Biology subjects

Greene, S.

Publications and source records attributed to Greene, S..

6 recordsLinked to original sources

bioRxiv2025

A parallelly distributed microscope and software system for scalable high-throughput multispectral 3D imaging

Recent advances in high throughput optical microscopy have achieved whole-organism scale imaging at diffraction-limited resolutions. Current microscopes, however, require making compromises between achieving the optimal resolution, imaging depth, multispectral capability, and data throughput due to limitations in optical design and data stream handling. We have created a parallel-line scanning confocal microscope (plSCM) which provides 1.1 Gigavoxels/second high speed imaging while achieving an optical resolution of [~]180 x 220 x 650 nm with 2 millimeters of imaging depth in 3 simultaneous spectral channels. To handle such a massive imaging data stream, we have engineered a scalable network-distributed image acquisition/processing framework (SNDiF), which allows continuous capture, real-time processing, and cluster storage of petabyte-scale single image datasets in days. Together, our parallelly distributed microscope and software system presents a general solution for large-scale high-throughput and high-resolution multicolor imaging which can operate for days at a time.

bioengineering↗

bioRxiv2024

Unique Signatures of Highly Constrained Genes Across Publicly Available Genomic Databases

Publicly available genomic databases and genetic constraint scores are crucial in understanding human population variation and the identification of variants that are likely to have a deleterious impact causing human disease. We utilized the one of largest publicly available databases, gnomAD, to determine genes that are highly constrained for only LoF, only missense, and both LoF/missense variants, identified their unique signatures, and explored their causal relationship with human conditions. Those genes were evaluated for unique patterns including their chromosomal location, tissue level expression, gene ontology analysis, and gene family categorization using multiple publicly available databases. Those highly constrained genes associated with human disease, we identified unique patterns of inheritance, protein size, and enrichment in distinct molecular pathways. In addition, we identified a cohort of highly constrained genes that are currently not known to cause human disease, that we suggest will be candidates to pursue as novel disease-associated genes. In summary, these insights not only elucidate biological pathways of highly constrained genes that expand our understanding of critical cellular proteins but also advance research in rare diseases.

genetics↗

bioRxiv2023

Progressive circuit hyperexcitability in mouse neocortical slice cultures with increasing duration of activity silencing

Forebrain neurons deprived of activity become hyperactive when activity is restored. Rebound activity has been linked to spontaneous seizures in vivo following prolonged activity blockade. Here we measured the time course of rebound activity and the contributing circuit mechanisms using calcium imaging, synaptic staining and whole cell patch clamp in organotypic slice cultures of mouse neocortex. Calcium imaging revealed hypersynchronous activity increasing in intensity with longer periods of deprivation. While activity partially recovered three days after slices were released from five days of deprivation, they were less able to recover after ten days of deprivation. However, even after the longer period of deprivation, activity patterns eventually returned to baseline levels. The degree of deprivation-induced rebound was age-dependent, with the greatest effects occurring when silencing began in the second week. Pharmacological blockade of NMDA receptors indicated that hypersynchronous rebound activity did not require Hebbian plasticity evoked. In single neuron recordings, input resistance roughly doubled with a concomitant increase in intrinsic excitability. Synaptic imaging of pre- and postsynaptic proteins revealed dramatic reductions in the number of presumptive synapses with a larger effect on inhibitory than seen in excitatory synapses. Putative excitatory synapses colocalizing PSD-95 and Bassoon declined by 39% and 56% following five and ten days of deprivation, but presumptive inhibitory synapses colocalizing gephyrin and VGAT declined by 55% and 73% respectively. The results suggest that with prolonged deprivation, a progressive reduction in synapse number is accompanied by a shift in the balance between excitation and inhibition and increased cellular excitability. SIGNIFICANCE STATEMENTWhen cortical activity is silenced during development, lifelong seizures often result. Here we explored whether these seizures result from overcompensation of homeostatic recovery mechanisms. Prior work showed that neurons briefly deprived of the ability to fire action potentials compensate by becoming more excitable, increasing synaptic drive and intrinsic excitability. We found that prolonged silencing of cortex produced a profound loss of synapse density, especially for inhibitory synapses, pointing to a circuit unable to maintain excitatory/inhibitory balance. These results show that homeostatic responses, which are normally restorative, can result in maladaptive circuit configurations when brought to extremes.

neuroscience↗

bioRxiv2023

Prolonged activity-deprivation causes pre- and postsynaptic compensatory plasticity at neocortical excitatory synapses

Homeostatic plasticity stabilizes firing rates of neurons, but the pressure to restore low activity rates can significantly alter synaptic and cellular properties. Most previous studies of homeostatic readjustment to complete activity silencing in rodent forebrain have examined changes after two days of deprivation, but it is known that longer periods of deprivation can produce adverse effects. To better understand the mechanisms underlying these effects and to address how presynaptic as well as postsynaptic compartments change during homeostatic plasticity, we subjected mouse cortical slice cultures to a more severe five-day deprivation paradigm. We developed and validated a computational framework to measure the number and morphology of presynaptic and postsynaptic compartments from super resolution light microscopy images of dense cortical tissue. Using these tools, combined with electrophysiological miniature excitatory postsynaptic current measurements, and synaptic imaging at the electron microscopy level, we assessed the functional and morphological results of prolonged deprivation. Excitatory synapses were strengthened both presynaptically and postsynaptically. Surprisingly, we also observed a decrement in the density of excitatory synapses, both as measured from colocalized staining of pre- and postsynaptic proteins in tissue, and from the number of dendritic spines. Overall, our results suggest that cortical networks deprived of activity progressively move towards a smaller population of stronger synapses. SIGNIFICANCE STATEMENTBlocking activity in neocortical slice cultures produced coordinated pre and postsynaptic changes at excitatory synapses. Functional and structural assays suggest that deprivation results in fewer excitatory synapses, but each is strengthened both pre- and postsynaptically. This may contribute to the emergence of epileptiform activity.

neuroscience↗

bioRxiv2020

Process-based model of fish incubation survival for designing reservoir operations: a case study for Sacramento River winter-run Chinook salmon

Allocating reservoir flows to meet societal and ecosystem needs under increasing demands for water and increasing climatic variability presents challenges to resource managers. Often, regulated rivers have been operated to meet flow and temperature compliance points that mimic historical patterns. Because it is difficult to assess if this approach is efficient or equitable, new more process-based approaches to regulation are being advanced. This paper describes such an approach with a model of egg incubation survival of Sacramento River winter-run Chinook salmon (SRWRC, Oncorhynchus tshawytscha). Thermal mortality only occurs in a critical window around egg hatching when the embryo is most sensitive to temperature stress. The duration of the critical window has significant implications for Shasta Reservoir operations that are designed to control temperature during SRWRC incubation. Previous operations sought to maintain a low temperature over the entire incubation period. However, model analysis suggests that targeting cold water directly to the critical egg hatching stage provides higher survival while requiring less cold water resources. The calibrated model is publicly accessible through a web interface connected to real-time river and fish databases and a river temperature forecast model. The system is an example of the next step of river management that integrates databases with hydrological and process-based biological models for real-time analysis and for forecasting effects of river operations on the environment.

ecology↗

bioRxiv2020

Tumor control via targeting PD-L1 with chimeric antigen receptor modified NK cells

Failed T cell-based immunotherapies in the presence of genomic alterations in antigen presentations pathways may be overcome by NK cell-based immunotherapy. This approach may still be limited by the presence of immunosuppressive myeloid populations. Here we demonstrate that NK cells (haNKs) engineered to express a PD-L1 chimeric antigen receptor (CAR) haNKs killed a panel of human and murine head and neck cancer cells at low effector-to-target ratios in a PD-L1-dependent fashion. Treatment of syngeneic tumors resulted in CD8 and PD-L1-dependent tumor rejection or growth inhibition and a reduction in myeloid cells endogenously expressing high levels of PD-L1. Treatment of xenograft tumors resulted in PD-L1 dependent tumor growth inhibition. PD-L1 CAR haNKs reduced levels of macrophages and other myeloid cells endogenously expressing high PD-L1 in peripheral blood from patients with head and neck cancer. The clinical study of PD-L1 CAR haNKs is warranted.

immunology↗