Figure 1.

Soluble neuroligin 1 (NL1) ectodomain reduces cyclic adenosine monophosphate (cAMP) levels via activation of metabotropic glutamate receptor 2 (mGluR2). (A) Soluble NL1 reduces cAMP levels in neurons. Hippocampal neurons were grown for 7 days and subsequently treated with 8 μM forskolin and different concentrations of recombinant ecto-NL1 for 20 min. Glutamate was used as a positive control. Statistical significance was determined using Kruskal-Wallis test followed by the Dunn’s multiple comparison test *p = 0.0344, **p = 0.01; n = 3–7. (B) NL1 reduces cAMP levels via activation of mGluR2. Hippocampal neurons were grown for 7 days and subsequently treated for 20 min with 8 μM forskolin only (black bars) or forskolin and recombinant ecto-NL1 (gray bars) in the presence of non-competitive mGluR2 inhibitor, RO 64-5229 in indicated concentrations. Statistical significance was determined using Friedman’s test followed by the Dunn’s multiple comparison test *p = 0.0316, n = 4. (C) Soluble NL1 activates mGluR2 and decreases cAMP levels. HEK-293 cells were transfected with mGluR2-green fluorescent protein (GFP; black bars) or empty vector (gray bars) and then treated with 8 μM forskolin and recombinant ecto-NL1 for 20 min. Statistical significance was determined using Kruskal-Wallis test followed by the Dunn’s multiple comparison test *p = 0.0049; n = 5. (D) HEK-293 cells were double-transfected with mGluR2-GFP and NL1B-HA, and then treated with 8 μM forskolin only. Statistical significance was determined using Mann-Whitney test *p = 0.0286; n = 4. The results are expressed as a percentage ± SEM relative to the untreated control, which was set at 100%.