Fig. 3

Structural changes in the choanoderm during dissociation. A- C’’ Control conditions in natural sea water (1 h NSW): (A) Schematic view of a choanocyte chamber organized in a round shape with choanocyte surrounded by an apical microvilli (Mi) rich in actin and a flagellum (Fl) composed by α-tubulin. PhaE: PhaE lectin staining, Nu: Nucleus (A’) Confocal microscopic view (LM) of a choanocyte chamber, the classical round shape of the choanocyte chamber is visible (dotted orange line). The choanocyte (orange line) is recognizable with its typical collar of apical microvilli (blue arrow). The type IV collagen staining (in magenta) is lining the basal pole of choanocyte chambers (pink arrow). B LM focused view on the actin-rich cellular junction between choanocytes (yellow arrow). C Scanning Electron Microscopic (SEM) view of a choanocyte chamber with collar of apical microvilli (cyan arrow), and a basement membrane lining the choanocyte chamber (pink arrow). C’ Focus on the cell–cell junction (yellow arrow), basal lamina (pink arrow) and apical microvilli (cyan arrow). C’’ Choanocyte with its complete apical microvilli collar (cyan arrow) and complete basal lamina (pink arrow). D–F’’’ Choanocyte chambers observed one hour after incubation in calcium-magnesium-free sea water (1hD CMSFW): D The general architecture of the choanocyte chamber is still observable in LM (dotted orange line), immunostaining of type IV collagen is not visible anymore (dotted pink arrow), apical microvilli are disintegrated (dotted cyan arrow). E Focus (LM) on a choanocyte with its disintegrated apical microvilli (dotted cyan arrow) and numerous basal actin protrusions (red arrow). F SEM view of a choanocyte chamber. Choanocytes present vacuoles (black arrow), disintegrated apical microvilli (dotted cyan arrow) and protrusions (red arrow). The basal lamina is detached from the choanocytes (dotted pink arrow). F’ Focus (SEM) on the disintegration of the apical microvilli (cyan dotted arrow) and on the loss of cell–cell contact (dotted yellow arrow). F’’ Focus on basal actin protrusions at the basal pole of a choanocyte (SEM) (red arrow). F’’’ Focus on two choanocytes with a clear detachment from the basal lamina (dotted pink arrow). G–I’’ Choanocyte chambers observed four hours after incubation in calcium-magnesium-free sea water (4hD CMFSW): G LM view of a destructured choanocyte chamber (dotted orange line), immunostaining of type IV collagen is not visible and the general architecture of the choanocyte chamber is no more recognizable (yellow dotted line). H (LM) Focus on a choanocyte with numerous actin basal protrusions (red arrow). I SEM view of choanocyte showing actin protrusions (red arrow) and the whole destructuring of the basal lamina (dotted pink arrow). I’ (SEM) focus on the actin protrusions (red arrow). I’’ The basal lamina is completely lost at this time-point (SEM) (dotted pink arrow). For all confocal (LM) pictures: Staining in grey: Phalloidin, Green: PhaE lectin, Blue: DAPI, Magenta: type IV collagen. Additional picture of control condition in NSW is available in the Additional file 2: Figure S2B