Biology subjects

Holy, T. E.

Publications and source records attributed to Holy, T. E..

12 recordsLinked to original sources

bioRxiv2026

In vivo optical clearing of the mouse brain

Light microscopy is one of the most powerful tools for understanding living systems, but the opacity of tissue prevents visualization of all but superficial layers. Several methods to clarify tissue have been developed, but most require fixed specimens. To address the challenge of improving resolution in functioning neuronal circuits, we developed a biocompatible clearing agent, iodixanol-ACSF, which is capable of increasing the transparency of living neuronal tissue. Brain-cleared mice were motile and unimpaired on a variety of behavioral tasks, and extracellular recordings showed that many cellular and circuit phenomena were well-preserved. In live iodixanol-ACSF cleared mouse brain tissue, both transmission and cellular-resolution fluorescence microscopy indicate improvements of 150-200% in penetration depth with one-third to one-half the laser intensity when compared to untreated tissue. Our results show that iodixanol-ACSF clearing will enable deeper imaging and extend our understanding of neuronal circuit function.

neuroscience↗

bioRxiv2026

The Laminar Organization of a Decision Circuit in Orbitofrontal Cortex is Agnostic to the Variable Encoding Scheme

Economic choices are believed to depend on the orbitofrontal cortex (OFC). Work in primates and rodents indicates that neurons in OFC participate in computing and comparing subjective values, suggesting that different groups of cells constitute the building blocks of a decision circuit. In a recent study (Livi et al., 2025), we examined the laminar organization of this circuit in mice. We found that different decision variables were differentially represented in layer 2/3 (L2/3) and layer 5 (L5). Furthermore, the temporal dynamics of decision signals indicated a combination of feed-forward and feed-back across layers, and pointed to L5 as the locus for winner-take-all value comparison. Importantly, these results were obtained under the constraint that each neuron encoded a single variable. Here, we tested whether our results on laminar organization depended on the categorical framework. We applied LASSO regression to identify a minimal set of variables explaining each neurons activity. Even with approximately half of all neurons representing two or more variables, the layer specificity of decision variables was preserved. In addition, Granger Causality Analysis and activity profiles reached similar conclusions as for analyses conducted under the single-variable constraint. We conclude that the decision circuit in OFC exhibits a laminar architecture, independently of whether the representation of decision variables in this area is categorical or mixed.

neuroscience↗

bioRxiv2025

Laminar Architecture of a Decision Circuit in Orbitofrontal Cortex

During economic choice, different neurons in orbitofrontal cortex (OFC) encode individual offer values, the binary choice outcome, and the chosen value. Previous work suggests that these cell groups form a decision circuit, but the anatomical organization of this circuit is poorly understood. Using calcium imaging, we recorded from layer 2/3 (L2/3) and layer 5 (L5) of mice choosing between juice flavors. Decision variables were differentially represented across layers: juice-specific offer values and their spatial configuration were predominant in L2/3, while spatial offer values, chosen side, and chosen value were predominant in L5. Within each layer, functional cell groups were organized in clusters. The temporal dynamics of neural signals in the two layers indicated a combination of feed-forward and feed-back processes, and pointed to L5 as the locus for winner-take-all value comparison. These results reveal that economic decisions rely on a complex architecture distributed across layers of OFC.

neuroscience↗

bioRxiv2024

STARDUST: a pipeline for the unbiased analysis of astrocyte regional calcium dynamics

Calcium imaging has become a popular way to probe astrocyte activity, but few analysis methods holistically capture discrete calcium signals that occur across the astrocyte domain. Here, we introduce STARDUST, a pipeline for the Spatio-Temporal Analysis of Regional Dynamics & Unbiased Sorting of Transients from fluorescence recordings of astrocytes, and provide step-by-step guidelines. STARDUST yields fluorescence time- series from data-defined regions of activity and performs systematic signal detection and feature extraction, enabling the in-depth and unbiased study of astrocyte calcium signals. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/588196v2_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@c6a97aorg.highwire.dtl.DTLVardef@a0b0fdorg.highwire.dtl.DTLVardef@1c3dd73org.highwire.dtl.DTLVardef@1ffa9ec_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2024

The Representation of Decision Variables in Orbitofrontal Cortex is Longitudinally Stable

The computation and comparison of subjective values underlying economic choices rely on the orbitofrontal cortex (OFC). In this area, distinct groups of neurons encode the value of individual options, the binary choice outcome, and the chosen value. These variables capture both the input and the output of the choice process, suggesting that the cell groups found in OFC constitute the building blocks of a decision circuit. Here we show that this neural circuit is longitudinally stable. Using two-photon calcium imaging, we recorded from mice choosing between different juice flavors. Recordings of individual cells continued for up to 20 weeks. For each cell and each pair of sessions, we compared the activity profiles using cosine similarity, and we assessed whether the cell encoded the same variable in both sessions. These analyses revealed a high degree of stability and a modest representational drift. A quantitative estimate indicated this drift would not randomize the circuit within the animals lifetime.

neuroscience↗

bioRxiv2022

Polyphasic circadian neural circuits drive differential activities in multiple downstream rhythmic centers

Circadian clocks align various behaviors such as locomotor activity, sleep/wake, feeding, and mating to times of day that are most adaptive. How rhythmic information in pacemaker circuits is translated to neuronal outputs is not well understood. Here we used brain-wide, 24-hr in vivo calcium imaging in the Drosophila brain and searched for circadian rhythmic activity among identified clusters of dopaminergic (DA) and peptidergic neuroendocrine (PNE) neurons. Such rhythms were widespread and imposed by the PERIOD-dependent clock activity within the [~]150 cell circadian pacemaker network. The rhythms displayed either a Morning, an Evening, or Mid-Day phase. Different sub-groups of circadian pacemakers imposed neural activity rhythms onto different downstream non-clock neurons. Outputs from the canonical M and E pacemakers converged to regulate DA-PPM3 and DA-PAL neurons. E pacemakers regulate the Evening-active DA-PPL1 neurons. In addition to these canonical M and E oscillators, we present evidence for a third dedicated phase occurring at Mid-Day (MD): the l-LNv pacemakers present the MD activity peak and they regulate the MD-active DA-PPM1/2 neurons and three distinct PNE cell types. Thus, the Drosophila circadian pacemaker network is a polyphasic rhythm generator. It presents dedicated M, E and MD phases that are functionally transduced as neuronal outputs to organize diverse daily activity patterns in downstream circuits.

neuroscience↗

bioRxiv2021

Circadian pacemaker neurons display co-phasic rhythms in basal calcium level and in fast calcium fluctuations

Circadian pacemaker neurons in the Drosophila brain display daily rhythms in the levels of intracellular calcium. These calcium rhythms are driven by molecular clocks and are required for normal circadian behavior. To study their biological basis, we employed genetic manipulations in conjunction with in vivo light-sheet microscopy to measure calcium dynamics in individual pacemaker neurons over complete 24-hour periods. We found co-phasic daily rhythms in basal calcium levels and in high frequency calcium fluctuations. Further we found that the rhythms of basal calcium levels require the activity of the IP3R, a channel that mediates calcium fluxes from internal endoplasmic reticulum (ER) calcium stores. Independently, the rhythms of fast calcium fluctuations required the T-type voltage-gated calcium channel, a conductance that mediates extracellular calcium influx. These results suggest that Drosophila molecular clocks regulate IP3R and T-type channels to generate coupled rhythms in basal calcium and in fast calcium fluctuations, respectively. We propose that both internal and external calcium fluxes are essential for circadian pacemaker neurons to provide rhythmic outputs, and thereby regulate the activities of downstream brain centers.

neuroscience↗

bioRxiv2020

Spatial and Temporal Organization of Odor-evoked Responses in a Fly Olfactory Circuit: Inputs, Outputs and Idiosyncrasies

How is sensory information routed through different types of neurons within a circuit, and do equivalent circuits in different individuals follow similar organizational principles? We examined this issue in the fruit fly olfactory system. Odor-evoked signals from sensory neurons (ORNs) triggered neural responses that were patterned over space and time in cholinergic ePNs and GABAergic iPNs within the antennal lobe. The dendritic-axonal (I/O) response mapping was complex and diverse, and axonal organization was region-specific (mushroom body vs. lateral horn). In the lateral horn, feed-forward excitatory and inhibitory axonal projections matched odor tuning in a stereotyped, dorsal-lateral locus, but mismatched in most other locations. In the temporal dimension, ORN, ePN and iPN odor-evoked responses had similar encoding features, such as information refinement over time and divergent ON and OFF responses. Notably, analogous spatial and temporal coding principles were observed in all flies, and the latter emerged from idiosyncratic neural processing approaches. HighlightsO_LIConsistency and idiosyncrasy both exist in ORN, ePN and iPN functional maps C_LIO_LISignal transformations between different ePN compartments are complex and diverse C_LIO_LITemporal decorrelation between stimuli happens in all three neuronal populations C_LIO_LIOFF responses that are orthogonal to ON responses emerge after odor termination C_LI

neuroscience↗

bioRxiv2019

A molecular logic of sensory coding revealed by optical tagging of physiologically-defined neuronal types

Neural circuit analysis relies on having molecular markers for specific cell types. However, for a cell type identified only by its circuit function, the process of identifying markers remains laborious. Here, we report physiological optical tagging sequencing (PhOTseq), a technique for tagging and expression-profiling cells based on their functional properties. We demonstrate that PhOTseq is capable of selecting rare cell types and enriching them by nearly one hundred-fold. We applied PhOTseq to the challenge of mapping receptor-ligand pairings among vomeronasal pheromone-sensing neurons in mice. Together with in vivo ectopic expression of vomeronasal chemoreceptors, PhOTseq identified the complete combinatorial receptor code for a specific set of ligands, and revealed that the primary sequence of a chemoreceptor was an unexpectedly strong predictor of functional similarity.

neuroscience↗

bioRxiv2019

Neural Mechanisms of Economic Choices in Mice

Economic choices entail computing and comparing subjective values. Evidence from primates indicates that this behavior relies on the orbitofrontal cortex. Conversely, previous work in rodents provided conflicting results. Here we present a mouse model of economic choice behavior, and we show that the lateral orbital (LO) area is intimately related to the decision process. In the experiments, mice chose between different juices offered in variable amounts. Choice patterns closely resembled those measured in primates. Optogenetic inactivation of LO dramatically disrupted choices by inducing erratic changes of relative value and by increasing choice variability. Neuronal recordings revealed that different groups of cells encoded the values of individual options, the binary choice outcome and the chosen value. These groups match those previously identified in primates, except that the neuronal representation in mice is spatial (in monkeys it is good-based). Our results lay the foundations for a circuit-level analysis of economic decisions.

neuroscience↗

bioRxiv2018

Fast Objective Coupled Planar Illumination Microscopy

Among optical imaging techniques light sheet fluorescence microscopy stands out as one of the most attractive for capturing high-speed biological dynamics unfolding in three dimensions. The technique is potentially millions of times faster than point-scanning techniques such as two-photon microscopy. However current-generation light sheet microscopes are limited by volume scanning rate and/or camera frame rate. We present speed-optimized Objective Coupled Planar Illumination (OCPI) microscopy, a fast light sheet technique that avoids compromising image quality or photon efficiency. We increase volume scanning rate to 40 Hz for volumes up to 700 {micro}m thick and introduce Multi-Camera Image Sharing (MCIS), a technique to scale imaging rate by parallelizing acquisition across cameras. Finally, we demonstrate fast calcium imaging of the larval zebrafish brain and find a heartbeat-induced artifact that can be removed by filtering when the imaging rate exceeds 15 Hz. These advances extend the reach of fluorescence microscopy for monitoring fast processes in large volumes.

bioengineering↗

bioRxiv2018

Morning and Evening Circadian Pacemakers Independently Drive Premotor Centers via a Specific Dopamine Relay

Many animals exhibit morning and evening peaks of locomotor behavior. In Drosophila, previous studies identified two corresponding circadian neural oscillators: M (morning) cells which exhixbit a morning neural activity peak, and E (evening) cells which exhibit a corresponding evening peak of activity. Yet we know little of how these distinct circadian oscillators produce specific outputs that regulate pre-motor circuits to precisely control behavioral episodes. Here we show that the Ring Neurons of the Ellipsoid Body (EB-RNs), a defined pre-motor center, display a spontaneous in vivo neural activity rhythm, with peaks in the morning and in the evening. The two EB-RN activity peaks coincide with the major bouts of locomotor activity and result from independent activation by M and E cells, respectively. Further, M and E cells regulate EB-RNs via two identified dopaminergic neurons PPM3-EB, which project to the EB and which are normally co-active with EB-RNs. Blocking the dopaminergic modulation onto EB-RNs prevents the daily two-peak pattern of neural activity in the EB-RN and greatly impairs circadian locomotor activity. These in vivo findings establish the fundamental elements of a circadian neuronal output pathway: distinct circadian oscillators independently drive a common pre-motor center through the agency of specific dopaminergic interneurons.

neuroscience↗