Platynereis larval electrophysiology
Performing electrophysiological recordings on Platynereis larvae is a challenging task. It requires a dedicated electrophysiology setup with micromanipulators that we do not describe here. We just provide the specifics of the Platynereis experiments that were published in Conzelmann M. et al. Neuropeptides regulate swimming depth of Platynereis larvae. PNAS, 2011.
- For the recordings we used 29 to 33 hpf larvae. Larvae older than 48 hpf (reared at 18°C) start to show frequent muscle contractions that make the recordings very difficult.
- Larvae can be immobilized with a holding pipette with an opening of ∼30 μm. Larvae can be caught from the seawater bath at the recording apparatus, by applying gentle suction to the holding capillary (see movie).
- For recordings we use borosilicate capillaries (Science Products GB150F-8P) pulled with a Sutter P-1000 Flaming/Brown micropipette puller (Heka Elektronik) to 2–4 MΩ resistance.
- Platynereis larvae have a strong cuticle that can be softened by adding 0.002% trifluoromethanesulfonic acid to the seawater 10 min before the experiments. We found that this greatly facilitate penetration across the cuticle.
- For recordings we use 70 mM CsCl, 10 mM Hepes, 11 mM glucose, 10 mM glutathione, 5 mM EGTA, 500 mM aspartic acid, 5 mM ATP, and 0.1 mM GTP, pH 7.3 (set with CsOH).
- To record signals from the ciliary band, it is sufficient to place the recording pipette in close proximity to the ciliated cells, without penetrating the cuticle.
This video shows the catching of a Platynereis larva with
a holding capillary.
This video shows the penetration of the cuticle with a capillary.