Forskolin

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.

All chemical structures used in this study are depicted in Figure 1. Fentanyl citrate, morphine sulphate and DAMGO ([d‐Ala²,NMe‐Phe⁴,Gly‐ol⁵]‐enkephalin) were purchased from Sigma‐Aldrich, UK. Luciferin was purchased from Promega, UK. AP‐5 and forskolin was purchased from HelloBio, UK. The syntheses of MT‐45, 2F‐MT‐45, 3F‐MT‐45 and 4F‐MT‐45 have been described previously (McKenzie et al., 2018). 1‐Cyclohexylpiperazine (M17) was purchased from Fluorochem, UK. All other chemicals were from Sigma‐Aldrich, UK.

Drugs were bath applied. MK-801, forskolin, CGP55845, and picrotoxin were acquired from Hello Bio Princeton, NJ. CNQX, sulpiride, L-DOPA, were acquired from Sigma-Aldrich. Amphetamine was acquired from NIH NIDA and sumatriptan was acquired from Glaxo Wellcome Inc.