InteractiveFly: GeneBrief

KaiR1D: Biological Overview | References

Homeostatic signaling systems are thought to interface with other forms of plasticity to ensure flexible yet stable levels of neurotransmission. The role of neurotransmitter receptors in this process, beyond mediating neurotransmission itself, is not known. Through a forward genetic screen, the Drosophila kainate-type ionotropic glutamate receptor subunit DKaiR1D was identified as being required for the retrograde, homeostatic potentiation of synaptic strength. DKaiR1D is necessary in presynaptic motor neurons, localized near active zones, and confers robustness to the calcium sensitivity of baseline synaptic transmission. Acute pharmacological blockade of DKaiR1D disrupts homeostatic plasticity, indicating that this receptor is required for the expression of this process, distinct from developmental roles. Finally, this study demonstrates that calcium permeability through DKaiR1D is necessary for baseline synaptic transmission, but not for homeostatic signaling. It is proposed that DKaiR1D is a glutamate autoreceptor that promotes robustness to synaptic strength and plasticity with active zone specificity (Kiragasi, 2017).

The nervous system is endowed with potent and adaptive homeostatic signaling systems that maintain stable functionality despite the myriad changes that occur during neural development and maturation. The importance of homeostatic regulation in the nervous system is underscored by associations with a variety of neurological diseases, yet the genes and mechanisms involved remain enigmatic. A powerful model of presynaptic homeostatic plasticity has been established at the Drosophila neuromuscular junction (NMJ), a model glutamatergic synapse with molecular machinery that parallels central synapses in mammals. Here, genetic and pharmacological manipulations that reduce postsynaptic (muscle) glutamate receptor function trigger a trans-synaptic, retrograde feedback signal to the neuron that increases presynaptic release to precisely compensate for this perturbation. This process is referred to as 'presynaptic homeostatic potentiation' (PHP), because the expression mechanism requires a presynaptic increase in neurotransmitter release (Kiragasi, 2017).

In recent years, forward and candidate genetic approaches have revealed several new and unanticipated genes necessary for PHP expression. While perturbations to the glutamate receptors in muscle are crucial events in the induction of PHP, whether other ionotropic glutamate receptors (iGluRs) function in PHP or are even expressed at the Drosophila NMJ is unknown. Finally, although evidence has emerged that homeostatic modulation is synapse specific, no roles for neurotransmitter receptors or other factors have been found to enable the presynaptic tuning of release efficacy at individual synapses (Kiragasi, 2017).

This study has identified the kainate-type iGluR subunit DKaiR1D to be necessary for PHP expression at the Drosophila NMJ. DKaiR1D is necessary for the calcium sensitivity of baseline synaptic transmission, as well as for the acute and chronic expression of homeostatic potentiation. Recently, the functional reconstitution of DKaiR1D was achieved in heterologous cells, revealing that these receptors form homomeric calcium-permeable channels with atypical pharmacological properties compared to their vertebrate homologs (Li, 2016). This study has found that DKaiR1D is expressed in the nervous system and not the muscle, is present near presynaptic active zones, and is required specifically in motor neurons to enable the robustness of baseline neurotransmission and homeostatic plasticity. It is proposd that glutamate activates DKaiR1D at presynaptic release sites to translate autocrine activity into the robust stabilization of synaptic strength with active zone specificity (Kiragasi, 2017).

This unexpected role for iGluRs in sensing glutamate at presynaptic terminals indicates an autocrine mechanism that responds to glutamate release to adaptively modulate presynaptic activity at individual active zones (Kiragasi, 2017).

Glutamate receptors have diverse functions in modulating presynaptic excitability and short-term plasticity in addition to their established roles in postsynaptic excitation. Similar to what was observed with DKaiR1D at the Drosophila NMJ, rodent iGluRs also localize to presynaptic active zone, are activated by high concentrations of glutamate, and can modulate release during single action potentials (McGuinness, 2010; Pinheiro, 2007; Schmitz, 2000). This suggests conserved autocrine modulatory mechanisms shared between these systems (Kiragasi, 2017).

Rodent autoreceptors are known to modulate presynaptic activity on rapid timescales. In these cases, most of the impact on release is likely to derive from a calcium store-dependent mechanism, or from modulation of the action potential during the repolarization phase, when most of the calcium influx that drives vesicle release occurs (Schneggenburger, 2015). In a similar fashion, activation of presynaptic DKaiR1D during a single action potential could lead to a rapid additional source of presynaptic calcium influx from DKaiR1D itself and/or through modulation of presynaptic membrane potential to drive increased vesicle release. Voltage imaging at the Drosophila NMJ has found the half width of the action potential waveform to be ~5 ms (Ford, 2014), which is sufficient time to be modulated through such a mechanism. Therefore, dynamic changes in voltage or calcium influx at or near active zones could, in principle, drive additional vesicle release during a single action potential. This modulation may be restricted to nearby active zones and compartments relative to the site of glutamate release. Indeed, presynaptic kainate autoreceptors have the capacity to confer short-range, synapse-specific modulation to synaptic transmission (Scott, 2008), while presynaptic ligand-gated ion channels in C. elegans can also rapidly modulate synaptic transmission (Takayanagi-Kiya, 2016). Local activation of DKaiR1D could, therefore, subserve a powerful and flexible means of tuning presynaptic efficacy at or near individual release sites (Kiragasi, 2017).

How does DKaiR1D promote the expression of presynaptic homeostatic plasticity? In contrast to the role of DKaiR1D in baseline release, the data indicate that the DKaiR1D-dependent mechanism that drives PHP is calcium independent. This implies two changes to DKaiR1D functionality that are unique to homeostatic adaptation compared to baseline transmission. First, because presynaptic release is acutely potentiated following application of PhTx, the activity, levels, and/or localization of DKaiR1D receptors must change to acquire a novel influence on neurotransmitter release following PHP induction. The activity of synaptic glutamate receptors can change through associations with additional subunits and auxiliary factors such as Neto (Kim, 2012; Straub, 2011). Furthermore, various forms of plasticity are expressed through dynamic changes in the levels and localization of glutamate receptors trafficking between active zones and endosome pools or extra-synaptic membrane (Anggono, 2012; Kneussel, 2016; Yan, 2013). Indeed, when DKaiR1D is overexpressed in motor neurons, it rescues baseline transmission and PHP expression while localizing to heterogeneous puncta of varying distances relative to active zones. Notably, there is evidence that DKaiR1D interacts with other glutamate receptor subunits in vivo (Karuppudurai, 2014), which may contribute to the pharmacological differences observed in this study compared with the in vitro characterization (Li, 2016) and may also be targets of modulation during PHP (Kiragasi, 2017).

Second, calcium signaling through DKaiR1D differentially drives baseline release and homeostatic plasticity. Therefore, mechanisms distinct from calcium permeability of the channel must contribute to PHP expression. One possibility is that DKaiR1D signals through an undefined metabotropic mechanism during PHP, which might contribute to the ability of the calcium impermeable DKaiR1DR transgene, with reduced conductance (Li, 2016), to rescue PHP expression. Alternatively, an attractive possibility is that following PHP induction, glutamate released from nearby active zones may dynamically modulate the presynaptic membrane potential and/or action potential waveform to promote additional synaptic vesicle release. Indeed, small, sub-threshold depolarizations of the presynaptic resting potential, as small as 5 mV, are sufficient to induce a 2-fold increase in presynaptic release. The timescale of this activity could occur within a few milliseconds as discussed above, and studies at the Drosophila NMJ have revealed that glutamate is released from single synaptic vesicles over timescales of milliseconds. Interestingly, epithelial sodium channels (ENaCs) have been proposed to enable PHP expression through changes in the presynaptic membrane potential, and such a mechanism could be shared by DKaiR1D but gated by glutamate release at individual active zones. Thus, DKaiR1D may serve to homeostatically modulate presynaptic release through modulation of presynaptic voltage and, intriguingly, with active zone specificity (Kiragasi, 2017).

This characterization of DKaiR1D has revealed a role for presynaptic glutamate signaling in homeostatic plasticity. In the mammalian CNS, glutamate signaling drives the adaptive regulation of postsynaptic AMPA receptor insertion and removal, known as homeostatic scaling (Turrigiano, 2008). Further, kainate receptors were recently demonstrated to regulate postsynaptic homeostatic scaling (Yan, 2013). Together with the present study, these results demonstrate that glutamatergic signaling through kainate receptors orchestrate the potent and adaptive homeostatic control of synaptic strength on both sides of the synapse. Future studies will reveal the integration between synaptic glutamate signaling and other forces that modulate synaptic strength to enable robust, flexible, and stable neurotransmission (Kiragasi, 2017).

Neto-alpha controls synapse organization and homeostasis at the Drosophila neuromuscular junction

Glutamate receptor auxiliary proteins control receptor distribution and function, ultimately controlling synapse assembly, maturation, and plasticity. At the Drosophila neuromuscular junction (NMJ), a synapse with both pre- and postsynaptic kainate-type glutamate receptors (KARs), this study shows that the auxiliary protein Neto evolved functionally distinct isoforms to modulate synapse development and homeostasis. Using genetics, cell biology, and electrophysiology, this study demonstrates that Neto-α functions on both sides of the NMJ. In muscle, Neto-α limits the size of the postsynaptic receptor field. In motor neurons (MNs), Neto-α controls neurotransmitter release in a KAR (KaiR1D)-dependent manner. In addition, Neto-α is both required and sufficient for the presynaptic increase in neurotransmitter release in response to reduced postsynaptic sensitivity. This KAR-independent function of Neto-α is involved in activity-induced cytomatrix remodeling. It is proposed that Drosophila ensures NMJ functionality by acquiring two Neto isoforms with differential expression patterns and activities (Han, 2020).

Formation of functional synapses during development and their fine-tuning during plasticity and homeostasis relies on ion channels and their accessory proteins, which control where, when, and how the channels function. Auxiliary proteins are diverse transmembrane proteins that associate with channel complexes and mediate their properties, subcellular distribution, surface expression, synaptic recruitment, and associations with various synaptic scaffolds. Channel subunits have expanded and diversified during evolution to impart different channel biophysical properties, but whether auxiliary proteins have evolved to match channel diversity remains unclear (Han, 2020).

Ionotropic glutamate receptors (iGluRs) mediate neurotransmission at most excitatory synapses in the vertebrate CNS and at the neuromuscular junction (NMJ) of insects and crustaceans and include α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), N-methyl-D-aspartic acid receptors (NMDARs), and kainate receptors (KARs). Sequence analysis of the Drosophila genome identified 14 iGluRs genes that resemble vertebrate AMPARs, NMDARs, and KARs. The fly receptors have strikingly different ligand binding profiles; nonetheless, phylogenetic analysis indicates that two of the Drosophila genes code for AMPARs, two code for NMDARs, and 10 code for subunits of the KAR family, which is highly expanded in insects. In flies and vertebrates, AMPARs and KARs have conserved, dedicated auxiliary proteins. For example, AMPARs rely on Stargazin and its relatives to selectively modulate receptors' gating properties, trafficking, and interactions with scaffolds such as PSD-95-like membrane-associated guanylate kinases. Stargazin is also required for the functional reconstitution of invertebrate AMPARs. KARs are modulated by the Neto (Neuropilin and Tolloid-like) family of proteins, including vertebrate Neto1 and Neto2, C. elegans SOL-2/Neto (Wang et al., 2012), and Drosophila Neto. Neto proteins differentially modulate the gating properties of vertebrate KARs. A role for Neto in the biology of KARs in vivo has been more difficult to assess because of the low levels of KARs and Neto proteins. Nevertheless, vertebrate Netos modulate synaptic recruitment of selective KARs by association with synaptic scaffolds such as GRIP and PSD-95, and the PDZ binding domains of vertebrate KAR/Neto complexes are essential for basal synaptic transmission and long-term potentiation (LTP). Post-translational modifications regulate Neto activities in vitro, but the in vivo relevance of many of these observations remains unknown (Han, 2020).

Drosophila NMJ is an excellent genetic system to probe the repertoire of Neto functions. This glutamatergic synapse appears to rely exclusively on KARs, with one presynaptic and five postsynaptic subunits. Previous work has shown that Drosophila Neto is an obligatory auxiliary subunit of the postsynaptic KAR complexes: in the absence of Neto, postsynaptic KARs fail to cluster at synaptic sites and the animals die as paralyzed embryos. Heterologous reconstitution of postsynaptic KARs in Xenopus oocytes revealed that Neto is required for functional receptors. The fly NMJ contains two glutamate receptor (GluR) complexes (types A and B) with different subunit compositions (either GluRIIA or GluRIIB, plus GluRIIC, GluRIID, and GluRIIE) and distinct properties, regulation, and localization patterns. The postsynaptic response to the fusion of single synaptic vesicles (quantal size) is reduced for NMJs with type B receptors only, and the dose of GluRIIA and GluRIIB is a key determinant of quantal size. The fly NMJ is also a powerful model system to study homeostatic plasticity. Manipulations that decrease the responsiveness of postsynaptic GluR (leading to a decrease in quantal size) trigger a robust compensatory increase in presynaptic neurotransmitter release or quantal content (QC). This increase in QC restores evoked muscle responses to normal levels. A presynaptic KAR, KaiRID, has recently been implicated in basal neurotransmission and presynaptic homeostatic potentiation (PHP) at the larval NMJ (Kiragasi, 2017; Li, 2016). The role of KaiRID in modulation of basal neurotransmission resembles GluK2/GluK3 function as autoreceptors (Pinheiro, 2007). The role of KaiRID in PHP must be indirect, because a mutation that renders this receptor Ca2+ impermeable has no effect on the expression of presynaptic homeostasis (Kiragasi, 2017) (Han, 2020).

The fly NMJ reliance on KARs raises the possibility that Drosophila diversified and maximized its use of Neto proteins. Drosophila Neto encodes two isoforms (Neto-α and Neto-β) with distinct intracellular domains generated by alternative splicing. Both cytoplasmic domains are rich in phosphorylation sites and docking motifs, suggesting rich modulation of Neto/KAR distribution and function. Neto-β, the predominant isoform at the larval NMJ, mediates intracellular interactions that recruit PSD components and enables synaptic stabilization of selective receptor subtypes. Neto-α can rescue viability and receptor clustering defects of Neto null. However, the endogenous functions of Neto-α remain unknown (Han, 2020).

This study shows that Neto-α is key to synapse development and homeostasis and fulfills functions distinct from those of Neto-β. Using isoform-specific mutants and tissue-specific manipulations, it was found that loss of Neto-α in the postsynaptic muscle disrupts GluR fields and produces enlarged PSDs. Loss of presynaptic Neto-α disrupts basal neurotransmission and renders these NMJs unable to express PHP. The different functions of Neto-α were mapped to distinct protein domains and Neto-α was shown to be both required and sufficient for PHP, functioning as a bona fide effector for PHP. It is proposed that Drosophila ensured NMJ functionality by acquiring two Neto isoforms with differential expression patterns and activities (Han, 2020).

This study showed that Neto-α is required in both pre- and postsynaptic compartments for the proper organization and function of the Drosophila NMJ. In muscle, Neto-α limits the size of the postsynaptic receptor field; the PSDs are significantly enlarged in muscle where Neto-α has been perturbed. In MNs, Neto-α is required for two distinct activities: (1) modulation of basal neurotransmission in a KaiRID-dependent manner and (2) effector of presynaptic homeostasis response. This is an extremely rare example of a GluR auxiliary protein that modulates receptors on both sides of a particular synapse and plays a distinct role in homeostatic plasticity (Han, 2020).

Vertebrate KARs depend on Neto proteins for their distribution and function (Copits and Swanson, 2012). Because of their reliance on KARs, Drosophila Neto^null mutants have no functional NMJs (no postsynaptic KARs) and consequently die as paralyzed embryos. Previous work has shown that muscle expression of Neto-ΔCTD, or minimal Neto, at least partly rescues the recruitment and function of KARs at synaptic locations. This study reports that neuronal Neto-ΔCTD also rescues the KaiRID-dependent basal neurotransmission. Thus, Neto-ΔCTD, the part of Neto conserved from worms to humans, seems to represent the Neto core required for KAR modulatory activities (Han, 2020).

The intracellular parts of Neto proteins are highly divergent, likely reflecting the microenvironments in which different Neto proteins operate. Similar to mammalian Neto1 and Neto2, Drosophila Neto-α and Neto-β are differentially expressed in the CNS and have different intracellular domains that mediate distinct functions. These large intracellular domains are rich in putative phosphorylation sites and docking motifs and could further modulate the distribution and function of KARs or serve as signaling hubs and protein scaffolds. Post-translational modifications regulate vertebrate Neto activities in vitro, although the in vivo relevance of these changes remains unknown. The current data demonstrate that Neto-α and Neto-β could not substitute for each other. For example, Neto-β, but not Neto-α, controls the recruitment of PAK, a PSD component that stabilizes selective KAR subtypes at the NMJ, and ensures proper postsynaptic differentiation. Conversely, postsynaptic Neto-β alone cannot maintain a compact PSD size; muscle Neto-α is required for this function. Neto-β cannot fulfill presynaptic functions of Neto-α, presumably because is confined to the somato-dendritic compartment and cannot reach the synaptic terminals. Histology and western blot analyses indicate that Neto-α constitutes less than 1/10th of the net Neto at the Drosophila NMJ. These low levels impaired direct visualization of endogenous Neto-α. Several isoform-specific antibodies have been generated, but they could only detect Neto-α when overexpressed. Similar challenges have been encountered in the vertebrate Neto field (Han, 2020).

The two Neto isoforms are limiting in different synaptic compartments. Neto-β limits the recruitment and synaptic stabilization of postsynaptic KARs. In contrast, several lines of evidence indicate that Neto-α is limiting in MNs. First, overexpression of KaiRID cannot increase basal neurotransmission (Kiragasi, 2017); however, neuronal overexpression of Neto-ΔCTD increases basal neurotransmission, indicating that Neto, but not KaiRID, is limiting in the MNs. Second, neuronal overexpression of Neto-α exacerbates the PHP response to PhTx exposure and even rescues this response in KaiRID^null. These findings suggest that KaiRID's function during PHP is to help traffic and stabilize Neto-α, a low-abundance PHP effector. Similarly, studies in mammals reported that KARs trafficking in the CNS do not require Neto proteins; instead, KARs regulate the surface expression and stabilization of Neto1 and Neto2. Nonetheless, the KAR-mediated stabilization of Neto proteins at CNS synapses supports KAR distribution and function. In flies, KaiRID-dependent Neto-α stabilization at synaptic terminals ensures KAR-dependent function, normal basal neurotransmission, and Neto-α-specific activity as an effector of PHP (Han, 2020).

Previous studies showed that presynaptic KARs regulate neurotransmitter release; however, the site and mechanism of action of presynaptic KARs have been difficult to pin down. This study provides strong evidence for Neto activities at presynaptic terminals. First, Neto-α is both required and sufficient for PHP. It has been shown that the PhTx-induced expression of PHP occurs even when the MN axon is severed. In addition, the signaling necessary for PHP expression is restricted to postsynaptic densities and presynaptic boutons. Second, Neto-ΔCTD, but not Neto-β, rescued basal neurotransmission defects in Neto-αnull. Both variants contain the minimal Neto required for KAR modulation, but only Neto-ΔCTD can reach the presynaptic terminal, whereas Neto-β is restricted to the somato-dendritic compartment. This suggests that Neto-ΔCTD (or Neto-α), together with KaiRID, localizes at presynaptic terminals, where KaiRID could function as an autoreceptor. Finally, upon PhTx exposure, Neto-α enabled fast recruitment of Brp at the active zone. Multiple homeostasis paradigms trigger Brp mobilization, followed by remodeling of presynaptic cytomatrix. These localized activities support Neto-α functioning at presynaptic terminals (Han, 2020).

Rapid application of glutamate to outside-out patches from HEK cells transfected with KaiRID indicated that KaiRID forms rapidly desensitizing channels; addition of Neto increases the desensitization rates and open probability for this channel. Neto-α has a large intracellular domain (250 residues) rich in post-translational modification sites and docking motifs, including putative phosphorylation sites for Ca2+/calmodulin-dependent protein kinase II (CaMKII), protein kinase C (PKC), and protein kinase A (PKA). This intracellular domain may engage in finely tuned interactions that allow Neto-α to (1) further modulate the KaiRID properties and distribution in response to cellular signals and (2) function as an effector of presynaptic homeostasis in response to low postsynaptic GluR activity. Mammalian Neto1 and Neto2 are phosphorylated by multiple kinases in vitro (Lomash, 2017); CaMKII- and PKA-dependent phosphorylation of Neto2 restrict GluK1 targeting to synapses in vivo and in vitro. Similarly, Neto-α may function in a kinase-dependent manner to stabilize KaiRID and/or other presynaptic components. Second, Neto-α may recruit Brp or other presynaptic molecules that mediate activity-related changes in glutamate release at the fly NMJ. Besides Brp, several presynaptic components have been implicated in the control of PHP. They include (1) Cacophony (Cac), the α1 subunit of CaV2-type calcium channels and its auxiliary protein α2Δ-3, that control the presynaptic Ca2+ influx; (2) the signaling molecules Eph, Ephexin, and Cdc42 upstream of Cac; and (3) the BMP pathway components, Wit and Mad, required for retrograde BMP signaling. In addition, expression of PHP requires molecules that regulate vesicle release and the RRP size, such as RIM, Rab3-GAP, Dysbindin, and SNAP25 and Snapin. Recent studies demonstrated that trans-synaptic Semaphorin/Plexin interactions control synaptic scaling in cortical neurons in vertebrates and drive PHP at the fly NMJ (Orr, 2017). Neto-α may interact with one or several such presynaptic molecules and function as an effector of PHP. Future studies on what the Neto-α cytoplasmic domain binds to and how is it modulated by post-translational modifications should provide key insights into the understanding of molecular mechanisms of homeostatic plasticity (Han, 2020).

On the muscle side, Neto-α activities may include (1) engaging scaffolds that limit the PSD size and (2) modulating postsynaptic KAR distribution and function. For example, Neto-α may recruit trans-synaptic complexes such as Ten-a/Ten-m or Nrx/Nlgs that have been implicated in limiting the postsynaptic fields (Banovic, 2010; Mosca, 2012). In particular, DNlg3, like Neto-α, is present in both pre- and postsynaptic compartments and has similar loss-of function phenotypes, including smaller boutons with larger individual PSDs, and reduced EJP amplitudes (Xing, 2014). Neto-α may also indirectly interact with the Drosophila PSD-95 and Dlg and help establish the PSD boundaries. Fly Netos do not have PDZ binding domains, but the postsynaptic Neto/KAR complexes contain GluRIIC, a subunit with a class II PDZ binding domain. It has been reported that mutations that change the NMJ receptors' gating behavior alter their synaptic trafficking and distribution (Petzoldt, 2014). Neto-α could be key to these observations, because it may influence both receptor gating properties and ability to interact with synapse organizers (Han, 2020).

Phylogenetic analyses indicate that Neto-β is the ancestral Neto. In insects, Neto-β is predicted to control NMJ development and function, including recruitment of iGluRs and PSD components, and postsynaptic differentiation. Neto-α appears to be a rapidly evolving isoform present in higher Diptera. This large order of insects is characterized by a rapid expansion of the KAR branch to ten distinct subunits. Insect KARs have unique ligand binding profiles, strikingly different from vertebrate KARs. However, like vertebrate KARs, they all seem to be modulated by Neto proteins. It is speculated that the rapid expansion of KARs forced the diversification of the relevant accessory protein, Neto, and the extension of its repertoire. In flies, the Neto locus acquired an additional exon and consequently an alternative isoform with distinct expression profiles, subcellular distributions, and isoform-specific functions. It will be interesting to investigate how flies differentially regulate the expression and distribution of the two Neto isoforms and control their tissue- and synapse-specific functions. Mammals have five KAR subunits, three of which have multiple splice variants that confer rich regulation. In addition, mammalian Neto proteins have fairly divergent intracellular parts that presumably further integrate cell-specific signals and fine-tune KAR localization and function. In Diptera, KARs have relatively short C tails and thus limited signaling input, whereas Netos have long cytoplasmic domains that could function as scaffolds and signaling hubs. Consequently, most information critical for NMJ assembly and postsynaptic differentiation has been outsourced to the intracellular part of Neto-β. Neto-α-mediated intracellular interactions may also hold key insights into the mechanisms of homeostatic plasticity. This study reveals that Neto functions as a bona fide effector of presynaptic homeostasis (Han, 2020).

The auxiliary glutamate receptor subunit dSol-1 promotes presynaptic neurotransmitter release and homeostatic potentiation

Presynaptic glutamate receptors (GluRs) modulate neurotransmitter release and are physiological targets for regulation during various forms of plasticity. Although much is known about the auxiliary subunits associated with postsynaptic GluRs, far less is understood about presynaptic auxiliary GluR subunits and their functions. At the Drosophila neuromuscular junction, a presynaptic GluR, DKaiR1D, localizes near active zones and operates as an autoreceptor to tune baseline transmission and enhance presynaptic neurotransmitter release in response to diminished postsynaptic GluR functionality, a process referred to as presynaptic homeostatic potentiation (PHP). This study identified an auxiliary subunit that collaborates with DKaiR1D to promote these synaptic functions. This subunit, dSol-1, is the homolog of the Caenorhabditis elegans CUB (Complement C1r/C1s, Uegf, Bmp1) domain protein Sol-1. dSol-1 functions in neurons to facilitate baseline neurotransmission and to enable PHP expression, properties shared with DKaiR1D. Intriguingly, presynaptic overexpression of dSol-1 is sufficient to enhance neurotransmitter release through a DKaiR1D-dependent mechanism. Furthermore, dSol-1 is necessary to rapidly increase the abundance of DKaiR1D receptors near active zones during homeostatic signaling. Together with recent work showing the CUB domain protein Neto2 (see Drosophila Neto) is necessary for the homeostatic modulation of postsynaptic GluRs in mammals, these data demonstrate that dSol-1 is required for the homeostatic regulation of presynaptic GluRs. Thus, it is proposed that CUB domain proteins are fundamental homeostatic modulators of GluRs on both sides of the synapse (Kiragasi, 2020).

Synaptic strength is dynamically tuned during both Hebbian and homeostatic forms of plasticity. One major mechanism that achieves this modulation targets the abundance, localization, and/or functionality of ionotropic glutamate receptors (GluRs). For instance, the expression of long-term potentiation and depression requires bidirectional changes in the abundance of postsynaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors to adjust synaptic strength. Furthermore, some forms of homeostatic plasticity also tune the abundance of N-methyl-D-aspartate (NMDA), AMPA, and kainate receptors at postsynaptic densities to stabilize neuronal activity . Auxiliary subunits associated with GluRs are key factors that control GluR trafficking and dynamics during plasticity, where transmembrane AMPA receptor regulatory protein (TARP), Cornichon, and Neto subunits orchestrate AMPA and kainate receptor function and plasticity. Although much is known about the GluR subtypes and associated auxiliary subunits that regulate GluR trafficking, abundance, and functionality at postsynaptic densities during both Hebbian and homeostatic plasticity, far less is understood about these mechanisms at presynaptic release sites (Kiragasi, 2020).

Presynaptic autoreceptors have emerged as important regulators of neurotransmitter release at glutamatergic synapses. For example, presynaptic kainate receptors are present in hippocampal mossy fibers, where autocrine feedback facilitates neurotransmission during trains of activity. In addition, presynaptic NMDA receptors in the hippocampus mediate presynaptic inhibition in response to excess glutamate release as well as presynaptic facilitation following the induction of long-term potentiation. Furthermore, presynaptic metabotropic receptors play critical roles in various forms of plasticity and can bidirectionally tune presynaptic neurotransmitter release. Finally, ionotropic neurotransmitter receptors at presynaptic terminals can modulate release at neuromuscular junctions (NMJs) in Caenorhabditis elegans and Drosophila. While it is now clear that presynaptic autoreceptors are important bidirectional modulators of neurotransmitter release, how the levels, activity, and localization of these receptors are controlled to establish baseline function, and to what extent they are further modified during plasticity, remains unclear (Kiragasi, 2020).

A kainate-type ionotropic GluR, DKaiR1D, was previously shown to be necessary at the Drosophila NMJ for the expression of presynaptic homeostatic potentiation (PHP). PHP is a fundamental form of synaptic plasticity in which pharmacological and genetic challenges that diminish postsynaptic neurotransmitter receptor functionality trigger a transsynaptic retrograde signal that enhances presynaptic neurotransmitter release to precisely compensate for reduced postsynaptic excitability (19, 20). PHP has been observed at NMJs of Drosophila, rodents, and humans and was recently demonstrated to be rapidly expressed in the mammalian central nervous system. DKaiR1D was identified in a forward genetic screen to be required for the rapid expression of PHP at the fly NMJ. DKaiR1D receptors form homomers that are permeable to both sodium and calcium, localized near presynaptic release sites, and proposed to homeostatically regulate presynaptic voltage following autocrine activation by glutamate. This DKaiR1D-dependent enhancement in neurotransmitter output implies a rapid modulation in the abundance, functionality, and/or localization of these receptors must occur in the course of PHP induction. This regulation could, in principle, be achieved through interactions with auxiliary GluR subunits. However, the precise mechanisms that control DKaiR1D and enable robust and stable neurotransmission at baseline and during plasticity, and whether auxiliary factors are involved, are unknown (Kiragasi, 2020).

A candidate screen of Drosophila GluR modulators and auxiliary subunits was performed to identify potential functions in PHP expression. This effort has discovered an uncharacterized auxiliary GluR subunit that functions in neurons to promote neurotransmitter release and enable homeostatic potentiation. This factor, a homolog of the C. elegans auxiliary GluR subunit Sol-1 (Walker, 2006), contains multiple CUB domains and is structurally similar to the Neto/Sol-2 family of auxiliary GluR subunits. dSol-1 mutants essentially phenocopy DKaiR1D mutants in neurotransmission and PHP. Further experiments demonstrate that dSol-1 functions to homeostatically modulate presynaptic glutamate release by promoting the rapid accumulation of DKaiR1D receptors near active zones. Together, these data indicate that the interactions between CUB domain auxiliary subunits and their associated GluRs are fundamental physiological targets of homeostatic signaling (Kiragasi, 2020).

In the context of baseline neurotransmission, dSol-1 promotes release without measurably changing the abundance or localization of DKaiR1D receptors, indicating a functional role in modulating DKaiR1D activity. However, dSol-1 is necessary during PHP signaling to drive a rapid increase in DKaiR1D receptor abundance at presynaptic terminals. These findings define a CUB domain auxiliary GluR subunit as a central target for the presynaptic modulation of synaptic efficacy and homeostatic plasticity (Kiragasi, 2020).

Several lines of evidence suggest that dSol-1 enhances baseline neurotransmitter release by targeting DKaiR1D receptor functionality. First, dSol-1 promotes baseline neurotransmission in low extracellular Ca2+, a function shared with DKaiR1D. In addition, neurotransmitter release is reduced in dSol-1 mutants and enhanced by neuronal overexpression of dSol-1, indicating a capacity for dSol-1 expression levels to bidirectionally tune release. However, this potentiation in baseline transmission occurs without a significant increase in DKaiR1D receptor abundance, at least when both dSol-1 and DKaiR1D are overexpressed in motor neurons, suggesting a change in DKaiR1D functionality. Interestingly, in C. elegans, Sol-1 regulates GLR1 functionality by modulating channel gating, promoting the open state, and slowing sensitization, without an apparent change in glr1 expression. In heterologous cells, both Sol-1 and dSol-1 promote GLR1 function without altering expression levels. This indicates a potentially conserved function between sol-1 and dSol-1 to confer similar modulations to GluR functionality. In mammals, Neto auxiliary subunits selectively associate with kainate-subtype GluRs, while TARPs such as Stargazin associate with AMPA-type receptors. In Drosophila, Neto is an important auxiliary subunit for the postsynaptic GluRs at the NMJ, which, like DKaiR1D, are generally characterized as non-NMDA, kainate-type GluRs. However, in C. elegans, both Sol-1 and Sol-2/Neto form a complex together with the AMPA receptor subtype GLR1, suggesting some level of promiscuity, at least in invertebrates, between AMPA and kainate GluRs and their auxiliary subunits. One possibility is that at baseline states, a substantial proportion of DKaiR1D receptors do not interact with dSol-1. By increasing levels of dSol-1, more DKaiR1D receptors may become associated with dSol-1, perhaps leading to changes in gating properties that enhance DKaiR1D receptor function. However, the possibility cannot be ruled out that dSol-1 somehow regulates DKaiR1D through a more indirect mechanism. While DKaiR1D receptors can form homomers and traffic to the cell surface when expressed alone in heterologous cells, future in vitro studies will be needed to determine the precise role dSol-1 has in DKaiR1D receptor trafficking and/or functionality (Kiragasi, 2020).

dSol-1 enables PHP expression through a mechanism that is distinct from its role in baseline transmission, although both functions converge on DKaiR1D. The data suggest that PHP signaling leads to a rapid accumulation of DKaiR1D receptors near presynaptic release sites that requires dSol-1. This dSol-1-dependent increase in DKaiR1D levels may be a unique feature of homeostatic signaling in Drosophila, as there is no evidence for worm sol-1 to promote surface levels or changes in GLR1 localization in vivo or in vitro. Although the rapid increase in DKaiR1D receptor levels at synaptic terminals during PHP signaling is surprising, it is not unprecedented. DKaiR1D receptors are present near presynaptic release sites, and many other active zone components rapidly accumulate and/or remodel following PhTx application. An attractive possibility is that DKaiR1D receptors participate in this process of rapid active zone remodeling during PHP signaling. Mechanistically, new protein synthesis of DKaiR1D is unlikely to be involved, as PHP expression and active zone remodeling can occur without new translation. Recently, the lysosomal kinesin adaptor arl-8 and other axonal transport factors were identified to be necessary for the rapid increase in active zone components during PHP signaling. Thus, it is tempting to speculate that DKaiR1D receptors might be cotransported during PHP as part of this pathway. In vertebrates, auxiliary subunits traffic GluRs during synaptic plasticity, so dSol-1 may function similarly in delivering DKaiR1D receptors to the plasma membrane and/or to release sites during PHP signaling. Finally, it is possible that DKaiR1D receptors are constitutively degraded under baseline conditions, and that PHP signaling through dSol-1 inhibits this degradation. The role of protein degradation in PHP signaling has been recently studied in both pre- and postsynaptic compartments at the Drosophila NMJ. Interestingly, inhibition of proteasomal degradation in presynaptic compartments is capable of rapidly enhancing neurotransmission to levels similar to what is observed after overexpression of dSol-1. In both cases, no further increase in neurotransmitter release is observed after PhTx application. While there are apparently distinct roles for dSol-1 in baseline function and homeostatic plasticity, a common point of convergence is DKaiR1D (Kiragasi, 2020).

CUB domains define a structural motif in a large family of extracellular and plasma membrane-associated proteins present in invertebrates to humans. While the specific four extracellular CUB domains that define sol-1/dSol-1 are unique to invertebrates, genes containing multiple CUB domains (between two and eight) are present throughout vertebrate species and function in diverse processes including intercellular signaling, developmental patterning, inflammation, and tumor suppression. CUB domains mediate dimerization and binding to collagen-like regions; this interaction may be relevant to its role in promoting PHP, as the Drosophila collagen member Multiplexin is present in the extracellular matrix and has been proposed to be part of the homeostatic retrograde signaling system. This characterization of dSol-1 contrasts with what is known about another CUB domain auxiliary glutamate receptor in Drosophila, Neto-β. neto-β is highly expressed in the larval muscle, where it is clearly involved in the trafficking and/or stabilization of postsynaptic GluRs at the NMJ. In contrast, dSol-1 is exclusively expressed in the nervous system. Another interesting distinction is that while both dSol-1 and Neto contain multiple extracellular CUB domains and a single transmembrane domain, dSol-1 lacks any intracellular domain while two isoforms of Neto are expressed with one of two alternative intracellular C-terminal cytosolic domains, Neto-α or Neto-β. Neto-β is clearly the major isoform and performs the key functions in controlling postsynaptic GluR levels and composition, while neto-α was recently proposed to function in motor neurons with DKaiR1D and to be necessary for PHP. Interestingly, the C. elegans receptor GLR1 requires the auxiliary subunits Sol-1, Stargazin, and Neto/Sol-2 for functionality in vivo and in vitro. It is therefore possible that Drosophila Neto-α and/or Stargazin-like interact with both dSol-1 and DKaiR1D. In mammals, there is a large body of evidence demonstrating that presynaptic GluRs, including kainate receptors, modulate presynaptic function. While postsynaptic kainate receptors in mammals associate with the CUB domain auxiliary GluR subunit Neto2 to regulate synaptic function and homeostatic plasticity, to what extent Neto2 or other auxiliary subunits function with presynaptic kainate receptors remains enigmatic. This study indicates that CUB domain proteins may be fundamental modulators of GluRs in synaptic function and plasticity on both sides of the synapse (Kiragasi, 2020).

Novel functional properties of Drosophila CNS glutamate receptors

Phylogenetic analysis reveals AMPA, kainate, and NMDA receptor families in insect genomes, suggesting conserved functional properties corresponding to their vertebrate counterparts. However, heterologous expression of the Drosophila kainate receptor DKaiR1D and the AMPA receptor DGluR1A revealed novel ligand selectivity at odds with the classification used for vertebrate glutamate receptor ion channels (iGluRs). DKaiR1D forms a rapidly activating and desensitizing receptor that is inhibited by both NMDA and the NMDA receptor antagonist AP5; crystallization of the KaiR1D ligand-binding domain reveals that these ligands stabilize open cleft conformations, explaining their action as antagonists. Surprisingly, the AMPA receptor DGluR1A shows weak activation by its namesake agonist AMPA and also by quisqualate. Crystallization of the DGluR1A ligand-binding domain reveals amino acid exchanges that interfere with binding of these ligands. The unexpected ligand-binding profiles of insect iGluRs allows classical tools to be used in novel approaches for the study of synaptic regulation (Li, 2016). Video Abstract

Glutamate is the major excitatory neurotransmitter in the vertebrate CNS; its actions are mediated largely via three classes of ionotropic glutamate receptors (iGluRs) named AMPA, kainate, and NMDA receptors. The classification of iGluRs into AMPA, kainate, and NMDA receptors was based on the efforts of medicinal chemists who identified subtype selective heterocyclic amino acids such as AMPA, kainate, and quisqualate and amino acid analogs such as NMDA and 2(R)-amino-5-phosphonopentanoic acid (D-AP5) that act as agonists and antagonists. This work was so successful that the selective action of NMDA and D-AP5 formed the corner stone on which the role of NMDA receptors in synaptic plasticity was established (Li, 2016).

Subsequent cloning of insect iGluRs, which revealed sequence similarity with their vertebrate AMPA, kainate, and NMDA receptor counterparts, suggests that the same series of ligands can be used to investigate their role in CNS function. However, with the exception of the neuromuscular junction (NMJ) of larval Drosophila and the NMJ of adult locusts, the small size and inaccessibility of insect neurons has to date challenged characterization of the functional properties of native insect iGluRs. Sequence analysis of the Drosophila genome identified 14 iGluR genes that resemble vertebrate AMPA, kainate, and NMDA receptors. Transcript profiling revealed that nine of these iGluRs are expressed in the brain, with five expressed at the neuromuscular junction. Very little is known about the structure and functional properties of Drosophila iGluRs and only recently was a functional reconstitution achieved for recombinant Drosophila NMJ iGluRs (Han, 2015). As a result, iGluRs are understudied in model organisms like Drosophila for which powerful genetic techniques have otherwise yielded numerous insights into the molecular neurobiology of synapse development and function (Li, 2016).

Four presumptive Drosophila kainate receptors (Clumsy, DKaiR1C, DKaiR1D, and CG11155) are functionally required for spectral preference behavior and are thought to mediate excitatory synaptic transmission from the second-order neuron Dm8 to the third-order neuron Tm5c (Karuppudurai, 2014). The eye-enriched kainate receptor (EKAR) is expressed in photoreceptors, receiving feedback glutamatergic signals from amacrine cells, but so far, in vitro reconstitution has not been achieved for any of these presumptive kainate receptors. Instead, functional analysis of their role in CNS glutamatergic circuits relies solely on chronic inactivation using genetic mutants and RNAi-mediated knockdown. This study combined electrophysiological, biochemical, and crystallographic analyses to determine receptor activity and ligand specificity of a Drosophila kainate receptor DKaiR1D and a Drosophila AMPA receptor DGluR1A. DKaiR1D was found to form functional homomeric channels in HEK cells and oocytes with pharmacological properties distinct from vertebrate and Drosophila NMJ iGluRs. Crystal structures of DKaiR1D ligand-binding dimer complexes with glutamate, NMDA, and AP5 revealed that only glutamate triggers domain closure and that NMDA and AP5 are antagonists. DGluR1A receptors respond weakly to AMPA and quisqualate; the crystal structure of DGluR1A revealed that the binding of these ligands is hindered by steric occlusion. Thus, despite structural and sequence similarity between insect and vertebrate iGluRs, insect iGluRs do not conform to the pharmacology-based classification of vertebrate iGluRs. However, the agonist/antagonist binding properties of insect iGluRs we report here provide a new approach for acute inactivation/activation in vivo and for dissecting their functions in complex neural circuits (Li, 2016).

This study found that DGluR1A and DKaiR1D, similar to vertebrate GluA1-4 AMPA and GluK1-3 kainate receptor subunits, form homomeric calcium-permeable channels. Based on sequence alignments and the lack of RNA-editing of Drosophila iGluRs mRNA at their Q/R sites, it is likely that most insect iGluRs are calcium permeable and that they are inhibited by endogenous cytoplasmic polyamines and by spider venom polyamine toxins. It is noted that homomeric DKaiR1D has a very fast desensitization rate, while for DGluR1A, fit has not yet been possible to achieve sufficient expression to allow recording from outside-out patches with rapid perfusion. Structural analyses revealed that DKaiR1D LBD dimers contain conserved Na+ ion binding sites characteristic of vertebrate kainate receptors, but these appear to not strongly modulate the activation or desensitization of KaiR1D, perhaps because the Cl- binding site found in vertebrate kainate receptors is absent in insect kainate receptors. Sequence analysis revealed that this separation of Na+ and Cl- binding sites in KaiR1D subunits occurs in all insect species examined. Structure-aided sequence analysis also reveals that in the other three groups of fly kainate receptors, different combinations of amino acid substitutions destroy or significantly weaken both the Na+ and Cl− binding sites. Thus, the allosteric modulation by both anions and cations that is characteristic of vertebrate kainate receptors is uncoupled in insect kainate receptors and for the majority of cases both ion binding sites are eliminated (Li, 2016).

Previous phylogenetic studies suggest that most bilateria, including insects, worms, and vertebrates, have three major classes of cation-selective iGluRs, corresponding to vertebrate AMPA, kainate, and NMDA receptors. The current analysis reveals that in insects, the kainate receptor family is expanded into four groups, while a prior phylogenetic analysis revealed that in Mollusca the AMPA receptor family is expanded (Alberstein, 2015). At the neuromuscular junction of Drosophila and the locust Schistocerca gregaria, iGluRs have been extensively studied, in part serving as a surrogate model for CNS iGluRs. Interestingly, this study found that late in evolution, in higher Diptera, the five Drosophila NMJ iGluR subunits, GluRIIA-E, were derived from two separate kainate receptor subtypes, KaiR1C and Clumsy. Thus, despite their unique obligate heterotetrameric subunit stoichiometry and insensitivity to kainate (Han, 2015), fly NMJ iGluRs evolved from ancestral kainate-sensitive iGluRs, and it is likely that in other insect species iGluRs related to KaiR1C and Clumsy may function in both the CNS and NMJ (Li, 2016).

Although phylogenetic analysis supports classification of Drosophila and other insect iGluRs into the familiar AMPA, kainate, and NMDA receptor families, the current results reveal unexpected differences in their ligand-binding properties. The most dramatic change was the conversion of NMDA from an agonist for vertebrate NMDA receptors to an antagonist for Drosophila KaiR1D, a kainate receptor that is also inhibited by both isomers of AP5, while D-AP5, but not L-AP5 acts as a potent vertebrate NMDA receptor antagonist. The crystal structures solved in this study for the DKaiR1D LBD establish that NMDA and AP5 inhibit activation of DKaiR1D by stabilizing an open cleft conformation, similar to the action of competitive antagonists for vertebrate iGluRs from each of the three major families. In addition, NMDA triggered separation of the upper lobes of the DKaiR1D LBD dimer assembly, a conformational change that occurs during desensitization of vertebrate AMPA and kainate receptors, may in addition contribute to the inhibitory action of NMDA on DKaiR1D (Li, 2016).

AMPA receptors were initially identified by and named in response to their activation by quisqualic acid, a glutamate bioisostere that is nonselective and which activates all of the major vertebrate iGluR subtypes, in addition to acting as a potent agonist for G protein-coupled glutamate receptors. Prior to the cloning of GluA1–4 subunits, the so-called quisqualate receptors were renamed AMPA receptors, following the synthesis of AMPA and the discovery that it was a highly selective agonist, without activity at kainate, NMDA, or G protein-coupled glutamate receptors. These serendipitous events in the history of the development of selective ligands for iGluR subtypes were strongly reinforced when a large family of vertebrate iGluR subunits were cloned, and it was discovered that these encoded discrete families of iGluR subtypes, each with high sequence identity, the ligand-binding properties of which corresponded to the familiar AMPA, kainate, and NMDA receptor subtypes. The current experiments reveal an unexpected breakdown of the classification scheme for Drosophila and most likely other insect species iGluRs (Li, 2016).

With the plethora of genetic tools and advanced connectome analyses, Drosophila has emerged as a key model organism for studying the circuit basis of behavior. It is now evident that like vertebrates, glutamatergic synapses are abundantly utilized in fly CNS circuits. Functional and structural analyses revealed that Drosophila iGluRs have agonist and antagonist selectivity very different from those of vertebrates, indicating that sequence and structural homology does not confer conserved pharmacological properties. However, the unique pharmacology of Drosophila iGluRs reported in this study has proven of use to reveal the role of KaiR1D in presynaptic homeostasis. It is envisioned that appropriate use of pharmacological tools in combination with powerful fly genetics will greatly aid studies of complex neural circuits in Drosophila (Li, 2016).

Search PubMed for articles about Drosophila DKaiR1D

Alberstein, R., Grey, R., Zimmet, A., Simmons, D. K. and Mayer, M. L. (2015). Glycine activated ion channel subunits encoded by ctenophore glutamate receptor genes. Proc Natl Acad Sci U S A 112(44): E6048-6057. PubMed ID: 26460032

Anggono, V. and Huganir, R. L. (2012). Regulation of AMPA receptor trafficking and synaptic plasticity. Curr Opin Neurobiol 22(3): 461-469. PubMed ID: 22217700

Banovic, D., Khorramshahi, O., Owald, D., Wichmann, C., Riedt, T., Fouquet, W., Tian, R., Sigrist, S. J. and Aberle, H. (2010). Drosophila neuroligin 1 promotes growth and postsynaptic differentiation at glutamatergic neuromuscular junctions. Neuron 66(5): 724-738. PubMed ID: 20547130

Ford, K. J. and Davis, G. W. (2014). Archaerhodopsin voltage imaging: synaptic calcium and BK channels stabilize action potential repolarization at the Drosophila neuromuscular junction. J Neurosci 34(44): 14517-14525. PubMed ID: 25355206

Han, T. H., Dharkar, P., Mayer, M. L. and Serpe, M. (2015). Functional reconstitution of Drosophila melanogaster NMJ glutamate receptors. Proc Natl Acad Sci U S A 112(19): 6182-6187. PubMed ID: 25918369

Han, T. H., Vicidomini, R., Ramos, C. I., Wang, Q., Nguyen, P., Jarnik, M., Lee, C. H., Stawarski, M., Hernandez, R. X., Macleod, G. T. and Serpe, M. (2020). Neto-alpha controls synapse organization and homeostasis at the Drosophila neuromuscular junction. Cell Rep 32(1): 107866. PubMed ID: 32640231

Karuppudurai, T., Lin, T. Y., Ting, C. Y., Pursley, R., Melnattur, K. V., Diao, F., White, B. H., Macpherson, L. J., Gallio, M., Pohida, T. and Lee, C. H. (2014). A hard-wired glutamatergic circuit pools and relays UV signals to mediate spectral preference in Drosophila. Neuron 81(3): 603-615. PubMed ID: 24507194

Kim, Y.J., Bao, H., Bonanno, L., Zhang, B., and Serpe, M. (2012). Drosophila Neto is essential for clustering glutamate receptors at the neuromuscular junction. Genes Dev. 26: 974-987. PubMed ID: 22499592

Kiragasi, B., Wondolowski, J., Li, Y. and Dickman, D. K. (2017). A presynaptic glutamate receptor subunit confers robustness to neurotransmission and homeostatic potentiation. Cell Rep 19(13): 2694-2706. PubMed ID: 28658618

Kneussel, M. and Hausrat, T. J. (2016). Postsynaptic neurotransmitter receptor reserve pools for synaptic potentiation. Trends Neurosci 39(3): 170-182. PubMed ID: 26833258

Li, Y., Dharkar, P., Han, T. H., Serpe, M., Lee, C. H. and Mayer, M. L. (2016). Novel functional properties of Drosophila CNS glutamate receptors. Neuron 92(5): 1036-1048. PubMed ID: 27889096

Lomash, R. M., Sheng, N., Li, Y., Nicoll, R. A. and Roche, K. W. (2017). Phosphorylation of the kainate receptor (KAR) auxiliary subunit Neto2 at serine 409 regulates synaptic targeting of the KAR subunit GluK1. J Biol Chem 292(37): 15369-15377. PubMed ID: 28717010

McGuinness, L., Taylor, C., Taylor, R. D., Yau, C., Langenhan, T., Hart, M. L., Christian, H., Tynan, P. W., Donnelly, P. and Emptage, N. J. (2010). Presynaptic NMDARs in the hippocampus facilitate transmitter release at theta frequency. Neuron 68(6): 1109-1127. PubMed ID: 21172613

Kiragasi, B., Goel, P., Perry, S., Han, Y., Li, X. and Dickman, D. (2020). The auxiliary glutamate receptor subunit dSol-1 promotes presynaptic neurotransmitter release and homeostatic potentiation. Proc Natl Acad Sci U S A 117(41): 25830-25839. PubMed ID: 32973097

Mosca, T. J., Hong, W., Dani, V. S., Favaloro, V. and Luo, L. (2012). Trans-synaptic Teneurin signalling in neuromuscular synapse organization and target choice. Nature 484(7393): 237-241. PubMed ID: 22426000

Orr, B. O., Gorczyca, D., Younger, M. A., Jan, L. Y., Jan, Y. N. and Davis, G. W. (2017). Composition and control of a Deg/ENaC channel during presynaptic homeostatic plasticity. Cell Rep 20(8): 1855-1866. PubMed ID: 28834749

Petzoldt, A. G., Lee, Y. H., Khorramshahi, O., Reynolds, E., Plested, A. J., Herzel, H. and Sigrist, S. J. (2014). Gating characteristics control glutamate receptor distribution and trafficking in vivo. Curr Biol 24(17): 2059-2065. PubMed ID: 25131677

Pinheiro, P. S., Perrais, D., Coussen, F., Barhanin, J., Bettler, B., Mann, J. R., Malva, J. O., Heinemann, S. F. and Mulle, C. (2007). GluR7 is an essential subunit of presynaptic kainate autoreceptors at hippocampal mossy fiber synapses. Proc Natl Acad Sci U S A 104(29): 12181-12186. PubMed ID: 17620617

Schmitz, D., Frerking, M. and Nicoll, R. A. (2000). Synaptic activation of presynaptic kainate receptors on hippocampal mossy fiber synapses. Neuron 27(2): 327-338. PubMed ID: 10985352

Schneggenburger, R. and Rosenmund, C. (2015). Molecular mechanisms governing Ca(2+) regulation of evoked and spontaneous release. Nat Neurosci 18(7): 935-941. PubMed ID: 26108721

Scott, R., Lalic, T., Kullmann, D. M., Capogna, M. and Rusakov, D. A. (2008). Target-cell specificity of kainate autoreceptor and Ca2+-store-dependent short-term plasticity at hippocampal mossy fiber synapses. J Neurosci 28(49): 13139-13149. PubMed ID: 19052205

Straub, C., Hunt, D. L., Yamasaki, M., Kim, K. S., Watanabe, M., Castillo, P. E. and Tomita, S. (2011). Distinct functions of kainate receptors in the brain are determined by the auxiliary subunit Neto1. Nat Neurosci 14(7): 866-873. PubMed ID: 21623363

Takayanagi-Kiya, S., Zhou, K. and Jin, Y. (2016). Release-dependent feedback inhibition by a presynaptically localized ligand-gated anion channel. Elife 5. PubMed ID: 27782882

Turrigiano, G. G. (2008). The self-tuning neuron: synaptic scaling of excitatory synapses. Cell 135(3): 422-435. PubMed ID: 18984155

Walker, C. S., Francis, M. M., Brockie, P. J., Madsen, D. M., Zheng, Y. and Maricq, A. V. (2006). Conserved SOL-1 proteins regulate ionotropic glutamate receptor desensitization. Proc Natl Acad Sci U S A 103(28): 10787-10792. PubMed ID: 16818875

Xing, G., Gan, G., Chen, D., Sun, M., Yi, J., Lv, H., Han, J. and Xie, W. (2014). Drosophila neuroligin3 regulates neuromuscular junction development and synaptic differentiation. J Biol Chem 289(46): 31867-31877. PubMed ID: 25228693

Yan, D., Yamasaki, M., Straub, C., Watanabe, M. and Tomita, S. (2013). Homeostatic control of synaptic transmission by distinct glutamate receptors. Neuron 78(4): 687-699. PubMed ID: 23719165

date revised: 20 December 2020

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