Reduced Expression of the Vesicular Acetylcholine Transporter and Neurotransmitter Content Affects Synaptic Vesicle Distribution and Shape in Mouse Neuromuscular Junction

Abstract

In vertebrates, nerve muscle communication is mediated by the release of the neurotransmitter acetylcholine packed inside synaptic vesicles by a specific vesicular acetylcholine transporter (VAChT). Here we used a mouse model (VAChT KDHOM) with 70% reduction in the expression of VAChT to investigate the morphological and functional consequences of a decreased acetylcholine uptake and release in neuromuscular synapses. Upon hypertonic stimulation, VAChT KDHOM mice presented a reduction in the amplitude and frequency of miniature endplate potentials, FM 1–43 staining intensity, total number of synaptic vesicles and altered distribution of vesicles within the synaptic terminal. In contrast, under electrical stimulation or no stimulation, VAChT KDHOM neuromuscular junctions did not differ from WT on total number of vesicles but showed altered distribution. Additionally, motor nerve terminals in VAChT KDHOM exhibited small and flattened synaptic vesicles similar to that observed in WT mice treated with vesamicol that blocks acetylcholine uptake. Based on these results, we propose that decreased VAChT levels affect synaptic vesicle biogenesis and distribution whereas a lower ACh content affects vesicles shape.

Muscle atrophy is associated with cervical spinal motoneuron loss in BACHD mouse model for Huntington's disease

“…For ultrastructure analyses, we used the protocol previously described by our research group (Rodrigues et al, 2013). Briefly, mice were anesthetized with ketamine/xylazine (0.1 mL/20 g), left ventricular transcardiac perfused with ice-cold modified Karnovsky fixative (4% paraformaldehyde and 2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer at 4°C) and maintained in this solution for at least 24 h at 4°C.…”

Drosophila melanogaster as a genetic model system to study neurotransmitter transporters

“…In particular, electrophysiological analysis of adult dVAChT heterozygotes suggest that during periods of sustained vesicle release, at least one circuit in the adult CNS fails to maintain normal levels of ACh release (Kitamoto et al, 2000). This phenomenon may be related to changes in ACh release observed during periods of sustained release at the cholinergic NMJ of frogs (Naves and Van der Kloot, 1996; Van der Kloot, 2003) and in VAChT knock-down mice (Lima et al, 2010; Rodrigues et al, 2013). The mechanisms underlying these changes may include the presence of variable numbers of VAChT molecules per SV and a consequent reduction in the amount of transmitter stored in each vesicle (Prado et al, 2013).…”

Variants in SLC18A3 , vesicular acetylcholine transporter, cause congenital myasthenic syndrome

“…12 Two mouse models of reduced SLC18A3 expression, a knockout mouse and a knockdown mouse, have previously been described and provide strong support for SLC18A3 as a genetic cause of presynaptic congenital myasthenic syndrome. [13][14][15][16] Homozygous SLC18A3 (VAChT , with deletion of a regulatory element within the 59 untranslated region, had a 65% to 70% reduction in VAChT expression. Motor performance and grip strength were impaired, and mice were not able to maintain long periods of physical activity.…”