scholarly journals Degenerative changes in the structure of neuromuscular junctions of Manduca sexta during metamorphosis

1992 ◽  
Vol 167 (1) ◽  
pp. 119-154
Author(s):  
M. B. Rheuben
Keyword(s):  
Manduca Sexta ◽  
Muscle Fiber ◽  
Nerve Terminal ◽  
Structural Changes ◽  
Neuromuscular Junctions ◽  
Motor Nerve ◽  
Freeze Fracture ◽  
Outer Region ◽  
Peripheral Region ◽  
Phagocytic Cells

During the degenerative processes that precede and accompany metamorphosis of the larval mesothoracic dorsal longitudinal muscles of Manduca sexta, the motor nerves and neuromuscular junctions undergo a variety of structural changes that are largely secondary to the changing morphologies of their respective glia. In the central region of the main motor nerve, the multiple layers of glial processes surrounding each of the large axons withdraw, leaving them apposed. In the peripheral region of the main motor nerve and in the secondary and tertiary nerve branches supplying the muscle, the outer glial processes of the nerve sheath and those that loosely wrap accompanying small neurosecretory axons all swell. Phagocytic cells and cells of unknown function invade the outer region of the nerve. In the neuromuscular junctions, the glial cells withdraw their processes from a complicated interdigitation with processes from the muscle fiber and from their relationship with the nerve terminal. As degeneration proceeds, this allows a greater area of contact between each nerve terminal and the muscle fiber. Within each junction there is a mixture of both functional and non-functional regions and active zones, as determined by both thin-section and freeze-fracture observations. No correlation was found between the degree of degeneration of a neuromuscular junction and its association with a particular muscle fiber or its position on the fiber relative to the origin or insertion.

Degenerative changes in the function of neuromuscular junctions of Manduca sexta during metamorphosis

1992 ◽  
Vol 167 (1) ◽  
pp. 61-89
Author(s):  
I. M. Sonea ◽  
M. B. Rheuben
Keyword(s):  
Manduca Sexta ◽  
Muscle Fiber ◽  
Neuromuscular Junctions ◽  
Motor Nerve ◽  
Normal Function ◽  
Degenerative Changes ◽  
Resting Potential ◽  
Morphological Evidence ◽  
Low Frequencies ◽  
Cable Properties

In Manduca sexta the decline in neuromuscular function during metamorphic degeneration was compared in two muscles which differed characteristically with regard to pre- and postsynaptic physiological properties. In both muscles, morphological evidence indicated that a significant number of the active zones within the population of neuromuscular junctions on a given fiber were nonfunctional. Nevertheless, the degenerating nerve terminals were able to produce an above-threshold excitatory junction potential (EJP) which was facilitated in a manner characteristic of the muscle being observed. Abnormal findings during the early stages of degeneration included a larger than normal EJP, a decline in EJP amplitude over a 20 min period even with low frequencies of stimulation, an increase in EJP duration, a decline in muscle fiber resting potential amplitude with age, a decrease or disappearance of post-tetanic potentiation and long-term facilitation, and an increased likelihood that the motor nerve would fail to conduct a stimulus. The two muscles were qualitatively similar but quantitatively different with regard to these degenerative changes. It is suggested that this combination of relatively normal function with abnormal properties might be associated with the withdrawal of glial processes from the neuromuscular junctions, changes in the cable properties associated with shrivelling of the muscle fibers, and a decline in the metabolic functions supporting both muscle fiber resting potentials and those underlying transmitter synthesis, mobilization and release.

scholarly journals Endocytosis of synaptic vesicle membrane at the frog neuromuscular junction.

1984 ◽  
Vol 98 (2) ◽  
pp. 685-698 ◽  
Author(s):  
T M Miller ◽  
J E Heuser
Keyword(s):  
Plasma Membrane ◽  
Synaptic Vesicle ◽  
Thin Section ◽  
Nerve Terminal ◽  
Motor Nerve ◽  
Freeze Fracture ◽  
Frog Neuromuscular Junction ◽  
Vesicle Membrane ◽  
Coated Pits ◽  
Active Zones

Frog nerve-muscle preparations were quick-frozen at various times after a single electrical stimulus in the presence of 4-aminopyridine (4-AP), after which motor nerve terminals were visualized by freeze-fracture. Previous studies have shown that such stimulation causes prompt discharge of 3,000-6,000 synaptic vesicles from each nerve terminal and, as a result, adds a large amount of synaptic vesicle membrane to its plasmalemma. In the current experiments, we sought to visualize the endocytic retrieval of this vesicle membrane back into the terminal, during the interval between 1 s and 2 min after stimulation. Two distinct types of endocytosis were observed. The first appeared to be rapid and nonselective. Within the first few seconds after stimulation, relatively large vacuoles (approximately 0.1 micron) pinched off from the plasma membrane, both near to and far away from the active zones. Previous thin-section studies have shown that such vacuoles are not coated with clathrin at any stage during their formation. The second endocytic process was slower and appeared to be selective, because it internalized large intramembrane particles. This process was manifest first by the formation of relatively small (approximately 0.05 micron) indentations in the plasma membrane, which occurred everywhere except at the active zones. These indentations first appeared at 1 s, reached a peak abundance of 5.5/micron2 by 30 s after the stimulus, and disappeared almost completely by 90 s. Previous thin-section studies indicate that these indentations correspond to clathrin-coated pits. Their total abundance is comparable with the number of vesicles that were discharged initially. These endocytic structures could be classified into four intermediate forms, whose relative abundance over time suggests that, at this type of nerve terminal, endocytosis of coated vesicles has the following characteristics: (a) the single endocytotic event is short lived relative to the time scale of two minutes; (b) earlier forms last longer than later forms; and (c) a single event spends a smaller portion of its lifetime in the flat configuration soon after the stimulus than it does later on.

scholarly journals Freeze-fracture studies of frog neuromuscular junctions during intense release of neurotransmitter. II. Effects of electrical stimulation and high potassium.

1979 ◽  
Vol 81 (1) ◽  
pp. 178-192 ◽  
Author(s):  
B Ceccarelli ◽  
F Grohovaz ◽  
W P Hurlbut
Keyword(s):  
Nerve Terminal ◽  
Neuromuscular Junctions ◽  
Freeze Fracture ◽  
Presynaptic Membrane ◽  
High Potassium ◽  
Unique Distribution ◽  
Active Zones ◽  
Nerve Muscle ◽  
Indirect Stimulation ◽  
Quantal Release Of Neurotransmitter

Frog cutaneous pectoris nerve muscle preparations were studied by the freeze-fracture technique under the following conditions: (a) during repetitive indirect stimulation for 20 min, 10/s; (b) during recovery from this stimulation; and (c) during treatment with 20 mM K+. Indirect stimulation causes numerous dimples or protuberances to appear on the presynaptic membrane of nerve terminal, and most are located near the active zones. Deep infoldings of the axolemma often develop between the active zones. Neither the number nor the distribution of dimples, protuberances, of infoldings changes markedly during the first minute of recovery. The number of dimples, protuberances, and infoldings is greatly reduced after 10 min of recovery. Since endocytosis proceeds vigorously during the recovery periods, we conclude that endocytosis occurs mostly at the active zones, close to the sites of exocytosis. 20 mM K+ also causes many dimples or protuberances to appear on the axolemma of the nerve terminal but they are distributed almost uniformly along the presynaptic membrane. Experiments with horseradish peroxidase (HRP) show that recycling of synaptic vesicles occurs in 20 mM K+. This recycling is not accompanied by changes in the number of coated vesicles. Since both exocytosis and endocytosis occur in 20 mM K+, it is difficult to account for this unique distribution. However, we suggest that K+ causes dimples or protuberances to appear between the active zones because it activates latent sites of exocytosis specified by small numbers of large intramembrane particles located between active zones. The activation of latent release sites may be related to the complex effects that K+ has on the quantal release of neurotransmitter.

scholarly journals Glutamate at the Vertebrate Neuromuscular Junction: From Modulation to Neurotransmission

Cells ◽  
2019 ◽  
Vol 8 (9) ◽  
pp. 996 ◽  
Author(s):  
Colombo ◽  
Francolini
Keyword(s):  
Neuromuscular Junction ◽  
Signaling Pathways ◽  
Muscle Fiber ◽  
Neuromuscular Junctions ◽  
Motor Nerve ◽  
Vertebrate Development ◽  
Motor Nerve Ending ◽  
Downstream Signaling ◽  
Synaptic Neurotransmission ◽  
Synaptic Receptors

Although acetylcholine is the major neurotransmitter operating at the skeletal neuromuscular junction of many invertebrates and of vertebrates, glutamate participates in modulating cholinergic transmission and plastic changes in the last. Presynaptic terminals of neuromuscular junctions contain and release glutamate that contribute to the regulation of synaptic neurotransmission through its interaction with pre- and post-synaptic receptors activating downstream signaling pathways that tune synaptic efficacy and plasticity. During vertebrate development, the chemical nature of the neurotransmitter at the vertebrate neuromuscular junction can be experimentally shifted from acetylcholine to other mediators (including glutamate) through the modulation of calcium dynamics in motoneurons and, when the neurotransmitter changes, the muscle fiber expresses and assembles new receptors to match the nature of the new mediator. Finally, in adult rodents, by diverting descending spinal glutamatergic axons to a denervated muscle, a functional reinnervation can be achieved with the formation of new neuromuscular junctions that use glutamate as neurotransmitter and express ionotropic glutamate receptors and other markers of central glutamatergic synapses. Here, we summarize the past and recent experimental evidences in support of a role of glutamate as a mediator at the synapse between the motor nerve ending and the skeletal muscle fiber, focusing on the molecules and signaling pathways that are present and activated by glutamate at the vertebrate neuromuscular junction.

Degenerative changes in the muscle fibers of Manduca sexta during metamorphosis

1992 ◽  
Vol 167 (1) ◽  
pp. 91-117 ◽  
Author(s):  
M. B. Rheuben
Keyword(s):  
Manduca Sexta ◽  
Basal Lamina ◽  
Muscle Fiber ◽  
Structural Changes ◽  
Muscle Fibers ◽  
Resting Potential ◽  
Ultrastructural Changes ◽  
Intimate Contact ◽  
Cross Sectional ◽  
Dense Granules

The ultrastructural changes associated with the early stages of degeneration of the larval mesothoracic muscle fibers of Manduca sexta were examined during the prepupal period and on the first day after ecdysis. Over this 5 day period, the muscle fibers decrease in cross-sectional area but increase in apparent surface area compared to the dimensions of early fifth-instar fibers. Large numbers of electron-dense granules or droplets are formed and extruded from the muscle cytoplasm into the hemolymph; this process may account for some of the decrease in muscle fiber mass and may represent a developmental mechanism for recycling nutrients. As the fibers shrink, the thick basal lamina is thrown into folds. Phagocytic hemocytes (granulocytes) congregate in clusters over the surface of the degenerating fibers and appear to remove specifically the basal lamina. The timely removal of the thick larval basal lamina may be essential for subsequent fusion of myoblasts to the residual larval myofibers. The contractile elements within the degenerating muscle fibers become disorganized but are not dysfunctional at the end of the first 12 h after the pupal ecdysis. Tracheoles withdraw from intimate contact with each muscle fiber in its clefts and T-tubules and associate in groups adjacent to it. Mitochondria appear to be degenerating. These structural changes are concurrent with a previously observed decline in resting potential and suggest that a significant change in the electrical properties of the muscle fibers should be expected as well.

Quantal Secretion and Nerve-Terminal Cable Properties at Neuromuscular Junctions in an Amphibian (Bufo marinus)

1999 ◽  
Vol 81 (3) ◽  
pp. 1135-1146 ◽  
Author(s):  
G. T. Macleod ◽  
L. Farnell ◽  
W. G. Gibson ◽  
M. R. Bennett
Keyword(s):  
Nerve Terminal ◽  
Neuromuscular Junctions ◽  
Motor Nerve ◽  
Current Injection ◽  
Bufo Marinus ◽  
Motor Nerve Terminal ◽  
Terminal Branch ◽  
Quantal Release ◽  
Cable Properties ◽  
Test Electrode

Quantal secretion and nerve-terminal cable properties at neuromuscular junctions in an amphibian ( Bufo marinus). The effect of a conditioning depolarizing current pulse (80–200 μs) on quantal secretion evoked by a similar test pulse at another site was examined in visualized motor-nerve terminal branches of amphibian endplates ( Bufo marinus). Tetrodotoxin (200 nM) and cadmium (50 μM) were used to block voltage-dependent sodium and calcium conductances. Quantal release at the test electrode was depressed at different distances (28–135 μm) from the conditioning electrode when the conditioning and test pulses were delivered simultaneously. This depression decreased when the interval between conditioning and test current pulses was increased, until, at an interval of ∼0.25 ms, it was negligible. At no time during several thousand test-conditioning pairs, for electrodes at different distances apart (28–135 μm) on the same or contiguous terminal branches, did the electrotonic effects of quantal release at one electrode produce quantal release at the other. Analytic and numerical solutions were obtained for the distribution of transmembrane potential at different sites along terminal branches of different lengths for current injection at a point on a terminal branch wrapped in Schwann cell, in the absence of active membrane conductances. Solutions were also obtained for the combined effects of two sites of current injection separated by different time delays. This cable model shows that depolarizing current injections of a few hundred microseconds duration produce hyperpolarizations at ∼30 μm beyond the site of current injection, with these becoming larger and occurring at shorter distances the shorter the terminal branch. Thus the effect of a conditioning depolarizing pulse at one site on a subsequent test pulse at another more than ∼30 μm away is to substantially decrease the absolute depolarization produced by the latter, provided the interval between the pulses is less than a few hundred microseconds. It is concluded that the passive cable properties of motor nerve terminal branches are sufficient to explain the effects on quantal secretion by a test electrode depolarization of current injections from a spatially removed conditioning electrode.

Focal adhesions to the synaptic matrix at mouse neuromuscular junctions

1989 ◽  
Vol 47 ◽  
pp. 944-945
Author(s):  
Norman Robbins ◽  
Joseph Polak
Keyword(s):  
Soleus Muscle ◽  
Nerve Terminal ◽  
Neuromuscular Junctions ◽  
Motor Nerve ◽  
Focal Adhesions ◽  
Polyclonal Antiserum ◽  
Motor Nerve Terminal ◽  
Synaptic Contacts ◽  
The Matrix ◽  
Synaptic Remodelling

Studies of identified synapses in living animals have revealed ongoing synaptic remodelling in the mature nervous system. We recently demonstrated the existence of filopodia and lamellipodia (structures associated with nerve outgrowth in developing or tissue culture systems) at the mature mouse neuromuscular junction (NMJ) . Evidence of motor nerve terminal retraction, regeneration and perisynaptic outgrowth indicated that even established synaptic contacts were not permanent. It now appears that variation in the stability of synaptic contacts, not the amount of nerve terminal outgrowth,is the basis for synaptic remodelling and extension. Although it has been documented that regenerating motor nerve terminals adhere to former synaptic matrix,and that the matrix contains information for inducing and aligning presynaptic and postsynaptic specializations,it was not clear whether the synaptic matrix was uniformly adhesive, or formed specialized adhesive foci with the preterminal membrane. To determine if there are specific sites of synaptic adhesion we utilized hypertonic fixatives to induce shrinkage at the NMJ.Male mice were anaesthetized, the soleus muscle was exposed, and the overlying skin was retracted to create a pool into which either 2% glutaraldehyde in HEPES Krebs (normal fixative) or 2% glutaraldehyde in HEPES Krebs with a 2X NaCl concentration (hypertonic fixative) was added continually for 30 minutes. The surface layer of each muscle was then removed and submerged in the same fixative for an additional 1.5 hours. Tissue was washed in buffer (pH 7.2) and immersed in cholinesterase stain for 5 minutes to localize endplate regions . Endplate regions were cut into 1 mm blocks, washed in buffer (pH 7.2) and post-fixed for 1 hour in HEPES Krebs buffered 2% osmium tetroxide (pH 7.2). Following fixation, specimens were dehydrated in a graded ethanol series and embedded in Polybed 812 (Polysciences). Mouse soleus muscle was stained for immunocytochemical localization of actin with a rabbit polyclonal antiserum against chicken gizzard actin (from Dr. James Lessard, University of Cincinnati) as previously described .

scholarly journals Low-frequency Neuromuscular Depression Is a Consequence of a Reduction in Nerve Terminal Ca2+ Currents at Mammalian Motor Nerve Endings

2013 ◽  
Vol 119 (2) ◽  
pp. 326-334
Author(s):  
Eugene M. Silinsky
Keyword(s):  
Nerve Terminal ◽  
Nerve Stimulation ◽  
Neurotransmitter Release ◽  
Neuromuscular Junctions ◽  
Motor Nerve ◽  
Neuromuscular Block ◽  
Low Frequency ◽  
Neuromuscular Blocking ◽  
Train Of Four ◽  
Train Of Four Monitoring

Abstract Background: The decline in voluntary muscle contraction during low-frequency nerve stimulation is used clinically to assess the type and degree of neuromuscular block. The mechanism underlying this depression is unknown. Methods: Simultaneous electrophysiological measurements of neurotransmitter release and prejunctional Ca2+ currents were made at mouse neuromuscular junctions to evaluate the hypothesis that decreases in nerve terminal Ca2+ currents are responsible for low-frequency depression. Results: Under conditions generally used to measure Ca2+ currents at the neuromuscular junction, increasing the frequency of nerve stimulation briefly from 0.017 to 0.1–1 Hz caused a simultaneous reduction in the release of the neurotransmitter acetylcholine to 52.2 ± 4.4% of control and the Ca2+ current peak to 75.4 ± 2.0% of control (P < 0.001, n = 5 experiments for both measurements, mean ± SEM for all data). In conditions used for train-of-four monitoring (4 stimuli, 2 Hz), neurotransmitter release declined to 42.0 ± 1.0% of control and the Ca2+ current peak declined to 75.8 ± 3.3% of control between the first and fourth stimulus (P < 0.001, n = 7 experiments for both measurements). Depression in acetylcholine release during train-of-four protocols also occurred in the absence of neuromuscular-blocking drugs. Discussion: The results demonstrate that neuromuscular depression during train-of-four monitoring is due to a decline in nerve terminal Ca2+ currents, hence reducing the release of acetylcholine. As similar processes may come into play at higher stimulation frequencies, agents that antagonize the decline in Ca2+ currents could be used to treat conditions in which neuromuscular depression can be debilitating.

How to sample the entire length of a single muscle fiber quick-frozen after electrical point stimulation for high resolution EM

1992 ◽  
Vol 50 (1) ◽  
pp. 776-777
Author(s):  
Joachim R. Sommer ◽  
Teresa High ◽  
Betty Scherer ◽  
Isaiah Taylor ◽  
Rashid Nassar
Keyword(s):  
Skeletal Muscle ◽  
High Resolution ◽  
Action Potential ◽  
Muscle Fiber ◽  
Freeze Fracture ◽  
Freeze Substitution ◽  
Electrical Field Stimulation ◽  
Skeletal Muscle Fiber ◽  
Freeze Dried ◽  
Quick Freezing

We have developed a model that allows the quick-freezing at known time intervals following electrical field stimulation of a single, intact frog skeletal muscle fiber isolated by sharp dissection. The preparation is used for studying high resolution morphology by freeze-substitution and freeze-fracture and for electron probe x-ray microanlysis of sudden calcium displacement from intracellular stores in freeze-dried cryosections, all in the same fiber. We now show the feasibility and instrumentation of new methodology for stimulating a single, intact skeletal muscle fiber at a point resulting in the propagation of an action potential, followed by quick-freezing with sub-millisecond temporal resolution after electrical stimulation, followed by multiple sampling of the frozen muscle fiber for freeze-substitution, freeze-fracture (not shown) and cryosectionmg. This model, at once serving as its own control and obviating consideration of variances between different fibers, frogs etc., is useful to investigate structural and topochemical alterations occurring in the wake of an action potential.

Gap junctions in the prothoracic glands of the tobacco hornworm, Manduca sexta

1992 ◽  
Vol 50 (1) ◽  
pp. 706-707
Author(s):  
Ji-da Dai ◽  
M. Joseph Costello ◽  
Lawrence I. Gilbert
Keyword(s):  
Gap Junctions ◽  
Small Molecules ◽  
Manduca Sexta ◽  
Freeze Fracture ◽  
Cell Communication ◽  
Cellular Level ◽  
Thin Sections ◽  
Prothoracic Glands ◽  
Polyhydroxylated Steroids ◽  
Stimulation Of

Insect molting and metamorphosis are elicited by a class of polyhydroxylated steroids, ecdysteroids, that originate in the prothoracic glands (PGs). Prothoracicotropic hormone stimulation of steroidogenesis by the PGs at the cellular level involves both calcium and cAMP. Cell-to-cell communication mediated by gap junctions may play a key role in regulating signal transduction by controlling the transmission of small molecules and ions between adjacent cells. This is the first report of gap junctions in the PGs, the evidence obtained by means of SEM, thin sections and freeze-fracture replicas.

Sign in / Sign up

Export Citation Format

Share Document