Threatening stimuli elicit a sequential cardiac pattern in arthropods.

In order to cope with the challenges of living in dynamic environments, animals rapidly adjust their behaviors in coordination with different physiological responses. Here, we studied whether threatening visual stimuli evoke different heart rate patterns in arthropods and whether these patterns are related with defensive behaviors. We identified two sequential phases of crab's cardiac response that occur with a similar timescale to that of the motor arrest and later escape response. The first phase was modulated by low salience stimuli and persisted throughout spaced stimulus presentation. The second phase was modulated by high-contrast stimuli and reduced by repetitive stimulus presentation. The overall correspondence between cardiac and motor responses suggests that the first cardiac response phase might be related to motor arrest while the second to the escape response. We show that in the face of threat arthropods coordinate their behavior and cardiac activity in a rapid and flexible manner.

Polarized object detection in crabs: a two-channel system.

Many animal species take advantage of polarization vision for vital tasks such as orientation, communication and contrast enhancement. Previous studies have suggested that decapod crustaceans use a two-channel polarization system for contrast enhancement. Here, we characterize the polarization contrast sensitivity in a grapsid crab. We estimated the polarization contrast sensitivity of the animals by quantifying both their escape response and changes in heart rate when presented with polarized motion stimuli. The motion stimulus consisted of an expanding disk with an 82 deg polarization difference between the object and the background. More than 90% of animals responded by freezing or trying to avoid the polarized stimulus. In addition, we co-rotated the electric vector (e-vector) orientation of the light from the object and background by increments of 30 deg and found that the animals' escape response varied periodically with a 90 deg period. Maximum escape responses were obtained for object and background e-vectors near the vertical and horizontal orientations. Changes in cardiac response showed parallel results but also a minimum response when e-vectors of object and background were shifted by 45 deg with respect to the maxima. These results are consistent with an orthogonal receptor arrangement for the detection of polarized light, in which two channels are aligned with the vertical and horizontal orientations. It has been hypothesized that animals with object-based polarization vision rely on a two-channel detection system analogous to that of color processing in dichromats. Our results, obtained by systematically varying the e-vectors of object and background, provide strong empirical support for this theoretical model of polarized object detection.

An integrated calcium imaging processing toolbox for the analysis of neuronal population dynamics.

The development of new imaging and optogenetics techniques to study the dynamics of large neuronal circuits is generating datasets of unprecedented volume and complexity, demanding the development of appropriate analysis tools. We present a comprehensive computational workflow for the analysis of neuronal population calcium dynamics. The toolbox includes newly developed algorithms and interactive tools for image pre-processing and segmentation, estimation of significant single-neuron single-trial signals, mapping event-related neuronal responses, detection of activity-correlated neuronal clusters, exploration of population dynamics, and analysis of clusters' features against surrogate control datasets. The modules are integrated in a modular and versatile processing pipeline, adaptable to different needs. The clustering module is capable of detecting flexible, dynamically activated neuronal assemblies, consistent with the distributed population coding of the brain. We demonstrate the suitability of the toolbox for a variety of calcium imaging datasets. The toolbox open-source code, a step-by-step tutorial and a case study dataset are available at https://github.com/zebrain-lab/Toolbox-Romano-et-al.

Functional Interactions between Newborn and Mature Neurons Leading to Integration into Established Neuronal Circuits.

From development up to adulthood, the vertebrate brain is continuously supplied with newborn neurons that integrate into established mature circuits. However, how this process is coordinated during development remains unclear. Using two-photon imaging, GCaMP5 transgenic zebrafish larvae, and sparse electroporation in the larva's optic tectum, we monitored spontaneous and induced activity of large neuronal populations containing newborn and functionally mature neurons. We observed that the maturation of newborn neurons is a 4-day process. Initially, newborn neurons showed undeveloped dendritic arbors, no neurotransmitter identity, and were unresponsive to visual stimulation, although they displayed spontaneous calcium transients. Later on, newborn-labeled neurons began to respond to visual stimuli but in a very variable manner. At the end of the maturation period, newborn-labeled neurons exhibited visual tuning curves (spatial receptive fields and direction selectivity) and spontaneous correlated activity with neighboring functionally mature neurons. At this developmental stage, newborn-labeled neurons presented complex dendritic arbors and neurotransmitter identity (excitatory or inhibitory). Removal of retinal inputs significantly perturbed the integration of newborn neurons into the functionally mature tectal network. Our results provide a comprehensive description of the maturation of newborn neurons during development and shed light on potential mechanisms underlying their integration into a functionally mature neuronal circuit.

The Emergence of the Spatial Structure of Tectal Spontaneous Activity Is Independent of Visual Inputs.

The brain is spontaneously active, even in the absence of sensory stimulation. The functionally mature zebrafish optic tectum shows spontaneous activity patterns reflecting a functional connectivity adapted for the circuit's functional role and predictive of behavior. However, neither the emergence of these patterns during development nor the role of retinal inputs in their maturation has been characterized. Using two-photon calcium imaging, we analyzed spontaneous activity in intact and enucleated zebrafish larvae throughout tectum development. At the onset of retinotectal connections, intact larvae showed major changes in the spatiotemporal structure of spontaneous activity. Although the absence of retinal inputs had a significant impact on the development of the temporal structure, the tectum was still capable of developing a spatial structure associated with the circuit's functional roles and predictive of behavior. We conclude that neither visual experience nor intrinsic retinal activity is essential for the emergence of a spatially structured functional circuit.

Sustained Rhythmic Brain Activity Underlies Visual Motion Perception in Zebrafish.

Following moving visual stimuli (conditioning stimuli, CS), many organisms perceive, in the absence of physical stimuli, illusory motion in the opposite direction. This phenomenon is known as the motion aftereffect (MAE). Here, we use MAE as a tool to study the neuronal basis of visual motion perception in zebrafish larvae. Using zebrafish eye movements as an indicator of visual motion perception, we find that larvae perceive MAE. Blocking eye movements using optogenetics during CS presentation did not affect MAE, but tectal ablation significantly weakened it. Using two-photon calcium imaging of behaving GCaMP3 larvae, we find post-stimulation sustained rhythmic activity among direction-selective tectal neurons associated with the perception of MAE. In addition, tectal neurons tuned to the CS direction habituated, but neurons in the retina did not. Finally, a model based on competition between direction-selective neurons reproduced MAE, suggesting a neuronal circuit capable of generating perception of visual motion.

Perceptual learning effect on decision and confidence thresholds.

Practice can enhance of perceptual sensitivity, a well-known phenomenon called perceptual learning. However, the effect of practice on subjective perception has received little attention. We approach this problem from a visual psychophysics and computational modeling perspective. In a sequence of visual search experiments, subjects significantly increased the ability to detect a "trained target". Before and after training, subjects performed two psychophysical protocols that parametrically vary the visibility of the "trained target": an attentional blink and a visual masking task. We found that confidence increased after learning only in the attentional blink task. Despite large differences in some observables and task settings, we identify common mechanisms for decision-making and confidence. Specifically, our behavioral results and computational model suggest that perceptual ability is independent of processing time, indicating that changes in early cortical representations are effective, and learning changes decision criteria to convey choice and confidence.

Spontaneous neuronal network dynamics reveal circuit's functional adaptations for behavior.

Spontaneous neuronal activity is spatiotemporally structured, influencing brain computations. Nevertheless, the neuronal interactions underlying these spontaneous activity patterns, and their biological relevance, remain elusive. Here, we addressed these questions using two-photon calcium imaging of intact zebrafish larvae to monitor the neuron-to-neuron spontaneous activity fine structure in the tectum, a region involved in visual spatial detection. Spontaneous activity was organized in topographically compact assemblies, grouping functionally similar neurons rather than merely neighboring ones, reflecting the tectal retinotopic map despite being independent of retinal drive. Assemblies represent all-or-none-like sub-networks shaped by competitive dynamics, mechanisms advantageous for visual detection in noisy natural environments. Notably, assemblies were tuned to the same angular sizes and spatial positions as prey-detection performance in behavioral assays, and their spontaneous activation predicted directional tail movements. Therefore, structured spontaneous activity represents "preferred" network states, tuned to behaviorally relevant features, emerging from the circuit's intrinsic non-linear dynamics, adapted for its functional role.