The proposed aims build on investigations during the previous funding period on the identification of a single ligand-receptor pair, EphB1-ephrinB2 that directs repulsion of ventrotemporal (VT) retinal ganglion cell (RGC) growth cones at the optic chiasm midline. EphB1, expressed strictly on VT RGCs, interacts with the ligand ephrinB2 on radial glia at the chiasm midline to direct formation of the uncrossed projection. Mechanisms of midline crossing that guide RGC axons across the optic chiasm midline remain unclear. We have identified NrCAM as necessary for the establishment of the late crossed projection from VT retina. NrCAM is also expressed in RGCs outside the VT crescent.
In Aim 1. a, the function of two other CAMs, TAG- 1 and Neurofascin, expressed similarly to NrCAM, will be investigated for their involvement in crossing the midline. In addition, we localized Sema6D at the chiasm midline and its receptor, Plexin-A1, is expressed in all crossing RGCs. In vitro, Sema6D inhibits the growth of RGCs that project across the midline.
In Aim 1. b., the actions of Sema6D and Plexin-A1 in mediating the crossed RGC projection will be studied. CAMs have been shown to collaborate with semaphorins in other crossing systems. This issue will be addressed in Aim 1.c. Transgenic mice, culture assays, and methods for gene delivery (electroporation in utero and ex utero) developed in our lab will be used to provide a comprehensive view of the crossing process in the chiasm. The formation of eye-specific terminations in the dLGN is a direct consequence of retinal axon decussation in the optic chiasm. In our studies on the uncrossed projection, we asked how aberrant midline decussation affects specific innervation patterns in the dLGN. In EphB1 gain- and loss-of-function models, RGCs misproject to the wrong side of the midline yet terminate in appropriate eye-specific zones. These results implicate molecular "tags" for eye-specific innervation. As a foundation for further analysis, in Aim 2.a., we will chronicle the ingrowth, arbor formation and segregation of mouse retinogeniculate axons at the single-fiber level, as this information is lacking.
In Aim 2. b., we will identify molecules that may implement eye-specific innervation of the dLGN, by 1) localization of candidates studied in Aim 1, and 2) transcriptional profiling of specific tissue regions.
In Aim 2. c. and d., we will then determine the role of candidate molecules by perturbing them and by altering neural activity. The phenomenon of coordinate action by multiple guidance families such as Semaphorins and CAMs, presents a new level of analysis of axon navigation in higher vertebrate retinal pathways, where divergence is controlled by two distinct molecular systems. The proposed experiments thus address fundamental questions on how the binocular pathways develop.

Public Health Relevance

This research aims to understand how growing retinal ganglion cells from each eye converge at the X- shaped optic chiasm, and then diverge toward targets on the same and opposite side of the brain. Proper binocular vision is dependent on a normal distribution of retinal axons at the optic chiasm, and if altered, reduced visual acuity and depth perception ensue. These studies use this model to investigate the molecular factors that help guide axons to their appropriate route and synaptic targets in the brain.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY012736-24
Application #
8500287
Study Section
Central Visual Processing Study Section (CVP)
Program Officer
Steinmetz, Michael A
Project Start
1999-07-01
Project End
2014-06-30
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
24
Fiscal Year
2013
Total Cost
$394,337
Indirect Cost
$149,407
Name
Columbia University (N.Y.)
Department
Pathology
Type
Schools of Medicine
DUNS #
621889815
City
New York
State
NY
Country
United States
Zip Code
10032
Bhansali, Punita; Rayport, Ilana; Rebsam, Alexandra et al. (2014) Delayed neurogenesis leads to altered specification of ventrotemporal retinal ganglion cells in albino mice. Neural Dev 9:11
Assali, Ahlem; Gaspar, Patricia; Rebsam, Alexandra (2014) Activity dependent mechanisms of visual map formation--from retinal waves to molecular regulators. Semin Cell Dev Biol 35:136-46
Kuwajima, Takaaki; Sitko, Austen A; Bhansali, Punita et al. (2013) ClearT: a detergent- and solvent-free clearing method for neuronal and non-neuronal tissue. Development 140:1364-8
Roffler-Tarlov, Suzanne; Liu, Jin Hong; Naumova, Elena N et al. (2013) L-Dopa and the albino riddle: content of L-Dopa in the developing retina of pigmented and albino mice. PLoS One 8:e57184
Rebsam, Alexandra; Bhansali, Punita; Mason, Carol A (2012) Eye-specific projections of retinogeniculate axons are altered in albino mice. J Neurosci 32:4821-6
Petros, Timothy J; Bryson, J Barney; Mason, Carol (2010) Ephrin-B2 elicits differential growth cone collapse and axon retraction in retinal ganglion cells from distinct retinal regions. Dev Neurobiol 70:781-94
Rebsam, Alexandra; Petros, Timothy J; Mason, Carol A (2009) Switching retinogeniculate axon laterality leads to normal targeting but abnormal eye-specific segregation that is activity dependent. J Neurosci 29:14855-63
Petros, Timothy J; Rebsam, Alexandra; Mason, Carol A (2009) In utero and ex vivo electroporation for gene expression in mouse retinal ganglion cells. J Vis Exp :
Petros, Timothy J; Shrestha, Brikha R; Mason, Carol (2009) Specificity and sufficiency of EphB1 in driving the ipsilateral retinal projection. J Neurosci 29:3463-74
Lee, Ramee; Petros, Timothy J; Mason, Carol A (2008) Zic2 regulates retinal ganglion cell axon avoidance of ephrinB2 through inducing expression of the guidance receptor EphB1. J Neurosci 28:5910-9

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