Mitochondrial Transfer Between Inner Retinal Neurons

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This abstract was presented today, April 26th at the 2023 Association for Research in Vision and Opthalmology (ARVO) meetings in New Orleans, Louisiana by Selena Wirthlin, Crystal Sigulinsky, James Anderson, and Bryan William Jones.

Full resolution version here.

Purpose
Intercellular mitochondrial transfer has been reported across a variety of cells and tissues under both physiological and pathological conditions. Such transfer has shown broad therapeutic potential. The effectiveness of this therapy, however, is limited by a lack of understanding of the cellular and molecular mechanisms. Here, the ultrastructural features of mitochondrial transfer between inner retinal neurons discovered through retinal connectomics analysis is shown.

Methods
Retinal Connectome 2 (RC2) was built by automated transmission electron microscopy at ultrastructural (2nm/pixel) resolution. RC2 is a 0.25mm diameter volume of retina obtained from a 5-month-old female C57BL/6J mouse. The Viking application was used to visualize and annotate inter- and intracellular features of interest in the connectome.

Results
Exploration of RC2 revealed material transfer between apposing neural processes within the OFF subliminal of the inner plexiform layer. The transferred material can be defined as a mitochondria, confirmed by the presence of crustae. At the transfer site, a short, electron-dense 140-nm diameter tube with a curved cap tightly associated with the inner mitochondrial membrane of one neuritis extends into a vacuole within the apposing neuritis formed by the plasma membranes of the two cells. Thin cytoskeletal components consistent with actin microfilaments extend into the mitochondrion. Morphology and synaptology of the acceptor cell confirm it is an Aii amacrine cell, while preliminary findings suggest the donor cell is a type of ON/OFF ganglion cell.

Conclusions
These findings demonstrate active mitochondrial transfer between different classes of endogenous inner retinal neurons and suggests it may represent an important component of tissue homeostasis in the retina. Features of this transfer differ from previously reported mitochondrial transfer between photoreceptors upon transplantation, which may indicate cell type- or context-dependent differences in the cellular or molecular mechanisms. Our findings demonstrate active mitochondrial transfer between different classes of endogenous inner retinal neurons and suggest it may represent an important component of tissue homeostasis in the retina. Features of this transfer differ from previous reports by the Wallace and Pearson groups of material transfer between photoreceptors upon transplantation through tunneling nanotubes (Ortin- Martinez et al., 2021; Kalargyrou et al., 2021), which may indicate cell type- or context-dependent differences in the cellular or molecular mechanisms. Understanding these mechanisms could serve as a catalyst for development of novel therapeutics for disease in the retina and beyond.

Müller Cell Connectomics In Health And Disease

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This talk was presented today, April 25th at the 2023 Association for Research in Vision and Opthalmology (ARVO) meetings in New Orleans, Louisiana by Rebecca Pfeiffer as part of an ARVO Minisymposium Bryan William Jones organized.

Abstract: Muller cells are a critical component of retinal function and rapidly change metabolically and morphologically in retinal disease. Of Muller cell functions, many require close physical relationships between the Muller cell and the synapses of the neurons they support. Despite this required neuro-glial relationship, little is known about the direct contacts between Muller cells and synapses in healthy or diseased retinas. In order to address this, I use a connectomics/pathoconnectomics approach to reconstruct Muller cells and their neighboring synapses. The retinas evaluated are from a healthy rabbit, retinal connectome 1 (RC1), and from the P347L rabbit model of retinitis pigmentosa, retinal pathoconnectome 1 (RPC1). Preliminary data demonstrate an increase in endfoot entanglement in RPC1 when compared with RC1, and direct synaptic contact analysis of both connectomes is ongoing.

Species-Specific Connectivity In The Aii Connectome

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This talk was presented today, April 25th at the 2023 Association for Research in Vision and Opthalmology (ARVO) meetings in New Orleans, Louisiana by Crystal Sigulinsky as part of an ARVO Minisymposium organized by Bryan William Jones.

Abstract: Biomedical research relies heavily on animal models to study human disease and develop therapeutics. Understanding the architectural diversity in neural networks between humans and these model species is essential for choosing a relevant study model and interpreting conflicting results. Using comparative connectomics, we sought to map and compare the local neural network architecture of rabbit and mouse retinal Aii amacrine cells. This specialized narrow-field, multistratified, glycinergic interneuron has critical feedforward and feedback roles in both the photopic and scotopic retinal networks spanning the ON and OFF pathways, making it an ideal candidate for investigating species-specific differences in retinal networks. High-resolution, serial-section transmission electron microscopy (TEM) volumes of rabbit (RC1: female, 13- month, Dutch Belted) and mouse (RC2: female, 5-month, C57BL/6J) retinal tissue provided spatially-registered synaptic maps of Aii connectivity at directly comparable resolution and completeness. These reveal that despite species-specific morphologies, gross synaptology and compartmentalization appear conserved. Yet, rabbit and mouse Aii cells diverge in the weighting of their partnerships, most notably in their coupling profiles. Opposing biases in gap junction partnerships and their respective sizing rules indicate a greater relative output by mouse Aii cells to ON pathways than in rabbit. However, a unique topological conformation for a subset of conventional presynapses formed by Aii cell lobular dendrites with species-specific features and prevalence may influence signal output to specific partner classes within the OFF pathway and either nullify or exacerbate this difference in ON/OFF output. Additionally, rabbit Aii cells in RC1 showed greater Aii-Aii coupling than in mouse, which may suggest greater signal-to-noise compensation. Lastly, preliminary data suggest mouse Aii cells receive greater excitatory, but not inhibitory input/feedback from the OFF pathway than in rabbit. Together these data indicate that precise neural circuit architectures diverge between species and require detailed, comprehensive mapping to begin to dissect potential influence on signal flow.

Structural Motifs Of Excitatory Synapses In The Mammalian Retina

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This abstract was presented today, April 24th at the 2023 Association for Research in Vision and Opthalmology (ARVO) meetings in New Orleans, Louisiana by Taylor Otterness, Crystal Sigulinsky, James Anderson, and Bryan William Jones.

Full resolution version here.

Purpose
Connectivity within the nervous system is precise and disruptions lead to degraded performance and disease, yet the rules that govern connectivity remain unknown. Recent efforts reveal that different types of cone bipolar cells in the neural retina show preferences in the selection and frequency of presynaptic structure types used for signal transmission. However, it is not yet known how these differences are related to the quantity or type of postsynaptic partner. We used Retinal Connectome 1 (RC1) to analyze the synaptic output of rabbit CBb6 cells, a type of ON cone bipolar cell that forms excitatory synapses via diverse presynaptic structure types, to identify patterns in how these cells interact with their postsynaptic partners.

Methods
RC1 is a 0.25 mm diameter volume sampled from mid-peripheral retina of a 13 month old female Dutch-Belted rabbit, serially sectioned at 70 nm, and imaged at ultrastructural resolution (2nm/px) using transmission electron microscopy. Postsynaptic partners of CBb6 cell 6156’s presynaptic structures were annotated using the Viking Viewer for Connectomics. Statistical analyses were conducted in Microsoft Excel and investigated further with 3D rendering and graph visualization of connectivity.

Results
The factors tracked for comparison included presynaptic structure type, target number, and postsynaptic partner type. Multiribbon synapses of CBb6 cell 6156 trended towards having a greater number of output partners, with a greater proportion of dyads than monads. Despite this, triads and quadrads were only found opposing single ribbon synapses. As the different presynaptic structure types may differ in the strength of neurotransmitter release (ribbonless < single ribbon < multiribbon), these findings are inconsistent with scaling of output to the number of postsynaptic targets. Both amacrine cells (AC) and ganglion cells (GC) are postsynaptic partners of 6156. However, single ribbon and ribbonless structures appear biased towards AC only targets, while multiribbon synapses appear biased toward mixed AC and GC targets.

Conclusions
Target type relationships appear more important than the number of targets in determining presynaptic structure type in CBb6. Future efforts will examine size differences of postsynaptic structures and presynaptic ribbon size, and even compare across bipolar cell classes, in order to provide further insight on the connectivity rules underlying excitatory synapses.

Distinctive Synaptic Structural Motifs Link Excitatory Retinal Interneurons To Diverse Postsynaptic Partner Types

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We have a new manuscript from the lab in Cell Reports, Distinctive Synaptic Structural Motifs Link Excitatory Retinal Interneurons To Diverse Postsynaptic Partner Types. This manuscript is in collaboration with the first author, Wan-Qing Yu @wanqing_yu, then co-authors Rachael Swanstrom, Crystal L. Sigulinsky @CSigulinsky, Richard M. Ahlquist, along with Sharm Knecht, myself Bryan W. Jones @BWJones. David M. Berson, and Rachel O. Wong. The PDF is here.

Abstract:
Neurons make converging and diverging synaptic connections with distinct partner types. Whether synapses involving separate partners demonstrate similar or distinct structural motifs is not yet well understood. We thus used serial electron microscopy in mouse retina to map output synapses of cone bipolar cells (CBCs) and compare their structural arrangements across bipolar types and postsynaptic partners. Three presynaptic configurations emerge—single-ribbon, ribbonless, and multiribbon synapses. Each CBC type exploits these arrangements in a unique combination, a feature also found among rabbit ON CBCs. Though most synapses are dyads, monads and triads are also seen. Altogether, mouse CBCs exhibit at least six motifs, and each CBC type uses these in a stereotypic pattern. Moreover, synapses between CBCs and particular partner types appear biased toward certain motifs. Our observations reveal synaptic strategies that diversify the output within and across CBC types, potentially shaping the distinct functions of retinal microcircuits.

Current Perspective on Retinal Remodeling: Implications for Therapeutics

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We have a new paper out of the lab, a perspectives paper on Retinal Remodeling: Implications for Therapeutics. (pdf here).

Authors are Rebecca L. Pfeiffer @BeccaPfeiffer19, and Bryan W. Jones @BWJones.

Abstract: The retinal degenerative diseases retinitis pigmentosa and age-related macular degeneration are a leading cause of irreversible vision loss. Both present with progressive photoreceptor degeneration that is further complicated by processes of retinal remodeling. In this perspective, we discuss the current state of the field of retinal remodeling and its implications for vision-restoring therapeutics currently in development. Here, we discuss the challenges and pitfalls retinal remodeling poses for each therapeutic strategy under the premise that understanding the features of retinal remodeling in totality will provide a basic framework with which therapeutics can interface. Additionally, we discuss the potential for approaching therapeutics using a combined strategy of using diffusible molecules in tandem with other vision-restoring therapeutics. We end by discussing the potential of the retina and retinal remodeling as a model system for more broadly understanding the progression of neurodegeneration across the central nervous system.

Snowbird Neuroscience Symposium 2022

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Lab Members Bryan Jones, Rebecca Pfeiffer and graduate students Megan Croom and Qwynn Landfield, and undergraduate Taylor Otterness attended this year’s annual Snowbird Neuroscience Symposium after two years of the pandemic.  It was good to see people in person again, presenting their science and having an opportunity for the trainees to engage in the practice of presenting their work. Qwynn Landfield was one of the student organizers this year and asked me to provide some photographs for the program that you can see here.

One Connectome Finished, Another Pathoconnectome Begins

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We finished sectioning and capturing a massive new retinal connectome that we are going to be so excited to announce at some point in the not too distant future.  Effective immediately, we are also starting on a brand new pathoconnectome that we will be powering through over the next little while. Thanks to the team of people who make this happen, shown in this post are Jia-Hui Yang, Matt Berardy, and Rebecca Pfeiffer. More photos here.