Tygon

To perform Aiptasia imaging experiments, Aiptasia larvae were loaded directly from culture wells (∼100 μL) using 1/16″ tubing (Tygon, Fisher) attached to a 1 mL syringe (Plastipak, BD) with a 23 G luer-lock connector (McMaster-Carr). Lines were hung from the syringe pump vertically for 2 minutes to concentrate the motile larvae and connected upstream of the inlet stopcock. A step-by-step loading and operation protocol relative to each trial is provided in Supplemental Information. In trials with DCMU treatment, seawater containing 25 μM DCMU (Diuron, Sigma) was introduced instead of normal seawater after larval loading.

An individual slice was transferred to a recording chamber where it was suspended on nylon mesh grid, completely submerged in standard aCSF and perfused at 6 ml/min, to allow all round perfusion and thus reducing the risk of hypoxia. All perfusion solutions were bubbled with 95% O₂ and 5% CO₂ and maintained at around 32°C. All tubing used had low gas permeability (Tygon; Fisher Scientific, Loughborough, United Kingdom). In order to record synaptic responses, a concentric tungsten bipolar metal stimulating electrode was placed in stratum radiatum in the CA1 region to stimulate afferent Schaffer collaterals/commissural fibers, at 15 s intervals (100 μs pulses). A single aCSF-filled glass microelectrode was positioned in the CA1 dendritic region to record evoked field excitatory postsynaptic potentials (fEPSPs; filtered at1 Hz-3 kHz). WinLTP® software was used to control stimulus parameters and acquisition, as well as to record and analyze the slope of fEPSPs (Anderson et al., 2012 (link)).