FIGURE SUMMARY
Title

An amputation resets positional information to a proximal identity in the regenerating zebrafish caudal fin

Authors
Azevedo, A.S., Sousa, S., Jacinto, A., and Saude, L.
Source
Full text @ BMC Dev. Biol.

The bifurcation position is distalized with repeated amputations. (A) The same caudal fin before amputation and after 15 and 27 amputations. (B) Schematic representation of the bifurcation distalization with the repeated amputations. (C) Number of segments formed in the 3rd dorsal ray between the base of the fin and the bifurcation after consecutive amputations. The caudal fin was subjected to three amputations every month and this was repeated 10 times. During the first 15 amputations, the third consecutive amputation (the last before allowing the fin to completely regenerate) was done three segments below the most proximal bony ray bifurcation (near the bifurcation). In the next 12 cuts, the third consecutive amputation was done 4 segments distally from the base of the fin (near the base of the fin). (D) 3rd dorsal ray segment length before any amputation and after 24 amputations. Asterisk marks the bifurcation position. The fins were allowed to regenerate for 4 weeks between each round of three consecutive amputations.

The expression pattern ofshhduring regeneration does not change with the proximal-distal level or the number of amputations.(A-D) Caudal fins were amputated at 4 segments from the base of the fin (near the base of the fin) once or twice and shh expression was determined at 3 or 4 days following the amputation. (E-H) Caudal fins were amputated at 1 segment below the bifurcation (near the bifurcation) once or twice and shh expression was determined at 3 or 4 days following the amputation. (I) Caudal fins with no bifurcations were amputated near the base of the fin and shh expression was examined at 4 days following the amputation. (J) Top view of the caudal fin shown in I. (K-V) Caudal fins of 2.2shh:gfp:ABC#15 transgenic fish were amputated near the bifurcation and analyzed at different time-points after amputation by a double immunostaining with anti-GFP (green) (K-O) and anti-Zns5 (red) (P-T) antibodies. (U,V) merge of 1 dpa + 18 hpa (U) and 2 dpa (V). Dashed line represents amputation plane. hpa: hours-post-amputation; dpa: days-post-amputation. Between the first and the second amputation, fins were allowed to regenerate for 2 weeks.

Fgf signalling does not play a role in the determination of the proximal-distal level where the bifurcation will form. (A) Heat-shock protocol following an amputation. Transgenic hsp70:dn-fgfr1 fish were amputated at 1 segment proximal to the bifurcation (near the bifurcation) and heat-shocked at 35°C for 1 hour, during 6 days, starting at day 2 post amputation. (B) Picture of a 7 days-post-amputation regenerated caudal fin of hsp70:dn-fgfr1 transgenic fish with or without the heat-shock protocol. (C) Number of segments formed in the 3rd dorsal ray between the base of the fin and the bifurcation after one amputation near the bifurcation in heat-shocked and non-heat-shocked siblings. Dashed line represents amputation plane.

Acknowledgments
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