Differentiation of Voltage- and Ligand-gated Currents in Early and Late Retinal Progenitors

Abstract

Abstract: : Purpose: The generation of retinal cells involves two different progenitor populations. The early retinal progenitors give rise to retinal ganglion cells (RGCs), cone photoreceptors and horizontal cells between E 11 and E 16 (early neurogenesis) and the late retinal progenitors generate the majority of rod photoreceptors, amacrine cells, bipolar cells and Muller glia from E 18 onward (late neurogenesis). The early and late progenitors are distinct in terms of gene expression and in their ability to differentiate into neurons and glia (Jackson et al., 2002, ARVO abstract). We were interested in knowing whether early and late retinal progenitors are distinct physiologically in terms of voltage- and ligand-gated currents under differentiating culture conditions. Methods: Cell dissociates from E 14 and E 18 retina were cultured in the presence of EGF and /or FGF2 to obtain neurospheres. To promote differentiation, neurospheres were plated on poly-D-lysine and laminin coated cover slips and cultured in the presence of 1 % FBS for 14 days. Whole cell recordings were carried out at two-day intervals as previously described (Ahmad et al, 1999, Brain Research, Jun 12:831:1-10). Results: Sodium (I_Na) and potassium (I_K) currents were detected in both early and late retinal progenitors on the second day in culture. While the majority of early and late progenitors displayed I_K, only 22 % of these cells displayed I_Na. The proportion of cells with I_Na increased with culture time. The response of early and late progenitors to NMDA and KA was distinct; while early progenitors showed no response to NMDA, a small proportion (~12%) of late retinal progenitors displayed NMDA-evoked currents on day 2. However, with time the early retinal progenitors acquired NMDA responsiveness similar to their late counterpart. In contrast to NMDA, KA-responsiveness was observed in cells belonging to both progenitor populations on day 2, however, more early retinal progenitors displayed KA-evoked currents than late retinal progenitors (~75 % Vs 22 %). In general, a trend of an increase in the number of Ka and NMDA-responsive cells with time was observed in both cell-populations. Conclusions: The early and late retinal progenitors are distinct in their ability to generate neural and specific retinal cell types. Preliminary results suggest that they may also be distinct in the acquisition of functional characteristics as defined by voltage- and ligand-gated channels.

Keywords: retinal development • proliferation • electrophysiology: non-clinical