ambenonium dichloride  (Tocris)

Journal: NPJ Parkinson's Disease

Article Title: Postsynaptic adaptations in direct pathway muscarinic M4-receptor signaling follow the temporal and regional pattern of dopaminergic degeneration

doi: 10.1038/s41531-025-01047-3

Figure Lengend Snippet: A Schematics of AAV9.hSyn.ACh3.0 injection into DSt of WT mice and cartoon schematic showing the principle of GRAB ACh3.0 sensor function and the bath application of ACh with ambenonium. B Time courses of GRAB ACh3.0 fluorescence change (mean frame) following bath application of 100 µM ACh and 100 nM ambenonium for DLS and DMS (left). Summary data for maximal ΔF/F (center). Representative images and surface plots showing the maximal changes in GRAB ACh3.0 fluorescence across striatal regions (right) (DLS n = 10, N = 5; DMS n = 7, N = 4; unpaired t test p = 0.5967). C Schematics of AAV9.hSyn.ACh3.0 and saline or 6-OHDA (LD or HD) injections into DSt and MFB in WT mice (top). Cartoon depicting how ACh released from ChIs is detected by GRAB ACh3.0 expressed in striatal cells (bottom). D Representative images and surface plots of GRAB ACh3.0 fluorescence changes (mean frame) in DLS (top) and DMS (bottom) following application of a single electrical stimulus (25 µA, 0.5 ms) in the center of the striatal region of interest (left). Quantification of ΔF/F is shown on the bar charts (right) (DLS: Saline n = 32, N = 9; 6-OHDA LD n = 27, N = 6; 6-OHDA HD n = 38, N = 9; Kruskal-Wallis p = 0.2419 / DMS: Saline n = 38, N = 9; 6-OHDA LD n = 28, N = 6; 6-OHDA HD n = 42, N = 9; Kruskal-Wallis p = 0.0019; Dunn’s post hoc test). E Representative 2P-point scanning traces and quantification of fluorescence changes of GRAB ACh3.0 (DLS: Saline n = 28, N = 6; 6-OHDA LD n = 15, N = 4; 6-OHDA HD n = 34, N = 5; Kruskal-Wallis p = 0.1903 / DMS: Saline n = 31, N = 7; 6-OHDA LD n = 15, N = 4; 6-OHDA HD n = 27, N = 5; Kruskal-Wallis p = 0.467). F Representative 2P-point scanning traces and summary data of GRAB ACh3.0 fluorescence changes following paired-pulse stimulation (P1 + P2), with the digitally subtracted P2 component shown on gray (DLS: Saline n = 26, N = 6; 6-OHDA LD n = 15, N = 4; 6-OHDA HD n = 30, N = 5; Kruskal-Wallis p = 0.2159 / DMS: Saline n = 31, N = 7; 6-OHDA LD n = 15, N = 4; 6-OHDA HD n = 28, N = 5; Kruskal-Wallis p = 0.5913). Summary data is mean ± SEM. Extended statistical data is provided in Supplementary Table . n: number of cells, N: number of mice; n.s. p > 0.05; * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: Picrotoxin, MK-801, DNQX, DHβE, SCH 23390, sulpiride, acetylcholine, oxotremorine M, and ambenonium were purchased from Tocris Bioscience.

Techniques: Injection, Fluorescence, Saline

Journal: NPJ Parkinson's Disease

Article Title: Postsynaptic adaptations in direct pathway muscarinic M4-receptor signaling follow the temporal and regional pattern of dopaminergic degeneration

doi: 10.1038/s41531-025-01047-3

Figure Lengend Snippet: A Schematics of Cre-dependent GIRK2-encoding virus and saline or 6-OHDA (LD or HD) injections into the DSt and MFB from D1-Cre mice respectively (left). Cartoon of ChI-dMSN synapse depicting the bath application of ambenonium to block ACh hydrolysis by AChE (right). B Representative traces of electrically evoked M4-IPSCs (left) before (Basal) and after ( + Amb) bath application of ambenonium (10 nM) in DLS (top) and DMS (bottom). Quantification of net charge as absolute values (center) and normalized to control (right) (DLS: Saline n = 16, N = 4; 6-OHDA LD n = 20, N = 8; 6-OHDA HD n = 15, N = 5; Wilcoxon tests p < 0.0001 for absolute values and Kruskal-Wallis p = 0.4736 for ratio / DMS: Saline n = 10, N = 6; 6-OHDA LD n = 8, N = 6; 6-OHDA HD n = 13, N = 8; Wilcoxon test p = 0.002 for saline, paired t test p = 0.0082 for 6-OHDA LD and paired t test p = 0.0003 for 6-OHDA HD in absolute values and One-Way ANOVA p = 0.1687 for ratio). Summary data is mean ± SEM. Extended statistical data is provided in Supplementary Table . n: number of cells, N: number of mice; n.s. p > 0.05; * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: Picrotoxin, MK-801, DNQX, DHβE, SCH 23390, sulpiride, acetylcholine, oxotremorine M, and ambenonium were purchased from Tocris Bioscience.

Techniques: Virus, Saline, Blocking Assay, Control