Biology subjects

Cruz, B.

Publications and source records attributed to Cruz, B..

7 recordsLinked to original sources

bioRxiv2025

Aeon: an open-source platform to study the neural basis of ethological behaviours over naturalistic timescales

Ethological behaviours are a powerful tool for neuroscience since they leverage the robust neural computations shaped by the species evolution to study the neural basis of cognitive functions. However, such behaviours are often transitory and dependent on factors that vary over space, time and number of individuals, making them difficult to capture with standard laboratory tasks. Here we present Aeon, an open-source platform designed for continuous, long-term study of self-guided behaviours in multiple mice and simultaneous recording of brain activity within large, customizable habitats. By integrating specialized modules for navigation, nesting and sleeping, escaping, foraging, and social interaction, Aeon enables the expression of key ethological behaviours while achieving experimental control and multi-dimensional quantifications from sub-millisecond to month-long durations. Its software architecture ensures robust data acquisition via many synchronized data streams and delivers a new standardised, unified data format that yields seamless, integrated analysis pipelines. Using assays such as digging-to-threshold and social foraging, Aeon reveals how mice adapt strategies in a changing environment and in response to conspecifics. Aeon bridges ecological relevance with rigorous experimental control to advance our understanding of how neural circuit activity gives rise to a range of highly conserved and adaptive behaviours.

neuroscience↗

bioRxiv2023

Molecular understanding of anthocyanin biosynthesis activated by PAP1 in engineered red Artemisia annua cells and regulation of 2, 4-dichlorophenoxyacetic acid

Artemisia annua is an effective antimalarial medicinal crop. We have established anthocyanin-producing red cell cultures from this plant with the overexpression of Production of Anthocyanin Pigment 1 (PAP1) encoding a R2R3MYB transcription factor. To understand the molecular mechanism by which PAP1 activated the entire anthocyanin pathway, we mined the genomic sequences of A. annua and obtained eight promoters of the anthocyanin pathway genes. Sequence analysis identified four types of AC cis-elements from six promoters, the MYB response elements (MRE) bound by PAP1. In addition, six promoters were determined to have at least one G-Box cis-element. Eight promoters were cloned for activity analysis. Duel luciferase assays showed that PAP1 significantly enhanced the promoting activity of seven promoters, indicating that PAP1 turned on the biosynthesis of anthocyanins via the activation of these pathway gene expression. To understand how 2,4-dichlorophenoxyacetic acid (2,4-D), an auxin, regulates the PAP1-activated anthocyanin biosynthesis, five different concentrations (0, 0.05, 0.5, 2.5, and 5 M) were tested to characterize anthocyanin production and profiles. The resulting data showed that the concentrations tested decreased the fresh weight of callus growth, anthocyanin levels, and the production of anthocyanins per petri dish. HPLC-qTOF-MS/MS based profiling showed that these concentrations did not alter anthocyanin profiles. Real time RT-PCR was completed to characterize the expression PAP1 and four representative pathway genes. The results showed that the five concentrations reduced the expression levels of the constitutive PAP1 transgene and three pathway genes significantly and eliminated chalcone synthase gene in expression either significantly or slightly. These data indicate that the constitutive PAP1 expression depends on gradients added in the medium. Based on these findings, the regulation of 2,4-D is discussed for anthocyanin engineering in red cells of A. annua. ConclusionPromoters of eight anthocyanin pathway genes were cloned. Four types of AC cis-elements were identified from six promoters and G-Box elements were also determined from six promoters. PAP1 enhanced the activity of eight promoters. 2,4-D downregulated the expression of the constitutive PAP1 transgene leading to the decrease the biosynthesis of anthocyanins.

plant biology↗

bioRxiv2021

Aberrant hippocampal transmission and behavior in mice with a stargazin mutation linked to intellectual disability

Mutations linked to neurodevelopmental disorders, such as intellectual disability (ID), are frequently found in genes that encode for proteins of the excitatory synapse. Transmembrane AMPA receptor regulatory proteins (TARPs) are AMPA receptor auxiliary proteins that regulate crucial aspects of receptor function. Here, we investigate an ID-associated mutant form of the TARP family member stargazin. Molecular dynamics analyses showed that the stargazin V143L variant weakens the overall interface of the AMPAR:stargazin complex and hinders the stability of the complex. Knock-in mice for the V143L stargazin mutation manifest cognitive and social deficits and hippocampal synaptic transmission defects. In the hippocampus of stargazin V143L mice, CA1 neurons show impaired spine maturation in basal dendrites, and synaptic ultrastructural alterations. These data demonstrate a causal role for mutated stargazin in the pathogenesis of ID and highlight its role in the development and function of hippocampal synapses.

neuroscience↗

bioRxiv2019

Estradiol promotes and progesterone reduces anxiety-like behavior produced by nicotine withdrawal in rats

The present study assessed sex differences and the role of ovarian hormones in the behavioral effects of nicotine withdrawal. Study 1 compared physical signs, anxiety-like behavior, and corticosterone levels in male, intact female, and ovariectomized (OVX) female rats during nicotine withdrawal. Estradiol (E2) and progesterone levels were also assessed in intact females that were tested during different phases of the 4-day estrous cycle. Study 2 assessed the role of ovarian hormones in withdrawal by comparing the same measures in OVX rats that received vehicle, E2, or E2+progesterone prior to testing. Briefly, rats received a sham surgery or an ovariectomy procedure. Fifteen days later, rats were prepared with a pump that delivered nicotine for 14 days. On the test day, rats received saline or the nicotinic receptor antagonist, mecamylamine to precipitate withdrawal. Physical signs and anxiety-like behavior were assessed on the elevated plus maze (EPM) and light-dark transfer (LDT) tests. During withdrawal, intact females displayed greater anxiety-like behavior and corticosterone levels as compared to male and OVX rats. Females tested in estrus (when E2 is relatively low) displayed less anxiety-like behavior and corticosterone versus all other phases. Anxiety-like behavior and corticosterone were positively correlated with E2 and negatively correlated with progesterone. Intact females displaying high E2/low progesterone displayed greater anxiety-like behavior and corticosterone as compared to females displaying low E2/high progesterone. Lastly, OVX-E2 rats displayed greater anxiety-like behavior than OVX-E2+progesterone rat. These data suggest that E2 promotes and progesterone reduces anxiety-like behavior produced by withdrawal.

animal behavior and cognition↗

bioRxiv2019

Optimized double emulsion flow cytometry with high-throughput single droplet isolation

Droplet microfluidics has made large impacts in diverse areas such as enzyme evolution, chemical product screening, polymer engineering, and single-cell analysis. However, while droplet reactions have become increasingly sophisticated, phenotyping droplets by a fluorescent signal and sorting them to isolate variants-of-interest remains a field-wide bottleneck. Here, we present an optimized double emulsion workflow, sdDE-FACS, that enables high-throughput phenotyping, selection, and sorting of droplets using standard flow cytometers. Using a 130 m nozzle, we demonstrate robust post-sort recovery of intact droplets, with little to no shear-induced droplet breakage, at high sort frequency (12-14 kHz) across two industry-standard FACS instruments. We report the first quantitative plate statistics for double emulsion droplet isolation and demonstrate single droplet recovery with >70% efficiency. In addition, we establish complete downstream recovery of nucleic acids from single, sorted double emulsion droplets, an advance in droplet sorting comparable with the capabilities of single-cell FACS. This work resolves several hurdles in the field of high-throughput droplet analysis and paves the way for a variety of new droplet assays, including rare variant isolation and multiparameter single-cell analysis, marrying the full power of flow cytometry with droplet microfluidics.

bioengineering↗

bioRxiv2018

Live imaging of Aiptasia larvae, a model system for studying coral bleaching, using a simple microfluidic device

Coral reefs, and their associated diverse ecosystems, are of enormous ecological importance. In recent years, coral health has been severely impacted by environmental stressors brought on by human activity and climate change, threatening the extinction of several major reef ecosystems. Reef damage is mediated by a process called coral bleaching where corals, sea anemones, and other cnidarians lose their photosynthetic algal symbionts (genus Symbiodinium) upon stress induction, resulting in drastically decreased host energy harvest and, ultimately, coral death. The mechanism by which this critical cnidarian-algal symbiosis is lost remains poorly understood. Here, we report Traptasia, a simple microfluidic device with multiple traps designed to isolate and image individual live larvae of Aiptasia, a sea anemone model organism, and their algal symbionts over extended time courses. Aiptasia larvae are ~100 m in length, deformable, and highly motile, posing particular challenges for long-term imaging. Using a trap design optimized via fluid flow simulations and polymer bead loading tests, we trapped Aiptasia larvae containing algal symbionts and demonstrated stable imaging for >10 hours. We visualized algal migration within Aiptasia larvae and observed algal expulsion under an environmental stressor. To our knowledge, this device is the first to enable live imaging of cnidarian larvae and their algal symbionts and, in further implementation, could provide important insights into the cellular mechanisms of coral bleaching under different environmental stressors. The device is simple to use, requires minimal external equipment and no specialized training to operate, and can easily be adapted to study a variety of large, motile organisms.

ecology↗

bioRxiv2017

An Open-Source, Programmable Pneumatic Setup for Operation and Automated Control of Single- and Multi-Layer Microfluidic Devices

Microfluidic technologies have been used across diverse disciplines (e.g. high-throughput biological measurement, fluid physics, laboratory fluid manipulation) but widespread adoption has been limited due to the lack of openly disseminated resources that enable non-specialist labs to make and operate their own devices. Here, we report the open-source build of a pneumatic setup capable of operating both single and multilayer (Quake-style) microfluidic devices with programmable scripting automation. This setup can operate both simple and complex devices with 48 device valve control inputs and 18 sample inputs, with modular design for easy expansion, at a fraction of the cost of similar commercial solutions. We present a detailed step-by-step guide to building the pneumatic instrumentation, as well as instructions for custom device operation using our software, Geppetto, through an easy-to-use GUI for live on-chip valve actuation and a scripting system for experiment automation. We show robust valve actuation with near real-time software feedback and demonstrate use of the setup for high-throughput biochemical measurements on-chip. This open-source setup will enable specialists and novices alike to run microfluidic devices easily in their own laboratories.\n\nO_TBL View this table:\norg.highwire.dtl.DTLVardef@d25ba2org.highwire.dtl.DTLVardef@7dfd9eorg.highwire.dtl.DTLVardef@1959573org.highwire.dtl.DTLVardef@186dceaorg.highwire.dtl.DTLVardef@111af70_HPS_FORMAT_FIGEXP M_TBL O_TABLECAPTIONSpecifications table\n\nC_TABLECAPTION C_TBL

bioengineering↗