Biology subjects

Clark, A. M.

Publications and source records attributed to Clark, A. M..

11 recordsLinked to original sources

bioRxiv2025

Laminar-specific control of response gain and orientation-tuning byparvalbumin-expressing inhibitory interneurons in primate visual cortex

Understanding the role of the different types of inhibitory interneurons in cortical computations is central to elucidating how the neocortex processes sensory information. The emergence of orientation tuning in primate primary visual cortex (V1) is a canonical model for studying how cortical sensory circuits and inhibitory interneurons compute relevant stimulus features. The selective feedforward convergence of non-orientation-selective thalamic afferents establishes initial orientation tuning in the granular V1 input layer. As signals propagate through the cortical microcircuit, orientation tuning sharpens in extra-granular layers, yet the underlying mechanisms and the contribution of specific inhibitory neuron subtypes within V1 remain unresolved. To study the role of the largest cortical inhibitory neuron subclass, parvalbumin-expressing (PV+) interneurons, in this V1 computation, we combined laminar extracellular recordings with bidirectional optogenetic manipulations of PV+ cells in marmoset V1. Our results reveal a striking laminar specificity: in granular layers, PV+ cells implement divisive/ multiplicative linear gain control, whereas in extra-granular layers they exert tuned nonlinear suppression that enhances orientation tuning. Computational modeling suggests that PV+ neurons can control gain by changing a neurons spiking threshold, and orientation tuning by changing a neurons input noise, which regulates the neurons input-output function. Our findings reconcile divergent results from previous rodent studies and establish a framework for understanding layer-dependent inhibitory computations in the primate cortex.

neuroscience↗

bioRxiv2025

Distinct growth regimes govern crowding in foveal and extrafoveal vision

Visual crowding--where object recognition is impaired by nearby stimuli--is a well-documented phenomenon in the visual periphery, thought to reflect fundamental processes of grouping and segmentation. A hallmark of crowding is its spatial extent increasing linearly with eccentricity, as described by Boumas law, reflecting retinal convergence and cortical magnification. Although crowding is typically studied in the periphery--and Boumas law predicts no crowding at the center of gaze--it occurs even foveally. Within the foveola, the central 1 deg of the visual field, characterized by peak visual acuity and largely one-to-one connectivity between photoreceptors and ganglion cells, the Bouma law breaks and the extent of crowding is expected to remain constant with eccentricity. Yet, whether crowding varies at this fine spatial scale remains unknown. To investigate this, we combined high-resolution eye tracking with a gaze-contingent display system to precisely localize gaze and measure crowding thresholds at multiple foveolar eccentricities. Our results reveal that crowding does increase linearly within the foveola, but at a rate significantly slower--approximately 3.5 times-than in the extrafovea. By showing that even within the highest-acuity region of the visual field, spatial integration zones are not fixed but change with eccentricity these findings challenge the view of the central fovea as a spatially uniform processing zone and demonstrate that crowding at this scale follows a distinct regime. SignificanceWhen we view objects in clutter, recognition declines--a phenomenon known as crowding. This effect increases with distance from the center of gaze, following Boumas law, which predicts little or no crowding at the foveas center. As this law breaks at the center of gaze, it has been hypothesized that around this region, crowding remains constant with eccentricity. Using high-resolution eyetracking, we show that even within the central 0.4{degrees} of the visual field, crowding increases steadily with minute stimulus offsets from the point of fixation. This growth is much slower than in peripheral vision, revealing a distinct crowding regime in the foveola, likely reflecting cortical and retinal differences. Our findings offer new insights into the fine-scale organization of spatial vision.

neuroscience↗

bioRxiv2025

Subfoveal scotomas trigger fine-scale fixation reorganization: insights from retinal imaging and retinal-contingent stimulation

Fine spatial vision relies on the foveola, the 1-degree retinal region with highest cone density. Despite its importance, the relationship between retinal anatomy, fixational behavior, and visual perception in the foveola is not fully understood. Using an Adaptive Optics Scanning Light Ophthalmoscope for high-resolution retinal imaging and stimulation, we studied the effect of a simulated subfoveolar ({approx}0.03 degrees2) scotoma on fine spatial vision and fixation behavior in healthy observers. Our findings show that the visuomotor system adapts to the scotoma with striking precision by shifting the preferred locus of fixation in a systematic fashion by minute ({approx}5 arcmin) amounts to bring stimuli into a region of visibility. These results reveal an unprecedented level of fine-scale plasticity in the human visuomotor system. Interestingly, this new retinal locus of fixation is characterized by lower cone density among those surrounding the scotoma, indicating that factors beyond spatial sampling maximization influence these fine-scale adjustments.

neuroscience↗

bioRxiv2025

Cortical Response to Acute Implantation of the Utah Optrode Array in Macaque Cortex

Optogenetics has transformed the study of neural circuit function, but limitations in its application to species with large brains, such as non-human primates (NHPs), remain. A major challenge in NHP optogenetics is delivering light to sufficiently large volumes of deep neural tissue with high spatiotemporal precision, without simultaneously affecting superficial tissue. To overcome these limitations, we recently developed and tested in vivo in NHP cortex, the Utah Optrode Array (UOA). This is a 10x10 array of penetrating glass shanks, tiling a 4x4mm2 area, bonded to interleaved needle-aligned and interstitial {micro}LED arrays, which allows for independent photostimulation of deep and superficial brain tissue. Here, we investigate the acute biological response to UOA implantation in NHP cortex, with the goal of optimizing device design for reduced insertion trauma and subsequent chronic response. To this goal, we systematically vary UOA shank diameter, surface texture, tip geometry, and insertion pressure, and assess their effects on astrocytes, microglia, and neuronal viability, following acute implantation. We find that UOAs with shanks of smaller diameter, smooth surface texture and round tips cause the least damage. Higher insertion pressures have limited effects on the inflammatory response, but lead to greater tissue compression. Our results highlight the importance of balancing shank diameter, tip geometry, and insertion pressure in UOA design for preserving tissue integrity and improving long-term UOA performance and biocompatibility.

bioengineering↗

bioRxiv2024

Identification of the MRTFA/SRF pathway as a critical regulator of quiescence in cancer.

Chemoresistance is a major driver of cancer deaths. One understudied mechanism of chemoresistance is quiescence. We used single cell culture to identify, retrieve, and RNA-Seq profile primary quiescent ovarian cancer cells (qOvCa). We found that many qOvCa differentially expressed genes are transcriptional targets of the Myocardin Related Transcription Factor/Serum Response Factor (MRTF/SRF) pathway. We also found that genetic disruption of MRTF-SRF interaction, or an MRTF/SRF inhibitor (CCG257081) impact qOvCa gene expression and induce a quiescent state in cancer cells. Suggesting a broad role for this pathway in quiescence, CCG257081 treatment induced quiescence in breast, lung, colon, pancreatic and ovarian cancer cells. Furthermore, CCG081 (i) maintained a quiescent state in patient derived breast cancer organoids and, (ii) induced tumor growth arrest in ovarian cancer xenografts. Together, these data suggest that MRTF/SRF pathway is a critical regulator of quiescence in cancer and a possible therapeutic target. SignificanceQuiescence is a critical driver of chemoresistance. The MRFT-SRF pathway regulates cancer cell quiescence and inhibiting the MRTF-SRF pathway can prevent the outgrowth of quiescent cancer cells and improve cancer outcomes.

cancer biology↗

bioRxiv2024

Hope for Others: Research Results from the University of Pittsburgh Rapid Autopsy Program for Breast Cancer

Breast cancer affects 1/8 of women throughout their lifetimes, with over 90% of cancer deaths being caused by metastasis. However, metastasis poses unique challenges to research, as complex changes in the microenvironment in different metastatic sites and difficulty obtaining tissue for study hinder the ability to examine in depth the changes that occur during metastasis. Rapid autopsy programs thus fill a unique need in advancing metastasis research. Here, we describe our protocol and processes for establishing and improving the US-based Hope for OTHERS (Our Tissue Helping Enhance Research and Science) program for organ donation in metastatic breast cancer. As of August 2024, we consented 114 patients and performed 37 autopsies, from which we collected 551 unique metastatic frozen tumor samples, 1244 FFPE blocks, 90 longitudinal liquid biopsy samples and developed 14 patient-derived organoid and 8 patient-derived xenograft models. We report in-depth clinical and histopathological information and discuss extensive new research and novel findings in patient outcomes, metastatic phylogeny, and factors in successful living model development. Our results reveal key logistical and protocol improvements that are uniquely beneficial to certain programs based on identifiable features, such as working closely with patient advocates, methods to rescue RNA quality in cases where tissue quality may degrade due to time delays, as well as guidelines and future expansions of our program. Statement of SignificanceRapid autopsy programs are unique research settings with huge potential for studying metastatic cancer, however, they have complex research challenges. Our work provides a valuable resource in advancing this field of research.

cancer biology↗

bioRxiv2022

High-resolution eye-tracking via digital imaging of Purkinje reflections

Reliably measuring eye movements and determining where the observer looks are fundamental needs in vision science. A classical approach to achieve high-resolution oculomotor measurements is the so-called Dual-Purkinje-Image (DPI) method, a technique that relies on the relative motion of the reflections generated by two distinct surfaces in the eye, the cornea and the back of the lens. This technique has been traditionally implemented in fragile and difficult to operate analog devices, which have remained exclusive use of specialized oculomotor laboratories. Here we describe progress on the development of a digital DPI, a system that builds on recent advances in digital imaging to enable fast, highly precise eye-tracking without the complications of previous analog devices. This system integrates an optical setup with no moving components with a digital imaging module and dedicated software on a fast processing unit. Data from both artificial and human eyes demonstrate sub-arcminute resolution at 1 Khz. Furthermore, when coupled with previously developed gaze-contingent calibration methods, this system enables localization of the line of sight within a few arcminutes.

neuroscience↗

bioRxiv2022

Disruption of CFAP418 interaction with lipids causes abnormal membrane-associated cellular processes in retinal degenerations

Syndromic ciliopathies and retinal degenerations are large heterogeneous groups of genetic diseases. CFAP418 is a causative gene of both disorders, and its protein sequence is evolutionarily conserved. However, the pathogenic mechanism caused by CFAP418 mutations is largely unknown. Here, we employed affinity purification coupled with mass spectrometry and quantitative lipidomic, proteomic, and phosphoproteomic approaches to address the molecular function of CFAP418 in mouse retinas. We showed that CFAP418 bound to lipid metabolism precursor phosphatidic acid (PA) and mitochondrion-specific lipid cardiolipin but did not form a tight and static complex with proteins. Loss of Cfap418 led to membrane lipid imbalance and protein-membrane association alteration, which subsequently caused mitochondrial defects and membrane remodeling abnormalities in multiple vesicular trafficking pathways. Loss of Cfap418 also increased the activity of PA-binding protein kinase C. Our results indicate that membrane lipid imbalance is a new pathological mechanism underlying syndromic ciliopathies and retinal degenerations, which is associated with other known causative genes for these diseases, such as RAB28 and BBS genes.

cell biology↗

bioRxiv2022

Individual differences in eye drift predict visual acuity

Visual acuity is commonly assumed to be determined by the eye optics and spatial sampling in the retina. Unlike a camera, however, the eyes are never stationary during the acquisition of visual information, a jittery motion known as ocular drift, incessantly displaces stimuli over many photoreceptors. Previous studies have shown that acuity is impaired in the absence of retinal image motion caused by eye drift. However, the relation between individual drift characteristics and acuity remains unknown. Here we show that (a) healthy emmetropes exhibit a large variability in their amount of drift; and (b) that these differences profoundly affect the structure of spatiotemporal signals to the retina. We further show that (c) the spectral distribution of the resulting luminance modulations strongly correlates with individual visual acuity; and (d) that natural inter-trial fluctuations in the amount of drift modulate acuity. As a consequence, in healthy emmetropes acuity can be predicted from the motor behavior elicited by a simple fixation task, without directly measuring it. These results shed new light on how oculomotor behavior contributes to fine spatial vision. SignificanceHealthy humans can visually resolve extremely fine patterns, in some cases with the relevant features spanning less than a single photoreceptor on the retina. This accomplishment is particularly remarkable considering that the eyes are never stationary. Ocular drift--a motion that eludes human awareness--shifts the stimulus across many photoreceptors during the acquisition of visual information. Here we show that visual acuity depends on ocular drift. Natural variations in the amount of drift are associated with acuity both within and across subjects, so that individual acuity limits can be directly inferred from the amount of motion during fixation on a marker. Results closely follow the strength of the luminance modulations caused by ocular drift, providing support to long-standing dynamic theories of visual acuity.

neuroscience↗

bioRxiv2022

An Optrode Array for Spatiotemporally Precise Large-Scale Optogenetic Stimulation of Deep Cortical Layers in Non-human Primates

Optogenetics has transformed studies of neural circuit function, but remains challenging to apply in large brains, such as those of non-human primates (NHPs). A major challenge is delivering intense, spatiotemporally precise, patterned photostimulation across large volumes in deep tissue. Such stimulation is critical, for example, to modulate selectively deep-layer corticocortical feedback projections. To address this unmet need, we have developed the Utah Optrode Array (UOA), a 10x10 glass needle waveguide array fabricated atop a novel opaque optical interposer then bonded to an electrically addressable LED array. In vivo experiments with the UOA demonstrated large-scale, spatiotemporally precise, activation of deep circuits in monkey cortex. Specifically, the UOA permitted both focal (confined to single layers/columns), and widespread (multiple layers/columns) optogenetic activation of deep layer neurons, simply by varying the number of activated LEDs and/or the irradiance. Thus, the UOA represents a powerful optoelectronic device for targeted manipulation of deep-layer circuits in NHP models.

neuroscience↗

bioRxiv2021

Interferon-γ Increases Sensitivity to Chemotherapy and Provides Immunotherapy Targets in Models of Metastatic Castration-Resistant Prostate Cancer

Interferon-{gamma} (IFN{gamma}) is a cytokine with limited evidence of benefit in cancer clinical trials to date. However, it could potentially play a role in potentiating anti-tumor immunity in the immunologically "cold" metastatic castration-resistant prostate cancer (mCRPC) by inducing antigen presentation pathways and concurrently providing targets for immune checkpoint blockade therapy. Moreover, it could additionally increase sensitivity to chemotherapy based on its pleiotropic effects on cell phenotype. Here, we show that IFN{gamma} treatment induced expression of major histocompatibility class-I (MHC-I) genes and PD-L1 in prostate cancer cells in vitro. Furthermore, IFN{gamma} treatment led to a decrease in E-cadherin expression with a consequent increase in sensitivity to chemotherapy in vitro. In an in vivo murine tumor model of spontaneous metastatic prostate cancer, IFN{gamma} systemic pretreatment upregulated the expression of HLA-A and decreased E-cadherin expression in the primary tumor, and more importantly in the metastatic site led to increased apoptosis and limited micrometastases in combination with paclitaxel treatment compared to diffuse metastatic disease in control and monotherapy treatment groups. These findings suggest that IFN{gamma} may be useful in combinatorial regimens to induce sensitivity to immunotherapy and chemotherapy in hepatic metastases of mCRPC.

cancer biology↗