N-ethylmaleimide Sensitive Fusion Research Articles

Synaptophysin chaperones the assembly of 12 SNAREpins under each ready-release vesicle

The synaptic vesicle protein Synaptophysin (Syp) has long been known to form a complex with the Vesicle associated soluble N-ethylmaleimide sensitive fusion protein attachment receptor (v-SNARE) Vesicle associated membrane protein (VAMP), but a more specific molecular function or mechanism of action in exocytosis has been lacking because gene knockouts have minimal effects. Utilizing fully defined reconstitution and single-molecule measurements, we now report that Syp functions as a chaperone that determines the number of SNAREpins assembling between a ready-release vesicle and its target membrane bilayer. Specifically, Syp directs the assembly of 12 ± 1 SNAREpins under each docked vesicle, even in the face of an excess of SNARE proteins. The SNAREpins assemble in successive waves of 6 ± 1 and 5 ± 2 SNAREpins, respectively, tightly linked to oligomerization of and binding to the vesicle Ca++ sensor Synaptotagmin. Templating of 12 SNAREpins by Syp is likely the direct result of its hexamer structure and its binding of VAMP2 dimers, both of which we demonstrate in detergent extracts and lipid bilayers.

Rhodojaponin VI indirectly targets Cav2.2 channels via N-ethylmaleimide-sensitive fusion protein to alleviate neuropathic pain.

Neuropathic pain is a chronic disease that severely afflicts the life and emotional status of patients, but currently available treatments are often ineffective. Novel therapeutic targets for the alleviation of neuropathic pain are urgently needed. Rhodojaponin VI, a grayanotoxin from Rhododendron molle, showed remarkable antinociceptive efficacy in models of neuropathic pain, but its biotargets and mechanisms are unknown. Given the reversible action of rhodojaponin VI and the narrow range over which its structure can be modified, we perforwmed thermal proteome profiling of the rat dorsal root ganglion to determine the protein target of rhodojaponin VI. N-Ethylmaleimide-sensitive fusion (NSF) was confirmed as the key target of rhodojaponin VI through biological and biophysical experiments. Functional validation showed for the first time that NSF facilitated trafficking of the Cav2.2 channel to induce an increase in Ca2+ current intensity, whereas rhodojaponin VI reversed the effects of NSF. In conclusion, rhodojaponin VI represents a unique class of analgesic natural products targeting Cav2.2 channels via NSF.

PRG-1 prevents neonatal stimuli-induced persistent hyperalgesia and memory dysfunction via NSF/Glu/GluR2 signaling

SummaryNeonatal repetitive noxious stimuli (RNS) has been shown to cause long-term harmful effects on nociceptive processing, learning, and memory which persist until adulthood. Plasticity-related gene 1 (PRG-1) regulates synaptic plasticity and functional reorganization in the brain during neuronal development. In this study, neonatal RNS rats were established by repetitive needle pricks to neonatal rats on all four feet to model repetitive pain exposure in infants. Neonatal RNS caused thermal hyperalgesia, mechanical allodynia, learning, and memory impairments which manifested in young rats and persisted until adulthood. Hippocampal PRG-1/N-ethylmaleimide sensitive fusion protein (NSF) interaction was determined to be responsible for the RNS-induced impairment via enhanced extracellular glutamate release and AMPAR GluR2 trafficking deficiency in a cell-autonomous manner. These pathways likely act synergistically to cause changes in dendritic spine density. Our findings suggest that PRG-1 prevents the RNS-induced hyperalgesia, learning, and memory impairment by regulating synaptic plasticity via NSF/Glu/GluR2 signaling.

Tau target identification reveals NSF-dependent effects on AMPA receptor trafficking and memory formation.

Microtubule-associated protein tau is a central factor in Alzheimer's disease and other tauopathies. However, the physiological functions of tau are unclear. Here, we used proximity-labelling proteomics to chart tau interactomes in primary neurons and mouse brains in vivo. Tau interactors map onto pathways of cytoskeletal, synaptic vesicle and postsynaptic receptor regulation and show significant enrichment for Parkinson's, Alzheimer's and prion disease. We find that tau interacts with and dose-dependently reduces the activity of N-ethylmaleimide sensitive fusion protein (NSF), a vesicular ATPase essential for AMPA-type glutamate receptor (AMPAR) trafficking. Tau-deficient (tau-/- ) neurons showed mislocalised expression of NSF and enhanced synaptic AMPAR surface levels, reversible through the expression of human tau or inhibition of NSF. Consequently, enhanced AMPAR-mediated associative and object recognition memory in tau-/- mice is suppressed by both hippocampal tau and infusion with an NSF-inhibiting peptide. Pathologic mutant tau from mouse models or Alzheimer's disease significantly enhances NSF inhibition. Our results map neuronal tau interactomes and delineate a functional link of tau with NSF in plasticity-associated AMPAR-trafficking and memory.

AQP2 trafficking in health and diseases: an updated overview.

Renal collecting duct principal cells play a key role in controlling body water balance. Principal cells express the water channels AQP2, AQP3, and AQP4 that mediate renal water reabsorption. AQP3 and AQP4 are expressed at the basolateral membrane constitutively. Conversely, AQP2 is localized in intracellular vesicles and translocates to the plasma membrane under vasopressin action. Stimulation with vasopressin activates the cAMP/PKA signal transduction pathway that induces the redistribution of AQP2 from an intracellular pool to the apical plasma membrane. AQP2 trafficking and function depend on multiple post-translational modifications. Moreover, several proteins control different steps activated by the vasopressin stimulation that triggers the redistribution of the AQP2 vesicles. A-kinase anchoring proteins (AKAPs) together with phosphodiesterases and adenylate cyclases play crucial roles in modulating local changes of cAMP. Soluble N-ethylmaleimide sensitive fusion factor attachment protein receptors (SNARE), cytoskeletal proteins, and the small GTPases of the Rho family regulate the fusion and the endocytotic retrieval of AQP2 vesicles. Abnormal vasopressin signaling and altered AQP2 expression or trafficking can lead to disorders characterized by deregulated mechanisms controlling water homeostasis. This review provides updated data on the molecular signals regulating vasopressin-induced AQP2 trafficking in health and disease.

Lipid⁻Protein Interactions in Niemann⁻Pick Type C Disease: Insights from Molecular Modeling.

The accumulation of lipids in the late endosomes and lysosomes of Niemann–Pick type C disease (NPCD) cells is a consequence of the dysfunction of one protein (usually NPC1) but induces dysfunction in many proteins. We used molecular docking to propose (a) that NPC1 exports not just cholesterol, but also sphingosine, (b) that the cholesterol sensitivity of big potassium channel (BK) can be traced to a previously unappreciated site on the channel’s voltage sensor, (c) that transient receptor potential mucolipin 1 (TRPML1) inhibition by sphingomyelin is likely an indirect effect, and (d) that phosphoinositides are responsible for both the mislocalization of annexin A2 (AnxA2) and a soluble NSF (N-ethylmaleimide Sensitive Fusion) protein attachment receptor (SNARE) recycling defect. These results are set in the context of existing knowledge of NPCD to sketch an account of the endolysosomal pathology key to this disease.

Discovery of the First Germline-Restricted Gene by Subtractive Transcriptomic Analysis in the Zebra Finch, Taeniopygia guttata

Developmentally programmed genome rearrangements are rare in vertebrates, but have been reported in scattered lineages including the bandicoot, hagfish, lamprey, and zebra finch (Taeniopygia guttata) [1]. Inthe finch, a well-studied animal model for neuroendocrinology and vocal learning [2], one such programmed genome rearrangement involves a germline-restricted chromosome, or GRC, which is found in germlines of both sexes but eliminated from mature sperm [3, 4]. Transmitted only through the oocyte, it displays uniparental female-driven inheritance, and early in embryonic development is apparently eliminated from all somatic tissue in both sexes [3, 4]. The GRC comprises the longest finch chromosome at over 120 million base pairs [3], and previously the only known GRC-derived sequence was repetitive and non-coding [5]. Because the zebra finch genome project was sourced from male muscle (somatic) tissue [6], the remaining genomic sequence and protein-coding content of the GRC remain unknown. Here we report the first protein-coding gene from the GRC: a member of the α-soluble N-ethylmaleimide sensitive fusion protein (NSF) attachment protein (α-SNAP) family hitherto missing from zebra finch gene annotations. In addition to the GRC-encoded α-SNAP, we find an additional paralogous α-SNAP residing in the somatic genome (a somatolog)-makingthe zebra finch the first example in which α-SNAP is not a single-copy gene. We show divergent, sex-biased expression for the paralogs and also thatpositive selection is detectable across the birdα-SNAP lineage, including the GRC-encoded α-SNAP. This study presents the identification and evolutionary characterization of the first protein-coding GRC gene in any organism.

Agnoprotein Is an Essential Egress Factor during BK Polyomavirus Infection.

BK polyomavirus (BKPyV; hereafter referred to as BK) causes a lifelong chronic infection and is associated with debilitating disease in kidney transplant recipients. Despite its importance, aspects of the virus life cycle remain poorly understood. In addition to the structural proteins, the late region of the BK genome encodes for an auxiliary protein called agnoprotein. Studies on other polyomavirus agnoproteins have suggested that the protein may contribute to virion infectivity. Here, we demonstrate an essential role for agnoprotein in BK virus release. Viruses lacking agnoprotein fail to release from host cells and do not propagate to wild-type levels. Despite this, agnoprotein is not essential for virion infectivity or morphogenesis. Instead, agnoprotein expression correlates with nuclear egress of BK virions. We demonstrate that the agnoprotein binding partner α-soluble N-ethylmaleimide sensitive fusion (NSF) attachment protein (α-SNAP) is necessary for BK virion release, and siRNA knockdown of α-SNAP prevents nuclear release of wild-type BK virions. These data highlight a novel role for agnoprotein and begin to reveal the mechanism by which polyomaviruses leave an infected cell.

The Benefits of Botulinum Neurotoxin Treatment in a Multitude of Medical Conditions

Botulinum neurotoxins (BoNTs) are responsible for botulism in humans and vertebrates, being one of the six most catastrophic potential bioterrorism agents. This are ~150 kDa proteins, assembled as a ~50 kDa light chain (LC) and a ~100 kDa heavy chain (HC). The LC acts like a zinc metalloproteinase that cleaves three proteins in neurons, members of the SNARE (Soluble N-ethylmaleimide sensitive fusion attachment protein receptors) family: VAMP (vesicle-associated membrane protein) / synaptobrevin, SNAP-25 (synaptosomal-associated protein 25) and syntaxin. After cleavage of any of this proteins, neurotransmission is blocked and flaccid paralysis of the muscle is installed. This extraordinary restricted tropism for the cholinergic presynaptic membrane makes this drug unique regarding its toxicity, pharmacological and therapeutic use. Taking into consideration the potential of this substance, this paper aims to summarize the most relevant data regarding the mechanism of actions and its main clinical applications, in order to improve medical practice. Therefore, we presented the mechanism of action in order to understand its usage in different pathologies, such as dystonias, spasticity, nephrologic and urologic conditions, cosmetic use, depression, gastroenterologic and proctologic diseases, dermatologic conditions, pathologies specific to plastic surgery and also the role of BoNT therapy in pain management. It is well documented in the literature that important discoveries have been made through recent experimental and clinical studies. Even so, there is still much to learn about the therapeutic action of this drug in terms of molecular and pathophysiological mechanisms, in order to benefit of the whole healing potential of this amazing toxin.

Visualizing Intracellular SNARE Trafficking by Fluorescence Lifetime Imaging Microscopy.

Soluble N-ethylmaleimide sensitive fusion protein (NSF) attachment protein receptor (SNARE) proteins are key for membrane trafficking, as they catalyze membrane fusion within eukaryotic cells. The SNARE protein family consists of about 36 different members. Specific intracellular transport routes are catalyzed by specific sets of 3 or 4 SNARE proteins that thereby contribute to the specificity and fidelity of membrane trafficking. However, studying the precise function of SNARE proteins is technically challenging, because SNAREs are highly abundant and functionally redundant, with most SNAREs having multiple and overlapping functions. In this protocol, a new method for the visualization of SNARE complex formation in live cells is described. This method is based on expressing SNARE proteins C-terminally fused to fluorescent proteins and measuring their interaction by Förster resonance energy transfer (FRET) employing fluorescence lifetime imaging microscopy (FLIM). By fitting the fluorescence lifetime histograms with a multicomponent decay model, FRET-FLIM allows (semi-)quantitative estimation of the fraction of the SNARE complex formation at different vesicles. This protocol has been successfully applied to visualize SNARE complex formation at the plasma membrane and at endosomal compartments in mammalian cell lines and primary immune cells, and can be readily extended to study SNARE functions at other organelles in animal, plant, and fungal cells.

Visualizing Intracellular SNARE Trafficking by Fluorescence Lifetime Imaging Microscopy

Soluble N-ethylmaleimide sensitive fusion protein (NSF) attachment protein receptor (SNARE) proteins are key for membrane trafficking, as they catalyze membrane fusion within eukaryotic cells. The SNARE protein family consists of about 36 different members. Specific intracellular transport routes are catalyzed by specific sets of 3 or 4 SNARE proteins that thereby contribute to the specificity and fidelity of membrane trafficking. However, studying the precise function of SNARE proteins is technically challenging, because SNAREs are highly abundant and functionally redundant, with most SNAREs having multiple and overlapping functions. In this protocol, a new method for the visualization of SNARE complex formation in live cells is described. This method is based on expressing SNARE proteins C-terminally fused to fluorescent proteins and measuring their interaction by Förster resonance energy transfer (FRET) employing fluorescence lifetime imaging microscopy (FLIM). By fitting the fluorescence lifetime histograms with a multicomponent decay model, FRET-FLIM allows (semi-)quantitative estimation of the fraction of the SNARE complex formation at different vesicles. This protocol has been successfully applied to visualize SNARE complex formation at the plasma membrane and at endosomal compartments in mammalian cell lines and primary immune cells, and can be readily extended to study SNARE functions at other organelles in animal, plant, and fungal cells.

Ultrastructure and immunohistochemical characterization of proteins concerned with the secretory machinery in goat ceruminous glands.

The expression of soluble N-ethylmaleimide sensitive fusion attachment protein receptor (SNARE) proteins in apocrine glands has not been fully elucidated. In addition to performing ultrastructural observation of the ceruminous glands in goats, our study focuses on the demonstration of β-defensins, SNARE proteins and Rab3D in these glands with the use of immunohistochemical methods. The secretory cells were equipped with two types of vesicles, Golgi apparatus and abundant rough endoplasmic reticulum (ER). Additionally, in some of them, the characteristic concentric structures composed of rough ER were observed in their circum- and infranuclear parts. The expression of phosphorylated inositol requiring enzyme 1rrwas also detected. These findings may indicate their ability to produce numerous secretory proteins and the maintenance of homeostasis in the glandular cells. Furthermore, β-defensins were demonstrated as products of the ceruminous glands. The present investigation also revealed the presence of SNARE proteins and Rab3D. It is suggested that these proteins are concerned with the secretory machinery of this gland type.

BAIAP3, a C2 domain-containing Munc13 protein, controls the fate of dense-core vesicles in neuroendocrine cells.

Dense-core vesicle (DCV) exocytosis is a SNARE (soluble N-ethylmaleimide-sensitive fusion attachment protein receptor)-dependent anterograde trafficking pathway that requires multiple proteins for regulation. Several C2 domain-containing proteins are known to regulate Ca2+-dependent DCV exocytosis in neuroendocrine cells. In this study, we identified others by screening all (∼139) human C2 domain-containing proteins by RNA interference in neuroendocrine cells. 40 genes were identified, including several encoding proteins with known roles (CAPS [calcium-dependent activator protein for secretion 1], Munc13-2, RIM1, and SYT10) and many with unknown roles. One of the latter, BAIAP3, is a secretory cell-specific Munc13-4 paralog of unknown function. BAIAP3 knockdown caused accumulation of fusion-incompetent DCVs in BON neuroendocrine cells and lysosomal degradation (crinophagy) of insulin-containing DCVs in INS-1 β cells. BAIAP3 localized to endosomes was required for Golgi trans-Golgi network 46 (TGN46) recycling, exhibited Ca2+-stimulated interactions with TGN SNAREs, and underwent Ca2+-stimulated TGN recruitment. Thus, unlike other Munc13 proteins, BAIAP3 functions indirectly in DCV exocytosis by affecting DCV maturation through its role in DCV protein recycling. Ca2+ rises that stimulate DCV exocytosis may stimulate BAIAP3-dependent retrograde trafficking to maintain DCV protein homeostasis and DCV function.

Presynaptic PICK1 facilitates trafficking of AMPA-receptors between active zone and synaptic vesicle pool

Previous studies have indicated that presynaptic α-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptors (AMPARs) contribute to the regulation of neurotransmitter release. In hippocampal synapses, the presynaptic surface expression of several AMPAR subunits, including GluA2, is regulated in a ligand-dependent manner. However, the molecular mechanisms underlying the presynaptic trafficking of AMPARs are still unknown. Here, using bright-field immunocytochemistry, western blots, and quantitative immunogold electron microscopy of the hippocampal CA1 area from intact adult rat brain, we demonstrate the association of AMPA receptors with the presynaptic active zone and with small presynaptic vesicles, in Schaffer collateral synapses in CA1 of the hippocampus. Furthermore, we show that GluA2 and protein interacting with C kinase 1 (PICK1) are colocalized at presynaptic vesicles. Similar to postsynaptic mechanisms, overexpression of either PICK1 or pep2m, which inhibit the N-ethylmaleimide sensitive fusion protein (NSF)-GluA2 interaction, decreases the concentration of GluA2 in the presynaptic active zone membrane. These data suggest that the interacting proteins PICK1 and NSF act as regulators of presynaptic GluA2-containing AMPAR trafficking between the active zone and a vesicle pool that may provide the basis of presynaptic components of synaptic plasticity.

Exocytosis in non-neuronal cells.

Exocytosis is the process by which stored neurotransmitters and hormones are released via the fusion of secretory vesicles with the plasma membrane. It is a dynamic, rapid and spatially restricted process involving multiple steps including vesicle trafficking, tethering, docking, priming and fusion. For many years great steps have been undertaken in our understanding of how exocytosis occurs in different cell types, with significant focus being placed on synaptic release and neurotransmission. However, this process of exocytosis is an essential component of cell signalling throughout the body and underpins a diverse array of essential physiological pathways. Many similarities exist between different cell types with regard to key aspects of the exocytosis pathway, such as the need for Ca(2+) to trigger it or the involvement of members of the N-ethyl maleimide-sensitive fusion protein attachment protein receptor protein families. However, it is also equally clear that non-neuronal cells have acquired highly specialized mechanisms to control the release of their own unique chemical messengers. This review will focus on several important non-neuronal cell types and discuss what we know about the mechanisms they use to control exocytosis and how their specialized output is relevant to the physiological role of each individual cell type. These include enteroendocrine cells, pancreatic β cells, astrocytes, lactotrophs and cytotoxic T lymphocytes. Non-neuronal cells have acquired highly specialized mechanisms to control the release of unique chemical messengers, such as polarised fusion of insulin granules in pancreatic β cells targeted towards the vasculature (top). This review discusses mechanisms used in several important non-neuronal cell types to control exocytosis, and the relevance of intermediate vesicle fusion pore states (bottom) and their specialized output to the physiological role of each cell type. These include enteroendocrine cells, pancreatic β cells, astrocytes, lactotrophs and cytotoxic T lymphocytes. This article is part of a mini review series on Chromaffin cells (ISCCB Meeting, 2015).

Intracellular Trafficking, Localization, and Mobilization of Platelet-Borne Thiol Isomerases.

Thiol isomerases facilitate protein folding in the endoplasmic reticulum, and several of these enzymes, including protein disulfide isomerase and ERp57, are mobilized to the surface of activated platelets, where they influence platelet aggregation, blood coagulation, and thrombus formation. In this study, we examined the synthesis and trafficking of thiol isomerases in megakaryocytes, determined their subcellular localization in platelets, and identified the cellular events responsible for their movement to the platelet surface on activation. Immunofluorescence microscopy imaging was used to localize protein disulfide isomerase and ERp57 in murine and human megakaryocytes at various developmental stages. Immunofluorescence microscopy and subcellular fractionation analysis were used to localize these proteins in platelets to a compartment distinct from known secretory vesicles that overlaps with an inner cell-surface membrane region defined by the endoplasmic/sarcoplasmic reticulum proteins calnexin and sarco/endoplasmic reticulum calcium ATPase 3. Immunofluorescence microscopy and flow cytometry were used to monitor thiol isomerase mobilization in activated platelets in the presence and absence of actin polymerization (inhibited by latrunculin) and in the presence or absence of membrane fusion mediated by Munc13-4 (absent in platelets from Unc13d(Jinx) mice). Platelet-borne thiol isomerases are trafficked independently of secretory granule contents in megakaryocytes and become concentrated in a subcellular compartment near the inner surface of the platelet outer membrane corresponding to the sarco/endoplasmic reticulum of these cells. Thiol isomerases are mobilized to the surface of activated platelets via a process that requires actin polymerization but not soluble N-ethylmaleimide-sensitive fusion protein attachment receptor/Munc13-4-dependent vesicular-plasma membrane fusion.

LRRK2 phosphorylates pre-synaptic N-ethylmaleimide sensitive fusion (NSF) protein enhancing its ATPase activity and SNARE complex disassembling rate.

BackgroundLrrk2, a gene linked to Parkinson’s disease, encodes a large scaffolding protein with kinase and GTPase activities implicated in vesicle and cytoskeletal-related processes. At the presynaptic site, LRRK2 associates with synaptic vesicles through interaction with a panel of presynaptic proteins.ResultsHere, we show that LRRK2 kinase activity influences the dynamics of synaptic vesicle fusion. We therefore investigated whether LRRK2 phosphorylates component(s) of the exo/endocytosis machinery. We have previously observed that LRRK2 interacts with NSF, a hexameric AAA+ ATPase that couples ATP hydrolysis to the disassembling of SNARE proteins allowing them to enter another fusion cycle during synaptic exocytosis. Here, we demonstrate that NSF is a substrate of LRRK2 kinase activity. LRRK2 phosphorylates full-length NSF at threonine 645 in the ATP binding pocket of D2 domain. Functionally, NSF phosphorylated by LRRK2 displays enhanced ATPase activity and increased rate of SNARE complex disassembling. Substitution of threonine 645 with alanine abrogates LRRK2-mediated increased ATPase activity.ConclusionsGiven that the most common Parkinson’s disease LRRK2 G2019S mutation displays increased kinase activity, our results suggest that mutant LRRK2 may impair synaptic vesicle dynamics via aberrant phosphorylation of NSF.Electronic supplementary materialThe online version of this article (doi:10.1186/s13024-015-0066-z) contains supplementary material, which is available to authorized users.

Lower brain-derived neurotrophic factor levels associated with worsening fatigue in prostate cancer patients during repeated stress from radiation therapy

Objectives. Fatigue during cancer treatment is associated with depression. Neurotrophic factors play a major role in depression and stress and might provide insight into mechanisms of fatigue. This study investigated the association between plasma concentrations of three neurotrophic factors (BDNF, brain-derived neurotrophic factor; GDNF, glial-derived neurotrophic factor; and SNAPIN, soluble N-ethylmaleimide sensitive fusion attachment receptor-associated protein) and initial fatigue intensification during external beam radiation therapy (EBRT) in euthymic non-metastatic prostate cancer men. Methods. Fatigue, as measured by the 13-item Functional Assessment of Cancer Therapy-Fatigue (FACT-F), and plasma neurotrophic factors were collected at baseline (prior to EBRT) and mid-EBRT. Subjects were categorized into fatigue and no fatigue groups using a > 3-point change in FACT-F scores between the two time points. Multiple linear regressions analysed the associations between fatigue and neurotrophic factors. Results. FACT-F scores of 47 subjects decreased from baseline (43.95 ± 1.3) to mid-EBRT (38.36 ± 1.5, P < 0.001), indicating worsening fatigue. SNAPIN levels were associated with fatigue scores (rs = 0.43, P = 0.005) at baseline. A significant decrease of BDNF concentration (P = 0.008) was found in fatigued subjects during EBRT (n = 39). Conclusions. Baseline SNAPIN and decreasing BDNF levels may influence worsening fatigue during EBRT. Further investigations are warranted to confirm their role in the pathophysiology and therapeutics of fatigue.

Astrocytic vesicles and gliotransmitters: Slowness of vesicular release and synaptobrevin2-laden vesicle nanoarchitecture

Neurotransmitters released at synapses activate neighboring astrocytes, which in turn, modulate neuronal activity by the release of diverse neuroactive substances that include classical neurotransmitters such as glutamate, GABA or ATP. Neuroactive substances are released from astrocytes through several distinct molecular mechanisms, for example, by diffusion through membrane channels, by translocation via plasmalemmal transporters or by vesicular exocytosis. Vesicular release regulated by a stimulus-mediated increase in cytosolic calcium involves soluble N-ethyl maleimide-sensitive fusion protein attachment protein receptor (SNARE)-dependent merger of the vesicle membrane with the plasmalemma. Up to 25 molecules of synaptobrevin 2 (Sb2), a SNARE complex protein, reside at a single astroglial vesicle; an individual neuronal, i.e. synaptic, vesicle contains ∼70 Sb2 molecules. It is proposed that this paucity of Sb2 molecules in astrocytic vesicles may determine the slow secretion. In the present essay we shall overview multiple aspects of vesicular architecture and types of vesicles based on their cargo and dynamics in astroglial cells.

Quantum interactive dualism: From Beck and Eccles tunneling model of exocytosis to molecular biology of SNARE zipping

We commence this review by outlining the challenges faced by physical theories of consciousness and briefly describe the two main approaches based on classical or quantum mechanics. Next, we provide a detailed exposition of the motivation, the theoretical construction and experimental falsification of the celebrated model due to Beck and Eccles concerning mind-brain interaction purported to operate at the sites of neurotransmitter release in the brain. Finally, we propose our own model of a vibrationally assisted quantum tunneling mechanism involving a Davydov soliton propagating along the hydrogen bonds in the protein four-α-helix bundle of the SNARE complex (soluble NSF attachment protein receptor; NSF, N-ethylmaleimide sensitive fusion proteins) that drives synaptic vesicle fusion. We also discuss the possible experimental tests that could falsify our model. Since erasure of consciousness by volatile anesthetics results from binding to the hydrophobic core of the SNARE four-α-helix bundle, our model is well suited to support quantum interactive dualism. Biomedical Reviews 2014; 25: 15-24.