Figure S3.

BC Axons Can Form a Sublaminated Neuropil in the Absence of Partner Neurons, Related to Figure 2

(A) Cryosections from 5 dpf ath5:GAP-RFP transgenic stained with anti-PKC (labeling ON-BCs) and HuC/D (labeling RGCs and ACs).

(B) ath5^−/−ptf1a^−/−;ptf1a MO retinas lack HuC/D positive RGCs and ACs (remaining AC - arrowhead), but an ath5:GAP-RFP/PKC-labeled IPL still forms.

(C) In the ath5:GAP-RFP;ath5^−/−ptf1a^−/−;ptf1a MO retinas, the ath5:GAP-RFP label is expressed by a subpopulation of BCs in the INL, which are distinct from the population of BCs labeled by the anti-PKC (cell #1 is ath5:GAP-RFP-positve, while cell #2 is PKC-positive).

(D) Higher magnification inset of the WT IPL showing the targeting of PKC axon terminals to 3 sublaminae in the basal half of the IPL. This is shown quantitatively by three peaks in the line intensity profile in region (i).

(E) The AC/RGC-free IPL shows some degree of disorganization. Yet, in some areas the PKC signal clearly segregates basal to the ath5:GAP-RFP signal in a stratified pattern (ii and iii). In other areas this is not the case (iv).

(F) Line intensity profiles were normalized for intensity of relative apical/basal position along the IPL, and plotted as a single graph. This demonstrates the basal accumulation of PKC-labeled axons typical of the WT retina (n = 30 measurements, from 10 sections), as well and the basal enrichment of the PKC signal and the apical enrichment of the ath5:GAP-RFP signal in the ath5:GAP-RFP;ath5^−/−ptf1a^−/−;ptf1a MO retinas (n = 54 measurements, from 18 sections), are is reflected in the shift of the trendline. Note that ath5:GAP-RFP signal quantification is not shown for WT, as this transgene is expressed by RGCs and ACs in this context.