MINI REVIEW published: 30 April 2021 doi: 10.3389/fnmol.2021.638858 Extrasynaptic Communication Francisco F. De-Miguel 1,2 *, Carolina Leon-Pinzon 1 , Susana G. Torres-Platas 1 , Vanessa del-Pozo 1 , Guillermo A. Hernández-Mendoza 1 , Dilia Aguirre-Olivas 1 , Bruno Méndez 1 , Sharlen Moore 1 , Celeste Sánchez-Sugía 1 , Marco Antonio García-Aguilera 1 , Alejandro Martínez-Valencia 3 , Guillermo Ramírez-Santiago 4 and J. Miguel Rubí 5 1 Instituto de Fisiología Celular-Neurociencias, Universidad Nacional Autónoma de México, México City, Mexico, 2 Centro de Ciencias de la Complejidad, Universidad Nacional Autónoma de México, México City, Mexico, 3 Posgrado en Ciencias Físicas, Universidad Nacional Autónoma de México, México City, Mexico, 4 Instituto de Matemáticas, Universidad Nacional Autónoma de México, Juriquilla, Mexico, 5 Facultat de Fisica, Universitat de Barcelona, Barcelona, Spain Streams of action potentials or long depolarizations evoke a massive exocytosis of transmitters and peptides from the surface of dendrites, axons and cell bodies of different neuron types. Such mode of exocytosis is known as extrasynaptic for occurring without utilization of synaptic structures. Most transmitters and all peptides can be released extrasynaptically. Neurons may discharge their contents with relative independence from the axon, soma and dendrites. Extrasynaptic exocytosis takes fractions of a second in Edited by: varicosities or minutes in the soma or dendrites, but its effects last from seconds to Michele Papa, University of Campania Luigi hours. Unlike synaptic exocytosis, which is well localized, extrasynaptic exocytosis is Vanvitelli, Italy diffuse and affects neuronal circuits, glia and blood vessels. Molecules that are liberated Reviewed by: may reach extrasynaptic receptors microns away. The coupling between excitation Eckart D. Gundelfinger, and exocytosis follows a multistep mechanism, different from that at synapses, but Leibniz Institute for Neurobiology (LG), Germany similar to that for the release of hormones. The steps from excitation to exocytosis Jiefu Li, have been studied step by step for the vital transmitter serotonin in leech Retzius Howard Hughes Medical Institute (HHMI), United States neurons. The events leading to serotonin exocytosis occur similarly for the release *Correspondence: of other transmitters and peptides in central and peripheral neurons. Extrasynaptic Francisco F. De-Miguel exocytosis occurs commonly onto glial cells, which react by releasing the same or ffernand@ifc.unam.mx other transmitters. In the last section, we discuss how illumination of the retina evokes Received: 07 December 2020 extrasynaptic release of dopamine and ATP. Dopamine contributes to light-adaptation; Accepted: 01 April 2021 ATP activates glia, which mediates an increase in blood flow and oxygenation. A proper Published: 30 April 2021 understanding of the workings of the nervous system requires the understanding of Citation: extrasynaptic communication. De-Miguel FF, Leon-Pinzon C, Torres-Platas SG, del-Pozo V, Keywords: serotonin, extrasynaptic release, modulation, nerve cell communication, somatic exocytosis, glia Hernández-Mendoza GA, Aguirre-Olivas D, Méndez B, Moore S, Sánchez-Sugía C, INTRODUCTION García-Aguilera MA, Martínez-Valencia A, Our view of the workings of the nervous system have been dominated by four threads of Ramírez-Santiago G and Rubí JM (2021) Extrasynaptic Communication. fundamental evidence: First, Cajal defined nerve circuits as networks of neurons connected in Front. Mol. Neurosci. 14:638858. stereotyped manner, forming transmission lines for specific information processing. Second, doi: 10.3389/fnmol.2021.638858 physiologists such as Helmholtz, Hodgkin and Huxley showed that nerve impulses spread Frontiers in Molecular Neuroscience | www.frontiersin.org 1 April 2021 | Volume 14 | Article 638858 De-Miguel et al. Extrasynaptic Communication along axons at ∼300 km/h. Third, Sherrington, Katz, Kuffler the gain’’ of the circuitry for aggression by acting all along the and Eccles demonstrated that synapses transmit information neuronal circuit. in ∼0.5 ms. Fourth, plasticity adapts synaptic transmission to variations in the ongoing pattern of electrical activity. Such Cellular Basis of Extrasynaptic Exocytosis conceptual framework explains how a table tennis player detects Extrasynaptic exocytosis has been studied in central and the trajectory and velocity of a ball approaching at 50 Km/h and peripheral neurons. The similitude among the mechanism that in ∼0.3 ms and coordinates his whole-body motion to send it links excitation with exocytosis suggests a widely conserved back to an opposite corner of the table. In the games, such cycles mechanism, similar to that in gland cells but remarkably different may occur twice per second! from that for synaptic release (Sun and Poo, 1987; Huang et al., This review article deals with a parallel form of 2012; Hirasawa et al., 2015; Ludwig and Stern, 2015; Hökfelt et al., communication: streams of electrical impulses or long 2018; Quentin et al., 2018). Most transmitters and all peptides depolarizations promote massive liberation of signaling have been shown to be released extrasynaptically (Trueta and molecules from certain neurons. Release occurs without De-Miguel, 2012), and neurons may release more than one use of synaptic structures, therefore, it is named extrasynaptic. type of substance (Burnstock, 2012; Nusbaum et al., 2017; Molecules that are released extrasynaptically from the soma, Hökfelt et al., 2018). dendrites and axon modulate the responses of entire neuronal circuits from seconds to days (Trueta and De-Miguel, 2012). Extrasynaptic Exocytosis From Different Such form of communication may explain why a table tennis Neuronal Compartments player is defeated after being left by his fiancée. His reduced An evolutionary feature shared by neurons that release concentration, motivation and attention make him react poorly extrasynaptically is that small numbers innervate the nervous during the game. A hypothesis gaining increasing support is system extensively, and produce a wide variety of effects. For that ranges of physiological concentrations of extracellular example, ∼235,000 serotonergic neurons in humans (Baker signaling molecules modulate the responses of whole neuronal et al., 1990) project from the raphe nuclei to the entire central circuits; concentrations below or above produce pathologies nervous system. Neurons releasing catecholamines, acetylcholine (Calabresi et al., 2015; Del-Bel and De-Miguel, 2018; Pál, 2018; or peptides exist in similar small numbers (Zetler, 1970; Quentin et al., 2018). Mouton et al., 1994; Nair-Roberts et al., 2008; Li et al., Our focus here is exclusively on molecules that are released by 2018). Such extensive innervation is complemented by the exocytosis. Other substances such as nitric oxide or cannabinoids neuronal capability to release differentially from the soma, are released by diffusion across the plasma membrane (Del-Bel dendrites and axon. A well-known example is the release and De-Miguel, 2018); nucleic acids and proteins are released of the peptides vasopressin or oxytocin from magnocellular encapsulated inside vesicles that flow extracellularly (Colombo hypothalamic neurons (Ludwig and Stern, 2015). et al., 2014; Mendolesi, 2018). Those forms of release also follow The axons of magnocellular neurons bear rosaries of increases in electrical activity. varicosities that release extrasynaptically on the spread of action A good example as to how extrasynaptic exocytosis exerts potentials; their terminals discharge peptide onto the blood its effects comes from studies of aggression in lobsters by flow (Acher and Chauvet, 1954; Du Vigneaud, 1954). During Kravitz and his colleagues (Kravitz, 1988; Huber et al., 1997). lactation, suckling evokes oxytocin axonal release but dendritic An encounter between two lobsters triggers aggression. Lobsters release is delayed. However, dendritic release is locally evoked approximate to each other displaying their powerful claws and on activation of extrasynaptic NMDA receptors (de Kock et al., urinating on each other. The episodic encounters, initially lasting 2004; Tobin et al., 2012), as it also happens in dendrites of raphe seconds, decrease their strength and duration as one lobster neurons (Colgan et al., 2012). becomes dominant and the other submissive. The aggressive posture is evoked by a systemic injection of serotonin; an Discovery of Extrasynaptic injection of octopamine reproduces the submissive posture Communication (Livingstone et al., 1980). Serotonin that had been released extrasynaptically was The serotonergic A1 neurons in the abdominal ganglia discovered by Dalstrom and Fuxe in the 60s using the Falck- of lobsters innervate the ganglia and project branches to Hillarp technique, by which exposure to formaldehyde vapors the circulation. Command neurons that evoke tail flipping transforms the monoamines serotonin, dopamine or adrenaline during aggression evoke serotonin release from the A1 neurons. into fluorescent derivatives (Fuxe et al., 2007; Borroto-Escuela Serotonin that is released in the ganglia lowers the firing et al., 2015). Brain sections of raphe nuclei contained serotonin- threshold of central neurons; the serotonin discharged to the derived fluorescence surrounding the fluorescent cell bodies, circulation increases motoneuron transmission and strengthens distantly from the axonal release sites. Similar observations muscle contractions and heart beat (Glusman and Kravitz, made in dopaminergic neurons, plus the fact that peptides can 1982; Hörner et al., 1997; Hernández-Falcón et al., 2005). In be released far away from their receptors led to the concept of experiments in which serotonergic neurons were depleted of volume transmission by Fuxe and his colleagues, meaning that serotonin by systemic injection of 5–7 dihydroxytryptamine, molecules act on receptors located distantly from the release sites aggression still occurred. However, the strength and duration (Borroto-Escuela et al., 2015). It was later shown that axons of of the encounters lacked modulation. Therefore, serotonin ‘‘sets neurons releasing monoamines, acetylcholine, ATP and peptides Frontiers in Molecular Neuroscience | www.frontiersin.org 2 April 2021 | Volume 14 | Article 638858 De-Miguel et al. Extrasynaptic Communication bear vesicle arrangements but scarce presynaptic active zones. actin-myosin transport systems prevent somatic exocytosis from Therefore, most exocytosis occurs extrasynaptically (Hökfelt, Retzius and magnocellular neurons (Tobin and Ludwig, 2007; 1968; Umbriaco et al., 1995; Contant et al., 1996; Descarries et al., De-Miguel et al., 2012; Noguez et al., 2019). 1996; Descarries and Mechawar, 2000; Burnstock, 2012). Stimulation and the activation of the transport are linked by a chain of events: first, stimulation promotes calcium Somatic Release of Serotonin entry through L-type channels (Trueta et al., 2003), which The vast diversity and distribution of serotonin functions, the are advantageous, for their slow inactivation sustains calcium small numbers of serotonergic neurons and synapses, and the entry along trains of impulses or long depolarizations. Second, extraordinary chemical properties of serotonin explain why increasing the stimulation frequency increases the amplitude of extrasynaptic serotonergic communication has been widely the intracellular calcium transient (Leon-Pinzon et al., 2014). studied. Serotonin that is released from the cell body and Third, calcium activates ryanodine receptors, inducing calcium- dendrites of raphe neurons has been detected distantly by dependent calcium release (Trueta et al., 2004; Leon-Pinzon et al., voltammetry, based on its redox properties (Bunin and 2014), as in substantia nigra of dopaminergic and magnocellular Wightman, 1998). Moreover, serotonin exocytosis has been neurons (Ludwig et al., 2002; Patel et al., 2009). Fourth, a calcium detected by amperometric electrodes apposed onto the soma tsunami floods the cell body and invades the mitochondria, of leech Retzius neurons (Bruns et al., 2000), or in the soma which respond by producing ATP. Fifth, ATP sets in motion the and dendrites of raphe neurons by multiphoton excitation kinesin- and myosin-dependent vesicle transport (Figure 1; De- (Kaushalya et al., 2008; Colgan et al., 2012; Sarkar et al., 2012; Miguel et al., 2012). Maity and Maiti, 2018). The mechanism for somatic exocytosis of serotonin has been Energy Cost of the Vesicle Transport studied step by step in Retzius neurons (De-Miguel et al., 2012). Application of thermodynamic theory to the kinetics of Their large (60–80 µm) soma contains ‘‘astronomic’’ numbers exocytosis predicts that three variables determine the latency of dense-core vesicles loaded with serotonin (Coggeshall, from stimulation to the onset of exocytosis (De-Miguel et al., 1972). Most vesicles rest distantly from the plasma membrane. 2012 ): the traveling distance to the plasma membrane (0.2–6.0 However, electron microscopy and fluorescence of FM dyes, µm), the velocity of the transport (15–90 nm/s) and the number which stain the intravesicular membrane during exo-endocytosis of vesicles carried per cluster (90 to >1,000). Upon arrival at (Hoopmann et al., 2012), indicate that vesicles move massively the plasma membrane, vesicles fuse at a 0.5–4.0 s−1 rate, which to the plasma membrane following trains of action potentials reflects the transport velocity. For example, exocytosis from but not individual impulses (Trueta et al., 2003). The formation 1,000 vesicles at a 4 s−1 rate lasts 250 s. The energy expenses of of fluorescent spots beneath the soma surface indicates that the transport, calculated from the work of the motors, range from fusion of dense-core vesicles occurs in preferential sites. The 10–200 ATP molecules per vesicle fused, depending on the same development of FM fluorescent spots reflects the kinetics of variables (De-Miguel et al., 2012). release by clusters of vesicles; the number of fluorescent spots How thermodynamically-efficient is the use of ATP during is a measure of the amount of release. Such experiments gave the vesicle transport? An answer has been obtained also from unexpected results: First, exocytosis starts seconds after the end the application of thermodynamic theory (Noguez et al., 2019). of stimulation. Second, exocytosis lasts hundreds of seconds Surprisingly, the largest thermodynamic efficiency value is 6.2%, (Trueta et al., 2003), as hormone release from gland cells which is lower than the 20% efficiency of a contemporary car does (Thorn et al., 2016). Similar results have been obtained running on a highway. The remaining energy is dissipated as from cholinergic, dopaminergic, noradrenergic and peptidergic heat along the path, owing to friction forces. The origin of neurons (Sun and Poo, 1987; Huang and Neher, 1996; Jaffe et al., such friction was predicted from the distribution of efficiency 1998; Puopolo et al., 2001; Bao et al., 2003; Kaushalya et al., 2008; values along the traveling pathway. The lowest values correlate Huang et al., 2012; Ludwig and Stern, 2015). with the penetration of vesicles to the actin cortex and their In Retzius neurons and magnocellular neurons it is the passage between endoplasmic reticulum layers. Both essential frequency of the action potentials, not their number, what contributors to the transport increase the energy cost by being determines the amount of release. The maximum release occurs frictive obstacles. Such a phenomenon adds energy cost to the at 20 impulses per second (Dreifuss et al., 1971; Leon-Pinzon modulation of neuronal circuits. et al., 2014), but may be enhanced by alternate periods of stimulation and rest (Dutton and Dyball, 1979). Calcium and Exocytosis Measurements of the intracellular calcium dynamics with Vesicle Transport fluorescent dyes unveiled that by the time vesicles arrive at The delay between stimulation and exocytosis reflects the vesicle the plasma membrane, the intracellular calcium concentration transport to the plasma membrane (De-Miguel et al., 2012). The has returned to resting levels except in the soma shell large calcium transient that develops upon the stimulation train (Leon-Pinzon et al., 2014). Such peripheral calcium elevation is essential for the vesicle transport. In electron micrographs, drives the fusion of vesicles as they arrive at the plasma vesicles remain at rest upon stimulation with extracellular membrane. Voltage clamp measurements failed to detect any magnesium substituting for calcium to block calcium entry. transmembrane calcium flow following the train of impulses. Moreover, experimental perturbations of the tubulin-kinesin or Instead, the peripheral calcium transient was reproduced Frontiers in Molecular Neuroscience | www.frontiersin.org 3 April 2021 | Volume 14 | Article 638858 De-Miguel et al. Extrasynaptic Communication FIGURE 1 | Schematic representation of the mechanism for somatic exocytosis of serotonin in Retzius neurons. (A) Electrical activity sets in motion the transport of dense core vesicles (dcv) to the plasma membrane. In response to a train of action potentials, L-type calcium channels (LCach) open. Calcium entry activates ryanodine receptors (RyR) in the endoplasmic reticulum (er) and produces calcium-induced calcium release. The amplified calcium wave invades the soma; in the mitochondria (mit), calcium stimulates the synthesis of ATP, which activates kinesin motors (km) and vesicle transport along microtubules (mt). (B) Vesicles enter the actin cortex and myosin motors (MyM) carried by the vesicles couple to actin filaments and contribute to the transport. Release is maintained by a positive feedback loop in which the serotonin that is released activates 5-HT2 receptors (5-HT2R). Activation of phospholipase C (PLC) produces IP3 which acts on its receptors (IP3 R) to activate calcium release. Such calcium maintains exocytosis going on until the last vesicles fuse (Adapted from Leon-Pinzon et al., 2014). by iontophoretic serotonin application to activate membrane which are unusual at synapses (Witkovsky et al., 2009). receptors. Conclusive evidence that the peripheral calcium Moreover, the synaptic calcium sensors synaptotagmins transient depends on serotonin that had been released came 1 and 2 are substituted by the more appropriate high from experiments in which both the peripheral calcium affinity synaptotagmins 4 and 7 (Mendez et al., 2011), transient and somatic exocytosis were prevented by blocking since the fusion of dense-core vesicles depends on distant serotonergic 5HT2 receptors with methysergide, or by blocking calcium sources. the activation of phospholipase C (PLC) with U-73122 before stimulation of somatic exocytosis (Leon-Pinzon et al., 2014). Dense-Core Vesicle Recycling Therefore, the calcium that sustains exocytosis is released by The vesicle recycling has been studied by adding the marker peripheral endoplasmic reticulum upon a serotonin-mediated peroxidase to the extracellular fluid before stimulation (Trueta IP3 production and activation of IP3 receptors. Similar et al., 2012). Sections for electron microscopy incubated with observations made in peptidergic dorsal root ganglion and anti-peroxidase antibody coupled to gold particles, showed magnocellular neurons (Bao et al., 2003; Ludwig and Stern, peroxidase inside dense-core vesicles, newly-formed small 2015) point to another general principle: the long-lasting (∼40 nm) clear vesicles, and inside multivesicular bodies exocytosis is sustained by a feedback loop. Transmitter that containing both vesicle types (Figure 1). Multivesicular bodies is liberated activates autoreceptors coupled to phospholipase are transported retrogradely and their content is recycled in the C; IP3 production evokes intracellular calcium release; calcium perikaryon to form new vesicles. promotes exocytosis; the released substance maintains the cycle. Termination of the loop follows the fusion of the last vesicles in Synaptic vs. Extrasynaptic Exocytosis the clusters (Figure 1). Studies in Retzius neurons permit a comparison of the amounts of transmitter liberated from synaptic and extrasynaptic vesicle Molecules Catalyzing Exocytosis pools. The formation of specific synapses between identified Synapses contain a calcium-sensitive molecular complex leech neurons in culture allowed John Nicholls and his that drives vesicle fusion upon local calcium elevations colleagues (Nicholls and Kuffler, 1990) to examine the fine (Südhof, 2013). In the soma and dendrites of dopaminergic mechanisms of transmission. At synapses, the fusion of clear neurons, antibodies recognize isoforms of the fusion vesicles is calcium-dependent. Amperometric records show that complex components VAMP-2, SNAP25, and syntaxin, a vesicle liberates ∼4,700 serotonin molecules (Bruns and Frontiers in Molecular Neuroscience | www.frontiersin.org 4 April 2021 | Volume 14 | Article 638858 De-Miguel et al. Extrasynaptic Communication Jahn, 1995). Synapses display short-term plasticity (Stewart The difference between synaptic and somatic exocytosis et al., 1989); for example, 10 impulses at 20-Hz evoke rapid is more drastic. Electron microscopy and FM dye staining facilitation followed by depression, along which ∼60 quanta of vesicles indicate that a 20-Hz train evokes fusion of get released. ∼40,000 vesicles from ∼80 release sites, each vesicle cluster A common form of extrasynaptic exocytosis of transmitters carrying on an average 500 vesicles (Del-Bel and De-Miguel, and peptides occurs from dense-core vesicles surrounding 2018). By assuming that ∼90,000 serotonin molecules integrate synaptic active zones (Hökfelt et al., 2018). The differences a quantum, a 10-impulse train at 20-Hz would trigger release of between synaptic and perisynaptic exocytosis are schematized ∼3.6 billion molecules. Moreover, the long thick axon discharges in Figure 2. The presynaptic boutons of cultured Retzius serotonin from clear and dense-core vesicles in undetermined neurons contain clear synaptic vesicles and dense-core amounts. It is predictable that the huge amount of serotonin perisynaptic vesicles all filled with serotonin (Kuffler being released from a pair of Retzius cells in each ganglion et al., 1987; Bruns and Jahn, 1995). Perisynaptic release suffices to modulate behavior (Willard, 1981). increases along stimulation trains and produces large quantal amperometric spikes upon release of ∼90,000 serotonin Transmitter Spillover molecules (Bruns and Jahn, 1995). Amazingly, three dense-core Glutamate and GABA, the conventional transmitters at synapses, vesicles release about the same number of serotonin act extrasynaptically upon spillover from the synaptic cleft molecules as the 60 synaptic vesicles that fuse along a when synaptic release increases (Isaacson et al., 1993; Rusakov 20-Hz train. and Kullmann, 1998; DiGregorio et al., 2002). Spillover- FIGURE 2 | Synaptic and perisynaptic exocytosis. (A) Low frequency stimulation activates presynaptic P or N type calcium channels and evokes fusion of synaptic vesicles in the active zone. Transmitter liberated into the synaptic cleft activates post-synaptic receptors. (B) Increasing the stimulation frequency increases synaptic exocytosis and produces transmitter spillover, which acts on extrasynaptic receptors in adjacent glia and neuronal processes. (C) In the presence of perisynaptic dense-core vesicles, a low stimulation frequency evokes mostly synaptic release. (D) Increasing the stimulation frequency produces summation of calcium currents flowing through L type channels. Dense core vesicles are transported to the plasma membrane and fuse in presynaptic regions of the terminal. Transmitter that has been released acts on extrasynaptic receptors in the pre-and post-synaptic terminals, and also in adjacent glial and neuronal processes. Clear and dense core vesicles may have the same or different transmitters. Dense core vesicles also may contain peptides. Frontiers in Molecular Neuroscience | www.frontiersin.org 5 April 2021 | Volume 14 | Article 638858 De-Miguel et al. Extrasynaptic Communication mediated transmission occurs through activation of low-affinity extrapolation, the magnetic resonance images may be a product extrasynaptic receptors in neighboring cells (Pál, 2018). In of extrasynaptic communication. addition, astroglia and microglia sense and release glutamate (Pál, 2018). CONCLUSIONS Glia as Mediator of Extrasynaptic 1. Extrasynaptic exocytosis is common in the nervous system. It Communication may occur differentially from the soma, dendrites and axon, Glia are common counterparts for extrasynaptic exocytosis. Glial allowing neurons to produce multiple effects. cell membranes respond to transmitters and transport many 2. Synaptic and extrasynaptic exocytosis coexist in the same of them (Marcaggi and Attwell, 2004; Káradóttir et al., 2005; neurons. Verkhratsky et al., 2009). In response to transmitters such as 3. Neurotransmitters and peptides are released extrasynaptically. glutamate, serotonin and ATP, networks of electrically-coupled 4. Synaptic transmission is punctual; extrasynaptic transmission astrocytes propagate calcium transients (Munsch and Deitmer, is diffuse. Substances released extrasynaptically act via volume 1992; Metea and Newman, 2006; Verkhratsky et al., 2009). In transmission at variable distances and with different time return, glia releases the same or other transmitters, peptides courses. and proteins (Billups and Attwell, 1996; Henneberger et al., 5. Extrasynaptic communication integrates the activity of 2010; Igelhorst et al., 2015). Observations like these have led neurons, glia and blood vessels. to the hypothesis of tripartite synapses, in which glia reacts 6. Other forms of extrasynaptic neurotransmission occur upon to transmitters that spillover and in return modulate synaptic diffusive release of molecules, such as gases and cannabinoids; activity (Perea et al., 2009; Corkrum et al., 2020). vesicles are released loaded with cocktails of molecules. 7. Understanding the functioning of the nervous system Extrasynaptic Integration of Retinal requires understanding of its modulation by extrasynaptic Responses to Light communication. The retina provides a clear example of the integrative roles of extrasynaptic communication at the cellular level. A light spot shone onto a dark-adapted retina, evokes visual processing AUTHOR CONTRIBUTIONS and extrasynaptic release of transmitters from amacrine cells (Hirasawa et al., 2015; Newman, 2015). Dopamine contributes FD-M wrote the first version of the manuscript. All authors to light adaptation by uncoupling electrical synapses and by contributed to the article and approved the submitted version. acting directly on neurons at every level of the visual processing (Piccolino et al., 1984; Witkovsky, 2004; Zhang et al., 2011). Solid FUNDING evidence about glia as mediator between extrasynaptic exocytosis and the regulation of blood flow has been contributed by This work was supported by Human Frontier Science Program Newman and his colleagues (Newman, 2015). ATP released from (HFSP; Ref RGP0060/2019); Consejo Nacional de Ciencia y Amacrine cells activates Muller cells, the main type of retinal Tecnología (CONACyT; grant 130031); and Dirección General glia. In response, Muller cells synthesize and release factors that de Asuntos del Personal Académico (DGAPA; grant IN210317) increase blood flow and oxygenation of the illuminated area. By from the Universidad Nacional Autónoma de México to FD-M. 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Neurotransmitter release: the last millisecond in the Copyright © 2021 De-Miguel, Leon-Pinzon, Torres-Platas, del-Pozo, Hernández- life of a synaptic vesicle. Neuron 80, 675–690. doi: 10.1016/j.neuron.2013. Mendoza, Aguirre-Olivas, Méndez, Moore, Sánchez-Sugía, García-Aguilera, 10.022 Martínez-Valencia, Ramírez-Santiago and Rubí. This is an open-access article Sarkar, B., Das, A. K., Arumugam, S., Kaushalya, S. K., Bandyopadhyay, A., distributed under the terms of the Creative Commons Attribution License (CC BY). Balaji, J., et al. (2012). The dynamics of somatic exocytosis in monoaminergic The use, distribution or reproduction in other forums is permitted, provided the neurons. Front. Physiol. 3:414. doi: 10.3389/fphys.2012.00414 original author(s) and the copyright owner(s) are credited and that the original Stewart, R. R., Adams, W. B., and Nicholls, J. G. (1989). Presynaptic calcium publication in this journal is cited, in accordance with accepted academic practice. currents and facilitation of serotonin release at synapses between cultured leech No use, distribution or reproduction is permitted which does not comply with neurones. J. Exp. Biol. 144, 1–12. these terms. Frontiers in Molecular Neuroscience | www.frontiersin.org 8 April 2021 | Volume 14 | Article 638858