Publications by authors named "Gregory W Schwartz"

32 Publications

The vascular landscape of human cancer.

J Clin Invest 2021 Jan;131(2)

Department of Medicine.

Tumors depend on a blood supply to deliver oxygen and nutrients, making tumor vasculature an attractive anticancer target. However, only a fraction of patients with cancer benefit from angiogenesis inhibitors. Whether antiangiogenic therapy would be more effective if targeted to individuals with specific tumor characteristics is unknown. To better characterize the tumor vascular environment both within and between cancer types, we developed a standardized metric - the endothelial index (EI) - to estimate vascular density in over 10,000 human tumors, corresponding to 31 solid tumor types, from transcriptome data. We then used this index to compare hyper- and hypovascular tumors, enabling the classification of human tumors into 6 vascular microenvironment signatures (VMSs) based on the expression of a panel of 24 vascular "hub" genes. The EI and VMS correlated with known tumor vascular features and were independently associated with prognosis in certain cancer types. Retrospective testing of clinical trial data identified VMS2 classification as a powerful biomarker for response to bevacizumab. Thus, we believe our studies provide an unbiased picture of human tumor vasculature that may enable more precise deployment of antiangiogenesis therapy.
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http://dx.doi.org/10.1172/JCI136655DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7810485PMC
January 2021

An offset ON-OFF receptive field is created by gap junctions between distinct types of retinal ganglion cells.

Nat Neurosci 2021 01 23;24(1):105-115. Epub 2020 Nov 23.

Departments of Ophthalmology and Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA.

In the vertebrate retina, the location of a neuron's receptive field in visual space closely corresponds to the physical location of synaptic input onto its dendrites, a relationship called the retinotopic map. We report the discovery of a systematic spatial offset between the ON and OFF receptive subfields in F-mini-ON retinal ganglion cells (RGCs). Surprisingly, this property does not come from spatially offset ON and OFF layer dendrites, but instead arises from a network of electrical synapses via gap junctions to RGCs of a different type, the F-mini-OFF. We show that the asymmetric morphology and connectivity of these RGCs can explain their receptive field offset, and we use a multicell model to explore the effects of receptive field offset on the precision of edge-location representation in a population. This RGC network forms a new electrical channel combining the ON and OFF feedforward pathways within the output layer of the retina.
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http://dx.doi.org/10.1038/s41593-020-00747-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7769921PMC
January 2021

Reversed Neurovascular Coupling on Optical Coherence Tomography Angiography Is the Earliest Detectable Abnormality before Clinical Diabetic Retinopathy.

J Clin Med 2020 Oct 31;9(11). Epub 2020 Oct 31.

Department of Ophthalmology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA.

Diabetic retinopathy (DR) has traditionally been viewed as either a microvasculopathy or a neuropathy, though neurovascular coupling deficits have also been reported and could potentially be the earliest derangement in DR. To better understand neurovascular coupling in the diabetic retina, we investigated retinal hemodynamics by optical coherence tomography angiography (OCTA) in individuals with diabetes mellitus (DM) but without DR (DM no DR) and mild non-proliferative DR (mild NPDR) compared to healthy eyes. Using an experimental design to monitor the capillary responses during transition from dark adaptation to light, we examined 19 healthy, 14 DM no DR and 11 mild NPDR individuals. We found that the only structural vascular abnormality in the DM no DR group was increased superficial capillary plexus (SCP) vessel density (VD) compared to healthy eyes, while mild NPDR eyes showed significant vessel loss in the SCP at baseline. There was no significant difference in inner retinal thickness between the groups. During dark adaptation, the deep capillary plexus (DCP) VD was lower in mild NPDR individuals compared to the other two groups, which may leave the photoreceptors more susceptible to ischemia in the dark. When transitioning from dark to ambient light, both diabetic groups showed a qualitative reversal of VD trends in the SCP and middle capillary plexus (MCP), with significantly decreased SCP at 5 min and increased MCP VD at 50 s compared to healthy eyes, which may impede metabolic supply to the inner retina during light adaptation. Mild NPDR eyes also demonstrated DCP dilation at 50 s and 5 min and decreased adjusted flow index at 5 min in light. Our results show altered neurovascular responses in all three macular vascular plexuses in diabetic subjects in the absence of structural neuronal changes on high resolution imaging, suggesting that neurovascular uncoupling may be a key mechanism in the early pathogenesis of DR, well before the clinical appearance of vascular or neuronal loss.
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http://dx.doi.org/10.3390/jcm9113523DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692675PMC
October 2020

Caffeine Delays Retinal Neurovascular Coupling during Dark to Light Adaptation in Healthy Eyes Revealed by Optical Coherence Tomography Angiography.

Invest Ophthalmol Vis Sci 2020 04;61(4):37

,.

Purpose: The purpose of this study was to investigate the acute effects of caffeine on retinal hemodynamics during dark to light adaptation using optical coherence tomography angiography (OCTA).

Methods: Thirteen healthy individuals (13 eyes) underwent OCTA imaging after dark adaptation and at repeated intervals during the transition to ambient light in two imaging sessions: control and after ingesting 200 mg of caffeine. We analyzed the parafoveal vessel density (VD) and adjusted flow index (AFI) of the superficial capillary plexus (SCP), middle capillary plexus (MCP), and deep capillary plexus (DCP), as well as the vessel length density (VLD) of the SCP. After adjusting for age, refractive error, and scan quality, we compared parameters between control and caffeine conditions.

Results: In the dark, MCP VD decreased significantly after caffeine (-2.63 ± 1.28%). During the transition to light, initially, DCP VD increased (12.55 ± 2.52%), whereas SCP VD decreased (-2.09 ± 0.91%) significantly with caffeine compared to control. By 15 minutes in light, DCP VD reversed and was significantly decreased (-5.45 ± 2.62%), whereas MCP VD increased (4.65 ± 1.74%). There were no differences in AFI or VLD.

Conclusions: We show that, overall, caffeine causes a trend of delayed vascular response in all three macular capillary plexuses in response to ambient light. Whereas the MCP is constricted in the dark, during the transition from dark to light, there is initially delay followed by prolonged constriction of the DCP and constriction followed by slow dilation of the SCP. We posit that these delayed vascular responses may present potential risk of capillary ischemia.
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http://dx.doi.org/10.1167/iovs.61.4.37DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7401906PMC
April 2020

Trib1 regulates T cell differentiation during chronic infection by restraining the effector program.

J Exp Med 2020 05;217(5)

Department of Pathology and Laboratory Medicine, Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA.

In chronic infections, the immune response fails to control virus, leading to persistent antigen stimulation and the progressive development of T cell exhaustion. T cell effector differentiation is poorly understood in the context of exhaustion, but targeting effector programs may provide new strategies for reinvigorating T cell function. We identified Tribbles pseudokinase 1 (Trib1) as a central regulator of antiviral T cell immunity, where loss of Trib1 led to a sustained enrichment of effector-like KLRG1+ T cells, enhanced function, and improved viral control. Single-cell profiling revealed that Trib1 restrains a population of KLRG1+ effector CD8 T cells that is transcriptionally distinct from exhausted cells. Mechanistically, we identified an interaction between Trib1 and the T cell receptor (TCR) signaling activator, MALT1, which disrupted MALT1 signaling complexes. These data identify Trib1 as a negative regulator of TCR signaling and downstream function, and reveal a link between Trib1 and effector versus exhausted T cell differentiation that can be targeted to improve antiviral immunity.
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http://dx.doi.org/10.1084/jem.20190888DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7201917PMC
May 2020

TooManyCells identifies and visualizes relationships of single-cell clades.

Nat Methods 2020 04 2;17(4):405-413. Epub 2020 Mar 2.

Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA, USA.

Identifying and visualizing transcriptionally similar cells is instrumental for accurate exploration of the cellular diversity revealed by single-cell transcriptomics. However, widely used clustering and visualization algorithms produce a fixed number of cell clusters. A fixed clustering 'resolution' hampers our ability to identify and visualize echelons of cell states. We developed TooManyCells, a suite of graph-based algorithms for efficient and unbiased identification and visualization of cell clades. TooManyCells introduces a visualization model built on a concept intentionally orthogonal to dimensionality-reduction methods. TooManyCells is also equipped with an efficient matrix-free divisive hierarchical spectral clustering different from prevalent single-resolution clustering methods. TooManyCells enables multiresolution and multifaceted exploration of single-cell clades. An advantage of this paradigm is the immediate detection of rare and common populations that outperforms popular clustering and visualization algorithms, as demonstrated using existing single-cell transcriptomic data sets and new data modeling drug-resistance acquisition in leukemic T cells.
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http://dx.doi.org/10.1038/s41592-020-0748-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7439807PMC
April 2020

Serine substitutions are linked to codon usage and differ for variable and conserved protein regions.

Sci Rep 2019 11 21;9(1):17238. Epub 2019 Nov 21.

Drexel School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, USA.

Serine is the only amino acid that is encoded by two disjoint codon sets (TCN & AGY) so that a tandem substitution of two nucleotides is required to switch between the two sets. We show that these codon sets underlie distinct substitution patterns at positions subject to purifying and diversifying selections. We found that in humans, positions that are conserved among ~100 vertebrates, and thus subjected to purifying selection, are enriched for substitutions involving serine (TCN, denoted S'), proline, and alanine, (S'PA). In contrast, the less conserved positions are enriched for serine encoded with AGY codons (denoted S″), glycine and asparagine, (GS″N). We tested this phenomenon in the HIV envelope glycoprotein (gp120), and the V-gene that encodes B-cell receptors/antibodies. These fast evolving proteins both have hypervariable positions, which are under diversifying selection, closely adjacent to highly conserved structural regions. In both instances, we identified an opposite abundance of two groups of serine substitutions, with enrichment of S'PA in the conserved positions, and GS″N in the hypervariable regions. Finally, we analyzed the substitutions across 60,000 individual human exomes to show that, when serine has a specific functional constraint of phosphorylation capability, S' codons are 32-folds less prone than S″ to substitutions to Threonine or Tyrosine that could potentially retain the phosphorylation site capacity. Combined, our results, that cover evolutionary signals at different temporal scales, demonstrate that through its encoding by two codon sets, serine allows for the existence of alternating substitution patterns within positions of functional maintenance versus sites of rapid diversification.
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http://dx.doi.org/10.1038/s41598-019-53452-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6872785PMC
November 2019

Molecular signatures of retinal ganglion cells revealed through single cell profiling.

Sci Rep 2019 10 31;9(1):15778. Epub 2019 Oct 31.

Emmune, Inc, 14155 U.S Highway 1, Juno Beach, FL, 33408, USA.

Retinal ganglion cells can be classified into more than 40 distinct subtypes, whether by functional classification or transcriptomics. The examination of these subtypes in relation to their physiology, projection patterns, and circuitry would be greatly facilitated through the identification of specific molecular identifiers for the generation of transgenic mice. Advances in single cell transcriptomic profiling have enabled the identification of molecular signatures for cellular subtypes that are only rarely found. Therefore, we used single cell profiling combined with hierarchical clustering and correlate analyses to identify genes expressed in distinct populations of Parvalbumin-expressing cells and functionally classified RGCs. RGCs were manually isolated based either upon fluorescence or physiological distinction through cell-attached recordings. Microarray hybridization and RNA-Sequencing were employed for the characterization of transcriptomes and in situ hybridization was utilized to further characterize gene candidate expression. Gene candidates were identified based upon cluster correlation, as well as expression specificity within physiologically distinct classes of RGCs. Further, we identified Prph, Ctxn3, and Prkcq as potential candidates for ipRGC classification in the murine retina. The use of these genes, or one of the other newly identified subset markers, for the generation of a transgenic mouse would enable future studies of RGC-subtype specific function, wiring, and projection.
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http://dx.doi.org/10.1038/s41598-019-52215-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6823391PMC
October 2019

Hemodynamic Response of the Three Macular Capillary Plexuses in Dark Adaptation and Flicker Stimulation Using Optical Coherence Tomography Angiography.

Invest Ophthalmol Vis Sci 2019 02;60(2):694-703

Department of Ophthalmology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States.

Purpose: To assess retinal microvascular reactivity during dark adaptation and the transition to ambient light and after flicker stimulation using optical coherence tomography angiography (OCTA).

Methods: Fifteen eyes of 15 healthy participants were dark adapted for 45 minutes followed by OCTA imaging in the dark-adapted state. After 5 minutes of normal lighting, subjects underwent OCTA imaging. Participants were then subjected to a flashing light-emitting diode (LED) light and repeat OCTA. Parafoveal vessel density and adjusted flow index (AFI) were calculated for superficial (SCP), middle (MCP), and deep capillary plexuses (DCP), and then compared between conditions after adjusting for age, refractive error, and scan quality. SCP vessel length density (VLD) was also evaluated. Between-condition capillary images were aligned and subtracted to identify differences. We then analyzed images from 10 healthy subjects during the transition from dark adaptation to ambient light.

Results: SCP vessel density was significantly higher while SCP VLD was significantly lower during ambient light and flicker compared to dark adaptation. There was a significant positive mean value for DCP "flicker minus dark or light," suggesting more visible vessels during flicker due to changes in flow, dilation, or vessel recruitment. We found a significant, transient increase in SCP and decrease in both MCP and DCP vessel density during the transition from dark to light.

Conclusions: We show evidence suggesting constriction of deeper vessels and dilation of large SCP vessels during the transition from dark to light. This contrasts to redistribution of blood flow to deeper layers during dark adaptation and flicker stimulation.
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http://dx.doi.org/10.1167/iovs.18-25478DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6383834PMC
February 2019

Oncogenic Notch Promotes Long-Range Regulatory Interactions within Hyperconnected 3D Cliques.

Mol Cell 2019 03 7;73(6):1174-1190.e12. Epub 2019 Feb 7.

Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Institute for Biomedical Informatics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA. Electronic address:

Chromatin loops enable transcription-factor-bound distal enhancers to interact with their target promoters to regulate transcriptional programs. Although developmental transcription factors such as active forms of Notch can directly stimulate transcription by activating enhancers, the effect of their oncogenic subversion on the 3D organization of cancer genomes is largely undetermined. By mapping chromatin looping genome-wide in Notch-dependent triple-negative breast cancer and B cell lymphoma, we show that beyond the well-characterized role of Notch as an activator of distal enhancers, Notch regulates its direct target genes by instructing enhancer repositioning. Moreover, a large fraction of Notch-instructed regulatory loops form highly interacting enhancer and promoter spatial clusters termed "3D cliques." Loss- and gain-of-function experiments show that Notch preferentially targets hyperconnected 3D cliques that regulate the expression of crucial proto-oncogenes. Our observations suggest that oncogenic hijacking of developmental transcription factors can dysregulate transcription through widespread effects on the spatial organization of cancer genomes.
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http://dx.doi.org/10.1016/j.molcel.2019.01.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6485942PMC
March 2019

A Self-Regulating Gap Junction Network of Amacrine Cells Controls Nitric Oxide Release in the Retina.

Neuron 2018 12 25;100(5):1149-1162.e5. Epub 2018 Oct 25.

Department of Ophthalmology, Northwestern University, Chicago, IL, USA; Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA; Department of Neurobiology, Weinberg College of Arts and Sciences, Northwestern University, Evanston, IL, USA. Electronic address:

Neuromodulators regulate circuits throughout the nervous system, and revealing the cell types and stimulus conditions controlling their release is vital to understanding their function. The effects of the neuromodulator nitric oxide (NO) have been studied in many circuits, including in the vertebrate retina, where it regulates synaptic release, gap junction coupling, and blood vessel dilation, but little is known about the cells that release NO. We show that a single type of amacrine cell (AC) controls NO release in the inner retina, and we report its light responses, electrical properties, and calcium dynamics. We discover that this AC forms a dense gap junction network and that the strength of electrical coupling in the network is regulated by light through NO. A model of the network offers insights into the biophysical specializations leading to auto-regulation of NO release within the network.
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http://dx.doi.org/10.1016/j.neuron.2018.09.047DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6317889PMC
December 2018

Receptive field center-surround interactions mediate context-dependent spatial contrast encoding in the retina.

Elife 2018 09 6;7. Epub 2018 Sep 6.

Department of Physiology and Biophysics, University of Washington, Seattle, United States.

Antagonistic receptive field surrounds are a near-universal property of early sensory processing. A key assumption in many models for retinal ganglion cell encoding is that receptive field surrounds are added only to the fully formed center signal. But anatomical and functional observations indicate that surrounds are added before the summation of signals across receptive field subunits that creates the center. Here, we show that this receptive field architecture has an important consequence for spatial contrast encoding in the macaque monkey retina: the surround can control sensitivity to fine spatial structure by changing the way the center integrates visual information over space. The impact of the surround is particularly prominent when center and surround signals are correlated, as they are in natural stimuli. This effect of the surround differs substantially from classic center-surround models and raises the possibility that the surround plays unappreciated roles in shaping ganglion cell sensitivity to natural inputs.
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http://dx.doi.org/10.7554/eLife.38841DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6185113PMC
September 2018

Classes of ITD Predict Outcomes in AML Patients Treated with FLT3 Inhibitors.

Clin Cancer Res 2019 01 4;25(2):573-583. Epub 2018 Sep 4.

Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania.

Purpose: Recurrent internal tandem duplication (ITD) mutations are observed in various cancers including acute myeloid leukemia (AML), where ITD mutations in tyrosine kinase receptor FLT3 are associated with poor prognostic outcomes. Several FLT3 inhibitors (FLT3i) are in clinical trials for high-risk -ITD-positive AML. However, the variability of survival following FLT3i treatment suggests that the mere presence of -ITD mutations might not guarantee effective clinical response. Motivated by the heterogeneity of -ITD mutations, we investigated the effects of -ITD structural features on the response of AML patients to treatment. We developed the HeatITup (HEAT diffusion for Internal Tandem dUPlication) algorithm to identify and quantitate ITD structural features including nucleotide composition. Using HeatITup, we studied the impact of ITD structural features on the clinical response to FLT3i and induction chemotherapy in -ITD-positive AML patients.

Results: HeatITup accurately identifies and classifies ITDs into newly defined categories of "typical" or "atypical" based on their nucleotide composition. A typical ITD's insert sequence completely matches the wild-type whereas an atypical ITD's insert contains nucleotides exogenous to the wild-type . Our analysis shows marked divergence between typical and atypical ITD mutation features. Furthermore, our data suggest that AML patients carrying typical -ITDs benefited significantly more from both FLT3i and induction chemotherapy treatments than patients with atypical -ITDs.

Conclusions: These results underscore the importance of structural discernment of complex somatic mutations such as ITDs in progressing toward personalized treatment of AML patients, and enable researchers and clinicians to unravel ITD complexity using the provided software..
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http://dx.doi.org/10.1158/1078-0432.CCR-18-0655DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6335170PMC
January 2019

Differential Integration of Transcriptome and Proteome Identifies Pan-Cancer Prognostic Biomarkers.

Front Genet 2018 15;9:205. Epub 2018 Jun 15.

Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA, United States.

High-throughput analysis of the transcriptome and proteome individually are used to interrogate complex oncogenic processes in cancer. However, an outstanding challenge is how to combine these complementary, yet partially disparate data sources to accurately identify tumor-specific gene products and clinical biomarkers. Here, we introduce inteGREAT for robust and scalable differential integration of high-throughput measurements. With inteGREAT, each data source is represented as a co-expression network, which is analyzed to characterize the local and global structure of each node across networks. inteGREAT scores the degree by which the topology of each gene in both transcriptome and proteome networks are conserved within a tumor type, yet different from other normal or malignant cells. We demonstrated the high performance of inteGREAT based on several analyses: deconvolving synthetic networks, rediscovering known diagnostic biomarkers, establishing relationships between tumor lineages, and elucidating putative prognostic biomarkers which we experimentally validated. Furthermore, we introduce the application of a clumpiness measure to quantitatively describe tumor lineage similarity. Together, inteGREAT not only infers functional and clinical insights from the integration of transcriptomic and proteomic data sources in cancer, but also can be readily applied to other heterogeneous high-throughput data sources. inteGREAT is open source and available to download from https://github.com/faryabib/inteGREAT.
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http://dx.doi.org/10.3389/fgene.2018.00205DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6018483PMC
June 2018

The dynamic receptive fields of retinal ganglion cells.

Prog Retin Eye Res 2018 11 23;67:102-117. Epub 2018 Jun 23.

Departments of Ophthalmology and Physiology, Feinberg School of Medicine, Northwestern University, United States. Electronic address:

Retinal ganglion cells (RGCs) were one of the first classes of sensory neurons to be described in terms of a receptive field (RF). Over the last six decades, our understanding of the diversity of RGC types and the nuances of their response properties has grown exponentially. We will review the current understanding of RGC RFs mostly from studies in mammals, but including work from other vertebrates as well. We will argue for a new paradigm that embraces the fluidity of RGC RFs with an eye toward the neuroethology of vision. Specifically, we will focus on (1) different methods for measuring RGC RFs, (2) RF models, (3) feature selectivity and the distinction between fluid and stable RF properties, and (4) ideas about the future of understanding RGC RFs.
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http://dx.doi.org/10.1016/j.preteyeres.2018.06.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6235744PMC
November 2018

Electrical synapses convey orientation selectivity in the mouse retina.

Nat Commun 2017 12 11;8(1):2025. Epub 2017 Dec 11.

Interdepartmental Neuroscience Program, Northwestern University, Chicago, IL, 60611, USA.

Sensory neurons downstream of primary receptors are selective for specific stimulus features, and they derive their selectivity both from excitatory and inhibitory synaptic inputs from other neurons and from their own intrinsic properties. Electrical synapses, formed by gap junctions, modulate sensory circuits. Retinal ganglion cells (RGCs) are diverse feature detectors carrying visual information to the brain, and receive excitatory input from bipolar cells and inhibitory input from amacrine cells (ACs). Here we describe a RGC that relies on gap junctions, rather than chemical synapses, to convey its selectivity for the orientation of a visual stimulus. This represents both a new functional role of electrical synapses as the primary drivers of feature selectivity and a new circuit mechanism for orientation selectivity in the retina.
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http://dx.doi.org/10.1038/s41467-017-01980-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5725423PMC
December 2017

An atlas of B-cell clonal distribution in the human body.

Nat Biotechnol 2017 Sep 21;35(9):879-884. Epub 2017 Aug 21.

Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.

B-cell responses result in clonal expansion, and can occur in a variety of tissues. To define how B-cell clones are distributed in the body, we sequenced 933,427 B-cell clonal lineages and mapped them to eight different anatomic compartments in six human organ donors. We show that large B-cell clones partition into two broad networks-one spans the blood, bone marrow, spleen and lung, while the other is restricted to tissues within the gastrointestinal (GI) tract (jejunum, ileum and colon). Notably, GI tract clones display extensive sharing of sequence variants among different portions of the tract and have higher frequencies of somatic hypermutation, suggesting extensive and serial rounds of clonal expansion and selection. Our findings provide an anatomic atlas of B-cell clonal lineages, their properties and tissue connections. This resource serves as a foundation for studies of tissue-based immunity, including vaccine responses, infections, autoimmunity and cancer.
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http://dx.doi.org/10.1038/nbt.3942DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5679700PMC
September 2017

Circuit Mechanisms of a Retinal Ganglion Cell with Stimulus-Dependent Response Latency and Activation Beyond Its Dendrites.

Curr Biol 2017 Feb 26;27(4):471-482. Epub 2017 Jan 26.

Department of Ophthalmology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA; Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA; Department of Neurobiology, Weinberg College of Arts and Sciences, Northwestern University, Evanston, IL 60208, USA. Electronic address:

Center-surround antagonism has been used as the canonical model to describe receptive fields of retinal ganglion cells (RGCs) for decades. We describe a newly identified RGC type in the mouse, called the ON delayed (OND) RGC, with receptive field properties that deviate from center-surround organization. Responding with an unusually long latency to light stimulation, OND RGCs respond earlier as the visual stimulus increases in size. Furthermore, OND RGCs are excited by light falling far beyond their dendrites. We unravel details of the circuit mechanisms behind these phenomena, revealing new roles for inhibition in controlling both temporal and spatial receptive field properties. The non-canonical receptive field properties of the OND RGC-integration of long temporal and large spatial scales-suggest that unlike typical RGCs, it may encode a slowly varying, global property of the visual scene.
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http://dx.doi.org/10.1016/j.cub.2016.12.033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5319888PMC
February 2017

Three Small-Receptive-Field Ganglion Cells in the Mouse Retina Are Distinctly Tuned to Size, Speed, and Object Motion.

J Neurosci 2017 01;37(3):610-625

Department of Ophthalmology,

Retinal ganglion cells (RGCs) are frequently divided into functional types by their ability to extract and relay specific features from a visual scene, such as the capacity to discern local or global motion, direction of motion, stimulus orientation, contrast or uniformity, or the presence of large or small objects. Here we introduce three previously uncharacterized, nondirection-selective ON-OFF RGC types that represent a distinct set of feature detectors in the mouse retina. The three high-definition (HD) RGCs possess small receptive-field centers and strong surround suppression. They respond selectively to objects of specific sizes, speeds, and types of motion. We present comprehensive morphological characterization of the HD RGCs and physiological recordings of their light responses, receptive-field size and structure, and synaptic mechanisms of surround suppression. We also explore the similarities and differences between the HD RGCs and a well characterized RGC with a comparably small receptive field, the local edge detector, in response to moving objects and textures. We model populations of each RGC type to study how they differ in their performance tracking a moving object. These results, besides introducing three new RGC types that together constitute a substantial fraction of mouse RGCs, provide insights into the role of different circuits in shaping RGC receptive fields and establish a foundation for continued study of the mechanisms of surround suppression and the neural basis of motion detection.

Significance Statement: The output cells of the retina, retinal ganglion cells (RGCs), are a diverse group of ∼40 distinct neuron types that are often assigned "feature detection" profiles based on the specific aspects of the visual scene to which they respond. Here we describe, for the first time, morphological and physiological characterization of three new RGC types in the mouse retina, substantially augmenting our understanding of feature selectivity. Experiments and modeling show that while these three "high-definition" RGCs share certain receptive-field properties, they also have distinct tuning to the size, speed, and type of motion on the retina, enabling them to occupy different niches in stimulus space.
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http://dx.doi.org/10.1523/JNEUROSCI.2804-16.2016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5242408PMC
January 2017

Nonlinear Spatiotemporal Integration by Electrical and Chemical Synapses in the Retina.

Neuron 2016 04 7;90(2):320-32. Epub 2016 Apr 7.

Department of Physiology and Biophysics and Howard Hughes Medical Institute, University of Washington, Seattle, WA 98195, USA. Electronic address:

Electrical and chemical synapses coexist in circuits throughout the CNS. Yet, it is not well understood how electrical and chemical synaptic transmission interact to determine the functional output of networks endowed with both types of synapse. We found that release of glutamate from bipolar cells onto retinal ganglion cells (RGCs) was strongly shaped by gap-junction-mediated electrical coupling within the bipolar cell network of the mouse retina. Specifically, electrical synapses spread signals laterally between bipolar cells, and this lateral spread contributed to a nonlinear enhancement of bipolar cell output to visual stimuli presented closely in space and time. Our findings thus (1) highlight how electrical and chemical transmission can work in concert to influence network output and (2) reveal a previously unappreciated circuit mechanism that increases RGC sensitivity to spatiotemporally correlated input, such as that produced by motion.
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http://dx.doi.org/10.1016/j.neuron.2016.03.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4840068PMC
April 2016

Cardinal Orientation Selectivity Is Represented by Two Distinct Ganglion Cell Types in Mouse Retina.

J Neurosci 2016 Mar;36(11):3208-21

Interdepartmental Neuroscience Program, Department of Ophthalmology, and Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611

Unlabelled: Orientation selectivity (OS) is a prominent and well studied feature of early visual processing in mammals, but recent work has highlighted the possibility that parallel OS circuits might exist in multiple brain locations. Although both classic and modern work has identified an OS mechanism in selective wiring from lateral geniculate nucleus (LGN) to primary visual cortex, OS responses have now been found upstream of cortex in mouse LGN and superior colliculus, suggesting a possible origin in the retina. Indeed, retinal OS responses have been reported for decades in rabbit and more recently in mouse. However, we still know very little about the properties and mechanisms of retinal OS in the mouse, including whether there is a distinct OS ganglion cell type, which orientations are represented, and what are the synaptic mechanisms of retinal OS. We have identified two novel types of OS ganglion cells in the mouse retina that are highly selective for horizontal and vertical cardinal orientations. Reconstructions of the dendritic trees of these OS ganglion cells and measurements of their synaptic conductances offer insights into the mechanism of the OS computation at the earliest stage of the visual system.

Significance Statement: Orientation selectivity (OS) is one of the most well studied computations in the brain and has become a prominent model system in various areas of sensory neuroscience. Although the cortical mechanism of OS suggested by Hubel and Wiesel (1962) has been investigated intensely, other OS cells exist upstream of cortex as early as the retina and the mechanisms of OS in subcortical regions are much less well understood. We identified two ON retinal ganglion cells (RGCs) in mouse that compute OS along the horizontal (nasal-temporal) and vertical (dorsoventral) axes of visual space. We show the relationship between dendritic morphology and OS for each RGC type and reveal new synaptic mechanisms of OS computation in the retina.
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http://dx.doi.org/10.1523/JNEUROSCI.4554-15.2016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4792935PMC
March 2016

An Amacrine Cell Circuit for Signaling Steady Illumination in the Retina.

Cell Rep 2015 Dec 17;13(12):2663-70. Epub 2015 Dec 17.

Department of Ophthalmology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, 60611, USA; Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, 60611, USA. Electronic address:

Decades of research have focused on the circuit connectivity between retinal neurons, but only a handful of amacrine cells have been described functionally and placed in the context of a specific retinal circuit. Here, we identify a circuit where inhibition from a specific amacrine cell plays a vital role in shaping the feature selectivity of a postsynaptic ganglion cell. We record from transgenically labeled CRH-1 amacrine cells and identify a postsynaptic target for CRH-1 amacrine cell inhibition in an atypical retinal ganglion cell (RGC) in mouse retina, the Suppressed-by-Contrast (SbC) RGC. Unlike other RGC types, SbC RGCs spike tonically in steady illumination and are suppressed by both increases and decreases in illumination. Inhibition from GABAergic CRH-1 amacrine cells shapes this unique contrast response profile to positive contrast. We show the existence and impact of this circuit, with both paired recordings and cell-type-specific ablation.
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http://dx.doi.org/10.1016/j.celrep.2015.11.062DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4698005PMC
December 2015

The synaptic and circuit mechanisms underlying a change in spatial encoding in the retina.

Neuron 2014 Apr;82(2):460-73

Department of Physiology and Biophysics and Howard Hughes Medical Institute, University of Washington, Seattle, WA 98195, USA. Electronic address:

Components of neural circuits are often repurposed so that the same biological hardware can be used for distinct computations. This flexibility in circuit operation is required to account for the changes in sensory computations that accompany changes in input signals. Yet we know little about how such changes in circuit operation are implemented. Here we show that a single retinal ganglion cell performs a different computation in dim light--averaging contrast within its receptive field--than in brighter light, when the cell becomes sensitive to fine spatial detail. This computational change depends on interactions between two parallel circuits that control the ganglion cell's excitatory synaptic inputs. Specifically, steady-state interactions through dendro-axonal gap junctions control rectification of the synapses providing excitatory input to the ganglion cell. These findings provide a clear example of how a simple synaptic mechanism can repurpose a neural circuit to perform diverse computations.
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http://dx.doi.org/10.1016/j.neuron.2014.02.037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4038266PMC
April 2014

Interplay of cell-autonomous and nonautonomous mechanisms tailors synaptic connectivity of converging axons in vivo.

Neuron 2014 Apr;82(1):125-37

Department of Biological Structure, University of Washington, Seattle, WA 98195-7420, USA. Electronic address:

Neurons receive input from diverse afferents but form stereotypic connections with each axon type to execute their precise functions. Developmental mechanisms that specify the connectivity of individual axons across populations of converging afferents are not well-understood. Here, we untangled the contributions of activity-dependent and independent interactions that regulate the connectivity of afferents providing major and minor input onto a neuron. Individual transmission-deficient retinal bipolar cells (BCs) reduced synapses with retinal ganglion cells (RGCs), but active BCs of the same type sharing the dendrite surprisingly did not compensate for this loss. Genetic ablation of some BC neighbors resulted in increased synaptogenesis by the remaining axons in a transmission-independent manner. Presence, but not transmission, of the major BC input also dissuades wiring with the minor input and with synaptically compatible but functionally mismatched afferents. Cell-autonomous, activity-dependent and nonautonomous, activity-independent mechanisms thus together tailor connectivity of individual axons among converging inner retinal afferents.
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http://dx.doi.org/10.1016/j.neuron.2014.02.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3990864PMC
April 2014

Trials and Tribulations with VH Replacement.

Front Immunol 2014 30;5:10. Epub 2014 Jan 30.

Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania , Philadelphia, PA , USA.

VH replacement (VHR) is a type of antibody gene rearrangement in which an upstream heavy chain variable gene segment (VH) invades a pre-existing rearrangement (VDJ). In this Hypothesis and Theory article, we begin by reviewing the mechanism of VHR, its developmental timing and its potential biological consequences. Then we explore the hypothesis that specific sequence motifs called footprints reflect VHR versus other processes. We provide a compilation of footprint sequences from different regions of the antibody heavy chain, and include data from the literature and from a high throughput sequencing experiment to evaluate the significance of footprint sequences. We conclude by discussing the difficulties of attributing footprints to VHR.
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http://dx.doi.org/10.3389/fimmu.2014.00010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3906580PMC
June 2014

Visual space is represented by nonmatching topographies of distinct mouse retinal ganglion cell types.

Curr Biol 2014 Feb 16;24(3):310-5. Epub 2014 Jan 16.

Department of Biological Structure, University of Washington, Seattle, WA 98195, USA. Electronic address:

The distributions of neurons in sensory circuits display ordered spatial patterns arranged to enhance or encode specific regions or features of the external environment. Indeed, visual space is not sampled uniformly across the vertebrate retina. Retinal ganglion cell (RGC) density increases and dendritic arbor size decreases toward retinal locations with higher sampling frequency, such as the fovea in primates and area centralis in carnivores [1]. In these locations, higher acuity at the level of individual cells is obtained because the receptive field center of a RGC corresponds approximately to the spatial extent of its dendritic arbor [2, 3]. For most species, structurally and functionally distinct RGC types appear to have similar topographies, collectively scaling their cell densities and arbor sizes toward the same retinal location [4]. Thus, visual space is represented across the retina in parallel by multiple distinct circuits [5]. In contrast, we find a population of mouse RGCs, known as alpha or alpha-like [6], that displays a nasal-to-temporal gradient in cell density, size, and receptive fields, which facilitates enhanced visual sampling in frontal visual fields. The distribution of alpha-like RGCs contrasts with other known mouse RGC types and suggests that, unlike most mammals, RGC topographies in mice are arranged to sample space differentially.
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http://dx.doi.org/10.1016/j.cub.2013.12.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3990865PMC
February 2014

Germline Amino Acid Diversity in B Cell Receptors is a Good Predictor of Somatic Selection Pressures.

Front Immunol 2013 8;4:357. Epub 2013 Nov 8.

Systems Immunology Laboratory, School of Biomedical Engineering, Science, and Health Systems, Drexel University , Philadelphia, PA , USA.

The diversity of the immune repertoire is important for the adaptive immune system's ability to detect pathogens. Much of this diversity is generated in two steps, first through the recombination of germline gene segments and second through hypermutation during an immune response. While both steps are to some extent based on the germline level repertoire of genes, the final structure and selection of specific receptors is at the somatic level. How germline diversity and selection relate to somatic diversity and selection has not been clear. To investigate how germline diversity relates to somatic diversity and selection, we considered the published repertoire of Ig heavy chain V genes taken from the blood of 12 individuals, post-vaccination against influenza, sequenced by 454 high-throughput sequencing. We here show that when we consider individual amino acid positions in the heavy chain V gene sequence, there exists a strong correlation between the diversity of the germline repertoire at a position and the number of B cell clones that change amino acids at that position. At the same time, we find that the diversity of amino acids used in the mutated positions is greater than in the germline, albeit still correlated to germline diversity. From these findings, we propose that while germline diversity and germline amino acid usage at a given position do not fully specify the amino acid mutant needed to promote survival of specific clones, germline diversity at a given position is a good indicator for the potential to survive after somatic mutation at that position. We would therefore suggest that germline diversity at each specific position is the better a priori model for the effects of somatic mutation and selection, than simply the division into complementarity determining and framework regions.
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http://dx.doi.org/10.3389/fimmu.2013.00357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3820969PMC
November 2013

Conserved variation: identifying patterns of stability and variability in BCR and TCR V genes with different diversity and richness metrics.

Phys Biol 2013 Jun 4;10(3):035005. Epub 2013 Jun 4.

School of Biomedical Engineering, Science and Health Systems Drexel University, Philadelphia, PA, USA.

The immune system can detect most invading pathogens. The potential for detection of pathogens is dependent on the somatic diversity of the immune repertoires. While it is known that this somatic diversity is carefully generated, it is unclear how the diversity is distributed in the different genes encoding receptors of immune cells. Utilizing different metrics for richness and diversity at the level of small sequence fragments, we present here an analysis of the entire known human germline repertoire as represented by the sequences from the ImMunoGeneTics database of immune receptors. We have developed a fragment sequence quantification analysis to track variation of repertoires with different degrees of precision. Somatic diversity has previously been functionally characterized mostly by division of the V gene sequences into the more conserved and invariant framework (FR) of the receptor and more varied complementarity determining regions (CDR), that interact with the antigen. We find that CDR and FR can be explicitly identified with our sequence fragment diversity quantification technique. In terms of diversity, CDR and FR are especially distinct in B cell V genes. T cell V genes show less of the CDR/FR periodicity but are more diverse overall. Our analysis further shows that there are other areas of diversity outside the CDR and FR that are found widely dispersed in T cell receptor V genes and more tightly focused in FR1 and FR3 in the B cell receptor V genes. The diversity we observe is not dependent on allelic differences nor is this diversity segregated by individual V gene families. We would thus expect that each individual exhibit a diversity equivalent to that of the entire potential repertoire.
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http://dx.doi.org/10.1088/1478-3975/10/3/035005DOI Listing
June 2013

Controlling gain one photon at a time.

Elife 2013 May 14;2:e00467. Epub 2013 May 14.

Department of Physiology and Biophysics , University of Washington , Seattle , United States ; Howard Hughes Medical Institute, University of Washington , Seattle , United States.

Adaptation is a salient property of sensory processing. All adaptational or gain control mechanisms face the challenge of obtaining a reliable estimate of the property of the input to be adapted to and obtaining this estimate sufficiently rapidly to be useful. Here, we explore how the primate retina balances the need to change gain rapidly and reliably when photons arrive rarely at individual rod photoreceptors. We find that the weakest backgrounds that decrease the gain of the retinal output signals are similar to those that increase human behavioral threshold, and identify a novel site of gain control in the retinal circuitry. Thus, surprisingly, the gain of retinal signals begins to decrease essentially as soon as background lights are detectable; under these conditions, gain control does not rely on a highly averaged estimate of the photon count, but instead signals from individual photon absorptions trigger changes in gain. DOI:http://dx.doi.org/10.7554/eLife.00467.001.
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http://dx.doi.org/10.7554/eLife.00467DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3654457PMC
May 2013

The spatial structure of a nonlinear receptive field.

Nat Neurosci 2012 Nov 23;15(11):1572-80. Epub 2012 Sep 23.

Department of Physiology and Biophysics, University of Washington, Seattle, Seattle, Washington, USA.

Understanding a sensory system implies the ability to predict responses to a variety of inputs from a common model. In the retina, this includes predicting how the integration of signals across visual space shapes the outputs of retinal ganglion cells. Existing models of this process generalize poorly to predict responses to new stimuli. This failure arises in part from properties of the ganglion cell response that are not well captured by standard receptive-field mapping techniques: nonlinear spatial integration and fine-scale heterogeneities in spatial sampling. Here we characterize a ganglion cell's spatial receptive field using a mechanistic model based on measurements of the physiological properties and connectivity of only the primary excitatory circuitry of the retina. The resulting simplified circuit model successfully predicts ganglion-cell responses to a variety of spatial patterns and thus provides a direct correspondence between circuit connectivity and retinal output.
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http://dx.doi.org/10.1038/nn.3225DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3517818PMC
November 2012