Inactivation of axonal EphA4 stimulates axon growth of nasal retinal ganglion cells

DI NAPOLI J.; FIORE L.; ALONSO C.; RODRÍGUEZ CELÍN A.; PASQUALE E.; CARRI N.; SCICOLONE G.

San Diego

Congreso; 40th Annual Meeting of Society for Neuroscience; 2010

Society for Neuroscience

The retinotectal system is useful to study topographic map formation because nasal retinal ganglion cells (RGC) connect to caudal tectum and temporal ones connect to rostral tectum. Eph receptor tyrosine kinases and their ligands, the ephrins, are expressed in complementary gradients in both, the retina and the tectum and guide retinotectal projections by producing bidirectional signaling. Ephrin-As located in the caudal tectum repel temporal axons by activating EphA3/A4. Ephrin-As expressed in nasal RGC activate EphA4 converting nasal RGC less sensitive to tectal ephrin-As. We demonstrated that tectal EphA3 stimulates axon growth of nasal RGC to caudal tectum. It is not known which molecule/s mediates this effect of EphA3. We postulated that EphA4 activation decreases axon growth and that tectal EphA3 increases axon growth by reducing EphA4 activation through competing with axonal EphA4 for axonal ephrin-As binding. Our objectives were to investigate whether: 1) the higher expression of retinal ephrin-As coincides with the RGC that grow shorter axons but increase their growth when binding to EphA3; 2) inactivation of axonal EphA4 mimics EphA3 effect on axon growth. Methods: We used retinal explants from 7 days-old chicken embryos (E7) to investigate the effect of inhibiting axonal EphA4 activity by using a peptide (KYL). We performed immunocytochemistry of retinal explants and paraffin sections from chicken eyes since E4 to first 10 postnatal days to investigate if the higher expression of these molecules correlates with the periods when connections are formed. Results: EphA4 is homogeneously expressed in RGC growth cones. Ephrin-As are highly expressed in nasal and moderately in temporal growth cones. These expression patterns are highest between E13 and E15 and decreases postnatally. Temporal RGC, which express lower levels of ephrin-As, grow significant longer axons than nasal ones. Inhibition of axonal EphA4 produces a significant increase in axongrowth of nasal RGC, which express higher levels of ephrin-As. Discussion: The fact that temporal RGC, which express lower levels of ephrin-As, grow longer axons suggests that EphA4 activation by coexpressed ephrin-As decreases axon growth. The fact that nasal RGC, which express higher levels of ephrin-As, grow longer axons with EphA4 inhibition and with EphA3 shows that EphA4 inhibition simulates axon growth produced by EphA3. This suggests that tectal EphA3 could stimulate axon growth of nasal RGC through binding with axonal ephrin-As and indirectly inhibiting EphA4 activity. The highest ephrin-A2 expression observed correlates with the ontogenic stage in which the topographic connections are formed.