology (Van Essen in NeuroImage 28:635-662, 2005) were performed. For each virtual subject (up to n = 160), the orientation and location of each single simulated dipole was randomly modified as a function of the variability of the cortical shape of a given point in the brain provided by the probabilistic atlas. The resulting activity was projected on the scalp, and topographical shifts were estimated. Then, different algorithms based on second order statistics (SOS) or higher order statistics were used to recover the simulated sources from sensor space information with group blind source separation (gBSS) procedures (based on UWSOBI or EFICA, respectively). As expected, the variability of orientation of the cortical surface across subjects was found to induce substantial variability in scalp potential maps, especially if the sources originate from the dorsolateral prefrontal cortex or the temporoparietal junction. These biases could be compensated for by increasing drastically the number of subjects included in the topographical analyses. By contrast, gBSS was found to be insensitive to inter-individual differences of neuroanatomy. Rather, the estimation of the spatial filters seems to be optimized for the population of interest. Thus, optimal performance of source separation and subsequent source localization did not require the inclusion of a large sample of subjects (n < 20), at least when applying SOS-based statistics that use source spectral diversity to identify and gather similar sources with variable location and orientation. The resulting conclusion that inter-individual neuroanatomical variability is not a major limitation to sensor-space gBSS methods provides boosting perspectives for this promising approach, especially for the detection and localization of task/population related neural sources.

Tracking markers of response inhibition in electroencephalographic data: why should we and how can we go beyond the N2 component?

Response inhibition is a pivotal component of executive control, which is especially difficult to assess. Indeed, it is a substantial challenge to gauge brain-behavior relationships because this function is precisely intended to suppress overt measurable behaviors. A further complication is that no single neuroimaging method has been found that can disentangle the accurate time-course of concurrent excitatory and inhibitory mechanisms. Here, we argue that this objective can be achieved with electroencephalography (EEG) on some conditions. Based on a systematic review, we emphasize that the standard event-related potential N2 (N200) is not an appropriate marker of prepotent response inhibition. We provide guidelines for assessing the cortical brain dynamics of response inhibition with EEG. This includes the combined use of inseparable data processing steps (source separation, source localization, and single-trial and time-frequency analyses) as well as the amendment of the classical experimental designs to enable the recording of different kinds of electrophysiological activity predicted by different models of response inhibition. We conclude with an illustration based on recent findings of how fruitful this approach can be.

The dorsal medial frontal cortex mediates automatic motor inhibition in uncertain contexts: evidence from combined fMRI and EEG studies.

Response inhibition is commonly thought to rely on voluntary, reactive, selective, and relatively slow prefrontal mechanisms. In contrast, we suggest here that response inhibition is achieved automatically, nonselectively, within very short delays in uncertain environments. We modified a classical go/nogo protocol to probe context-dependent inhibitory mechanisms. Because no single neuroimaging method can definitely disentangle neural excitation and inhibition, we combined fMRI and EEG recordings in healthy humans. Any stimulus (go or nogo) presented in an uncertain context requiring action restraint was found to evoke activity changes in the supplementary motor complex (SMC) with respect to a control condition in which no response inhibition was required. These changes included: (1) An increase in event-related BOLD activity, (2) an attenuation of the early (170 ms) event related potential generated by a single, consistent source isolated by advanced blind source separation, and (3) an increase in the evoked-EEG Alpha power of this source. Considered together, these results suggest that the BOLD signal evoked by any stimulus in the SMC when the situation is unpredictable can be driven by automatic, nonselective, context-dependent inhibitory activities. This finding reveals the paradoxical mechanisms by which voluntary control of action may be achieved. The ability to provide controlled responses in unpredictable environments would require setting-up the automatic self-inhibitory circuitry within the SMC. Conversely, enabling automatic behavior when the environment becomes predictable would require top-down control to deactivate anticipatorily and temporarily the inhibitory set.

Greater robustness of second order statistics than higher order statistics algorithms to distortions of the mixing matrix in blind source separation of human EEG: implications for single-subject and group analyses.

A mandatory assumption in blind source separation (BSS) of the human electroencephalogram (EEG) is that the mixing matrix remains invariant, i.e., that the sources, electrodes and geometry of the head do not change during the experiment. Actually, this is not often the case. For instance, it is common that some electrodes slightly move during EEG recording. This issue is even more critical for group independent component analysis (gICA), a method of growing interest, in which only one mixing matrix is estimated for several subjects. Indeed, because of interindividual anatomo-functional variability, this method violates the mandatory principle of invariance. Here, using simulated (experiments 1 and 2) and real (experiment 3) EEG data, we test how eleven current BSS algorithms undergo distortions of the mixing matrix. We show that this usual kind of perturbation creates non-Gaussian features that are virtually added to all sources, impairing the estimation of real higher order statistics (HOS) features of the actual sources by HOS algorithms (e.g., Ext-INFOMAX, FASTICA). HOS-based methods are likely to identify more components (with similar properties) than actual neurological sources, a problem frequently encountered by BSS users. In practice, the quality of the recovered signal and the efficiency of subsequent source localization are substantially impaired. Performing dimensionality reduction before applying HOS-based BSS does not seem to be a safe strategy to circumvent the problem. Second order statistics (SOS)-based BSS methods belonging to the less popular SOBI family class are much less sensitive to this bias.

An optimized iDISCO+ protocol for tissue clearing and 3D analysis of oxytocin and vasopressin cell network in the developing mouse brain.

Here, we present an optimized iDISCO+ protocol combining tissue clearing and light sheet microscopy to map the postnatal development of oxytocin and vasopressin neurons in mouse hypothalamus. We describe tissue preparation, immunostaining, clearing, and imaging. We then detail how to process the 3D cell dataset to analyze cell network using a point-based recording procedure that accurately maps neurons in the Allen brain atlas. This protocol can be applied to any neuronal population, in different brain regions and at different developmental stages. For complete details on the use and execution of this protocol, please refer to Soumier et al. (2021).1.

Assessing the allocation of attention during visual search using digit-tracking, a calibration-free alternative to eye tracking.

Digit-tracking, a simple, calibration-free technique, has proven to be a good alternative to eye tracking in vision science. Participants view stimuli superimposed by Gaussian blur on a touchscreen interface and slide a finger across the display to locally sharpen an area the size of the foveal region just at the finger's position. Finger movements are recorded as an indicator of eye movements and attentional focus. Because of its simplicity and portability, this system has many potential applications in basic and applied research. Here we used digit-tracking to investigate visual search and replicated several known effects observed using different types of search arrays. Exploration patterns measured with digit-tracking during visual search of natural scenes were comparable to those previously reported for eye-tracking and constrained by similar saliency. Therefore, our results provide further evidence for the validity and relevance of digit-tracking for basic and applied research on vision and attention.

Dissociation of early and late face-related processes in autism spectrum disorder and Williams syndrome.

BACKGROUND: Williams syndrome (WS) and Autism Spectrum Disorders (ASD) are neurodevelopmental conditions associated with atypical but opposite face-to-face interactions patterns: WS patients overly stare at others, ASD individuals escape eye contact. Whether these behaviors result from dissociable visual processes within the occipito-temporal pathways is unknown. Using high-density electroencephalography, multivariate signal processing algorithms and a protocol designed to identify and extract evoked activities sensitive to facial cues, we investigated how WS (N = 14), ASD (N = 14) and neurotypical subjects (N = 14) decode the information content of a face stimulus. RESULTS: We found two neural components in neurotypical participants, both strongest when the eye region was projected onto the subject's fovea, simulating a direct eye contact situation, and weakest over more distant regions, reaching a minimum when the focused region was outside the stimulus face. The first component peaks at 170 ms, an early signal known to be implicated in low-level face features. The second is identified later, 260 ms post-stimulus onset and is implicated in decoding salient face social cues. Remarkably, both components were found distinctly impaired and preserved in WS and ASD. In WS, we could weakly decode the 170 ms signal based on our regressor relative to facial features, probably due to their relatively poor ability to process faces' morphology, while the late 260 ms component was highly significant. The reverse pattern was observed in ASD participants who showed neurotypical like early 170 ms evoked activity but impaired late evoked 260 ms signal. CONCLUSIONS: Our study reveals a dissociation between WS and ASD patients and points at different neural origins for their social impairments.

Differential fate between oxytocin and vasopressin cells in the developing mouse brain.

Oxytocin (OXT) and arginine vasopressin (AVP), two neuropeptides involved in socio-emotional behaviors have been anatomically defined in the adult brain. Yet their spatial organization during postnatal development is not clearly defined. We built a developmental atlas using 3D imaging of cleared immunolabeled tissue over four early postnatal (P) stages, from birth (P0, P3, P7, P14) to young adulthood (≥P56). Our atlas-based mapping revealed that the number of OXT neurons doubles according to unique temporal dynamics in selective hypothalamic regions, namely, the periventricular and paraventricular nuclei, and in a novel location we named the antero-lateral preoptic. In the paraventricular nucleus, single-cell densities and fluorescence analysis demonstrated selective expansion of OXT cells in the antero-ventral division, whereas the postero-dorsal division contained cells present at birth. No changes were observed for AVP neurons. Our findings show the coexisting of innate and plastic OXT/AVP brain circuits probably triggered by environmental adaptation of the social brain.

Culture-free perceptual invariant for trustworthiness.

Humans beings decide to trust others selectively, often based on the appearance of a face. But how do observers deal with the wide variety of facial morphologies and, in particular, those outside their own familiar cultural group? Using reverse correlation, a data-driven approach to explore how individuals create internal representations without external biases, we studied the generation of trustworthy faces by French and Chinese participants (N = 160) within and outside their own cultural group. Participants selected the most trustworthy or attractive (control condition) face from two identical European or Asian descent faces that had been modified by different noise masks. A conjunction analysis to reveal facial features common to both cultures showed that Chinese and French participants unconsciously increased the contrast of the "pupil-iris area" to make the face appear more trustworthy. No significant effects common to both groups were found for the attraction condition suggesting that attraction judgements are dependent on cultural processes. These results suggest the presence of universal cross-cultural mechanisms for the construction of implicit first impressions of trust, and highlight the importance of the eyes area in this process.

Actionable Predictive Factors of Homelessness in a Psychiatric Population: Results from the REHABase Cohort Using a Machine Learning Approach.

BACKGROUND: There is a lack of knowledge regarding the actionable key predictive factors of homelessness in psychiatric populations. Therefore, we used a machine learning model to explore the REHABase database (for rehabilitation database-n = 3416), which is a cohort of users referred to French psychosocial rehabilitation centers in France. METHODS: First, we analyzed whether the different risk factors previously associated with homelessness in mental health were also significant risk factors in the REHABase. In the second step, we used unbiased classification and regression trees to determine the key predictors of homelessness. Post hoc analyses were performed to examine the importance of the predictors and to explore the impact of cognitive factors among the participants. RESULTS:  First, risk factors that were previously found to be associated with homelessness were also significant risk factors in the REHABase. Among all the variables studied with a machine learning approach, the most robust variable in terms of predictive value was the nature of the psychotropic medication (sex/sex relative mean predictor importance: 22.8, σ = 3.4). Post hoc analyses revealed that first-generation antipsychotics (15.61%; p < 0.05 FDR corrected), loxapine (16.57%; p < 0.05 FWER corrected) and hypnotics (17.56%; p < 0.05 FWER corrected) were significantly associated with homelessness. Antidepressant medication was associated with a protective effect against housing deprivation (9.21%; p < 0.05 FWER corrected). CONCLUSIONS: Psychotropic medication was found to be an important predictor of homelessness in our REHABase cohort, particularly loxapine and hypnotics. On the other hand, the putative protective effect of antidepressants confirms the need for systematic screening of depression and anxiety in the homeless population.

Face first impression of trustworthiness in Williams Syndrome: Dissociating automatic vs decision based perception.

Humans make rapid decisions of trustworthiness based on facial appearance. Williams Syndrome (WS) is a genetic disorder associated with hypersociability toward strangers, suggesting a disruption in trust assessment. We recorded eye-movements in neurotypically-developed (TD) participants (N = 21) and in patients with WS (N = 22) as we presented pairs of computer-generated faces, pre-rated as trustworthy or untrustworthy. In a spontaneous visual preference task, TD participants gazed significantly longer at trustworthy faces while patients with WS showed no preferences for either face-type. Next, in a decision task, participants selected the face that they judged consistent with one of three social descriptions relating to high levels of trust. Both groups gazed longer at trustworthy faces and made correct matches with the corresponding social descriptions. Improvements by patients with WS in the decision task were contingent on outcomes measured on the emotional NEPSY scale. We conclude that while spontaneous representation of trustworthiness is impaired in WS, top-down mechanisms of trust recognition appear partially preserved.

Digit-tracking as a new tactile interface for visual perception analysis.

Eye-tracking is a valuable tool in cognitive science for measuring how visual processing resources are allocated during scene exploration. However, eye-tracking technology is largely confined to laboratory-based settings, making it difficult to apply to large-scale studies. Here, we introduce a biologically-inspired solution that involves presenting, on a touch-sensitive interface, a Gaussian-blurred image that is locally unblurred by sliding a finger over the display. Thus, the user's finger movements provide a proxy for their eye movements and attention. We validated the method by showing strong correlations between attention maps obtained using finger-tracking vs. conventional optical eye-tracking. Using neural networks trained to predict empirically-derived attention maps, we established that identical high-level features hierarchically drive explorations with either method. Finally, the diagnostic value of digit-tracking was tested in autistic and brain-damaged patients. Rapid yet robust measures afforded by this method open the way to large scale applications in research and clinical settings.

Resting state oscillations suggest a motor component of Parkinson's Impulse Control Disorders.

OBJECTIVES: Impulse control disorders (ICDs) in Parkinson's disease (PD) have been associated with cognitive impulsivity and dopaminergic dysfunction and treatment. The present study tests the neglected hypothesis that the neurofunctional networks involved in motor impulsivity might also be dysfunctional in PD-ICDs. METHODS: We performed blind spectral analyses of resting state electroencephalographic (EEG) data in PD patients with and without ICDs to probe the functional integrity of all cortical networks. Analyses were performed directly at the source level after blind source separation. Discrete differences between groups were tested by comparing patients with and without ICDs. Gradual dysfunctions were assessed by means of correlations between power changes and clinical scores reflecting ICD severity (QUIP score). RESULTS: Spectral signatures of ICDs were found in the medial prefrontal cortex, the dorsal anterior cingulate and the supplementary motor area, in the beta and gamma bands. Beta power changes in the supplementary motor area were found to predict ICDs severity. CONCLUSION: ICDs are associated with abnormal activity within frequency bands and cortical circuits supporting the control of motor response inhibition. SIGNIFICANCE: These results bring to the forefront the need to consider, in addition to the classical interpretation based on aberrant mesocorticolimbic reward processing, the issue of motor impulsivity in PD-ICDs and its potential implications for PD therapy.

Removing deep brain stimulation artifacts from the electroencephalogram: Issues, recommendations and an open-source toolbox.

A major question for deep brain stimulation (DBS) research is understanding how DBS of one target area modulates activity in different parts of the brain. EEG gives privileged access to brain dynamics, but its use with implanted patients is limited since DBS adds significant high-amplitude electrical artifacts that can completely obscure neural activity measured using EEG. Here, we systematically review and discuss the methods available for removing DBS artifacts. These include simple techniques such as oversampling, antialiasing analog filtering and digital low-pass filtering, which are necessary but typically not sufficient to fully remove DBS artifacts when each is used in isolation. We also cover more advanced methods, including techniques tracking outliers in the frequency-domain, which can be effective, but are rarely used. The reason for that is twofold: First, it requires advanced skills in signal processing since no user friendly tool for removing DBS artifacts is currently available. Second, it involves fine-tuning to avoid over-aggressive filtering. We highlight an open-source toolbox incorporating most artifact removal methods, allowing users to combine different strategies.

Implicit preference for human trustworthy faces in macaque monkeys.

It has been shown that human judgements of trustworthiness are based on subtle processing of specific facial features. However, it is not known if this ability is a specifically human function, or whether it is shared among primates. Here we report that macaque monkeys (Macaca Mulatta and Macaca Fascicularis), like humans, display a preferential attention to trustworthiness-associated facial cues in computer-generated human faces. Monkeys looked significantly longer at faces categorized a priori as trustworthy compared to untrustworthy. In addition, spatial sequential analysis of monkeys' initial saccades revealed an upward shift with attention moving to the eye region for trustworthy faces while no change was observed for the untrustworthy ones. Finally, we found significant correlations between facial width-to-height ratio- a morphometric feature that predicts trustworthiness' judgments in humans - and looking time in both species. These findings suggest the presence of common mechanisms among primates for first impression of trustworthiness.

How components of facial width to height ratio differently contribute to the perception of social traits.

Facial width to height ratio (fWHR) is a morphological cue that correlates with sexual dimorphism and social traits. Currently, it is unclear how vertical and horizontal components of fWHR, distinctly capture faces' social information. Using a new methodology, we orthogonally manipulated the upper facial height and the bizygomatic width to test their selective effect in the formation of impressions. Subjects (n = 90) saw pair of faces and had to select the face expressing better different social traits (trustworthiness, aggressiveness and femininity). We further investigated how sex and fWHR components interact in the formation of these judgements. Across experiments, changes along the vertical component better predicted participants' ratings rather than the horizontal component. Faces with smaller height were perceived as less trustworthy, less feminine and more aggressive. By dissociating fWHR and testing the contribution of its components independently, we obtained a powerful and discriminative measure of how facial morphology guides social judgements.

Restoring consciousness with vagus nerve stimulation.

Patients lying in a vegetative state present severe impairments of consciousness [1] caused by lesions in the cortex, the brainstem, the thalamus and the white matter [2]. There is agreement that this condition may involve disconnections in long-range cortico-cortical and thalamo-cortical pathways [3]. Hence, in the vegetative state cortical activity is 'deafferented' from subcortical modulation and/or principally disrupted between fronto-parietal regions. Some patients in a vegetative state recover while others persistently remain in such a state. The neural signature of spontaneous recovery is linked to increased thalamo-cortical activity and improved fronto-parietal functional connectivity [3]. The likelihood of consciousness recovery depends on the extent of brain damage and patients' etiology, but after one year of unresponsive behavior, chances become low [1]. There is thus a need to explore novel ways of repairing lost consciousness. Here we report beneficial effects of vagus nerve stimulation on consciousness level of a single patient in a vegetative state, including improved behavioral responsiveness and enhanced brain connectivity patterns.