Investigating Synaptic Depression via O-GlcNAc and PKA

Forskolin (Hello Bio) and 1,9 DideoxyForskolin (Sigma-Aldrich) were prepared as a 50 mM stock in DMSO and stock was added to external solution for a final concentration of 50 μM. KT5720 (Hello Bio) was prepared as a 25 mM stock in DMSO and stock was added to external solution for a final concentration of 3 μM. SQ22536 (Tocris), 5α,21-pregnane-3α,21-diol-20-one (THDOC) (Sigma-Aldrich, mixed and sonicated), Progesterone (Sigma-Aldrich) was prepared as a 100 mM stock in DMSO and the stock was added to external solution for a final concentration of 100 μM, 10 μM, and 1 μM respectively. Jasplakinolide (Hello Bio) was prepared as a 1 mM stock in DMSO and stock was added to Cs-gluconate internal solution for a final concentration of 2 μM. Dynasore (Sigma-Aldrich) was prepared as a 100 mM stock in DMSO and stock was added to external solution for a final concentration of 80 μM. The actin stabilizer Jasplakinolide (2μM, Hello Bio)^{57 (link),58 (link)} was included in the pipet solution to prevent GABA_AR endocytosis. PKA dependent serine phosphorylation was triggered by bath application of the adenylate cyclase activator, Forskolin (50μM, Hello Bio)^{31 (link)} determine if the O-GlcNAc induced synaptic depression is prevented.