Figure 4.

Nicotine-induced increases in the number of functional glutamatergic synapses in cell culture. Rat hippocampal cultures were incubated 7 d (7d) in nicotine (Nic) with or without blockers (MLA, DHβE), rinsed to remove nicotine, and then incubated 1 h in MLA/DHβE. As a control, some cultures received nicotine only during the last hour of the 7 d incubation (1 h Nic). A, Patch-clamp recording of spontaneous mEPSCs in TTX and gabazine. B, Quantification of mEPSC frequency (15–20 cells from 5–8 cultures per condition). C, Images of cells in culture having received the indicated treatment, then incubated with MLA/DHβE for 1 h followed by labeling with a 1 min incubation in FM4–64 (red) plus 50 mm KCl to reveal active synapses on dendrites and spines of cells expressing GFP. Scale bar, 10 μm. D, Quantification of FM4–64-labeled puncta. Baseline control (7d Nic/noK): KCl omitted from FM4–64 incubation (n = 3–6 cultures; 5–7 cells/culture).