InteractiveFly: GeneBrief

Synaptotagmin 7: Biological Overview | References

Short-term synaptic plasticity is a fast and robust modification in neuronal presynaptic output that can enhance release strength to drive facilitation or diminish it to promote depression. The mechanisms that determine whether neurons display short-term facilitation or depression are still unclear. This study shows that the Ca(2+)-binding protein Synaptotagmin 7 (Syt7) determines the sign of short-term synaptic plasticity by controlling the initial probability of synaptic vesicle (SV) fusion. Electrophysiological analysis of Syt7 null mutants at Drosophila embryonic neuromuscular junctions demonstrate loss of the protein converts the normally observed synaptic facilitation response during repetitive stimulation into synaptic depression. In contrast, overexpression of Syt7 dramatically enhanced the magnitude of short-term facilitation. These changes in short-term plasticity were mirrored by corresponding alterations in the initial evoked response, with SV release probability enhanced in Syt7 mutants and suppressed following Syt7 overexpression. Indeed, Syt7 mutants were able to display facilitation in lower [Ca(2+)] where release was reduced. These data suggest Syt7 does not act by directly sensing residual Ca(2+) and argues for the existence of a distinct Ca(2+) sensor beyond Syt7 that mediates facilitation. Instead, Syt7 normally suppresses synaptic transmission to maintain an output range where facilitation is available to the neuron (Fujii, 2021).

Synaptotagmins (Syts) are a large family of Ca2+ binding proteins, with the Syt1 isoform functioning as the major Ca2+ sensor for synchronous synaptic vesicle (SV) fusion1. Ca2+ also controls presynaptic forms of short-term plasticity, with other Syt isoforms representing promising candidates to mediate these processes. Indeed, Synaptotagmin 7 (Syt7) has been reported to function in facilitation, a form of short-term plasticity that enhances synaptic transmission following consecutive action potentials. Facilitation is believed to be mediated by residual Ca2+ acting to enhance the number of SVs that are released during repetitive action potentials occurring within a short temporal window. Since Syt7 binds Ca2+ with high affinity (Wang, 2005) and slow kinetics (Hui, 2005), which match requirements for facilitation, the protein has been hypothesized to act as the Ca2+ sensor for this form of presynaptic plasticity. However, the role of Syt7 in facilitation is still unclear (Fujii, 2021).

Drosophila neuromuscular junctions (NMJs) provide an excellent system for testing the role of Syt7 in short-term synaptic plasticity. In particular, embryonic NMJs are highly plastic and allow stable recordings in high Ca2+ concentrations using the myosin heavy chain (Mhc) mutant background to prevent muscle contraction. Together with the lack of compensation that might occur at older synapses, these advantages provide highly reliable measurements of synaptic transmission. Indeed, analysis of Syt1 mutants at embryonic NMJs established this Syt isoform functions as the synchronous Ca2+ sensor for synaptic transmission. Although NMJs in many other species show depression, Drosophila embryonic NMJs are facilitative at physiological Ca2+ concentrations as shown here, similar to many mammalian central synapses such as those in the hippocampus. Using stable recordings from this facilitative and plastic synapse in Drosophila embryos, the absolute value of synaptic currents was quantified in Syt7 mutants and in animals overexpressing Syt7. While loss of Syt7 enhanced presynaptic output, overexpression of Syt7 suppressed release. These changes in the magnitude of presynaptic output were mirrored by changes in short-term presynaptic plasticity. High levels of Syt7 enabled robust facilitative responses while loss of Syt7 switched the normally facilitating synapse into one that displayed short-term depression. This work reveals that Syt7 normally reduces synaptic transmission to scale it to an appropriate range where facilitation is allowed, providing a bi-directional switch for short-term synaptic plasticity (Fujii, 2021).

The current study indicates Syt7 is indispensable for facilitation across the physiological range of Ca2+ concentrations at Drosophila embryonic NMJs as previously shown for mammalian preparations (Jackman, 2016). In the absence of Syt7, the normally facilitating embryonic NMJ now displays depression. Following Syt7 overexpression, facilitation is greatly enhanced. These data indicate the major reason for defective facilitation in Syt7^-/- mutants is due to loss of Syt7's ability to suppress release, which likely causes rapid SV depletion that is non-compatible with short-term synaptic facilitation. Likewise, overexpression of Syt7 reduces SV release and allows for enhanced facilitation. This role for Syt7 contrasts with current models proposed in mammals where Syt7 is hypothesized to bind residual Ca2+ to directly act as a facilitation Ca2+ sensor. Although the current data indicate Syt7 is not the primary Ca2+ sensor for facilitation, the possibility cannot be ruled out that Syt7 has dual roles in both suppressing and facilitating SV fusion as observed for Syt. If Syt7 has a dual role with C2A functioning for clamping and C2B for facilitation, the null mutant would lack both properties. It is possible the lack of clamping is the dominant phenotype, with any facilitative function being masked by SV depletion at higher Ca2+ concentrations. Thus, a Syt7-dependent component of facilitation cannot be ruled out. However, the presence of facilitation in Syt7^-/- mutants at lower [Ca2+] indicates there is a facilitation sensor besides Syt7 that monitors residual Ca2+ to directly activate this form of short-term plasticity. Although a role for residual maternally supplied Syt7 at the embryonic stage in Syt7^-/- mutants cannot be ruled out, there is no evidence from RNA profiling studies that indicate Syt7 is present at earlier stages of embryonic development prior to nervous system formation. Thus, it is unlikely residual Syt7 could sustain normal levels of facilitation as observed in low [Ca2+], consistent with the enhanced synaptic transmission observed across a broad [Ca2+] range in Syt7^-/- mutants. Given facilitation is also present in low [Ca2+] in Syt7^-/- mutants at the 3rd instar stage when any maternal contribution would be depleted, it is concluded that facilitation can occur in the complete absence of Syt7 under conditions where the initial response is reduced (Fujii, 2021).

A key advantage of the Drosophila embryonic NMJ preparation is the ability to unambiguously monitor the absolute baseline values of synaptic strength even in high [Ca2+] using the non-contracting Mhc mutant. In this regard, it is clear that synaptic transmission at Drosophila embryonic NMJs is much stronger in Syt7^-/- mutants at all Ca2+ concentrations tested. Moreover, the stronger transmission is due to higher release probability rather than an increased number of releasable SVs. Thus, the data predict that higher release probability leads to a lower facilitation ratio secondary to vesicle depletion. The precise mechanisms by which Syt7 suppresses SV release to enable facilitation will require further study. Beyond a potential clamping function for Syt7, the protein could alter local Ca2+ buffering or cause increased Ca2+ influx that could contribute to elevated SV release. Syt7 does not localize to SVs and may instead act from the plasma membrane or internal membrane compartments, allowing for several potential mechanisms for Syt7 to suppress release. Ca2+ binding to the C2A and C2B domains of Syt1 have been shown to have distinct functions in SV release, with C2B playing a dominant role in triggering SV fusion and C2A acting to clamp release. It is unclear if the C2A and C2B domains of Syt7 act similarly in Drosophila or have independent functions compared to Syt1. One possibility is that the C2A domain of Syt7 suppresses SV fusion and the C2B domain facilitates release, similar to Syt1. Structure function studies of Syt7 should help elucidate this biology in Drosophila, similar to prior studies of Syt1 function (Fujii, 2021).

As suppression of SV release by Syt7 is dose-dependent (Guan, 2020), increasing levels of Syt7 would elevate the ratio of facilitation. These results suggest the degree of facilitation across distinct neuronal populations may be set by Syt7 levels similar to a potentiometer. The analysis of Syt7^-/- nulls, heterozygotes and overexpression lines support such a model that changes release and short-term plasticity in a graded fashion. Depending on whether a synapse is facilitative or depressive, Syt7 expression could be modulated to gate plasticity to the level that most benefits the local circuit, similar to how Syt1 and Syt2 levels variably control release synchronicity across neuronal populations. Indeed, the squid giant synapse is facilitative only when Ca2+ is lowered from normal saline (artificial sea water), similar to Syt7^-/- mutants, suggesting synapses that normally depress may have reduced levels of Syt7. Indeed, recent evidence suggests that species-specific differences in presynaptic plasticity in rodents is linked to the levels of Syt7. In shrews, the levels of Syt7 are lower in hippocampal CA3 synapses and they show reduced presynaptic plasticity. In contrast, Syt7 levels are much higher in mice, with their CA3 output synapses displaying far greater forms of presynaptic plasticity. Drosophila adults and 3rd instar larvae also have less facilitative NMJs than embryonic NMJs. This difference may contribute to the distinct effects of Syt7 on clamping spontaneous SV release that is observed between embryonic and 3rd instar NMJs. Mammalian studies identified redundant functions for Syt1 and Syt7 in clamping spontaneous fusion at inhibitory synapses (Bacaj, 2013). While reductions in Syt7 levels alone did not increase spontaneous SV release, removal of both Syt1 and Syt7 enhanced mini frequency to a far greater level that loss of Syt1 alone. In addition, a Syt7 transgene was able to rescue the elevated miniature frequency in Syt1 mutants. These differences in clamping properties were attributed to an insufficient level of Syt7 expression compared to Syt1. Differences in the Syt1/Syt7 ratio between Drosophila 3rd instar and embryonic NMJs may also contribute to distinct effects on spontaneous SV clamping observed in Syt1 and Syt7 mutants at these distinct developmental stages. In conclusion, controlling expression level of Syt7 provides an attractive mechanism for activity-dependent presynaptic scaling of release probability as a homeostat for both presynaptic output and short-term facilitation, similar to postsynaptic scaling mechanisms previously described for chronic forms of synaptic plasticity (Fujii, 2021).

Drosophila Synaptotagmin 7 negatively regulates synaptic vesicle release and replenishment in a dosage-dependent manner

Synchronous neurotransmitter release is triggered by Ca(2+) binding to the synaptic vesicle protein Synaptotagmin 1, while asynchronous fusion and short-term facilitation is hypothesized to be mediated by plasma membrane-localized Synaptotagmin 7 (SYT7). This study generated mutations in Drosophila Syt7 to determine if it plays a conserved role as the Ca(2+) sensor for these processes. Electrophysiology and quantal imaging revealed evoked release was elevated 2-fold. Syt7 mutants also had a larger pool of readily-releasable vesicles, faster recovery following stimulation, and intact facilitation. Syt1/Syt7 double mutants displayed more release than Syt1 mutants alone, indicating SYT7 does not mediate the residual asynchronous release remaining in the absence of SYT1. SYT7 localizes to an internal membrane tubular network within the peri-active zone, but does not enrich at active zones. These findings indicate the two Ca(2+) sensor model of SYT1 and SYT7 mediating all phases of neurotransmitter release and facilitation is not applicable at Drosophila synapses (Guan, 2020).

To characterize the location and function of SYT7 in Drosophila the CRISPR-Cas9 system was used to endogenously label the protein and generate null mutations in the Syt7 locus. The findings indicate SYT7 acts as a negative regulator of SV release, active zone probability of release (AZ Pr), the readily releasable pool (RRP) size, and RRP refilling. The elevated P_r across the AZ population in Syt7 mutants provides a robust explanation for why defects in asynchronous release and facilitation are observed in the absence of the protein, as SYT7 levels set the baseline for the amount of evoked release. SYT7's presence on an internal tubular membrane network within the peri-AZ positions the protein to interface with the SV cycle at multiple points to regulate membrane trafficking. In addition, increased SV release in animals lacking both SYT1 and SYT7 indicate the full complement of Ca2+ sensors that mediate the distinct phases of SV release remain unknown (Guan, 2020).

Using a combination of synaptic physiology and imaging approaches, the findings indicate SYT7 acts to reduce SV recruitment and release. Minor defects in asynchronous release and facilitation were identified in Drosophila Syt7 mutants, as observed in mouse and zebrafish models. However, these defects are attrobited to reduced SV availability as a result of increased Pr in Syt7 mutants. Indeed, a key feature of facilitation is its critical dependence on initial Pr. Low Pr synapses increase SV fusogenicity as Ca2+ levels rise during paired-pulses or stimulation trains, resulting in short-term increases in Pr for SVs not recruited during the initial stimuli. In contrast, depression occurs at high Pr synapses due to the rapid depletion of fusion-capable SVs during the initial response. Prior quantal imaging at Drosophila NMJs demonstrated facilitation and depression can occur across different AZs within the same neuron, with high Pr AZs depressing and low Pr AZs facilitating. Given the elevated Pr in Syt7 mutants, the facilitation defects are likely related to differences in initial Pr and depletion of fusion-competent SVs available for release during the 2nd stimuli (Guan, 2020).

A similar loss of SVs due to elevated Pr in Syt7 mutants would reduce fusogenic SVs that are available during the delayed phase of the asynchronous response. Syt1; Syt7 double mutants continue to show asynchronous fusion and facilitation, demonstrating there must be other Ca2+ sensors that mediate these components of SV release. The predominant localization of endogenous SYT7 to an internal tubular membrane compartment at the peri-AZ also places the majority of the protein away from release sites where it would need to reside to directly activate SV fusion. As such, the data indicate SYT7 regulates SV release through a distinct mechanism from SYT1 (Guan, 2020).

It is also concluded that the remaining components of asynchronous fusion and facilitation must be mediated by an entirely different family of Ca2+-binding proteins than Synaptotagmins (or through Ca2+-lipid interactions). Of the seven Syt genes in the Drosophila genome, only 3 SYT proteins are expressed at the motor neuron synapses assayed in this study - SYT1, SYT4 and SYT7. For the remaining SYTs in the genome, SYT-α and SYT-β are expressed in neurosecretory neurons and function in DCV fusion. SYT12 and SYT14 lack Ca2+ binding residues in their C2 domains and are not expressed in motor neurons. In addition, SYT4 is found on exosomes and transferred to postsynaptic cells, where it participates in retrograde signaling. Syt1; Syt4 double mutants display the same SV fusion defects found in Syt1 mutants alone, indicating SYT4 cannot compensate for SYT1 function in SV release. As such, SYT1 and SYT7 are the only remaining SYT isoforms that could contribute to SV trafficking within Drosophila motor neuron terminals (Guan, 2020).

A prior study using a Syt7 exon-intron hairpin RNAi did not result in an increase in evoked release. Although a reduction in ectopic expression of SYT7 in muscles could be seen with Mhc-GAL4 driving the UAS-Syt7 RNAi, the anti-SYT7 antisera does not recognize the endogenous protein in neurons using immunocytochemistry, preventing a determination of presynaptic SYT7 levels following neuronal RNAi. To further examine this issue, western blot analysis was performed with this RNAi and compared those used in the current study. The results confirmed that the RNAi line failed to reduce endogenous GFP-tagged SYT7, although the two commercial RNAi lines used in the current study were highly effective. Based on these data, it is concluded that the previous Syt7 UAS-RNAi line was ineffective in knocking down endogenous SYT7. Given the Syt7^M1 and Syt7^M2 alleles result in early stop codons and lack SYT7 expression by western blot analysis and display elevated levels of fusion, the data indicate SYT7 normally acts to suppress SV release as demonstrated by electrophysiology and optical P_r imaging. SYT7 overexpression reduces SV release even more, further confirming that the levels of SYT7 set the baseline amount of SV fusion at Drosophila NMJ synapses (Guan, 2020).

Although the data indicate SYT7 is not the primary asynchronous or facilitation Ca2+ sensor in Drosophila, this study found it inhibits SV release in a dosage-sensitive manner. The reduction in SV release is not due to changes in the Ca2+ cooperativity of fusion or enhanced presynaptic Ca2+ entry, ruling out the possibility that SYT7 normally acts as a local Ca2+ buffer or an inhibitor of presynaptic voltage-gated Ca2+ channels. The reduction in release is also not due to altered endocytosis, as Syt7 mutants have a normal steady-state rate of SV cycling following depletion of the RRP. Instead, SYT7 regulates SV fusogenicity at a stage between SV endocytosis and fusion. Given the rapid enhanced refilling of the RRP observed in Syt7 mutants, and the suppression of RRP refilling following SYT7 overexpression, the data indicate SYT7 regulates releasable SVs in part through changes in SV mobilization to the RRP. Ca2+ is well known to control the replenishment rate of releasable SVs, with Calmodulin-UNC13 identified as one of several molecular pathways that accelerate RRP refilling in a Ca2+-dependent manner. The findings indicate SYT7 acts in an opposite fashion and slows RRP refilling, providing a Ca2+-dependent counter-balance for SV recruitment into the RRP. Although such an effect has not been described for mammalian SYT7, defects in RRP replenishment have been observed when both SYT1 and SYT7 are deleted or following high frequency stimulation trains (Guan, 2020).

SYT7's role in restricting SV fusion and RRP size also affects spontaneous release. Although Syt7 mutants alone do not show elevated mini frequency, DoubleNull mutants have a 2-fold increase in spontaneous release. Similar increases in spontaneous release were observed at mammalian synapses lacking both SYT7 and SYT1 (or SYT2), with the effect being attributed to a dual role in clamping fusion in the absence of Ca2+ (Luo, 2017; Turecek, 2019). The current results indicate the elevation in spontaneous release at Drosophila synapses is a result of an increase in releasable SVs rather than a clamping function for SYT7. Following overexpression of SYT7, there is a reduction in the number of fusogenic SVs available for both evoked and spontaneous release. The dosage-sensitivity of the various phenotypes indicate SYT7 abundance is a critical node in controlling SV release rate. Indeed, mammalian SYT7 levels are post-transcriptionally modulated by γ-secretase proteolytic activity and APP, linking it to SV trafficking defects in Alzheimer's disease (Guan, 2020).

How does SYT7 negatively regulate recruitment and fusion of SVs? The precise mechanism by which SYT7 reduces release and slows refilling of the RRP is uncertain given it is not enriched at sites of SV fusion. Although the possibility cannot be ruled out that a small fraction of the protein is found at AZs, SYT7 is predominantly localized to an internal membrane compartment at the peri-AZ where SV endocytosis and endosomal sorting occurs. SYT7 membrane tubules are in close proximity and could potentially interact with peri-AZs proteins, endosomes, lysosomes and the plasma membrane. Given its primary biochemical activity is to bind membranes in a Ca2+-dependent manner, SYT7 could mediate cargo or lipid movement across multiple compartments within peri-AZs. In addition, it is possible SYT7 tubules could form part of the poorly defined SV recycling endosome compartment. However, no change was observed in SV density or SV localization around AZs, making it unlikely SYT7 would be essential for endosomal trafficking of SVs. The best characterized regulator of the SV endosome compartment in Drosophila is the RAB35 GAP Skywalker (SKY). Although Sky mutations display some similarities to Syt7, including increased neurotransmitter release and larger RRP size, Syt7 lacks most of the well-described Sky phenotypes such as behavioral paralysis, FM1-43 uptake into discrete punctated compartments, cisternal accumulation within terminals and reduced SV density. In addition, no co-localization was found between SKY-GFP and SYT7RFP within presynaptic terminals (Guan, 2020).

By blocking SV refilling with bafilomycin, the findings indicate the fast recovery of the RRP can occur via enhanced recruitment from the reserve pool and does not require changes in endocytosis rate. The phosphoprotein Synapsin has been found to maintain the reserve SV pool by tethering vesicles to actin filaments at rest. Synapsin interacts with the peri-AZ protein Dap160/Intersectin to form a protein network within the peri-AZ that regulates clustering and release of SVs. Synapsin-mediated phase separation is also implicated in clustering SVs near release sites. SYT7 could similarly maintain a subset of SVs in a non-releasable pool and provide a dual mechanism for modulating SV mobilization. Phosphorylation of Synapsin and Ca2+ activation of SYT7 would allow multiple activity-dependent signals to regulate SV entry into the RRP. As such, SYT7 could play a key role in organizing membrane trafficking and protein interactions within the peri-AZ network by adding a Ca2+-dependent regulator of SV recruitment and fusogenicity (Guan, 2020).

Additional support for a role for SYT7 in regulating SV availability through differential SV sorting comes from recent studies on the SNARE complex binding protein CPX. Analysis of Drosophila Cpx mutants, which have a dramatic increase in minis, revealed a segregation of recycling pathways for SVs undergoing spontaneous versus evoked fusion. Under conditions where intracellular Ca2+ is low and SYT7 is not activated, spontaneously-released SVs do not transit to the reserve pool and rapidly return to the AZ for re-release. In contrast, SVs released during high frequency evoked stimulation when Ca2+ is elevated and SYT7 is engaged, re-enter the RRP at a much slower rate. This mechanism slows re-entry of SVs back into the releasable pool when stimulation rates are high and large numbers of SV proteins are deposited onto the plasma membrane at the same time, allowing time for endosomal sorting that might be required in these conditions. In contrast, SVs released during spontaneous fusion or at low stimulation rates would likely have less need for endosomal re-sorting. Given SYT7 restricts SV transit into the RRP, it provides an activity-regulated Ca2+-triggered switch for redirecting and retaining SVs in a non-fusogenic pool that could facilitate sorting mechanisms (Guan, 2020).

Beyond SV fusion, presynaptic membrane trafficking is required for multiple signaling pathways important for developmental maturation of NMJs. In addition, alterations in neuronal activity or SV endocytosis can result in synaptic undergrowth or overgrowth. No defect was found in synaptic bouton or AZ number, indicating SYT7 does not participate in membrane trafficking pathways that regulate synaptic growth and maturation. However, a decrease in T-bar area and presynaptic Ca2+ influx in Syt7 mutants was found. Although it is unclear how these phenotype arise, they may represent a form of homeostatic plasticity downstream of elevated synaptic transmission. There is also ample evidence that SV distance to Ca2+ channels plays a key role in defining the kinetics of SV release and the size of the RRP, suggesting a change in such coupling in Syt7 mutants might contribute to elevations in P_r and RRP refilling. Further studies will be required to precisely define how SYT7 controls the baseline level of SV release at synapses (Guan, 2020).

Neuromodulator release in neurons requires two functionally redundant calcium sensors

Neuropeptides and neurotrophic factors secreted from dense core vesicles (DCVs) control many brain functions, but the calcium sensors that trigger their secretion remain unknown. This study shows that in mouse hippocampal neurons, DCV fusion is strongly and equally reduced in synaptotagmin-1 (Syt1)- or Syt7-deficient neurons, but combined Syt1/Syt7 deficiency did not reduce fusion further. Cross-rescue, expression of Syt1 in Syt7-deficient neurons, or vice versa, completely restored fusion. Hence, both sensors are rate limiting, operating in a single pathway. Overexpression of either sensor in wild-type neurons confirmed this and increased fusion. Syt1 traveled with DCVs and was present on fusing DCVs, but Syt7 supported fusion largely from other locations. Finally, the duration of single DCV fusion events was reduced in Syt1-deficient but not Syt7-deficient neurons. In conclusion, two functionally redundant calcium sensors drive neuromodulator secretion in an expression-dependent manner. In addition, Syt1 has a unique role in regulating fusion pore duration (van Westen, 2021).

Presynaptic store-operated Ca(2+) entry drives excitatory spontaneous neurotransmission and augments endoplasmic reticulum stress

Store-operated calcium entry (SOCE) is activated by depletion of Ca(2+) from the endoplasmic reticulum (ER) and mediated by stromal interaction molecule (STIM) proteins. This study shows that in rat and mouse hippocampal neurons, acute ER Ca(2+) depletion increases presynaptic Ca(2+) levels and glutamate release through a pathway dependent on STIM2 and the synaptic Ca(2+) sensor synaptotagmin-7 (syt7). In contrast, synaptotagmin-1 (syt1) can suppress SOCE-mediated spontaneous release, and STIM2 is required for the increase in spontaneous release seen during syt1 loss of function. This study also demonstrate that chronic ER stress activates the same pathway leading to syt7-dependent potentiation of spontaneous glutamate release. During ER stress, inhibition of SOCE or syt7-driven fusion partially restored basal neurotransmission and decreased expression of pro-apoptotic markers, indicating that these processes participate in the amplification of ER-stress-related damage. Taken together, it is proposed that presynaptic SOCE links ER stress and augmented spontaneous neurotransmission, which may, in turn, facilitate neurodegeneration (Chanaday, 2021).

Synaptotagmin-7 deficiency induces mania-like behavioral abnormalities through attenuating GluN2B activity

Synaptotagmin-7 (Syt7) probably plays an important role in bipolar-like behavioral abnormalities in mice; however, the underlying mechanisms for this have remained elusive. Unlike antidepressants that cause mood overcorrection in bipolar depression, N-methyl-d-aspartate receptor (NMDAR)-targeted drugs show moderate clinical efficacy, for unexplained reasons. This study identified Syt7 single nucleotide polymorphisms (SNPs) in patients with bipolar disorder and demonstrated that mice lacking Syt7 or expressing the SNPs showed GluN2B-NMDAR dysfunction, leading to antidepressant behavioral consequences and avoidance of overcorrection by NMDAR antagonists. In human induced pluripotent stem cell (iPSC)-derived and mouse hippocampal neurons, Syt7 and GluN2B-NMDARs were localized to the peripheral synaptic region, and Syt7 triggered multiple forms of glutamate release to efficiently activate the juxtaposed GluN2B-NMDARs. Thus, while Syt7 deficiency and SNPs induced GluN2B-NMDAR dysfunction in mice, patient iPSC-derived neurons showed Syt7 deficit-induced GluN2B-NMDAR hypoactivity that was rescued by Syt7 overexpression. Therefore, Syt7 deficits induced mania-like behaviors in mice by attenuating GluN2B activity, which enabled NMDAR antagonists to avoid mood overcorrection (Wang, 2020).

Synaptotagmin-7 is a key factor for bipolar-like behavioral abnormalities in mice

The pathogenesis of bipolar disorder (BD) has remained enigmatic, largely because genetic animal models based on identified susceptible genes have often failed to show core symptoms of spontaneous mood cycling. However, pedigree and induced pluripotent stem cell (iPSC)-based analyses have implicated that dysfunction in some key signaling cascades might be crucial for the disease pathogenesis in a subpopulation of BD patients. It was hypothesized that the behavioral abnormalities of patients and the comorbid metabolic abnormalities might share some identical molecular mechanism. Hence, the expression was investigated of insulin/synapse dually functioning genes in neurons derived from the iPSCs of BD patients and the behavioral phenotype of mice with these genes silenced in the hippocampus. By these means,synaptotagmin-7 (Syt7) was identified as a candidate risk factor for behavioral abnormalities. Syt7 knockout (KO) mice were investigated and nocturnal manic-like and diurnal depressive-like behavioral fluctuations were observed in a majority of these animals, analogous to the mood cycling symptoms of BD. The Syt7 KO mice were investigated with clinical BD drugs including olanzapine and lithium, and it was found that the drug treatments could efficiently regulate the behavioral abnormalities of the Syt7 KO mice. To further verify whether Syt7 deficits existed in BD patients, the plasma samples of 20 BD patients were investigated, and the Syt7 mRNA level was significantly attenuated in the patient plasma compared to the healthy controls. It is therefore concluded that Syt7 is likely a key factor for the bipolar-like behavioral abnormalities (Shen, 2020).

Neuronal regulation of fast Synaptotagmin isoforms controls the relative contributions of synchronous and asynchronous release

Neurotransmitter release can be synchronous and occur within milliseconds of action potential invasion, or asynchronous and persist for tens of milliseconds. The molecular determinants of release kinetics remain poorly understood. It has been hypothesized that asynchronous release dominates when fast Synaptotagmin isoforms are far from calcium channels or when specialized sensors, such as Synaptotagmin 7, are abundant. This study tested these hypotheses for GABAergic projections onto neurons of the inferior olive, where release in different subnuclei ranges from synchronous to asynchronous. Surprisingly, neither of the leading hypotheses accounts for release kinetics. Instead, this study found that rapid Synaptotagmin isoforms are abundant in subnuclei with synchronous release but absent where release is asynchronous. Viral expression of Synaptotagmin 1 transforms asynchronous synapses into synchronous ones. Thus, the nervous system controls levels of fast Synaptotagmin isoforms to regulate release kinetics and thereby controls the ability of synapses to encode spike rates or precise timing (Turecek, 2019).

Synaptotagmin-7-mediated asynchronous release boosts high-fidelity synchronous transmission at a central synapse

Synchronous release triggered by Ca(2+) binding to synaptotagmin-1, -2, or -9 is thought to drive fast synaptic transmission, whereas asynchronous release induced by Ca(2+) binding to synaptotagmin-7 is thought to produce delayed synaptic signaling, enabling prolonged synaptic computations. However, it is unknown whether synaptotagmin-7-dependent asynchronous release performs a physiological function at fast synapses lacking a prolonged signaling mode, such as the calyx of Held synapse. This study shows at the calyx synapse that synaptotagmin-7-dependent asynchronous release indeed does not produce a prolonged synaptic signal after a stimulus train and does not contribute to short-term plasticity, but induces a steady-state, asynchronous postsynaptic current during stimulus trains. This steady-state postsynaptic current does not increase overall synaptic transmission but instead sustains reliable generation of postsynaptic spikes that are precisely time locked to presynaptic spikes. Thus, asynchronous release surprisingly functions, at least at some synapses, to sustain high-fidelity neurotransmission driven by synchronous release during high-frequency stimulus trains (Luo, 2017).

The calcium sensor synaptotagmin 7 is required for synaptic facilitation

It has been known for more than 70 years that synaptic strength is dynamically regulated in a use-dependent manner. At synapses with a low initial release probability, closely spaced presynaptic action potentials can result in facilitation, a short-term form of enhancement in which each subsequent action potential evokes greater neurotransmitter release. Facilitation can enhance neurotransmitter release considerably and can profoundly influence information transfer across synapses, but the underlying mechanism remains a mystery. One proposed mechanism is that a specialized calcium sensor for facilitation transiently increases the probability of release, and this sensor is distinct from the fast sensors that mediate rapid neurotransmitter release. Yet such a sensor has never been identified, and its very existence has been disputed. This study shows that synaptotagmin 7 (Syt7) is a calcium sensor that is required for facilitation at several central synapses. In Syt7-knockout mice, facilitation is eliminated even though the initial probability of release and the presynaptic residual calcium signals are unaltered. Expression of wild-type Syt7 in presynaptic neurons restored facilitation, whereas expression of a mutated Syt7 with a calcium-insensitive C2A domain did not. By revealing the role of Syt7 in synaptic facilitation, these results resolve a longstanding debate about a widespread form of short-term plasticity, and will enable future studies that may lead to a deeper understanding of the functional importance of facilitation (Jackman, 2016).

Synaptotagmin-1 and synaptotagmin-7 trigger synchronous and asynchronous phases of neurotransmitter release

In forebrain neurons, knockout of synaptotagmin-1 blocks fast Ca(2+)-triggered synchronous neurotransmitter release but enables manifestation of slow Ca(2+)-triggered asynchronous release. This study shows using single-cell PCR that individual hippocampal neurons abundantly coexpress two Ca(2+)-binding synaptotagmin isoforms, synaptotagmin-1 and synaptotagmin-7. In synaptotagmin-1-deficient synapses of excitatory and inhibitory neurons, loss of function of synaptotagmin-7 suppressed asynchronous release. This phenotype was rescued by wild-type but not mutant synaptotagmin-7 lacking functional Ca(2+)-binding sites. Even in synaptotagmin-1-containing neurons, synaptotagmin-7 ablation partly impaired asynchronous release induced by extended high-frequency stimulus trains. Synaptotagmins bind Ca(2+) via two C2 domains, the C2A and C2B domains. Surprisingly, synaptotagmin-7 function selectively required its C2A domain Ca(2+)-binding sites, whereas synaptotagmin-1 function required its C2B domain Ca(2+)-binding sites. These data show that nearly all Ca(2+)-triggered release at a synapse is due to synaptotagmins, with synaptotagmin-7 mediating a slower form of Ca(2+)-triggered release that is normally occluded by faster synaptotagmin-1-induced release but becomes manifest upon synaptotagmin-1 deletion (Gacaj, 2013).

Synaptotagmin VII is targeted to secretory organelles in PC12 cells, where it functions as a high-affinity calcium sensor

Synaptotagmin (syt) I is thought to act as a Ca2+ sensor that regulates neuronal exocytosis. Fifteen additional isoforms of syt have been identified, but their functions are less well understood. This study used PC12 cells to test the idea that different isoforms of syt impart cells with distinct metal (i.e., Ca2+, Ba2+, and Sr2+) requirements for secretion. These cells express syt's I and IX (syt IX sometimes referred to as syt V), which have low apparent metal affinities, at much higher levels than syt VII, which this study shows has a relatively high apparent affinity for metals. Syt I and VII partially colocalize on large dense core vesicles and that upregulation of syt VII produces a concomitant increase in the divalent cation sensitivity of catecholamine release from PC12 cells. Furthermore, RNA interference-mediated knockdown of endogenous syt VII reduced the metal sensitivity of release. These data support the hypothesis that the complement of syt's expressed by a cell, in conjunction with their metal affinity, determines the divalent cation sensitivity of exocytosis (Wang, 2005).

Three distinct kinetic groupings of the synaptotagmin family: candidate sensors for rapid and delayed exocytosis

Synaptotagmins (syts) are a family of membrane proteins present on a variety of intracellular organelles. In vertebrates, 16 isoforms of syt have been identified. The most abundant isoform, syt I, appears to function as a Ca2+ sensor that triggers the rapid exocytosis of synaptic vesicles from neurons. The functions of the remaining syt isoforms are less well understood. The cytoplasmic domain of syt I binds membranes in response to Ca2+, and this interaction has been proposed to play a key role in secretion. This study tested the Ca(2+)-triggered membrane-binding activity of the cytoplasmic domains of syts I-XII; eight isoforms tightly bound to liposomes that contained phosphatidylserine as a function of the concentration of Ca2+. The disassembly kinetics of Ca2+.syt.membrane complexes upon rapid mixing with excess Ca2+ chelator were compared; it was found that syts can be classified into three distinct kinetic groups. syts I, II, and III constitute the fast group; syts V, VI, IX, and X make up the medium group; and syt VII exhibits the slowest kinetics of disassembly. Thus, isoforms of syt, which have much slower disassembly kinetics than does syt I, might function as Ca2+ sensors for asynchronous release, which occurs after Ca2+ domains have collapsed. The temperature dependence of Ca2+.syt.membrane assembly and disassembly reactions were compared by using squid and rat syt I. These results indicate that syts have diverged to release Ca2+ and membranes with distinct kinetics (Hui, 2005).

Search PubMed for articles about Drosophila Syt7

Bacaj, T., Wu, D., Yang, X., Morishita, W., Zhou, P., Xu, W., Malenka, R. C. and Sudhof, T. C. (2013). Synaptotagmin-1 and synaptotagmin-7 trigger synchronous and asynchronous phases of neurotransmitter release. Neuron 80(4): 947-959. PubMed ID: 24267651

Chanaday, N. L., Nosyreva, E., Shin, O. H., Zhang, H., Aklan, I., Atasoy, D., Bezprozvanny, I. and Kavalali, E. T. (2021). Presynaptic store-operated Ca(2+) entry drives excitatory spontaneous neurotransmission and augments endoplasmic reticulum stress. Neuron 109(8): 1314-1332 e1315. PubMed ID: 33711258

Guan, Z., Quinones-Frias, M. C., Akbergenova, Y. and Littleton, J. T. (2020). Drosophila Synaptotagmin 7 negatively regulates synaptic vesicle release and replenishment in a dosage-dependent manner. Elife 9. PubMed ID: 32343229

Hui, E., Bai, J., Wang, P., Sugimori, M., Llinas, R. R. and Chapman, E. R. (2005). Three distinct kinetic groupings of the synaptotagmin family: candidate sensors for rapid and delayed exocytosis. Proc Natl Acad Sci U S A 102(14): 5210-5214. PubMed ID: 15793006

Fujii, T., Sakurai, A., Littleton, J. T. and Yoshihara, M. (2021). Synaptotagmin 7 switches short-term synaptic plasticity from depression to facilitation by suppressing synaptic transmission. Sci Rep 11(1): 4059. PubMed ID: 33603074

Jackman, S. L., Turecek, J., Belinsky, J. E. and Regehr, W. G. (2016). The calcium sensor synaptotagmin 7 is required for synaptic facilitation. Nature 529(7584): 88-91. PubMed ID: 26738595

Luo, F. and Sudhof, T. C. (2017). Synaptotagmin-7-Mediated Asynchronous Release Boosts High-Fidelity Synchronous Transmission at a Central Synapse. Neuron 94(4): 826-839 e823. PubMed ID: 28521135

Shen, W., Wang, Q. W., Liu, Y. N., Marchetto, M. C., Linker, S., Lu, S. Y., Chen, Y., Liu, C., Guo, C., Xing, Z., Shi, W., Kelsoe, J. R., Alda, M., Wang, H., Zhong, Y., Sui, S. F., Zhao, M., Yang, Y., Mi, S., Cao, L., Gage, F. H. and Yao, J. (2020). Synaptotagmin-7 is a key factor for bipolar-like behavioral abnormalities in mice. Proc Natl Acad Sci U S A 117(8): 4392-4399. PubMed ID: 32041882

Turecek, J. and Regehr, W. G. (2019). Neuronal Regulation of Fast Synaptotagmin Isoforms Controls the Relative Contributions of Synchronous and Asynchronous Release. Neuron 101(5): 938-949 e934. PubMed ID: 30733150

van Westen, R., Poppinga, J., Diez Arazola, R., Toonen, R. F. and Verhage, M. (2021). Neuromodulator release in neurons requires two functionally redundant calcium sensors. Proc Natl Acad Sci U S A 118(18). PubMed ID: 33903230

Wang, P., Chicka, M. C., Bhalla, A., Richards, D. A. and Chapman, E. R. (2005). Synaptotagmin VII is targeted to secretory organelles in PC12 cells, where it functions as a high-affinity calcium sensor. Mol Cell Biol 25(19): 8693-8702. PubMed ID: 16166648

Wang, Q. W., Lu, S. Y., Liu, Y. N., Chen, Y., Wei, H., Shen, W., Chen, Y. F., Fu, C. L., Wang, Y. H., Dai, A., Huang, X., Gage, F. H., Xu, Q. and Yao, J. (2020). Synaptotagmin-7 deficiency induces mania-like behavioral abnormalities through attenuating GluN2B activity. Proc Natl Acad Sci U S A 117(49): 31438-31447. PubMed ID: 33229564

date revised: 25 May 2021

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