Bryozoan and brachiopod development

Shipping live colonies


Colonies of Membranipora membranacea needed to be shipped from Bergen, Norway to Dresden, Germany. The first package contained around 10 pieces of kelp blades with colonies inside single transparent plastic bag filled with sea water (bubbled overnight) and sealed with tape. Bag was surrounded by icepacks to keep temperatures low. Upon arrival we realized that the plastic bag had ruptured during the trip and the colonies were out of the water for an unknown period of time. They were placed in a 50 L plastic tank in the cold room (4 °C) immediately. After 1h I checked under the scope and all the colonies were dead.



Our colleagues in Bergen prepared an emergency second shipment, this time with colonies placed inside three plastic bags with sea water (not bubbled) and less air to avoid water movement that could rip the plastic. Upon arrrival two of the inner plastic bags were broken, but colonies were still underwater. They were split in two groups to be kept at 4 °C and 12 °C and were acclimatized in 1:1 natural:artificial sea water for 1h. Then they were transferred to artificial sea water. Under the scope zooids were still moving. On the following morning, however, most of the colonies were dead or just-dead, but still with some normal embryos which I extracted as described below.



Acknowledgements: Annie Boddington and Aina Børve for collection and shipping from Bergen. Yu-Wen Hsieh and Daniele Soroldoni for preparing cultures and receiving the bryozoans in Dresden.

Sample preparation


I collected fertilized eggs by scrapping the surface of ripe zooids. Eggs were activated with 0.1 mM EDTA at 12 °C for 2 hours. The few embryos that started cleaving were transferred to a 4-well plate with 5µg/mL FM 4-64 solution in sea water. After 15 min I put the embryos in a round coverslip coated with poly-l-lysine and mounted on the sample holder of the Multi-modal scanned lightsheet SPIM (Meyers SPIM). I chose this scope because it has temperature control in the chamber (15 °C) and is adapted to small samples.

Imaging
Unfortunately, we only managed to acquire a small time-lapse from an embryo that was not healthy.






 * FIG: Gaussian vs si
 * MOV: stack gaussian vs si
 * MOV: timelapse

Multi-modal scanned lightsheet SPIM (Myers SPIM)



 * MOV: 3D model
 * FIG: larva 2 stainings


 * Zeiss
 * FIG: 7 angles + fused
 * MOV: fused 3d projection

Fixed brachiopod larvae

 * OpenSPIM
 * FIG: larva on openspim (L and T)
 * Zeiss
 * FIG: 7 angles + fused
 * MOV: fused 3d projection
 * FIG: tutorial on contentbased registration