McFarlane Lab

We study the development and maintenance of the neural circuits of the eye, as a mini-brain, which is easily accessible and separate from the rest of the central nervous system. My laboratory is interested how cell movements and position are controlled to: 1) give the eye its unique structure, 2) allow the appropriate connections to be made between retinal neurons, and 3) allow proper replacement of cells that are lost due to retinal injury. To do so, we use cutting-edge genetic and molecular approaches, combined with confocal imaging of cells in the living embryo. Our funded projects include:

1)   Understanding the regeneration of the retinal pigment epithelium (RPE)

The cells in your retina process visual inputs and send the information to the brain so that you can make sense of your visual world. The retinal pigment epithelium (RPE) is a thin layer of cells that covers the back of your retina, where it functions as a nursemaid for the photosensing cells of your retina, the photoreceptors: the RPE protects, provides structural and survival support, and keeps the light sensing pathways on-line. Because of the importance of the RPE in photoreceptor health, defects in the RPE lead to photoreceptor degeneration, and loss of sight. One approach to deal with diseases where photoreceptors degenerate, such as in Age-Related Macular Degeneration (AMD) and Retinitis Pigmentosa (RP), is to use stem cells to first generate and then transplant into the damaged retina the RPE cells that could block and/or delay photoreceptor degeneration. In this treatment paradigm, RPE cells have to migrate from the site of transplantation. Further, aberrant migration of RPE cells can occur in disease states where vision loss ensues, such as AMD. Thus, understanding what drives RPE migration and regeneration is important. Yet, we know little about what molecules control how RPE cells move and what drives their regeneration in vivo. Here, we use zebrafish as an experimental model, primarily because genetic mutagenesis is quick and effective, so we can study how specific proteins control RPE movement both in the embryo and after RPE or retinal injury. Further, the zebrafish model allows us to track RPE cells in real time in live embryos and larvae, by using a transgenic line where the RPE is engineered to express a fluorescent protein, providing unparalleled information about the consequences of gene disruption on RPE migration.

2)    Investigating Semaphorins as regulators of progenitor movement and position (Canadian Institutes of Health Research, 2021-2026; Lion’s Sight Centre Endowment Award, 2020-2022)

Retinal degenerative diseases such as glaucoma and diabetic retinopathy exhibit devastating vision loss, each as a result of the gradual death with time of specific types of eye cell. Each disease needs the affected cell types to be replaced, and for the new cells to integrate into functional vision circuits. Thus, we need to explore how different types of eye cell are made and in the right proportions. Importantly, we need to understand these events in the adult eye, because the blueprints likely differ from those for eye cell production in the fetus, from where most of our knowledge comes. The problem is that the mammalian eye is incapable of replacing new eye cells when they degenerate. This is not true in the eyes of adult fish, which retain two subpopulation of cells that allow the eye and retinal circuits to grow throughout life, and that in response to retinal injury are activated and replace lost cells. Importantly, these two classes of cells are also present in the mammalian adult retina, but do not respond to retinal damage by making new cells. The fish eye has the same main classes of eye cells, and with similar functions, as those in humans. Thus, we want to understand how new cells are generated in the fish adult eye, and how these pathways can be harnessed in repair programs after retinal cell loss. We have identified related proteins in the zebrafish eye, Semaphorin3fa and Semaphorin3fb, that our data indicate act on the two key cell populations, both as these cells continue to provide new cells for the ever-expanding eye visual circuits, and as they ramp up their activities in response to eye damage.

This project explores the roles of Sema3fs in both contexts with the hope of understanding how to transform the dormant cells of the human adult eye into progenitors like their fish counterparts, so that they too can replace specific eye cell types that get lost in each of the degenerative retinal diseases.

3)    Identifying the molecular mechanisms underlying light-dependent regulation of neural crest cell development

The peripheral nervous system is critical for mounting and coordinating an organism’s response to changes in its internal and external environment. Most cells of the peripheral nervous system -- including most peripheral neurons and glia -- are derived from neural crest cells (NCC).  The NCC progenitors are born at the dorsal edge of the forming neural tube, from which they delaminate, migrating to target sites where they differentiate to form target specific cell types. We understand only some of the steps of this fascinating process, including some of the molecules that control the induction and guidance of NCC progenitors; however, an important knowledge gap and the focus of this proposal, is how an organism’s environment can impinge on these intrinsic and external processes to impact NCC progenitor behaviour.

Pigmented skin cells (chromatophores) are involved in environmental responses related to camouflage and UV protection. Chromatophores also derive from NCC, and their development depends on many of the same molecular signals that control NCC-dependent peripheral neuron genesis. Their ready visibility in the skin of live embryos, and our recent demonstration that their genesis can be controlled by environmental light (Bertolesi et al., 2016), makes them a particularly good model for studying how environmental influences control the signalling events that regulate NCC development. We hypothesize that light perceived by the eye acts through a neural circuit to release systemic factors that control NCC progenitor proliferation and apoptosis. We are using the Xenopus laevis embryo model, readily amenable to surgical, molecular and pharmacological manipulations in vivo, to address this hypothesis.

This research program is at the forefront of understanding how the external environment can directly impact the developing nervous system, in that we have identified a natural signal, light, that controls NCC progenitor biology to adapt the organism to its milieu. We will study the neural circuitry and signalling events that control light-mediated changes in skin pigmentation using both in vitro and in vivo models. Skin pigmentation is readily assessed, and so the model is experimentally amenable, and will provide key foundational information that can be used to understand how the external environment can impact the processes that control the development of other NCC derived cell types.