Abstract
The aim of this study was to compare the acute effects of thrombin and brain-derived neurotrophic factor (BDNF) on spontaneous miniature endplate potentials (MEPPs) and multiquantal evoked endplate potentials (EPPs) in mouse neuromuscular junctions (NMJs) of m. diaphragma and m. EDL. Intracellular microelectrode recordings of MEPPs and EPPs were used to evaluate the changes in acetylcholine (ACh) release in mature and newly-formed mouse NMJs. Thrombin (1 nM) increased the amplitude of MEPPs and EPPs by 25–30% in mature and newly-formed NMJs. This effect was due to an enhanced loading of synaptic vesicles with ACh and increase of ACh quantal size, since it was fully prevented by blocking of vesicular ACh transporter. It was also prevented by tropomyosin-related kinase B (TrkB) receptors inhibitor ANA12. Exogenous BDNF (1 nM) mimicked thrombin effect and increased the amplitude of MEPPs and EPPs by 25–30%. It required involvement of protein kinase A (PKA) and mitogen-activated protein kinase (MEK1/2)-mediated pathway, but not phospholipase C (PLC). Blocking A2A adenosine receptors by ZM241385 abolished the effect of BDNF, whereas additional stimulation of A2A receptors by CGS21680 increased MEPP amplitudes, which was prevented by MEK1/2 inhibitor U0126. At mature NMJs, BDNF enhanced MEPPs frequency by 30–40%. This effect was selectively prevented by inhibition of PLC, but not PKA or MEK1/2. It is suggested that interrelated effects of thrombin/BDNF in mature and newly-formed NMJs are realized via enhancement of vesicular ACh transport and quantal size increase. BDNF-induced potentiation of synaptic transmission involves the functional coupling between A2A receptor-dependent active PKA and neurotrophin-triggered MAPK pathway, as well as PLC-dependent increase in frequency of MEPPs.
Highlights
In recent years, there has been a growing interest in muscle-derived factors—myokines, and their physiological activities
Blocking A2A adenosine receptors by ZM241385 abolished the effect of brain-derived neurotrophic factor (BDNF), whereas additional stimulation of A2A receptors by CGS21680 increased miniature endplate potentials (MEPPs) amplitudes, which was prevented by MEK1/2 inhibitor U0126
One of the canonical signaling pathways triggered by BDNF is the phosphoinositide pathway, we investigated which signaling pathways triggered by BDNF could be associated which starts with tropomyosin-related kinase B (TrkB)-mediated activation of phospholipase C (PLC) [29,30,31]
Summary
There has been a growing interest in muscle-derived factors—myokines, and their physiological activities. Myokines are a family of substances variable in structure, which can be released from skeletal muscle fibers and produce multiple systemic or local effects in organs and tissues [1,2,3,4]. The expression of proBDNF [6,7] and its release from skeletal muscle with subsequent regulatory action of its mature form on muscle receptors and metabolism was shown [5]. Skeletal muscle fibers may serve as a source of a growing number of myokines, and of active thrombin [8,9]. Prothrombin expression in skeletal muscle was reported already [10].
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