Synaptic transmission at the vertebrate neuromuscular junction (NMJ) occurs when acetylcholine (ACh) secreted by motor nerve terminals activate ACh receptors (AChRs) concentrated in the postsynaptic membrane of target muscle cells. AChRs are ligand-gated cation channels whose opening depolarizes the synaptic region of muscle to trigger muscle contraction. For my thesis research I studied the regulation of AChR activity by extrinsic and intrinsic signals in cultured Xenopus embryonic muscle cells using patch-clamp recordings and pharmacological and molecular methods. The results obtained have yielded three distinct and novel insights related to the synaptic function of AChRs....[ Read more ]

Synaptic transmission at the vertebrate neuromuscular junction (NMJ) occurs when acetylcholine (ACh) secreted by motor nerve terminals activate ACh receptors (AChRs) concentrated in the postsynaptic membrane of target muscle cells. AChRs are ligand-gated cation channels whose opening depolarizes the synaptic region of muscle to trigger muscle contraction. For my thesis research I studied the regulation of AChR activity by extrinsic and intrinsic signals in cultured Xenopus embryonic muscle cells using patch-clamp recordings and pharmacological and molecular methods. The results obtained have yielded three distinct and novel insights related to the synaptic function of AChRs.

First, elevation of extracellular proton concentrations was found to inhibit AChR activity by decreasing the apparent affinity of AChRs for ACh. This pH-effect was likely specific to the extracellular region of the receptor, was independent of membrane potential, and was not affected by the ion-permeation path of AChRs. Second, stretch-induced opening of a mechanosensitive channel (MSC) was detected in cultured Xenopus muscles. Increasing negative pressure enhanced the activity of the MSCs, which had conductance properties and gating kinetics comparable to that of AChRs and which were inhibited by AChR channel blockers, such as CPZ, d-tc and α-BTX. These findings support the hypothesis that the ligand-gated AChR channels are also mechanosensitive. Third, AChR channel opening was found to be enhanced by protein tyrosine kinase activity and suppressed by tyrosine phosphatase activity, and the use of specific chemical inhibitors further identified Src and Shp2 as candidate tyrosine kinase and phosphatase, respectively, involved in regulating AChR activity.

The findings of my thesis research, taken together, favor the view that multiple extra-and intra-cellular stimuli together modulate AChR channel function to tightly control synaptic transmission at the vertebrate NMJ