Regeneration of RIP axons in isolated CNS cultured in saline. A, Frequency of preparations in which RPeD1 and VD2/VD3 were retrograde labeled by backfilling the RIP nerve immediately after dissection of the CNS without nerve crush (no crush) or by backfilling the RIP nerve in crushed preparations after 2 d (crush, 2 days) or 7 d (crush, 7 days) in culture, respectively. Note that RPeD1 and VD2/VD3 are labeled in <40% of the preparations after 2 d in culture and that for both cell types this proportion did not significantly improve by keeping the preparations in culture for 7 d in culture. B, Frequency distribution of the number of retrograde-labeled RPA somata per CNS in preparations that were backfilled immediately after dissection without crushing the RIP nerve (no crush) and preparations in which the RIP nerve was crushed and backfilled after 2 d in culture (crush, 2 days) and 7 d (crush, 7 days) in culture, respectively.C, Average number of labeled RPA somata in the total 2 d data set (overall), a subset of the 2 d data in which neither RPeD1 nor VD2/VD3 was labeled (− −), and a subset of the 2 d data in which both RPeD1 and VD2/VD3 were labeled (+ +). ***p < 0.001.