1. A high-affinity, partial antagonist effect of 3,4-diaminopyridine mediates action potential broadening and enhancement of transmitter release at NMJs.
- Author
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Ojala KS, Ginebaugh SP, Wu M, Miller EW, Ortiz G, Covarrubias M, and Meriney SD
- Subjects
- Acetylcholine metabolism, Action Potentials drug effects, Action Potentials physiology, Animals, Calcium Channels, L-Type genetics, Calcium Channels, L-Type metabolism, Calcium Channels, N-Type genetics, Calcium Channels, N-Type metabolism, Dose-Response Relationship, Drug, Female, Gene Expression, Male, Mice, Microelectrodes, Neuromuscular Junction metabolism, Presynaptic Terminals metabolism, Rana pipiens, Shaw Potassium Channels antagonists & inhibitors, Shaw Potassium Channels genetics, Tissue Culture Techniques, Amifampridine pharmacology, Neuromuscular Agents pharmacology, Neuromuscular Junction drug effects, Potassium Channel Blockers pharmacology, Presynaptic Terminals drug effects, Shaw Potassium Channels metabolism
- Abstract
3,4-Diaminopyridine (3,4-DAP) increases transmitter release from neuromuscular junctions (NMJs), and low doses of 3,4-DAP (estimated to reach ∼1 μM in serum) are the Food and Drug Administration (FDA)-approved treatment for neuromuscular weakness caused by Lambert-Eaton myasthenic syndrome. Canonically, 3,4-DAP is thought to block voltage-gated potassium (Kv) channels, resulting in prolongation of the presynaptic action potential (AP). However, recent reports have shown that low millimolar concentrations of 3,4-DAP have an off-target agonist effect on the Cav1 subtype ("L-type") of voltage-gated calcium (Cav) channels and have speculated that this agonist effect might contribute to 3,4-DAP effects on transmitter release at the NMJ. To address 3,4-DAP's mechanism(s) of action, we first used the patch-clamp electrophysiology to characterize the concentration-dependent block of 3,4-DAP on the predominant presynaptic Kv channel subtypes found at the mammalian NMJ (Kv3.3 and Kv3.4). We identified a previously unreported high-affinity (1-10 μM) partial antagonist effect of 3,4-DAP in addition to the well-known low-affinity (0.1-1 mM) antagonist activity. We also showed that 1.5-μM DAP had no effects on Cav1.2 or Cav2.1 current. Next, we used voltage imaging to show that 1.5- or 100-μM 3,4-DAP broadened the AP waveform in a dose-dependent manner, independent of Cav1 calcium channels. Finally, we demonstrated that 1.5- or 100-μM 3,4-DAP augmented transmitter release in a dose-dependent manner and this effect was also independent of Cav1 channels. From these results, we conclude that low micromolar concentrations of 3,4-DAP act solely on Kv channels to mediate AP broadening and enhance transmitter release at the NMJ., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2021
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