Acetylcholine-Induced Inhibition of Presynaptic Calcium Signals and Transmitter Release in the Frog Neuromuscular Junction

Abstract

Acetylcholine (ACh), released from axonal terminals of motor neurons in neuromuscular junctions regulates the efficacy of neurotransmission through activation of presynaptic nicotinic and muscarinic autoreceptors. Receptor-mediated presynaptic regulation could reflect either direct action on exocytotic machinery or modulation of Ca2+ entry and resulting intra-terminal Ca2+ dynamics. We have measured free intra-terminal cytosolic Ca2+ ([Ca2+]i) using Oregon-Green 488 microfluorimetry, in parallel with voltage-clamp recordings of spontaneous (mEPC) and evoked (EPC) postsynaptic currents in post-junctional skeletal muscle fiber. Activation of presynaptic muscarinic and nicotinic receptors with exogenous acetylcholine and its non-hydrolized analog carbachol reduced amplitude of the intra-terminal [Ca2+]i transients and decreased quantal content (calculated by dividing the area under EPC curve by the area under mEPC curve). Pharmacological analysis revealed the role of muscarinic receptors of M2 subtype as well as d-tubocurarine-sensitive nicotinic receptor in presynaptic modulation of [Ca2+]i transients. Modulation of synaptic transmission efficacy by ACh receptors was completely eliminated by pharmacological inhibition of N-type Ca2+ channels. We conclude that ACh receptor-mediated reduction of Ca2+ entry into the nerve terminal through N-type Ca2+ channels represents one of possible mechanism of presynaptic modulation in frog neuromuscular junction.

Activation of Neuronal Nicotinic Receptors Inhibits Acetylcholine Release in the Neuromuscular Junction by Increasing Ca2+ Flux through Cav1 Channels

“…Thus, nicotine at a concentration of 10 µM exerts both postsynaptic and presynaptic inhibitory effects on the neuromuscular synapse, causing a decrease in the number of ACh quanta released in response to action potential. A similar decrease in the QC during the activation of cholinergic receptors has been noted earlier [17,20,21,32], however, these results were obtained on other preparations, in conditions of initially reduced QC, or in cut fiber preparations.…”

“…In contrast, activation of the M2-subtype caused inhibition of the ACh quanta release [14,15]. Both the M1-and M2-mediated mechanisms depend on calcium influx [14,[16][17][18].…”

Profiling Basal Forebrain Cholinergic Neurons Reveals a Molecular Basis for Vulnerability Within the Ts65Dn Model of Down Syndrome and Alzheimer’s Disease

“…Notably, Chrm2, encoding the muscarinic M2 cholinergic receptor and primarily localized to ChAT-positive cholinergic neurons (98,99), was downregulated along with downstream effectors Gnb5, which leads to downregulation of Mapk3, causing dampening of synaptic plasticity (100). In addition, presynaptic choline transferase is downregulated, along with Chrm2 and Gnb5 which block choline uptake and calcium to the presynaptic terminal (101). Conversely, Plcb2 and Prkcg, downstream of the muscarinic M1 cholinergic receptor, are upregulated, likely driving the increases seen in the calcium signaling pathway (102,103).…”

Amplification of Snake Venom Toxicity by Endogenous Signaling Pathways

“…Acute desensitization was one of the first autoregulatory aspects of nicotinic transmission at the neuromuscular junction described by Sir Bernard Katz in seminal studies of neuromuscular junction function [109]. The details of this have been revealed in the frog neuromuscular junction to involve the action of calcium-dependent negative feedback inhibition of neurotransmission mediated by pre-synaptic nicotinic and muscarinic autoreceptors [110]. At the same time that desensitization of post-junctional nicotinic receptors is occurring, depletion of synaptic vesicles develops, such that nerve depolarization does not accomplish neurotransmission.…”