Figure 3. Isoflurane reduces F-actin concentration in spines, and actin targeted drugs prevent isoflurane-induced spine loss.

Hippocampal neuron cultures transfected with eGFP were exposed to 95% air/5% CO₂ (Ctl) or 2 vol% isoflurane in 95% air/5% CO₂ at 37°C for 20 min and stained with phalloidin-Alexa 568 (A-representative images). Quantification of phalloidin fluorescence intensity shows that isoflurane reduced F-actin concentration (B) by Student unpaired t-test (****p<0.0001). Pretreatment with 0.1 µM jasplakinolide (C-representative images) or 0.1 µM cytochalasin D (E-representative images) for 2 min before exposure to 95% air/5% CO₂ (Ctl) or 2 vol% isoflurane in 95% air/5% CO₂ at 37°C for 20 min prevented isoflurane-induced spine loss (D,F) by one-way ANOVA with Tukey's post hoc test (****p<0.0001). Data are mean ± SEM; n = 30 to 40 dendritic ROIs (600–1260 spines) per experimental group for phalloidin staining and actin stabilizing drug treatments. Scale bar  = 1 µm for 3A and 5 µm for 3C,E.