5HT-Hi promotes growth cone repulsion through the 5-HT1b receptor. a Growth cone repulsion was measured in the presence of 5HT-Hi gradients (p = <0.0001) and CP-94253 gradients (p = 0.0018). Growth cone repulsion to 5HT-Hi was altered to levels indistinguishable from random growth (vehicle) with application of the selective antagonist to 5-HT1b, GR55562 (n = 16, p = <0.0001), by activating adenylate cyclase with forskolin (n = 8, p = 0.0003) and restoration of cAMP signals with Sp-cAMPs (n = 8, p = 0.0001). b Repulsion to 5HT-Hi was not perturbed when growth cones were treated with the 5-HT2a receptor antagonist ritanserin (n = 8, p = 0.002) or the 5-HT3 antagonist ondansetron (n = 10, p = 0.0003). Inhibition of ER calcium release with thapsigargin (n = 10, p = <0.0001) or calcium influx with nifedipine (n = 8, p = <0.0001) did not alter growth cone repulsion from 5HT-Hi. Turning angles were compared to vehicle and 5HT-Hi. c, d Axon extension was not perturbed by any pharmacological application (ns, p > 0.05). Turning angles were compared to vehicle and 5HT-Hi. (Kruskal–Wallis, Dunn’s multiple comparison test)