Quantal Release of Neurotransmitter and Evidence for Vesicular Release
Safina Rehman; Kynzie Lalliss; and Jim Hutchins
This is a first draft which has not been edited. If you have questions, or want to help in the writing or editing process, please contact the authors: safinarehman@mail.weber.edu or kynzielalliss@mail.weber.edu or jimhutchins@weber.edu
The Concept of a Synapse
Sherrington coined the name “synapse”
Dale and neurotransmitters
Loewi and Vagusstoff
Chemical Synapse
At the presynaptic terminal of a chemical synapse, voltage gated calcium channels open as a result of an action potential arriving after traveling down the neuron. Since there is no calcium in the cell, diffusion makes for the positively charged calcium outside to flow in. After calcium enters the presynaptic terminal, a process happens where neurotransmitters are released from synaptic vesicles which are tiny sac-like organelles that have a lipid bilayer membrane like cells do. This process occurs after calcium binds to SNARE proteins, which being attached to both membranes, twists and thereby fuses the two membranes together to the extent that the vesicle opens up and dumps the neurotransmitters all at once into the synaptic cleft.
Electron microscopy of the synapse (we have vesicles, but what are they for?)
Hypothesis that vesicles contain neurotransmitter and that neurotransmitter is released in same-size “bundles” called quanta
The Neuromuscular Junction
Circa 1950 physiologists knew that there were end-plate potentials in muscles (postsynaptic) at the NMJ
End-plate potentials are the NMJ equivalent of an EPSP
Cras eros ligula, pellentesque vitae rhoncus vitae, posuere ac lectus. Cras et ante vel urna facilisis sodales nec a justo. Aliquam vitae neque dui. Integer posuere imperdiet sem vitae vestibulum. Pellentesque quis metus sed mi semper dapibus at id magna. Donec augue sapien, eleifend eget blandit eget, ultrices sit amet leo. Nam sed pellentesque libero. Etiam pulvinar rutrum mattis. Vestibulum in aliquet eros. Sed ut risus ullamcorper, ullamcorper eros non, suscipit odio. Duis molestie, risus id suscipit semper, libero leo pharetra nibh, in dignissim libero metus a augue. Donec id augue ut velit semper rutrum. Praesent sem ligula, eleifend et sem a, vehicula efficitur felis. Ut elementum tempus sem. Praesent mollis placerat lacus, vel posuere mi cursus eget.
Donec pretium magna venenatis molestie tristique. Mauris placerat velit id felis blandit luctus. Etiam ac volutpat risus, at viverra dui. Aliquam erat volutpat. Cras vel arcu a sem dignissim tempus. Etiam sed scelerisque tellus. Sed luctus est facilisis risus tempus, non facilisis augue tincidunt. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos.
Proin mollis, magna in luctus gravida, metus dui feugiat lorem, nec convallis risus felis vel mi. Maecenas rhoncus volutpat tristique. Vestibulum ut sem non est finibus mattis. Suspendisse ultrices, erat vel placerat ullamcorper, enim lectus ornare eros, id tristique tortor justo sed magna. Curabitur mattis tristique elementum. Maecenas fringilla elit quis magna sollicitudin, eget tempus neque placerat. In volutpat metus in euismod elementum. Etiam faucibus ornare nunc sed auctor. Nullam id ultrices felis. Morbi finibus sapien vitae laoreet viverra. Integer sollicitudin est et nisi hendrerit pulvinar. Pellentesque habitant morbi tristique senectus et netus et malesuada fames ac turpis egestas.
Cras eros ligula, pellentesque vitae rhoncus vitae, posuere ac lectus. Cras et ante vel urna facilisis sodales nec a justo. Aliquam vitae neque dui. Integer posuere imperdiet sem vitae vestibulum. Pellentesque quis metus sed mi semper dapibus at id magna. Donec augue sapien, eleifend eget blandit eget, ultrices sit amet leo. Nam sed pellentesque libero. Etiam pulvinar rutrum mattis. Vestibulum in aliquet eros. Sed ut risus ullamcorper, ullamcorper eros non, suscipit odio. Duis molestie, risus id suscipit semper, libero leo pharetra nibh, in dignissim libero metus a augue. Donec id augue ut velit semper rutrum. Praesent sem ligula, eleifend et sem a, vehicula efficitur felis. Ut elementum tempus sem. Praesent mollis placerat lacus, vel posuere mi cursus eget.
Donec pretium magna venenatis molestie tristique. Mauris placerat velit id felis blandit luctus. Etiam ac volutpat risus, at viverra dui. Aliquam erat volutpat. Cras vel arcu a sem dignissim tempus. Etiam sed scelerisque tellus. Sed luctus est facilisis risus tempus, non facilisis augue tincidunt. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos.
Proin mollis, magna in luctus gravida, metus dui feugiat lorem, nec convallis risus felis vel mi. Maecenas rhoncus volutpat tristique. Vestibulum ut sem non est finibus mattis. Suspendisse ultrices, erat vel placerat ullamcorper, enim lectus ornare eros, id tristique tortor justo sed magna. Curabitur mattis tristique elementum. Maecenas fringilla elit quis magna sollicitudin, eget tempus neque placerat. In volutpat metus in euismod elementum. Etiam faucibus ornare nunc sed auctor. Nullam id ultrices felis. Morbi finibus sapien vitae laoreet viverra. Integer sollicitudin est et nisi hendrerit pulvinar. Pellentesque habitant morbi tristique senectus et netus et malesuada fames ac turpis egestas.
Vesicles
Exocytosis is the natural process through which molecules are transported from within a cell to the outside space of it. There are multiple different forms of exocytosis, but this chapter will focus on vesicular exocytosis. Vesicular exocytosis occurs when vesicles fuse with a cell’s plasma membrane to release their contents outside the cell. It is a form of active transport that allows a cell to move large molecules from within the cell to the exterior. This type of exocytosis is only found in bacteria and occurs through the involvement of lysosomes that help in membrane fusion.
A vesicle is a division within a cell that is formed by a lipid bilayer that separates its contents from the cell’s cytoplasm or extracellular fluid. These form naturally during processes of exocytosis, endocytosis, and the transportation of materials in the plasma membrane. Vesicles perform several different functions, mainly transporting materials and recycling waste. Additionally, they also absorb and dispose of toxic substances and pathogens within the cell to prevent cell damage and/or death.
Lorem ipsum dolor sit amet, consectetur adipiscing elit. Nam semper dui eget gravida varius. Ut varius risus nunc, in varius urna fermentum a. Sed risus odio, ullamcorper sit amet faucibus nec, condimentum sit amet arcu. Cras elit sem, semper et aliquet fringilla, dictum nec nibh. Interdum et malesuada fames ac ante ipsum primis in faucibus. Mauris vel nisl pretium, varius lorem cursus, cursus nulla. In eu nisi non augue bibendum posuere. Proin mollis eleifend mi, at vestibulum ipsum eleifend nec. Fusce id massa quis orci sollicitudin pulvinar. Donec ante dui, scelerisque sit amet velit et, iaculis pharetra orci. Morbi lobortis purus neque, et lacinia ipsum aliquam at.
Suspendisse auctor, sapien eget ultrices auctor, velit leo blandit tellus, iaculis volutpat felis neque vitae augue. Aenean eu enim in odio consectetur tristique. Sed massa est, condimentum et dui at, ultricies iaculis leo. Praesent nulla metus, pulvinar vehicula ex nec, dictum tempor libero. Nunc accumsan et quam sed porttitor. Sed risus ipsum, vestibulum a quam et, ultrices rutrum ex. Duis non felis eget nunc luctus suscipit non in massa. Etiam dictum eget ipsum ut pellentesque. In nec augue ac tellus finibus vestibulum. Suspendisse sit amet nulla erat. Aliquam interdum aliquet nulla, non sagittis felis rutrum a. Donec quis dolor mauris. Suspendisse at orci non urna cursus scelerisque. Duis in vehicula nibh. Aliquam mollis bibendum suscipit. Etiam posuere neque et enim vehicula, et commodo eros pretium.
Cras eros ligula, pellentesque vitae rhoncus vitae, posuere ac lectus. Cras et ante vel urna facilisis sodales nec a justo. Aliquam vitae neque dui. Integer posuere imperdiet sem vitae vestibulum. Pellentesque quis metus sed mi semper dapibus at id magna. Donec augue sapien, eleifend eget blandit eget, ultrices sit amet leo. Nam sed pellentesque libero. Etiam pulvinar rutrum mattis. Vestibulum in aliquet eros. Sed ut risus ullamcorper, ullamcorper eros non, suscipit odio. Duis molestie, risus id suscipit semper, libero leo pharetra nibh, in dignissim libero metus a augue. Donec id augue ut velit semper rutrum. Praesent sem ligula, eleifend et sem a, vehicula efficitur felis. Ut elementum tempus sem. Praesent mollis placerat lacus, vel posuere mi cursus eget.
Donec pretium magna venenatis molestie tristique. Mauris placerat velit id felis blandit luctus. Etiam ac volutpat risus, at viverra dui. Aliquam erat volutpat. Cras vel arcu a sem dignissim tempus. Etiam sed scelerisque tellus. Sed luctus est facilisis risus tempus, non facilisis augue tincidunt. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos.
Proin mollis, magna in luctus gravida, metus dui feugiat lorem, nec convallis risus felis vel mi. Maecenas rhoncus volutpat tristique. Vestibulum ut sem non est finibus mattis. Suspendisse ultrices, erat vel placerat ullamcorper, enim lectus ornare eros, id tristique tortor justo sed magna. Curabitur mattis tristique elementum. Maecenas fringilla elit quis magna sollicitudin, eget tempus neque placerat. In volutpat metus in euismod elementum. Etiam faucibus ornare nunc sed auctor. Nullam id ultrices felis. Morbi finibus sapien vitae laoreet viverra. Integer sollicitudin est et nisi hendrerit pulvinar. Pellentesque habitant morbi tristique senectus et netus et malesuada fames ac turpis egestas.
The presence of calcium ions (Ca2+) is important for exocytosis to occur. Several experiments have shown that an increase in extracellular or manipulation of intracellular calcium levels affects exocytosis, either inhibiting or promoting it. Studies using calcium chelators or channel blockers also provides evidence for vesicular exocytosis and its dependency on calcium ion levels.
Patch-clamp and Electrophysiological Studies
Patch-clamp and electrophysiological experiments in neurons have provided data that there are rapid changes in membrane potential that are associated with vesicle fusion and release. This study technique allows researchers to study ionic currents in cells, tissues, and patches of cell membrane, and directly measures membrane potentials. The dynamic nature of exocytosis is revealed with these studies, as well as evidence for the quantal release of neurotransmitters.
SNARE (soluble NSF attachments protein receptor) proteins, such as syntaxin, SNAP-25, and VAMP, are essential for the process of exocytosis and provide support for the concept on a molecular level. These proteins are the mediators for vesicle docking and fusion with a cell’s plasma membrane. When these proteins are tampered with and their function is disrupted, exocytosis is affected and inhibited.
The Quantal Hypothesis
In 1950 Bernard Katz formulated the quantal hypothesis, which is essential to our current understanding of how neurotransmitters are released from the presynaptic terminal. In this hypothesis, Katz proposed that neurotransmitters are released from presynaptic terminals not in a continuous flow, but in separate packets or “quanta”.
https://link.springer.com/referenceworkentry/10.1007/978-3-540-29678-2_2867
How do we know this?
Katz used a frog neuromuscular junction, where he was specifically looking at synapses used for motor neurons communicating with muscle fibers, to observe changes in electrical potential of the postsynaptic neuron. The changes to the postsynaptic neuron are called EPP (End Plate Potential), which represent the change in electrical potential of the post synaptic membrane as a result. He then reduced the extracellular calcium and stimulated the presynaptic cell by injecting a bit of current and just waited for spontaneous.
del Castillo and Katz 1954
https://link.springer.com/referenceworkentry/10.1007/978-3-540-29678-2_2867
https://pmc.ncbi.nlm.nih.gov/articles/PMC1363540/?page=10
https://pubmed.ncbi.nlm.nih.gov/38256/
Capturing Vesicles Mid-Fusion
Studies have found that vesicular exocytosis involves the formation of a temporary fusion pore. These pores span the membrane of both the vesicle and the plasma membranes and help to complete the fusion of the vesicle and cell membranes. Materials in a vesicle can also be released through these pores before the vesicle experiences full fusion with the membrane.
https://www.researchgate.net/publication/18816723_Evidence_for_Recycling_of_Synaptic_Vesicle_Membrane_During_Transmitter_Release_at_the_Frog_Neuromuscular_Junction/link/0deec51b632d2c1ce0000000/download
Specializations Associated with Vesicles
SNARE proteins
Mutations in these proteins cause alterations in synaptic release
? Low-calcium solutions slow the rate of neurotransmitter
Ribbon synapses
Electron Microscopy (EM) is a technique used to obtain high resolution images of biological and non-biological specimens. Researchers use this method to observe cells and their complex structures, utilizing electrons as the source of illumination to produce these detailed images. During studies of vesicles, observations made with EM provides direct evidence of vesicular exocytosis, as it shows detailed visualization of vesicles fusing with the cell membrane and releasing its contents into the extracellular space.
Lorem ipsum dolor sit amet, consectetur adipiscing elit. Proin tincidunt elit libero, a semper lectus porta at. Quisque et libero finibus, sagittis mauris in, auctor metus. Vivamus faucibus leo nunc, ac dignissim ligula tempor a. Vestibulum ligula leo, faucibus et justo ut, rhoncus accumsan neque. Nunc mollis quam ut interdum iaculis. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos. Morbi laoreet commodo est, vel fermentum sapien auctor eget. Praesent turpis metus, convallis consectetur felis quis, vulputate scelerisque turpis. Quisque eget vestibulum risus, in ultrices orci. Nulla vehicula mi vitae leo porttitor congue. Quisque finibus luctus tortor non efficitur. Fusce sit amet eros ex.
Maecenas elit tellus, consectetur quis purus vel, facilisis euismod arcu. Orci varius natoque penatibus et magnis dis parturient montes, nascetur ridiculus mus. Donec vitae pellentesque magna, id efficitur erat. Etiam eget nulla eget urna fringilla luctus dictum ac dolor. Curabitur eu justo congue, blandit velit auctor, luctus eros. Etiam vel risus et eros semper mollis. Curabitur non ultrices urna, quis viverra eros. Proin purus nibh, porttitor rhoncus ornare eget, interdum in lorem. Cras euismod non neque in volutpat. Morbi vel egestas odio, a pulvinar mi. Fusce in ligula quam. Fusce maximus lorem arcu, ut tempus leo ultrices vitae. Phasellus sit amet luctus velit. Proin rhoncus porttitor enim, non ullamcorper ex varius efficitur. Nullam aliquet porttitor convallis. Fusce tristique metus ex, dignissim pulvinar urna ornare placerat.
Praesent dapibus fringilla ante, et dapibus sapien porta et. Aliquam erat volutpat. Phasellus porttitor nulla in purus ornare, in sagittis lorem tincidunt. Ut dapibus elit ante, in consectetur odio scelerisque et. Suspendisse varius ligula mi, sed egestas turpis interdum ut. Nulla lorem nibh, feugiat non fermentum vitae, sodales id nulla. Donec rutrum luctus lectus, eget venenatis tellus aliquet non. Maecenas id dignissim mi. Vivamus imperdiet molestie ex, sed posuere ex. Integer mattis hendrerit bibendum. Nulla hendrerit tincidunt arcu sed tincidunt. Phasellus feugiat a nisi eget sodales. Suspendisse in pharetra odio, quis posuere magna. Nam pulvinar tincidunt tempor. Nam posuere est sit amet mi rhoncus mattis.
Nam in eros non libero rutrum malesuada. Vivamus id ex purus. Duis nec pellentesque metus. Curabitur iaculis tempus dolor a vestibulum. Sed rhoncus lectus tristique turpis vulputate varius. Mauris sagittis ipsum vel ipsum pulvinar tristique. Etiam elit tellus, rutrum eu est vel, condimentum mollis felis. Mauris ornare eros eu lorem imperdiet lacinia. Suspendisse arcu orci, sollicitudin vel nisl sed, tempus rhoncus augue. Nulla maximus sed ipsum ac bibendum. Ut posuere aliquet elit id bibendum. In suscipit vehicula ex ut interdum. Morbi mollis nisl venenatis lacus lacinia, sed dignissim enim vulputate. Integer aliquet libero eget dui vehicula rhoncus.
Maecenas quam neque, venenatis in dapibus vel, aliquam sit amet arcu. Quisque eu tellus vehicula quam euismod aliquam. Quisque porttitor metus eget neque euismod tristique. Morbi aliquet lectus eget tortor convallis, a maximus elit tristique. Aenean mattis arcu vel nisi tempor, ut rutrum ante mollis. Nulla molestie euismod magna tristique faucibus. Quisque ultricies eros ac massa finibus egestas. Proin a aliquet nunc. Nunc a pretium urna. Pellentesque habitant morbi tristique senectus et netus et malesuada fames ac turpis egestas. In sodales felis a massa egestas finibus. Maecenas sed venenatis massa.
Radioactive Labeling
Researchers have used labeling techniques, specifically radioactive labeling, to trace the release of molecules during exocytosis. When using this type of imaging technique, researchers can see the tagged neurotransmitters moving between the inside of the vesicle space to the outside extracellular space. Further studies using fluorescent and radioactive tagging allows for real-time observation of both vesicle movement and fusion with the plasma membrane.
Electrochemical Measurements
Electrochemical measurements, specifically amperometry, have been used to study vesicular movement and exocytosis. Amperometry measures the electrical currents that are generated when neurotransmitters are released from vesicles during exocytosis. The measurement of these currents is found by using a carbon fiber electrode in a solution. It applies a constant potential to collect data on changes in electrical currents in a cell.
https://pmc.ncbi.nlm.nih.gov/articles/PMC5748430/
https://pmc.ncbi.nlm.nih.gov/articles/PMC3064517/
https://www.sciencedirect.com/science/article/pii/S0959438896801206
Measurement of Membrane Capacitance
Lorem ipsum dolor sit amet, consectetur adipiscing elit. Nam semper dui eget gravida varius. Ut varius risus nunc, in varius urna fermentum a. Sed risus odio, ullamcorper sit amet faucibus nec, condimentum sit amet arcu. Cras elit sem, semper et aliquet fringilla, dictum nec nibh. Interdum et malesuada fames ac ante ipsum primis in faucibus. Mauris vel nisl pretium, varius lorem cursus, cursus nulla. In eu nisi non augue bibendum posuere. Proin mollis eleifend mi, at vestibulum ipsum eleifend nec. Fusce id massa quis orci sollicitudin pulvinar. Donec ante dui, scelerisque sit amet velit et, iaculis pharetra orci. Morbi lobortis purus neque, et lacinia ipsum aliquam at.
Suspendisse auctor, sapien eget ultrices auctor, velit leo blandit tellus, iaculis volutpat felis neque vitae augue. Aenean eu enim in odio consectetur tristique. Sed massa est, condimentum et dui at, ultricies iaculis leo. Praesent nulla metus, pulvinar vehicula ex nec, dictum tempor libero. Nunc accumsan et quam sed porttitor. Sed risus ipsum, vestibulum a quam et, ultrices rutrum ex. Duis non felis eget nunc luctus suscipit non in massa. Etiam dictum eget ipsum ut pellentesque. In nec augue ac tellus finibus vestibulum. Suspendisse sit amet nulla erat. Aliquam interdum aliquet nulla, non sagittis felis rutrum a. Donec quis dolor mauris. Suspendisse at orci non urna cursus scelerisque. Duis in vehicula nibh. Aliquam mollis bibendum suscipit. Etiam posuere neque et enim vehicula, et commodo eros pretium.
Cras eros ligula, pellentesque vitae rhoncus vitae, posuere ac lectus. Cras et ante vel urna facilisis sodales nec a justo. Aliquam vitae neque dui. Integer posuere imperdiet sem vitae vestibulum. Pellentesque quis metus sed mi semper dapibus at id magna. Donec augue sapien, eleifend eget blandit eget, ultrices sit amet leo. Nam sed pellentesque libero. Etiam pulvinar rutrum mattis. Vestibulum in aliquet eros. Sed ut risus ullamcorper, ullamcorper eros non, suscipit odio. Duis molestie, risus id suscipit semper, libero leo pharetra nibh, in dignissim libero metus a augue. Donec id augue ut velit semper rutrum. Praesent sem ligula, eleifend et sem a, vehicula efficitur felis. Ut elementum tempus sem. Praesent mollis placerat lacus, vel posuere mi cursus eget.
Donec pretium magna venenatis molestie tristique. Mauris placerat velit id felis blandit luctus. Etiam ac volutpat risus, at viverra dui. Aliquam erat volutpat. Cras vel arcu a sem dignissim tempus. Etiam sed scelerisque tellus. Sed luctus est facilisis risus tempus, non facilisis augue tincidunt. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos.
Proin mollis, magna in luctus gravida, metus dui feugiat lorem, nec convallis risus felis vel mi. Maecenas rhoncus volutpat tristique. Vestibulum ut sem non est finibus mattis. Suspendisse ultrices, erat vel placerat ullamcorper, enim lectus ornare eros, id tristique tortor justo sed magna. Curabitur mattis tristique elementum. Maecenas fringilla elit quis magna sollicitudin, eget tempus neque placerat. In volutpat metus in euismod elementum. Etiam faucibus ornare nunc sed auctor. Nullam id ultrices felis. Morbi finibus sapien vitae laoreet viverra. Integer sollicitudin est et nisi hendrerit pulvinar. Pellentesque habitant morbi tristique senectus et netus et malesuada fames ac turpis egestas.