ZD7288 (ICI D7288), a selective hyperpolarization-activated cyclic nucleotide-gated channel blocker, inhibits hippocampal synaptic plasticity.

Cultured hippocampal neurons were incubated with 1 μM Fura-2 acetoxy-methylol ester for 30 minutes at 37°C, washed three times with artificial cerebrospinal fluid (containing 140 mM NaCl, 5 mM KCl, 2 mM CaCl2, 1 mM MgCl2, 10 mM glucose and 10 mM hydroxyethyl piperazine ethanesulfonic acid, pH 7.3), and then incubated at room temperature in the dark for 30 minutes.?Fura-2 fluorescence was observed by a Ratio Vision digital fluorescence microscopy system.?Fluorescence signals were evoked by 340 and 380 nm excitation wavelengths and collected at 510 nm by TILLvisION 4.0 software.?The 340:380 nm fluorescence ratio was used to represent [Ca2+]i.?Peak calcium change was represented as the percentage increase from baseline.?Neurons were incubated in ZD7288 (25, 50 or 100 μM) or 8-Br-cAMP (5 or 50 μM) for 15 minutes prior to stimulation with 50 μM glutamate.?All experiments were repeated in triplicate, using different batches of cells across 4–5 dishes[1].