Figure 3. Opioid receptor (OR) signaling collectively constrains DG-CA3 proximal synapses.
(A) Cultured hippocampal neurons (>21 DIV) incubated for 24 hours under normal activity conditions (NT), hyperactivity (PTX) or inactivity (TTX) in the absence (top) or presence (bottom) of the broad-spectrum OR antagonist, naloxone. DG axon terminals and excitatory synapses were visualized with SPO (blue) and PSD-95 (red), respectively. (B) Representative proximal (<20 μm from soma) and distal (>80 μm) CA3 dendrites immunostained with SPO (blue) and PSD-95 (red). (C) Quantification of PSD-95 immunoreactivity in proximal (<20 μm from soma) and distal (>80 μm) CA3 dendrites. **P<0.01, ***P <0.001, one-way ANOVA and post hoc Dunnett’s comparison vs. NT. NS (P>0.05) vs. conditions indicated. (D) Quantification of SPO accumulation in presynaptic DG terminals opposite proximal (<20 μm from soma) and distal (>80 μm) CA3 dendrites. **P<0.01, ***P <0.001, ++P<0.01, one-way ANOVA and post hoc Bonferroni multiple comparison test. Data are expressed as mean±SEM from neuronal averages (n=10–20 neurons/group, 3 dendrites/neuron). Scale bar, 20 μm.