Biphasic calcium response is required for wound healing. (A) Time-lapse images show wave propagation in response to injury in control, calcium-free and 1 μM thapsigargin-treated Pax6
+/+ monolayer cultures. Removal of Ca2+ from extracellular medium prevented initial rise of [Ca2+]i in cells adjacent to wound but did not prevent propagation of a significantly less intense wave. Thapsigargin did not prevent rise of [Ca2+]i in cells immediately adjacent to wound edge but inhibited wave propagation (see Additional files 1, 2, 3). (B) Changes in fluorescence intensity in Pax6
+/+ cultures in Ca2+-free HBSS in time-lapse images. (C) Thapsigargin-treated cultures showed no significant difference in wave initiation (row 2, 6 seconds) compared with Pax6
+/+, but significantly less propagation (row 6) (*p = 0.014). (D) Rate of wound healing in Pax6
+/+ and Pax6
+/- cultures. Depletion of intracellular Ca2+ stores with thapsigargin (T) or incubation in calcium-free medium almost completely inhibited wound healing compared with untreated Pax6
+/+ controls (*, p < 0.001, n = 6). Scale bars = 50 μm.