Role of ATP-Dependent Calcium Regulation in Modulation of Drosophila Synaptic Thermotolerance

Maintenance of synaptic transmission requires regulation of intracellular Ca2+ in presynaptic nerve terminals; loss of this regulation at elevated temperatures may cause synaptic failure. Accordingly, we examined the thermosensitivity of presynaptic calcium regulation in Drosophila larval neuromuscular junctions, testing for effects of disrupting calcium clearance. Motor neurons were loaded with the ratiometric Ca2+ indicator Fura-dextran to monitor calcium regulation as temperature increased. Block of the Na+/Ca2+ exchanger or removal of extracellular Ca2+ prevented the normal temperature-induced increase in resting calcium. Conversely, two treatments that interfered with Ca2+ clearance—inactivation of the endoplasmic reticulum Ca2+-ATPase with thapsigargin and inhibition of the plasma membrane Ca2+-ATPase with high pH—significantly accelerated the temperature-induced rise in resting Ca2+ concentration and reduced the thermotolerance of synaptic transmission. Disrupting Ca2+-ATPase function by interfering with energy production also facilitated the temperature-induced rise in resting [Ca2+] and reduced thermotolerance of synaptic transmission. Conversely, fortifying energy levels with extra intracellular ATP extended the operating temperature range of both synaptic transmission and Ca2+ regulation. In each of these cases, Ca2+ elevations evoked by an electrical stimulation of the nerve (evoked Ca2+ responses) failed when resting Ca2+ remained >e 200 nM for several minutes. Failure of synaptic function was correlated with the release of intracellular calcium stores, and we provide evidence suggesting that release from the mitochondria disrupts evoked calcium responses and synaptic transmission. Thus the thermal limit of synaptic transmission may be directly linked to the stability of ATP-dependent mechanisms that regulate intracellular ion concentrations in the nerve terminal.

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Hyperthermic Preconditioning of Presynaptic Calcium Regulation in Drosophila

Journal of Neurophysiology ◽

10.1152/jn.01251.2007 ◽

2008 ◽

Vol 99 (5) ◽

pp. 2420-2430 ◽

Cited By ~ 20

Author(s):

M. K. Klose ◽

H. L. Atwood ◽

R. M. Robertson

Keyword(s):

Heat Shock ◽

Synaptic Transmission ◽

Motor Neurons ◽

Neuromuscular Junctions ◽

Calcium Influx ◽

Calcium Regulation ◽

Hsp70 Expression ◽

Presynaptic Calcium ◽

Hyperthermic Preconditioning ◽

The Stability

We examined the thermosensitivity of calcium regulation in Drosophila larval neuromuscular junctions, testing effects of prior heat shock and Hsp70 expression. Motor neurons were loaded with either the ratiometric indicator Fura-dextran or the nonratiometric indicator Oregon Green bis-( o-aminophenoxy)- N,N,N′,N′-tetraacetic acid to monitor parameters of calcium regulation as temperature increased. Nerve terminals treated to a prior heat shock, and those of transgenic flies expressing higher than normal levels of Hsp70, were better able to maintain near-normal resting calcium concentrations, calcium influx, and calcium clearance at higher temperatures. Synaptic transmission was also protected by prior heat shock and by higher than normal Hsp70 expression. Thus the thermal limit of synaptic transmission may be directly linked to the stability of calcium regulation.

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Activity dependent modulation of synaptic transmission by presynaptic calcium stores: A dichotomy of short-term depression and facilitation

BMC Neuroscience ◽

10.1186/1471-2202-14-s1-p351 ◽

2013 ◽

Vol 14 (S1) ◽

Author(s):

Suhita Nadkarni ◽

Thomas Bartol ◽

Herbert Levine ◽

Terrence Sejnowski

Keyword(s):

Synaptic Transmission ◽

Calcium Stores ◽

Short Term ◽

Presynaptic Calcium ◽

Activity Dependent

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The p75NTR neurotrophin receptor is required to organize the mature neuromuscular synapse by regulating synaptic vesicle availability

Acta Neuropathologica Communications ◽

10.1186/s40478-019-0802-7 ◽

2019 ◽

Vol 7 (1) ◽

Cited By ~ 2

Author(s):

Viviana Pérez ◽

Francisca Bermedo-Garcia ◽

Diego Zelada ◽

Felipe A. Court ◽

Miguel Ángel Pérez ◽

...

Keyword(s):

Neuromuscular Junction ◽

Motor Neurons ◽

Neuromuscular Junctions ◽

Neuronal Damage ◽

Neuromuscular Synapse ◽

Force Production ◽

Synaptic Function ◽

Neurotrophin Receptor ◽

Acetylcholinesterase Inhibition ◽

Pharmacological Intervention

Abstract The coordinated movement of organisms relies on efficient nerve-muscle communication at the neuromuscular junction. After peripheral nerve injury or neurodegeneration, motor neurons and Schwann cells increase the expression of the p75NTR pan-neurotrophin receptor. Even though p75NTR targeting has emerged as a promising therapeutic strategy to delay peripheral neuronal damage progression, the effects of long-term p75NTR inhibition at the mature neuromuscular junction have not been elucidated. We performed quantitative neuroanathomical analyses of the neuromuscular junction in p75NTR null mice by laser confocal and electron microscopy, which were complemented with electromyography, locomotor tests, and pharmacological intervention studies. Mature neuromuscular synapses of p75NTR null mice show impaired postsynaptic organization and ultrastructural complexity, which correlate with altered synaptic function at the levels of nerve activity-induced muscle responses, muscle fiber structure, force production, and locomotor performance. Our results on primary myotubes and denervated muscles indicate that muscle-derived p75NTR does not play a major role on postsynaptic organization. In turn, motor axon terminals of p75NTR null mice display a strong reduction in the number of synaptic vesicles and active zones. According to the observed pre and postsynaptic defects, pharmacological acetylcholinesterase inhibition rescued nerve-dependent muscle response and force production in p75NTR null mice. Our findings revealing that p75NTR is required to organize mature neuromuscular junctions contribute to a comprehensive view of the possible effects caused by therapeutic attempts to target p75NTR.

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Congenital myasthenic syndrome due to a TOR1AIP1 mutation: a new disease pathway for impaired synaptic transmission

Brain Communications ◽

10.1093/braincomms/fcaa174 ◽

2020 ◽

Vol 2 (2) ◽

Author(s):

Judith Cossins ◽

Richard Webster ◽

Susan Maxwell ◽

Pedro M Rodríguez Cruz ◽

Ravi Knight ◽

...

Keyword(s):

Membrane Protein ◽

Synaptic Transmission ◽

Muscle Weakness ◽

Nuclear Membrane ◽

Nerve Stimulation ◽

Neuromuscular Junctions ◽

Synaptic Function ◽

Inherited Disorders ◽

Repetitive Nerve Stimulation ◽

Nuclear Membrane Protein

Abstract Congenital myasthenic syndromes are inherited disorders characterized by fatiguable muscle weakness resulting from impaired signal transmission at the neuromuscular junction. Causative mutations have been identified in genes that can affect the synaptic function or structure. We identified a homozygous frameshift deletion c.127delC, p. Pro43fs in TOR1AIP1 in two siblings with limb-girdle weakness and impaired transmission at the neuromuscular synapse. TOR1AIP1 encodes the inner nuclear membrane protein lamin-associated protein 1. On muscle biopsy from the index case, lamin-associated protein 1 was absent from myonuclei. A mouse model with lamin-associated protein 1 conditionally knocked out in striated muscle was used to analyse the role of lamin-associated protein 1 in synaptic dysfunction. Model mice develop fatiguable muscle weakness as demonstrated by using an inverted screen hang test. Electromyography on the mice revealed a decrement on repetitive nerve stimulation. Ex vivo analysis of hemi-diaphragm preparations showed both miniature and evoked end-plate potential half-widths were prolonged which was associated with upregulation of the foetal acetylcholine receptor γ subunit. Neuromuscular junctions on extensor digitorum longus muscles were enlarged and fragmented, and the number of subsynaptic nuclei was significantly increased. Following these findings, electromyography was performed on cases of other nuclear envelopathies caused by mutations in LaminA/C or emerin, but decrement on repetitive nerve stimulation or other indications of defective neuromuscular transmission were not seen. Thus, this report highlights the first nuclear membrane protein in which defective function can lead to impaired synaptic transmission.

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Overexpression of tau results in defective synaptic transmission in Drosophila neuromuscular junctions

Biochemical Society Transactions ◽

10.1042/bst0340088 ◽

2006 ◽

Vol 34 (1) ◽

pp. 88-90 ◽

Cited By ~ 19

Author(s):

F. Chee ◽

A. Mudher ◽

T.A. Newman ◽

M. Cuttle ◽

S. Lovestone ◽

...

Keyword(s):

Synaptic Transmission ◽

Axonal Transport ◽

Motor Neurons ◽

Neuromuscular Junctions ◽

Clinical Symptoms ◽

Low Frequency ◽

Synaptic Dysfunction ◽

High Frequency Stimulation ◽

Low Frequency Stimulation ◽

Frequency Stimulation

Synaptic dysfunction is believed to be an early pathological change in neurodegenerative diseases and may cause the earliest clinical symptoms. We have used Drosophila to model a tauopathy in order to analyse the earliest neuronal and synaptic dysfunction. Our work has shown that overexpression of human tau (0N3R) in larval motor neurons causes a disruption of axonal transport and a morphological and functional disruption of NMJs (neuromuscular junctions). Tau-expressing NMJs are smaller with an abnormal structure. Despite abnormal morphology, tau-expressing NMJs retain synaptotagmin expression and can form active zones. Tau-expressing NMJs are functionally abnormal and exhibit disrupted vesicle cycling and synaptic transmission. At low-frequency stimulation (1 Hz), ESPs (evoked synaptic potentials) produced by tau-expressing motor neurons were indistinguishable from wild-type; however, following high-frequency stimulation (50 Hz), ESPs from tau-expressing NMJs were significantly decreased in amplitude. To investigate the mechanism underlying the change in ESPs, we analysed the relative numbers and distribution of mitochondria. This revealed that motor neurons expressing tau had a significant reduction in the number of detectable mitochondria in the pre-synaptic terminal. Our results demonstrate that tau overexpression results in synaptic dysfunction, associated with a reduced complement of functional mitochondria. These findings suggest that disruption of axonal transport and synaptic transmission may be key components of the pathogenic mechanism that underlie neuronal dysfunction in the early stages of tauopathies.

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Cul4 ubiquitin ligase cofactor DCAF12 promotes neurotransmitter release and homeostatic plasticity

The Journal of Cell Biology ◽

10.1083/jcb.201805099 ◽

2019 ◽

Vol 218 (3) ◽

pp. 993-1010 ◽

Cited By ~ 3

Author(s):

Lilian A. Patrón ◽

Kei Nagatomo ◽

David Tyler Eves ◽

Mays Imad ◽

Kimberly Young ◽

...

Keyword(s):

Glutamate Receptor ◽

Motor Neurons ◽

Ubiquitin Ligase ◽

Neurotransmitter Release ◽

Neuromuscular Junctions ◽

Synaptic Function ◽

Homeostatic Plasticity ◽

Independent Functions ◽

Synaptic Boutons

We genetically characterized the synaptic role of the Drosophila homologue of human DCAF12, a putative cofactor of Cullin4 (Cul4) ubiquitin ligase complexes. Deletion of Drosophila DCAF12 impairs larval locomotion and arrests development. At larval neuromuscular junctions (NMJs), DCAF12 is expressed presynaptically in synaptic boutons, axons, and nuclei of motor neurons. Postsynaptically, DCAF12 is expressed in muscle nuclei and facilitates Cul4-dependent ubiquitination. Genetic experiments identified several mechanistically independent functions of DCAF12 at larval NMJs. First, presynaptic DCAF12 promotes evoked neurotransmitter release. Second, postsynaptic DCAF12 negatively controls the synaptic levels of the glutamate receptor subunits GluRIIA, GluRIIC, and GluRIID. The down-regulation of synaptic GluRIIA subunits by nuclear DCAF12 requires Cul4. Third, presynaptic DCAF12 is required for the expression of synaptic homeostatic potentiation. We suggest that DCAF12 and Cul4 are critical for normal synaptic function and plasticity at larval NMJs.

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Synaptic Physiology and Mitochondrial Function in Crayfish Tonic and Phasic Motor Neurons

Journal of Neurophysiology ◽

10.1152/jn.1997.78.1.281 ◽

1997 ◽

Vol 78 (1) ◽

pp. 281-294 ◽

Cited By ~ 42

Author(s):

Peter V. Nguyen ◽

Leo Marin ◽

Harold L. Atwood

Keyword(s):

Synaptic Transmission ◽

Motor Neurons ◽

Mitochondrial Function ◽

Neuromuscular Junctions ◽

Mitochondrial Protein ◽

Synaptic Depression ◽

Oxidative Activity ◽

Protein Synthesis Inhibitor ◽

Mitochondrial Content ◽

Synaptic Physiology

Nguyen, Peter V., Leo Marin, and Harold L. Atwood. Synaptic physiology and mitochondrial function in crayfish tonic and phasic motor neurons. J. Neurophysiol. 78: 281–294, 1997. Phasic and tonic motor neurons of crustaceans differ strikingly in their junctional synaptic physiology. Tonic neurons generally produce small excitatory postsynaptic potentials (EPSPs) that facilitate strongly as stimulation frequency is increased, and normally show no synaptic depression. In contrast, phasic neurons produce relatively large EPSPs with weak frequency facilitation and pronounced depression. We addressed the hypothesis that mitochondrial function is an important determinant of the features of synaptic transmission in these neurons. Mitochondrial fluorescence was measured with confocal microscopy in phasic and tonic axons and terminals of abdominal and leg muscles after exposure to supravital mitochondrial fluorochromes, rhodamine-123 (Rh123) and 4-diethylaminostyryl-N-methylpyridinium iodide (4-Di-2-Asp). Mitochondria of tonic axons and neuromuscular junctions had significantly higher mean Rh123 and 4-Di-2-Asp fluorescence than in phasic neurons, indicating more accumulation of the fluorochromes. Mitochondrial membrane potential, which is responsible for Rh123 uptake and is related to mitochondrial oxidative activity (the production of ATP by oxidation of metabolic substrates), is likely higher in tonic axons. Electron microscopy showed that tonic axons contain approximately fivefold more mitochondria per μm2 cross-sectional area than phasic axons. Neuromuscular junctions of tonic axons also have a much higher mitochondrial content than those of phasic axons. We tested the hypothesis that synaptic fatigue resistance is dependent on mitochondrial function in crayfish motor axons. Impairment of mitochondrial function by uncouplers of oxidative phosphorylation, dinitrophenol or carbonyl cyanide m-chlorophenylhydrazone, or by the electron transport inhibitor sodium azide, led to marked synaptic depression of a tonic axon and accelerated depression of a phasic axon during maintained stimulation. Iodoacetate, an inhibitor of glycolysis, and chloramphenicol, a mitochondrial protein synthesis inhibitor, had no significant effects on either mitochondrial fluorescence or synaptic depression in tonic or phasic axons. Collectively, the results provide evidence that mitochondrial oxidative metabolism is important for sustaining synaptic transmission during maintained stimulation of tonic and phasic motor neurons. Tonic neurons have a higher mitochondrial content and greater oxidative activity; these features are correlated with their greater resistance to synaptic depression. Conversely, phasic neurons have a lower mitochondrial content, less oxidative activity, and greater synaptic fatigability.

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