Journal: Frontiers in Cellular Neuroscience

Article Title: Acetylcholine Receptor Stimulation Activates Protein Kinase C Mediated Internalization of the Dopamine Transporter

doi: 10.3389/fncel.2021.662216

Figure Lengend Snippet: DAT-mediated internalization by carbachol is dependent on both dynamin and clathrin. (A) In primary midbrain cultures, dynole (50 μM), an inhibitor of dynamin, prevented DAT internalization in response to both AMPH (10 μM) and carbachol (30 μM). These data further support the internalization of the transporter as the mechanism of loss of DAT function ( n = 3, * P ≤ 0.05 and ** P ≤ 0.01 by two-way ANOVA). (B) Pitstop II (30 μM), a clathrin inhibitor, prevents DAT internalization in response to carbachol. There was no affect effect of this clathrin inhibition on AMPH-mediated DAT internalization, consistent with prior studies demonstrating this as a clathrin-independent process ( n = 3, *** P ≤ 0.001 by two-way ANOVA). (C) TAT-fused peptides that were designed to interfere with GPCRs coupled to G Q , G S , or G 11 alpha subunits were applied to dopamine neurons in primary culture (70 μM). Subsequent treatment of these cultures with carbachol led to the loss of DAT-mediated DA transport under all conditions, except the TAT-G Q pre-treated cultures ( n = 5, * P ≤ 0.05 and ** P ≤ 0.01 by one-way ANOVA).

Article Snippet: Custom-designed TAT-peptides were ordered from LifeTein.

Techniques: Inhibition