Anatomy and Neuroscience - Research Publications
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ItemNo Preview AvailableFractalkine-induced microglial vasoregulation occurs within the retina and is altered early in diabetic retinopathy( 2020)Local blood flow control within the CNS is critical to proper function and is dependent on coordination between neurons, glia and blood vessels. Macroglia such as astrocytes and Müller cells, contribute to this neurovascular unit within the brain and retina, respectively. This study explored the role of microglia, the innate immune cell of the CNS, in retinal vasoregulation and highlights changes during early diabetes. Structurally, microglia were found to contact retinal capillaries and neuronal synapses. In the brain and retinal explants, the addition of fractalkine, the sole ligand for monocyte receptor Cx3cr1, resulted in capillary constriction at regions of microglial contact. This vascular regulation was dependent on microglial involvement, since mice lacking Cx3cr1, exhibited no fractalkine-induced constriction. Analysis of the microglial transcriptome identified several vasoactive genes, including angiotensinogen, a constituent of the renin-angiotensin system (RAS). Subsequent functional analysis showed that RAS blockade via candesartan, abolished microglial-induced capillary constriction. Microglial regulation was explored in a rat streptozotocin (STZ) model of diabetic retinopathy. Retinal blood flow was reduced after 4 weeks due to reduced capillary diameter and this was coincident with increased microglial association. Functional assessment showed loss of microglial-capillary response in STZ-treated animals and transcriptome analysis showed evidence of RAS pathway dysregulation in microglia. While candesartan treatment reversed capillary constriction in STZ-treated animals, blood flow remained decreased likely due to dilation of larger vessels. This work shows microglia actively participate in the neurovascular unit, with aberrant microglial-vascular function possibly contributing to the early vascular compromise during diabetic retinopathy.
Significance Statement
This work identifies a novel role for microglia, the innate immune cells of the CNS, in the local control of the retinal vasculature and identifies deficits early in diabetes. Microglia contact neurons and vasculature and express several vasoactive agents. Activation of microglial fractalkine-Cx3cr1 signalling leads to capillary constriction and blocking the renin-angiotensin system (RAS) with candesartan abolishes microglial-mediated vasoconstriction in the retina. In early diabetes, reduced retinal blood flow is coincident with capillary constriction, increased microglial-vessel association, loss of microglial-capillary regulation and altered microglial expression of the RAS pathway. While candesartan restores retinal capillary diameter early in diabetes, targeting of microglial-vascular regulation is required to prevent coincident dilation of large retinal vessels and reduced retinal blood flow. -
ItemPotential mechanisms of retinal ganglion cell type-specific vulnerability in glaucoma(WILEY, 2020-09)Glaucoma is a neurodegenerative disease characterised by progressive damage to the retinal ganglion cells (RGCs), the output neurons of the retina. RGCs are a heterogenous class of retinal neurons which can be classified into multiple types based on morphological, functional and genetic characteristics. This review examines the body of evidence supporting type-specific vulnerability of RGCs in glaucoma and explores potential mechanisms by which this might come about. Studies of donor tissue from glaucoma patients have generally noted greater vulnerability of larger RGC types. Models of glaucoma induced in primates, cats and mice also show selective effects on RGC types - particularly OFF RGCs. Several mechanisms may contribute to type-specific vulnerability, including differences in the expression of calcium-permeable receptors (for example pannexin-1, P2X7, AMPA and transient receptor potential vanilloid receptors), the relative proximity of RGCs and their dendrites to blood supply in the inner plexiform layer, as well as differing metabolic requirements of RGC types. Such differences may make certain RGCs more sensitive to intraocular pressure elevation and its associated biomechanical and vascular stress. A greater understanding of selective RGC vulnerability and its underlying causes will likely reveal a rich area of investigation for potential treatment targets.
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ItemOcular Phenotype of Relaxin Gene Knockout (Rln-/-) Mice(TAYLOR & FRANCIS INC, 2020-10-02)Purpose: To test if relaxin deficiency affects ocular structure and function we investigated expression of relaxin (Rln) and RXFP receptors (Rxfp1, Rxfp2), and compared ocular phenotypes in relaxin gene knockout (Rln-/- ) and wild type (Rln+/+ ) mice. Materials and Methods: Rln, Rxfp1 and Rxfp2 mRNA expression was detected in ocular tissues of Rln+/+ mice using RT-PCR. The eyes of 11 Rln-/- and 5 Rln+/+ male mice were investigated. Corneal and retinal thickness was assessed using optical coherence tomography. Intraocular pressure was measured using a rebound tonometer. Retinal, choroidal and sclera morphology and thickness were evaluated histologically. Eyes were collected and fixed for immunofluorescence staining or used for RNA extraction to evaluate mRNA expression using real-time PCR. Results: Rln mRNA was expressed only in the retina, whereas Rxfp1 transcripts were detected in the retina, cornea and sclera/choroid. Rxfp2 was only present in the cornea. None of these genes were expressed in the lacrimal gland, eyelid or lens. Intraocular pressure was higher and central cornea of Rln-/- mice was significantly thicker and had significantly larger endothelial cells and a lower endothelial cell density than Rln+/+ mice. Immunohistochemistry demonstrated no significant difference in AQP3 and AQP5 staining in the cornea or other regions between wildtype and Rln-/- mice. mRNA expression of Aqp4 was significantly higher in Rln-/- than in Rln+/+ corneas, whereas Col1a2, Mmp9, Timp1 and Timp2 were significantly decreased. Expression of Aqp1, Aqp4, Aqp5, Vim and Tjp1 was significantly decreased in Rln-/- compared to Rln+/+ uvea. No significant differences in these genes were detected in the retina. Retinal, choroidal and scleral thicknesses were not different and morphology appeared normal. Conclusion: The findings indicate that loss of Rln affects expression of several genes in the uvea and cornea and results in thicker corneas with altered endothelial cells. Many of the gene changes suggest alterations in extracellular matrix and fluid transport between cells.
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ItemTyro3 Contributes to Retinal Ganglion Cell Function, Survival and Dendritic Density in the Mouse Retina(FRONTIERS MEDIA SA, 2020-08-14)Retinal ganglion cells (RGCs) are the only output neurons of the vertebrate retina, integrating signals from other retinal neurons and transmitting information to the visual centers of the brain. The death of RGCs is a common outcome in many optic neuropathies, such as glaucoma, demyelinating optic neuritis and ischemic optic neuropathy, resulting in visual defects and blindness. There are currently no therapies in clinical use which can prevent RGC death in optic neuropathies; therefore, the identification of new targets for supporting RGC survival is crucial in the development of novel treatments for eye diseases. In this study we identify that the receptor tyrosine kinase, Tyro3, is critical for normal neuronal function in the adult mouse retina. The loss of Tyro3 results in a reduction in photoreceptor and RGC function as measured using electroretinography. The reduction in RGC function was associated with a thinner retinal nerve fiber layer and fewer RGCs. In the central retina, independent of the loss of RGCs, Tyro3 deficiency resulted in a dramatic reduction in the number of RGC dendrites in the inner plexiform layer. Our results show that Tyro3 has a novel, previously unidentified role in retinal function, RGC survival and RGC morphology. The Tyro3 pathway could therefore provide an alternative, targetable pathway for RGC protective therapeutics.