In retinal degeneration, death of photoreceptors causes blindness. Repair of the retina by transplanting photoreceptors has resulted in limited functional connectivity between transplanted and host neurons. We hypothesize that absence of appropriate biological cues, specifically positional (retinotopographic) cues, reduces synaptogenesis. Here we use micropatterning to test whether regional origin affects the early synaptic development of photoreceptors. Right and left retinas from salamanders were first labelled with dextran tetramethyl-rhodamine and fluorescein, respectively, bisected into nasal (N)/temporal (T) or dorsal (D)/ventral (V) halves, individually dissociated, mixed, and cultured for 1 week. Origin of cells was identified by the fluorescent label. Interactions between photoreceptors and neighboring (target) cells were assessed by the number of neuritic contacts with a presynaptic swelling (varicosity). Randomly-plated photoreceptors showed no preference for cellular origin, likely due to multiple potential interactions available to each cell. To reduce cell-cell interactions, culture substrate was patterned using a microfluidic device with 10 μm-wide channels separated by 200 μm, thus allowing only 1-2 targets per photoreceptor. In patterned cultures, 36.89% of N rod cells contacted T targets but only 27.42% of N rod cells contacted N targets; similarly 35.05% of T rod cells contacted N cells but only 17.08% contacted T cells. Thus, opposite regions were more permissive of contact. However, neither cone nor D/V rod cells showed preferences for positional origin of targets. In conclusion, micropatterning demonstrated that neuritic differentiation by rod cells depends on retinotopographic cues along the nasal/temporal plane, suggesting that transplanting rod cells of known positional origin will increase transplant success.
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