skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

Explore Research Products in the PAR
It may take a few hours for recently added research products to appear in PAR search results.


Title: A comparison of three different methods of eliciting rapid activity-dependent synaptic plasticity at the Drosophila NMJ
The Drosophila NMJ is a system of choice for investigating the mechanisms underlying the structural and functional modifications evoked during activity-dependent synaptic plasticity. Because fly genetics allows considerable versatility, many strategies can be employed to elicit this activity. Here, we compare three different stimulation methods for eliciting activity-dependent changes in structure and function at the Drosophila NMJ. We find that the method using patterned stimulations driven by a K+-rich solution creates robust structural modifications but reduces muscle viability, as assessed by resting potential and membrane resistance. We argue that, using this method, electrophysiological studies that consider the frequency of events, rather than their amplitude, are the only reliable studies. We contrast these results with the expression of CsChrimson channels and red-light stimulation at the NMJ, as well as with the expression of TRPA channels and temperature stimulation. With both these methods we observed reliable modifications of synaptic structures and consistent changes in electrophysiological properties. Indeed, we observed a rapid appearance of immature boutons that lack postsynaptic differentiation, and a potentiation of spontaneous neurotransmission frequency. Surprisingly, a patterned application of temperature changes alone is sufficient to provoke both structural and functional plasticity. In this context, temperature-dependent TRPA channel activation induces additional structural plasticity but no further increase in the frequency of spontaneous neurotransmission, suggesting an uncoupling of these mechanisms.  more » « less
Award ID(s):
1736019
PAR ID:
10341068
Author(s) / Creator(s):
; ;
Editor(s):
Kim, Hongkyun
Date Published:
Journal Name:
PLOS ONE
Volume:
16
Issue:
11
ISSN:
1932-6203
Page Range / eLocation ID:
e0260553
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. ; ; ; (, The Journal of Neuroscience)
    Neurotransmission is shaped by extracellular pH. Alkalization enhances pH-sensitive transmitter release and receptor activation, whereas acidification inhibits these processes and can activate acid-sensitive conductances in the synaptic cleft. Previous work has shown that the synaptic cleft can either acidify because of synaptic vesicular release and/or alkalize because of Ca2+extrusion by the plasma membrane ATPase (PMCA). The direction of change differs across synapse types. At the mammalian neuromuscular junction (NMJ), the direction and magnitude of pH transients in the synaptic cleft during transmission remain ambiguous. We set out to elucidate the extracellular pH transients that occur at this cholinergic synapse under near-physiological conditions and identify their sources. We monitored pH-dependent changes in the synaptic cleft of the mouse levator auris longus using viral expression of the pseudoratiometric probe pHusion-Ex in the muscle. Using mice from both sexes, a significant and prolonged alkalization occurred when stimulating the connected nerve for 5 s at 50 Hz, which was dependent on postsynaptic intracellular Ca2+release. Sustained stimulation for a longer duration (20 s at 50 Hz) caused additional prolonged net acidification at the cleft. To investigate the mechanism underlying cleft alkalization, we used muscle-expressed GCaMP3 to monitor the contribution of postsynaptic Ca2+. Activity-induced liberation of intracellular Ca2+in muscle positively correlated with alkalization of the synaptic cleft, whereas inhibiting PMCA significantly decreased the extent of cleft alkalization. Thus, cholinergic synapses of the mouse NMJ typically alkalize because of cytosolic Ca2+liberated in muscle during activity, unless under highly strenuous conditions where acidification predominates. SIGNIFICANCE STATEMENTChanges in synaptic cleft pH alter neurotransmission, acting on receptors and channels on both sides of the synapse. Synaptic acidification has been associated with a myriad of diseases in the central and peripheral nervous system. Here, we report that in near-physiological recording conditions the cholinergic neuromuscular junction shows use-dependent bidirectional changes in synaptic cleft pH—immediate alkalinization and a long-lasting acidification under prolonged stimulation. These results provide further insight into physiologically relevant changes at cholinergic synapses that have not been defined previously. Understanding and identifying synaptic pH transients during and after neuronal activity provides insight into short-term synaptic plasticity synapses and may identify therapeutic targets for diseases. 
    more » « less
  2. ; ; ; (, Biomolecules)
    The mouse neuromuscular junction (NMJ) has long been used as a model synapse for the study of neurotransmission in both healthy and disease states of the NMJ. Neurotransmission from these neuromuscular nerve terminals occurs at highly organized structures called active zones (AZs). Within AZs, the relationships between the voltage-gated calcium channels and docked synaptic vesicles govern the probability of acetylcholine release during single action potentials, and the short-term plasticity characteristics during short, high frequency trains of action potentials. Understanding these relationships is important not only for healthy synapses, but also to better understand the pathophysiology of neuromuscular diseases. In particular, we are interested in Lambert-Eaton myasthenic syndrome (LEMS), an autoimmune disorder in which neurotransmitter release from the NMJ decreases, leading to severe muscle weakness. In LEMS, the reduced neurotransmission is traditionally thought to be caused by the antibody-mediated removal of presynaptic voltage-gated calcium channels. However, recent experimental data and AZ computer simulations have predicted that a disruption in the normally highly organized active zone structure, and perhaps autoantibodies to other presynaptic proteins, contribute significantly to pathological effects in the active zone and the characteristics of chemical transmitters. 
    more » « less
  3. ; ; ; ; ; ; ; ; ; ; et al (, The Journal of Neuroscience)
    The dogma that the synaptic cleft acidifies during neurotransmission is based on the corelease of neurotransmitters and protons from synaptic vesicles, and is supported by direct data from sensory ribbon-type synapses. However, it is unclear whether acidification occurs at non–ribbon-type synapses. Here we used genetically encoded fluorescent pH indicators to examine cleft pH at conventional neuronal synapses. At the neuromuscular junction of femaleDrosophilalarvae, we observed alkaline spikes of over 1 log unit during fictive locomotionin vivo. Ex vivo, single action potentials evoked alkalinizing pH transients of only ∼0.01 log unit, but these transients summated rapidly during burst firing. A chemical pH indicator targeted to the cleft corroborated these findings. Cleft pH transients were dependent on Ca2+movement across the postsynaptic membrane, rather than neurotransmitter release per se, a result consistent with cleft alkalinization being driven by the Ca2+/H+antiporting activity of the plasma membrane Ca2+-ATPase at the postsynaptic membrane. Targeting the pH indicators to the microenvironment of the presynaptic voltage gated Ca2+channels revealed that alkalinization also occurred within the cleft proper at the active zone and not just within extrasynaptic regions. Application of the pH indicators at the mouse calyx of Held, a mammalian central synapse, similarly revealed cleft alkalinization during burst firing in both males and females. These findings, made at two quite different non–ribbon type synapses, suggest that cleft alkalinization during neurotransmission, rather than acidification, is a generalizable phenomenon across conventional neuronal synapses. SIGNIFICANCE STATEMENTNeurotransmission is highly sensitive to the pH of the extracellular milieu. This is readily evident in the neurological symptoms that accompany systemic acid/base imbalances. Imaging data from sensory ribbon-type synapses show that neurotransmission itself can acidify the synaptic cleft, likely due to the corelease of protons and glutamate. It is not clear whether the same phenomenon occurs at conventional neuronal synapses due to the difficulties in collecting such data. If it does occur, it would provide for an additional layer of activity-dependent modulation of neurotransmission. Our findings of alkalinization, rather than acidification, within the cleft of two different neuronal synapses encourages a reassessment of the scope of activity-dependent pH influences on neurotransmission and short-term synaptic plasticity. 
    more » « less
  4. ; ; (, The Journal of Neuroscience)
    NMDA-type glutamate receptors are heterotetrameric complexes composed of two GluN1 and two GluN2 subunits. The precise composition of the GluN2 subunits determines the channel's biophysical properties and influences its interaction with postsynaptic scaffolding proteins and signaling molecules involved in synaptic physiology and plasticity. The precise regulation of NMDAR subunit composition at synapses is crucial for proper synaptogenesis, neuronal circuit development, and synaptic plasticity, a cellular model of memory formation. In the forebrain during early development, NMDARs contain solely the GluN2B subunit, which is necessary for proper synaptogenesis and synaptic plasticity. In rodents, GluN2A subunit expression begins in the second postnatal week, replacing GluN2B-containing NMDARs at synapses in an activity- or sensory experience-dependent process. This switch in NMDAR subunit composition at synapses alters channel properties and reduces synaptic plasticity. The molecular mechanism regulating the switch remains unclear. We have investigated the role of activity-dependent internalization of GluN2B-containing receptors in shaping synaptic NMDAR subunit composition. Using molecular, pharmacological, and electrophysiological approaches in cultured organotypic hippocampal slices from rats of both sexes, we show that the process of incorporating GluN2A-containing NMDAR receptors requires activity-dependent internalization of GluN2B-containing NMDARs. Interestingly, blockade of GluN2A synaptic incorporation was associated with impaired potentiation of AMPA-mediated synaptic transmission, suggesting a potential coupling between the trafficking of AMPARs into synapses and that of GluN2A-containing NMDARs. These insights contribute to our understanding of the molecular mechanisms underlying synaptic trafficking of glutamate receptors and synaptic plasticity. They may also have implications for therapeutic strategies targeting NMDAR function in neurological disorders. 
    more » « less
  5. ; ; (, MRS Advances)
    Synaptic plasticity refers to activity-dependent synaptic strengthening or weakening between neurons. It is usually associated with homosynaptic plasticity, which refers to a synaptic junction controlled by interactions between specific neurons. Heterosynaptic plasticity, on the other hand, lacks this specificity. It involves much larger populations of synapses and neurons and can be associated with changes in synaptic strength due to nonlocal alterations in the ambient electrochemical environment. This paper presents specific examples demonstrating how variations in the ambient electrochemical environment of lipid membranes can impact the nonlinear dynamical behaviors of memristive and memcapacitive systems in droplet interface bilayers (DIBs). Examples include the use of pH as a modulatory factor that alters the voltage-dependent memristive behavior of alamethicin ion channels in DIB lipid bilayers, and the discovery of long-term potentiation (LTP) in a lipid bilayer-only system after application of electrical stimulation protocols. 
    more » « less
  • Citation Formats
  • MLA
  • APA
  • Chicago
  • BibTeX
  • Save / Share this Record
  • Send to Email