BDNF/TrkB signalling, in cooperation with muscarinic signalling, retrogradely regulates PKA pathway to phosphorylate SNAP-25 and Synapsin-1 at the neuromuscular junction.

Background: Protein kinase A (PKA) enhances neurotransmission at the neuromuscular junction (NMJ), which is retrogradely regulated by nerve-induced muscle contraction to promote Acetylcholine (ACh) release through the phosphorylation of molecules involved in synaptic vesicle exocytosis (SNAP-25 and Synapsin-1). However, the molecular mechanism of the retrograde regulation of PKA subunits and its targets by BDNF/TrkB pathway and muscarinic signalling has not been demonstrated until now. At the NMJ, retrograde control is mainly associated with BDNF/TrkB signalling as muscle contraction enhances BDNF levels and controls specific kinases involved in the neurotransmission.

Brain-derived neurotrophic factor signaling in the neuromuscular junction during developmental axonal competition and synapse elimination.

During the development of the nervous system, there is an overproduction of neurons and synapses. Hebbian competition between neighboring nerve endings and synapses performing different activity levels leads to their elimination or strengthening. We have extensively studied the involvement of the brain-derived neurotrophic factor-Tropomyosin-related kinase B receptor neurotrophic retrograde pathway, at the neuromuscular junction, in the axonal development and synapse elimination process versus the synapse consolidation.

Synaptic retrograde regulation of the PKA-induced SNAP-25 and Synapsin-1 phosphorylation.

Background: Bidirectional communication between presynaptic and postsynaptic components contribute to the homeostasis of the synapse. In the neuromuscular synapse, the arrival of the nerve impulse at the presynaptic terminal triggers the molecular mechanisms associated with ACh release, which can be retrogradely regulated by the resulting muscle contraction. This retrograde regulation, however, has been poorly studied.

TrkB signaling is correlated with muscular fatigue resistance and less vulnerability to neurodegeneration.

At the neuromuscular junction (NMJ), motor neurons and myocytes maintain a bidirectional communication that guarantees adequate functionality. Thus, motor neurons' firing pattern, which is influenced by retrograde muscle-derived neurotrophic factors, modulates myocyte contractibility. Myocytes can be fast-twitch fibers and become easily fatigued or slow-twitch fibers and resistant to fatigue.

Muscarinic Receptors in Developmental Axonal Competition at the Neuromuscular Junction.

In recent years, we have studied by immunohistochemistry, intracellular recording, and western blotting the role of the muscarinic acetylcholine receptors (mAChRs; M, M, and M subtypes) in the mammalian neuromuscular junction (NMJ) during development and in the adult. Here, we evaluate our published data to emphasize the mAChRs' relevance in developmental synaptic elimination and their crosstalk with other metabotropic receptors, downstream kinases, and voltage-gated calcium channels (VGCCs). The presence of mAChRs in the presynaptic membrane of motor nerve terminals allows an autocrine mechanism in which the secreted acetylcholine influences the cell itself in feedback.

Involvement of the Voltage-Gated Calcium Channels L- P/Q- and N-Types in Synapse Elimination During Neuromuscular Junction Development.

During the nervous system development, synapses are initially overproduced. In the neuromuscular junction (NMJ) however, competition between several motor nerve terminals and the synapses they made ends with the maturation of only one axon. The competitive signaling between axons is mediated by the differential activity-dependent release of the neurotransmitter ACh, co-transmitters, and neurotrophic factors.

PKA and PKC Balance in Synapse Elimination during Neuromuscular Junction Development.

During the development of the nervous system, synaptogenesis occurs in excess though only the appropriate connections consolidate. At the neuromuscular junction, competition between several motor nerve terminals results in the maturation of a single axon and the elimination of the others. The activity-dependent release of transmitter, cotransmitters, and neurotrophic factors allows the direct mutual influence between motor axon terminals through receptors such as presynaptic muscarinic ACh autoreceptors and the tropomyosin-related kinase B neurotrophin receptor.

Running and Swimming Differently Adapt the BDNF/TrkB Pathway to a Slow Molecular Pattern at the NMJ.

Physical exercise improves motor control and related cognitive abilities and reinforces neuroprotective mechanisms in the nervous system. As peripheral nerves interact with skeletal muscles at the neuromuscular junction, modifications of this bidirectional communication by physical activity are positive to preserve this synapse as it increases quantal content and resistance to fatigue, acetylcholine receptors expansion, and myocytes' fast-to-slow functional transition. Here, we provide the intermediate step between physical activity and functional and morphological changes by analyzing the molecular adaptations in the skeletal muscle of the full BDNF/TrkB downstream signaling pathway, directly involved in acetylcholine release and synapse maintenance.

The M muscarinic receptor, in association to M , regulates the neuromuscular PKA molecular dynamics.

Muscarinic acetylcholine receptor 1 subtype (M ) and muscarinic acetylcholine receptor 2 subtype (M ) presynaptic muscarinic receptor subtypes increase and decrease, respectively, neurotransmitter release at neuromuscular junctions. M involves protein kinase A (PKA), although the muscarinic regulation to form and inactivate the PKA holoenzyme is unknown. Here, we show that M signaling inhibits PKA by downregulating Cβ subunit, upregulating RIIα/β and liberating RIβ and RIIα to the cytosol.

The Impact of Kinases in Amyotrophic Lateral Sclerosis at the Neuromuscular Synapse: Insights into BDNF/TrkB and PKC Signaling.

Brain-derived neurotrophic factor (BDNF) promotes neuron survival in adulthood in the central nervous system. In the peripheral nervous system, BDNF is a contraction-inducible protein that, through its binding to tropomyosin-related kinase B receptor (TrkB), contributes to the retrograde neuroprotective control done by muscles, which is necessary for motor neuron function. BDNF/TrkB triggers downstream presynaptic pathways, involving protein kinase C, essential for synaptic function and maintenance.

Opposed Actions of PKA Isozymes (RI and RII) and PKC Isoforms (cPKCβI and nPKCε) in Neuromuscular Developmental Synapse Elimination.

Background: During neuromuscular junction (NMJ) development, synapses are produced in excess. By sensing the activity-dependent release of ACh, adenosine, and neurotrophins, presynaptic receptors prompt axonal competition and loss of the unnecessary axons. The receptor action is mediated by synergistic and antagonistic relations when they couple to downstream kinases (mainly protein kinases A and C (PKA and PKC)), which phosphorylate targets involved in axonal disconnection.

Running and swimming prevent the deregulation of the BDNF/TrkB neurotrophic signalling at the neuromuscular junction in mice with amyotrophic lateral sclerosis.

Nerve-induced muscle contraction regulates the BDNF/TrkB neurotrophic signalling to retrogradely modulate neurotransmission and protect the neuromuscular junctions and motoneurons. In muscles with amyotrophic lateral sclerosis, this pathway is strongly misbalanced and neuromuscular junctions are destabilized, which may directly cause the motoneuron degeneration and muscular atrophy observed in this disease. Here, we sought to demonstrate (1) that physical exercise, whose recommendation has been controversial in amyotrophic lateral sclerosis, would be a good option for its therapy, because it normalizes and improves the altered neurotrophin pathway and (2) a plausible molecular mechanism underlying its positive effect.

Overview of Impaired BDNF Signaling, Their Coupled Downstream Serine-Threonine Kinases and SNARE/SM Complex in the Neuromuscular Junction of the Amyotrophic Lateral Sclerosis Model SOD1-G93A Mice.

Amyotrophic lateral sclerosis (ALS) is a chronic neurodegenerative disease characterized by progressive motor weakness. It is accepted that it is caused by motoneuron degeneration leading to a decrease in muscle stimulation. However, ALS is being redefined as a distal axonopathy, in that neuromuscular junction dysfunction precedes and may even influence motoneuron loss.

nPKCε Mediates SNAP-25 Phosphorylation of Ser-187 in Basal Conditions and After Synaptic Activity at the Neuromuscular Junction.

Protein kinase C (PKC) and substrates like SNAP-25 regulate neurotransmission. At the neuromuscular junction (NMJ), PKC promotes neurotransmitter release during synaptic activity. Thirty minutes of muscle contraction enhances presynaptic PKC isoform levels, specifically cPKCβI and nPKCε, through retrograde BDNF/TrkB signaling.

Experimental myofascial trigger point creation in rodents.

Myofascial pain syndrome is one of the most common forms of muscle pain. In this syndrome, pain is originated by the so-called trigger points, which consists of a set of palpable contraction knots in the muscle. It has been proposed that a high, spontaneous neurotransmission may be involved in the generation of these contraction knots.

BDNF-TrkB Signaling Coupled to nPKCε and cPKCβI Modulate the Phosphorylation of the Exocytotic Protein Munc18-1 During Synaptic Activity at the Neuromuscular Junction.

Munc18-1, a neuron-specific member of the Sec1/Munc18 family, is involved in neurotransmitter release by binding tightly to syntaxin. Munc18-1 is phosphorylated by PKC on Ser-306 and Ser-313 which reduces the amount of Munc18-1 able to bind syntaxin. We have previously identified that PKC is involved in neurotransmitter release when continuous electrical stimulation imposes a moderate activity on the NMJ and that muscle contraction through TrkB has an important impact on presynaptic PKC isoforms levels, specifically cPKCβI and nPKCε.

Adenosine Receptors in Developing and Adult Mouse Neuromuscular Junctions and Functional Links With Other Metabotropic Receptor Pathways.

In the last few years, we have studied the presence and involvement in synaptogenesis and mature transmitter release of the adenosine autoreceptors (AR) in the mammalian neuromuscular junction (NMJ). Here, we review and bring together the previously published data to emphasize the relevance of these receptors for developmental axonal competition, synaptic loss and mature NMJ functional modulation. However, in addition to AR, activity-dependent mediators originating from any of the three cells that make the synapse (nerve, muscle, and glial cells) cross the extracellular cleft to generate signals in target metabotropic receptors.

Synaptic Activity and Muscle Contraction Increases PDK1 and PKCβI Phosphorylation in the Presynaptic Membrane of the Neuromuscular Junction.

Conventional protein kinase C βI (cPKCβI) is a conventional protein kinase C (PKC) isoform directly involved in the regulation of neurotransmitter release in the neuromuscular junction (NMJ). It is located exclusively at the nerve terminal and both synaptic activity and muscle contraction modulate its protein levels and phosphorylation. cPKCβI molecular maturation includes a series of phosphorylation steps, the first of which is mediated by phosphoinositide-dependent kinase 1 (PDK1).

Membrane Receptor-Induced Changes of the Protein Kinases A and C Activity May Play a Leading Role in Promoting Developmental Synapse Elimination at the Neuromuscular Junction.

Synapses that are overproduced during histogenesis in the nervous system are eventually lost and connectivity is refined. Membrane receptor signaling leads to activity-dependent mutual influence and competition between axons directly or with the involvement of the postsynaptic cell and the associated glial cell/s. Presynaptic muscarinic acetylcholine (ACh) receptors (subtypes mAChR; M, M and M), adenosine receptors (AR; A and A) and the tropomyosin-related kinase B receptor (TrkB), among others, all cooperate in synapse elimination.

Muscle Contraction Regulates BDNF/TrkB Signaling to Modulate Synaptic Function through Presynaptic cPKCα and cPKCβI.

The neurotrophin brain-derived neurotrophic factor (BDNF) acts via tropomyosin-related kinase B receptor (TrkB) to regulate synapse maintenance and function in the neuromuscular system. The potentiation of acetylcholine (ACh) release by BDNF requires TrkB phosphorylation and Protein Kinase C (PKC) activation. BDNF is secreted in an activity-dependent manner but it is not known if pre- and/or postsynaptic activities enhance BDNF expression at the neuromuscular junction (NMJ).

Presynaptic Membrane Receptors Modulate ACh Release, Axonal Competition and Synapse Elimination during Neuromuscular Junction Development.

During the histogenesis of the nervous system a lush production of neurons, which establish an excessive number of synapses, is followed by a drop in both neurons and synaptic contacts as maturation proceeds. Hebbian competition between axons with different activities leads to the loss of roughly half of the neurons initially produced so connectivity is refined and specificity gained. The skeletal muscle fibers in the newborn neuromuscular junction (NMJ) are polyinnervated but by the end of the competition, 2 weeks later, the NMJ are innervated by only one axon.

Synergistic Action of Presynaptic Muscarinic Acetylcholine Receptors and Adenosine Receptors in Developmental Axonal Competition at the Neuromuscular Junction.

The development of the nervous system involves the initial overproduction of synapses, which promotes connectivity. Hebbian competition between axons with different activities leads to the loss of roughly half of the overproduced elements and this refines connectivity. We used quantitative immunohistochemistry to investigate, in the postnatal day 7 (P7) to P9 neuromuscular junctions, the involvement of muscarinic receptors (muscarinic acetylcholine autoreceptors and the M1, M2, and M4 subtypes) and adenosine receptors (A1 and A2A subtypes) in the control of axonal elimination after the mouse levator auris longus muscle had been exposed to selective antagonists in vivo.

Presynaptic Muscarinic Acetylcholine Receptors and TrkB Receptor Cooperate in the Elimination of Redundant Motor Nerve Terminals during Development.

The development of the nervous system involves the overproduction of synapses but connectivity is refined by Hebbian activity-dependent axonal competition. The newborn skeletal muscle fibers are polyinnervated but, at the end of the competition process, some days later, become innervated by a single axon. We used quantitative confocal imaging of the autofluorescent axons from transgenic B6.

Presynaptic muscarinic acetylcholine autoreceptors (M1, M2 and M4 subtypes), adenosine receptors (A1 and A2A) and tropomyosin-related kinase B receptor (TrkB) modulate the developmental synapse elimination process at the neuromuscular junction.

Background: The development of the nervous system involves an initially exuberant production of neurons that make an excessive number of synaptic contacts. The initial overproduction of synapses promotes connectivity. Hebbian competition between axons with different activities (the least active are punished) leads to the loss of roughly half of the overproduced elements and this refines connectivity and increases specificity.