The Blind Woman Who Sees Rain, But Not Her Daughter’s Smile

“Once her belief was sanctified by science, her sight got better and better…”

I first read about blindsight back in high school, reading an essay by Oliver Sacks and was absolutely intrigued by the thought of seeing without sight. Now there is a wonderful vignette on NPR by Lulu Miller that talks about blindsight (be sure to listen to the story on All Things Considered).

Of course we know now that there are multiple visual pathways and each one of these visual pathways mediates a different aspect or derivative of a component of vision.  It turns out that vision is complex, more complex than simply carrying “visual pixels” to the brain to be mapped out topologically.  After photon capture by the photoreceptors of the eye and pre-processing operations in the circuitry of the retina, information is passed to the retinal ganglion cells or the output cells of the retina.  Each one of the 18-20 types of ganglion cells that project out of the retina through the optic nerve mediates different kinds of information.  Aside from projections to the primary visual cortex, some ganglion cells project to the lateral geniculate nucleus, others to the superior colliculus, the pretectum, the suprachiasmatic nucleus and the hypothalamus.  Each one of these projections out of the retina carries information relevant to a different feature of vision.  Some of these functions help control the size of your pupil limiting the amount of light that comes into the eye.  Other projections help you orient your head and eyes to the world around you while other projections still help you figure out what time of day and season it is.  Most of these meta-visual functions are not conscious, but play crucial roles in how we live our everyday lives and are only revealed when things in the visual system go awry like the man Oliver Sacks described who could catch a ball despite being completely and functionally blind.  Milena Channing’s experience with a stroke in her visual cortex reveals some of this unconscious aspect of seeing, ironically by causing blindness while preserving portions of the brain involved in motion detection.

 

The Judgment Of Solomon

I had the honor and privilege of attending a Lasker/IRRF Initiative’s plenary session on Restoring Vision to the Blind at Janelia Farm last month where Mr. Sanford D. Greenberg delivered an emotional and inspiring story of a time in his life where he lost his vision during his junior year at Columbia University.  The prospect of losing vision is absolutely and completely life altering for those affected as well as for those around the individual who has lost partial or complete sight.  Mr. Greenberg’s story in his words, reveals the raw emotion of blindness, the fear and angst as well as the compassion and love of those who travel through life alongside us.  At the end, there is also a surprise that will speak to aficionados of music and give some deeper insight into someone who has touched untold millions around the globe through their work and music.

This video is documentation of that event sent along for Webvision to share with you and the wider community by John Dowling the Chair of the Lasker/IRRF Initiative, and Janelia Farm who captured the video, the Lasker Foundation, the International Retinal Research Foundation and End Blindness by 2020.  We are grateful to Mr. Greenberg for sharing his story and allowing us to help spread his story and mission of ending blindness here on Webvision.

On a personal note: While I have friends who are blind or are going blind, Mr. Greenberg’s talk haunted me the night after I saw it, particularly because of the field of science I am engaged in.  Every scientist studying vision and diseases affecting vision should have the opportunity to spend time with those who have lost sight.  It is important for people in the sciences to sit down and talk with those affected by the disease they study.  I found this out this week after a meeting with a colleague who agreed to speak with a mutual friend who has Usher’s Syndrome.  When my colleague stated after the meeting that they had never actually sat down to talk with someone who has the disease that they study, I was initially surprised.  This is not uncommon though.  As scientists, not just in the vision sciences mind you, we obsess about the details of what we study and are absolutely driven by the work, but do not always look around and talk with people who’s diseases are the subjects of our studies.  This is fundamental to the process as it drives home the motivation for the long hours, late nights and frustrations with grant funding.  It forces an introspection and helps us to better communicate our work to a wider audience which is critical to science progress and funding.

 

 

Happy 90th Birthday Gerald Westheimer

Happy 90th birthday Gerald Westheimer

A very Happy 90th Birthday to Gerald Westheimer, Ph.D.  Professor of the Graduate School, Division of Neurobiology,  Molecular and Cell Biology, and Clinical Professor of Optometry Emeritus at the University of California, Berkeley.

Gerald to this day continues to contribute important papers to the literature and to celebrate his career, his friends at UC Berkeley have put together a website to help celebrate his birthday.  There is a link on Berkeley’s website with images and memorabilia from Gerald’s life that is worth a visit.  Be sure to leave a note to Gerald there as well.

 

 

Proposed NIH And NEI 2015 Budget

2015 HHS budget proposal

The new 2015 Budget proposal released by Health and Human Services for the NIH has been finalized.  This is a proposal released by the White House as a suggestion to Congress on how to distribute federal funds related to research.  As we know, this relationship has been contentious to say the least coming at the expense of research funding which has more or less been on a constant slide for the past decade.  Real research dollars, inflation adjusted are down almost 25% from a decade ago.

In the 2015 budget, it has been proposed that an additional $211 million dollars be assigned to the NIH for a total of $30.4 billion.  As expected, translational research is funded at higher amounts increasing the disparity of basic science vs. translational work with the NIAID, NCATS, NIMH, NIDDK and NINDS receiving the majority of this additional $211 million.  Of relevance to the vision community, there is good news and bad news.  The good news is that NEI funding is proposed to increase by $1 million over the 2014 appropriation.  The bad news is that, assuming a 1.4% rate of inflation, this additional funding does not keep up with inflation and we continue the decade long slide in research funding.

There may be other opportunities within the NIH budget for vision related research, not necessarily in the NEI portfolio including some of the $100 million devoted to the BRAIN initiative, though honestly… most of those dollars are already spoken for it seems.

There is some potential good news for pre-doctoral and post-doctoral students supported by NRSA funding mechanisms with a 2% increase in stipends proposed.

This is one of the most amazing times in biomedical research, especially in the vision community.  All the work I heard about at the 2014 ARVO meetings this week from investigators across the spectrum is inspiring and yet, we are facing so many frustrations with real shortfalls in research funding.

We, the science community and America at large, needs the partisan bickering to cease.  The President’s proposal in the Opportunity, Growth, and Security Initiative requests up to an additional $970 million in funding to the NIH to fund an addition 650 NIH grants and an additional 1000 NSF grants which are badly needed to stem the loss of talent in the science communities.  Rebuilding this countries infrastructure includes our intellectual capabilities.  Myopic budget approval processes that do not take into account scientific progress at both the basic and applied levels of scientific inquiry and investigation will lead to a continued reduction in the competitiveness of this country in academic and commercial venues.

 

Thy-1 Expressed In CNV And Increased In migrating Choroidal ECs In Human Neovascular AMD

Haibo Wang

This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by Haibo Wang, Yanchao Jiang and M. Elizabeth Hartnett.

Thy-1 regulates VEGF-induced choroidal endothelial cell migration 

Dept of Ophthalmology, John A. Moran Eye Center, The University of Utah-Salt Lake City. 

Purpose: Choroidal endothelial cell (CEC) activation and migration precede the development of choroidal neovascularization in neovascular AMD. Thy-1 is a cell surface protein expressed on different cells, including neurons and endothelial cells. As a glycosylphosphatidylinositol (GPI)-anchored glycoprotein, Thy-1 is located in lipid raft microdomains within the cell membrane, which brings Thy-1 into proximity of signaling molecules including cytoplasmic tyrosine kinases that can modulate adhesive and migratory events. In the retina, Thy-1 is well known as a retinal ganglion cell marker. Given the possibility that Thy-1 might be expressed in CECs, we addressed the hypothesis that upregulated Thy-1 in CECs by age-related stresses contributes to CEC migration.

Methods: Western blots of Thy-1 were determined in retinal pigment epithelial cells (RPE) and CECs. By real time quantitative PCR, Thy-1 mRNA was measured in CECs treated with vascular endothelial growth factor (VEGF) (20 ng/ml), CCL11 (100 ng/ml) or PBS for 24 hours, or in RPE/choroids from young (<40 yrs) and old (>60 yrs) donor eyes. Immunohistochemistry of Thy-1 was performed in posterior globe sections of human retina/RPE/choroids in the maculas of young and old donor eyes with or without AMD.  Colabeling with VE-cadherin was used to identify CECs. CECs transfected with Thy-1 siRNA or control siRNA were stimulated with VEGF, and CEC migration and phosphorylation of VEGF receptor 2 (VEGFR2) were measured. Statistics were performed using ANOVA.

Results:  Thy-1was highly expressed in CECs but not in RPE.  Thy-1 staining was not only detected in the retinal ganglion cell layer, but also in the choroid, and colocalized to a greater extent with VE-cadherin labeled CECs in sections from donors with AMD compared to age-matched controls without AMD.  Thy-1 mRNA was significantly increased in CECs treated with VEGF or CCL11 (p<0.05 vs. PBS) and greater in RPE/choroids from aged donor eyes (p<0.001 vs. young).  Knockdown of Thy-1 in CECs by siRNA transfection significantly inhibited VEGF-induced CEC migration (p<0.001) and VEGFR2 activation.

Conclusions: Thy-1is expressed in CECs and its expression is upregulated by stresses associated with neovascular AMD, including elderly age and increased VEGF.  Upregulated Thy-1 in CECs contributes to VEGF-induced VEGFR2 activation and CEC migration. Future studies into the potential role of Thy-1 in neovascular AMD are being considered. 

Müller Cell-derived VEGF164 Knockdown Reduces Retinal Neovascularization In A Rat Model Of Retinopathy Of Prematurity

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This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by  Deeksha Gambhir, Yanchao Jiang, George W. Smith, Zhihong Yang, Haibo Wang, Lori Fotheringham and M. Elizabeth Hartnett.

Full size poster available here.

Continue reading “Müller Cell-derived VEGF164 Knockdown Reduces Retinal Neovascularization In A Rat Model Of Retinopathy Of Prematurity”

A Study of PDZ Domain-Containing 7 (PDZD7) in the Mouse Retina

Powerpoint template for scientific poster

This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by  Junhuang Zou, Tihua Zheng and Jun Yang.

Full size poster can be downloaded here.

Purpose: PDZD7 is a newly identified modifier and contributor gene of Usher syndrome (USH). In the inner ear, PDZD7 colocalizes with GPR98, an USH2C protein, at ankle links in cochlear and vestibular hair cells. Therefore, PDZD7 is proposed to be a novel component of the USH2 complex, which is composed of the three known USH2 causative proteins. In this study, we investigated PDZD7 expression and its role in the organization of the USH2 complex in the retina.

Methods: The expression of Pdzd7 was examined at the mRNA and protein levels using RT-PCR, western blotting, and immunostaining assays. A Pdzd7 knockout mouse was generated by gene trapping and characterized phenotypically by immunostaining and electroretinogram.
Results: Five Pdzd7 splice variants were identified from 19 independent RT-PCR clones in adult mouse retinas. All of them are predicted as N-terminal but not full-length Pdzd7 isoforms. At the protein level, full-length Pdzd7 was found in the mouse retina during postnatal development. However, no Pdzd7 protein expression could be detected in adulthood by either western blotting or immunostaining. Pdzd7 knockout mice showed close to normal distribution of USH2A, GPR98 and WHRN at the periciliary membrane complex in photoreceptors. The knockout mice exhibited normal ERG responses at one month of age.

Conclusions: Despite the existence of multiple splice variants, PDZD7 expression at the protein level is very low in the retina. PDZD7 is not as important as WHRN in organizing the USH2 complex in photoreceptors.

A Synaptic Basis for Small World Network Design in the ON Inner Plexiform Layer of the Rabbit Retina

Bipolar cells_

This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by J Scott Lauritzen, Noah T. Nelson, Crystal L. Sigulinsky, Nathan Sherbotie, John Hoang, Rebecca L. PfeifferJames R. Anderson, Carl B. Watt, Bryan W. Jones and Robert E. Marc.

Purpose: Converging evidence suggests that large- and intermediate-scale neural networks throughout the nervous system exhibit small world’ design characterized by high local clustering of connections yet short path length between neuronal modules (Watts & Strogatz 1998 Nature; Sporns et al.2004 Trends in Cog Sci). It is suspected that this organizing principle scales to local networks (Ganmor et al. 2011 J Neurosci; Sporns 2006 BioSystems) but direct observation of synapses and local network topologies mediating small world design has not been achieved in any neuronal tissue. We sought direct evidence for synaptic and topological substrates that instantiate small world network architectures in the ON inner plexiform layer (IPL) of the rabbit retina. To test this we mined ≈ 200 ON cone bipolar cells (BCs) and ≈ 500 inhibitory amacrine cell (AC) processes in the ultrastructural rabbit retinal connectome (RC1).

Methods: BC networks in RC1 were annotated with the Viking viewer and explored via graph visualization of connectivity and 3D rendering (Anderson et al. 2011 J Microscopy). Small molecule signals embedded in RC1 e.g. GABA glycine and L-glutamate combined with morphological reconstruction and connectivity analysis allow for robust cell classification. MacNeil et al. (2004 J Comp Neurol) BC classification scheme used for clarity.

Results: Homocellular BC coupling (CBb3::CBb3 CBb4::CBb4 CBb5::CBb5) and within-class BC inhibitory networks (CBb3 → AC –| CBb3 CBb4 → AC –| CBb4 CBb5 → AC –| CBb5) in each ON IPL strata form laminar-specific functional sheets with high clustering coefficients. Heterocellular BC coupling (CBb3::CBb4 CBb4::CBb5 CBb3::CBb5) and cross-class BC inhibitory networks (CBb3 → AC –| CBb4 CBb4 → AC –| CBb3 CBb4 → AC –| CBb5 CBb5 → AC –| CBb4 CBb3 → AC –| CBb5 CBb5 → AC –| CBb3) establish short synaptic path lengths across all ON IPL laminae.

Conclusions: The retina contains a greater than expected number of synaptic hubs that multiplex parallel channels presynaptic to ganglion cells. The results validate a synaptic basis (ie. direct synaptic connectivity) and local network topology for the small world architecture indicated at larger scales providing neuroanatomical plausibility of this organization for local networks and are consistent with small world design as a fundamental organizing principle of neural networks on multiple spatial scales.

Support:  NIH EY02576 (RM), NIH EY015128 (RM), NSF 0941717 (RM), NIH EY014800 Vision Core (RM), RPB CDA (BWJ), Thome AMD Grant (BWJ).

The Role of NMDA Receptor Activity in Retinal Ganglion Cell Dendrite Development

Scientific poster example

 

This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by Eerik M. Elias, Ping Wang and Ning Tian.

Full size poster available here.

Purpose: To elucidate mechanisms underlying the dendrite developmental plasticity of retinal ganglion cells, we examined the role of glutamate receptors on retinal ganglion cell dendrite elongation and filopodia elimination.

Methods: We used the JamB genetically labeled subtype of RGCs as our working model. JamB-CreER:YFP ganglion cell dendritic arbors were imaged in whole mount retina using confocal microscopy. Dendrite length, area, branching, and filopodia number were traced and measured using Neurolucida. Visual inputs were blocked by dark-rearing pups after P5. Glutamatergic activity was blocked using daily intraocular injections of AP5 and CNQX from P9 to P13 or genetic ablation of the NMDA receptor in these RGCs.

Results: To test the role of visual inputs on dendrite development, we dark-reared mice from P5 to P30 and found a modest effect on filopodia elimination in JamB RGCs. Anticipating that spontaneous glutamatergic activity in the retina may also contribute to RGC filopodia elimination, we blocked spontaneous glutamatergic activity by daily intraocular injections of AP5 and CNQX from P9 to P13. This led to an increase in filopodia density due to decreased dendrite length but no change in filopodia number. We confirmed this result by examining NMDAR knockout JamB cells (JamB-CreER:YFP:Grin1-/-). As expected, Grin1-/- JamB RGCs have decreased dendrite outgrowth like the pharmacologic blockade. However, filopodia elimination in these cells was significantly decreased as well, suggesting that NMDA and non-NMDA glutamate receptors might regulate the RGC dendritic development in a differential manner. This effect was dramatic at P13. To test if this effect persists into adulthood, we examined Grin1-/- JamB RGCs at P30 and found that they are indistinguishable from wild-type JamB RGCs, suggesting that a compensatory mechanism exists to drive dendrite elongation and filopodia elimination in the absence of the NMDA receptor.

Conclusions: Our study demonstrated that ganglion cell dendrite outgrowth and pruning of filopodia require glutamatergic activity and visual input that act via NMDA and possibly non-NMDA glutamate receptors.

Suppressing Thyroid Hormone Signaling Preserves Cone Photoreceptors in Mouse Models of Retinal Degeneration

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This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by Xi-Qin Ding, Hongwei Ma, Arjun Thapa, Lynsie Morris, T M. Redmond and Wolfgang Baehr.

Full size poster available here.

Purpose: Cone phototransduction and survival of cones in the human macula is essential for color vision and for visual acuity. Progressive cone degeneration in age-related macular degeneration, Stargardt disease, and recessive cone dystrophies is a major cause of blindness. Thyroid hormone (TH) signaling which regulates cell proliferation, differentiation, and apoptosis plays a central role in cone opsin expression and patterning in the retina. Here, we investigated whether TH signaling affects cone viability in inherited retinal degeneration mouse models.

Methods: Rpe65-/- mice (a model of severe Leber congenital amaurosis or LCA) and Cpfl1mice (severe recessive achromatopsia) were used to determine whether suppressing TH signaling (with anti-thyroid treatment) reduces cone death. Further, Cngb3-/- mice (moderate achromatopsia) and Gucy2e-/- mice (moderate LCA) were used to determine whether stimulating TH signaling (with triiodothyronine (T3) treatment) deteriorates cones. The serum T3 levels were analyzed by ELISA. Cone and rod survival were evaluated by examining cone density and expression levels of cone specific proteins using immunohistochemical and biochemical approaches, and by examining morphological integrity of the retinas.

Results: Cone density increased about 6-fold in Rpe65-/- and cpfl1 mice following anti-thyroid treatment and decreased about 40% inCngb3-/- and Gucy2e-/- mice following T3 treatment. Anti-thyroid treatment did not affect rod survival, manifested as unchanged outer nuclear layer (ONL) thickness and the number of nuclei in ONL. However, T3 treatment significantly reduced ONL thickness and the number of nuclei in ONL in Cngb3-/- andGucy2e-/- mice.
Conclusions: With multiple retinal degeneration mouse models, we demonstrate that TH signaling regulates photoreceptor viability in degenerating retinas. Suppressing TH signaling protects cones whereas stimulating TH signaling has a negative effect on both cones and rods. The findings of this study provide new insights into cone preservation and therapeutic interventions.

Metabolic Changes Associated With Müller Cells In A Transgenic Rabbit Model Of Retinal Degeneration

Retina RLP

This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by  Rebecca L. Pfeiffer, Bryan W. Jones and Robert E. Marc.

Purpose: Müller cells play a central role in retinal metabolism via the glutamate cycle. During retinal degeneration Müller cells are among the first to demonstrate changes, reflected in alterations of metabolic signatures and morphology. The timing, extent and regulation of these changes is not fully characterized. To address this issue, we evaluated Müller cell metabolic phenotypes at multiple stages of retinal remodeling.

Methods: Samples were collected post-mortem from both WT and P347L rabbits. The retinas were then divided into fragments, fixed in buffered aldehydes, and embedded in epoxy resins. Tissues were sectioned at 200nm followed by classification with computational molecular phenotyping (CMP) using an array of small and macromolecular signatures (aspartate (D), glutamate (E), glycine (G), glutamine (Q), glutathione (J), GABA (yy), taurine (T), CRALBP, Glutamine Synthetase (GS), and GFAP). Levels of amino acid or protein were quantified by selecting a region of interest either within the Müller cell population or surrounding neurons and evaluating the intensity of the signal within that region.

Results: CMP reveals overall decreases in GS levels over the course of degeneration. Of notable importance, we saw that in regions of near complete photoreceptor loss neighboring Müller cells may express independent variation in metabolic signatures of E, Q, and GS. Also observed in these Müller cells, ratios of GS:E and GS:Q are not consistent with the ratios seen in WT retina. These results are inconsistent with the current models of both E to Q metabolism and microenvironment regulation of Müller cell phenotypes.

Conclusions: These observations indicate two conclusions. First, although the degenerate state of the retina is the likely trigger inducing Müller cells to express altered metabolic signatures, the rate at which the metabolic state changes is not purely a product of the surrounding environment, but also a stochastic change within individual Müller cells. Second, although it is commonly accepted that GS is the primary enzyme which converts Q to E as part of the glutamate cycle, in degenerate retina alternative pathways may be utilized following decrease in GS.

Support:  NIH EY02576 (RM), NIH EY015128 (RM), NSF 0941717 (RM), NIH EY014800 Vision Core (RM), RPB CDA (BWJ), Thome AMD Grant (BWJ).

Retinitis Pigmentosa 2 Protein Regulates Transport Of Isoprenylated Proteins To Photoreceptor Outer Segments

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This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by Houbin Zhang, Li Jiang, Christin Hanke and Wolfgang Baehr.

Full size poster can be downloaded here.

Purpose: X-linked retinitis pigmentosa (XLRP) is a devastating form of retinal degeneration, manifesting early in life with symptoms of night blindness, visual field defects, and decreased visual function. In-vitro, RP2 functions as a GAP for the small GTPase ARL3, a GDI displacement factor (GDF). Mutations in the Rp2 gene account for approximately one quarter of all XLRPs. The purpose of this study was to investigate the consequences of RP2 deletion and identify mechanisms causative of XLRP.

Methods: Intracellular localization of RP2 in photoreceptors was determined by neonatal electroporation of an RP2-EGFP expression vector. An Rp2 knockout mouse was generated using a EUCOMM ES cell line containing a gene trap in intron 1. The knockout mice were characterized by Western blot, immunocytochemistry, and electroretinography (ERG).

Results: RP2-eGFP was localized to the plasma membrane of inner segments, axons and synaptic termini in photoreceptors, but not in outer segments. The Rp2 gene knockout mice were viable and developed normally. Ablation of Rp2 gene expression led to slowly progressing degeneration of cone and rod photoreceptors as indicated by ERG recordings. Scotopic a-wave and photopic b wave amplitudes were reduced as early as one month of age in the knockout mice. The Rp2Y/- ERG amplitudes were further reduced at 6 months of age. Trafficking of transmembrane phototransduction proteins, including cone opsins, to Rp2Y/- photoreceptors outer segments was normal up to 14 months of age. While targeting of transducin α and βγ to the Rp2Y/- outer segments was not affected in the knockout, transport of rod and cone PDE6 as well as GRK1 to outer segments was impeded.

Conclusions: RP2 is distributed to plasma membrane of inner segments and synaptic termini in photoreceptors. RP2 is not essential for trafficking cone opsins and transducin to photoreceptor outer segments, but regulates transport of isoprenylated proteins to photoreceptor outer segments. Our results suggest that RP2/ARL3 may allosterically release prenylated proteins from their soluble complex with PDE6D and unload them to donor membranes (e.g., TGN vesicles). ). In the Rp2 knockout, this process is impeded.

Arl3 Rod-Specific Knockout Displays RP-Like Photoreceptor Degeneration

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This abstract was presented today at the 2014 Association for Research in Vision and Opthalmology (ARVO) meetings in Orlando, Florida by Christin Hanke, Houbin Zhang, Cecilia D. Gerstner, Jeanne M. Frederick AND Wolfgang Baehr.

Full size poster can be downloaded here.

Purpose: Arf-like protein 3 (Arl3) localizes predominantly in the photoreceptor inner segment. Germline Arl3 knockout mice do not survive beyond PN 21 and display multiple organ ciliary defects as well as retinal regeneration (Schrick et al., (2006). Am. J. Pathol. 168, 1288-1298). We therefore generated rod-specific Arl3 knockouts to elucidate the role of Arl3 in transport of photoreceptor membrane-associated proteins.

Methods: Knockouts containing a gene trap in intron 1 of the Arl3 gene were generated using a EUCOMM cell line. Breeding with Flp mice, followed by mating with iCre75+ mice, generated rod-specific knockouts. Photoreceptor function and retina morphology of wild-type (WT) and mutant mice were analyzed by confocal microscopy, ERG and immunohistochemistry. An Arl3-specific polyclonal antibody (Ab) was generated using a full-length recombinant Arl3 polypeptide expressed in bacteria.

Results: Western blot of WT retina with anti-Arl3-Ab identified a 20 kDa protein, which was significantly reduced in two month-old mutant (Arl3flox/flox;iCre75+) retina. Immunohistochemistry revealed Arl3 localization predominantly in the inner segments of WT photoreceptor cells. Arl3 immunoreactivity was absent in homozygous rod knockouts, but still present in cones and the inner retina. Scotopic and photopic ERGs of rod knockout and WT mice at PN15 had comparable amplitudes suggesting normal phototransduction. Retina histology of PN15 knockout mice was comparable to WT. One month-old Arl3flox/flox;iCre75+ mice showed reduced (80-90%) scotopic, but normal photopic ERG responses. In retinas of two month-old knockout mice, scotopic ERGs were extinguished, whereas cone ERGs were highly attenuated. Retinas of one month-old homozygous knockout mice had 4-5 rows of nuclei in the ONL, and only one row in two month-old mice. Immunohistochemistry of PN 15 and one month-old retina sections revealed that rhodopsin transport, as shown by rho1D4 labeling of ROS, is normal. Rhodopsin was undetectable in two month-old conditional knockout mice due to complete photoreceptor degeneration.

Conclusions: Rod-specific knockout of Arl3 revealed rapidly progressing photoreceptor degeneration, with knockout mice being completely blind at two months of age. Outer segment development appeared to be unimpaired by Arl3 deletion and rod photoreceptor function was normal at P14.

Flying to ARVO 2014

Flying to ARVO 2014

We are on our way to ARVO, 2014 in Orlando, Florida to participate in the largest gathering of vision scientists and clinicians in the world.  It’s the annual meeting of researchers presenting and discussing all things vision and ophthalmology and a large group from the Moran Eye Center will be going.

Over the next few days, you will see some of our research abstracts appear here as the presentations are completed at ARVO.  We hope that it will give some insight into the work that goes on here at the Moran Eye Center and our passion for understanding vision and what goes wrong in blinding diseases.

If you are going to be at ARVO and want to meet up, or want to come to the big soirée on Monday night (open bar) celebrating Wolfgang Baehr’s Proctor Medal Award, leave us a comment here or send a Tweet to @BWJones.  We might even be able to work you into the Moran social on Monday the 2nd…

Look forward to seeing you there.