Homeostatic synaptic plasticity: local and global mechanisms for stabilizing neuronal function

Neural circuits must maintain stable function in the face of many plastic challenges, including changes in synapse number and strength, during learning and development. Recent work has shown that these destabilizing influences are counterbalanced by homeostatic plasticity mechanisms that act to stabilize neuronal and circuit activity. One such mechanism is synaptic scaling, which allows neurons to detect changes in their own firing rates through a set of calcium-dependent sensors that then regulate receptor trafficking to increase or decrease the accumulation of glutamate receptors at synaptic sites. Additional homeostatic mechanisms may allow local changes in synaptic activation to generate local synaptic adaptations, and network-wide changes in activity to generate network-wide adjustments in the balance between excitation and inhibition. The signaling pathways underlying these various forms of homeostatic plasticity are currently under intense scrutiny, and although dozens of molecular pathways have now been implicated in homeostatic plasticity, a clear picture of how homeostatic feedback is structured at the molecular level has not yet emerged. On a functional level, neuronal networks likely use this complex set of regulatory mechanisms to achieve homeostasis over a wide range of temporal and spatial scales.     

Mechanisms underlying the rapid induction and sustained expression of synaptic homeostasis

Homeostatic signaling systems are thought to interface with the mechanisms of neural plasticity to achieve stable yet flexible neural circuitry. However, the time course, molecular design, and implementation of homeostatic signaling remain poorly defined. Here we demonstrate that a homeostatic increase in presynaptic neurotransmitter release can be induced within minutes following postsynaptic glutamate receptor blockade. The rapid induction of synaptic homeostasis is independent of new protein synthesis and does not require evoked neurotransmission, indicating that a change in the efficacy of spontaneous quantal release events is sufficient to trigger the induction of synaptic homeostasis. Finally, both the rapid induction and the sustained expression of synaptic homeostasis are blocked by mutations that disrupt the pore-forming subunit of the presynaptic Ca(V)2.1 calcium channel encoded by cacophony. These data confirm the presynaptic expression of synaptic homeostasis and implicate presynaptic Ca(V)2.1 in a homeostatic retrograde signaling system.     

Presynaptic LTP and LTD of Excitatory and Inhibitory Synapses

Ubiquitous forms of long-term potentiation (LTP) and depression (LTD) are caused by enduring increases or decreases in neurotransmitter release. Such forms or presynaptic plasticity are equally observed at excitatory and inhibitory synapses and the list of locations expressing presynaptic LTP and LTD continues to grow. In addition to the mechanistically distinct forms of postsynaptic plasticity, presynaptic plasticity offers a powerful means to modify neural circuits. A wide range of induction mechanisms has been identified, some of which occur entirely in the presynaptic terminal, whereas others require retrograde signaling from the postsynaptic to presynaptic terminals. In spite of this diversity of induction mechanisms, some common induction rules can be identified across synapses. Although the precise molecular mechanism underlying long-term changes in transmitter release in most cases remains unclear, increasing evidence indicates that presynaptic LTP and LTD can occur in vivo and likely mediate some forms of learning.At several excitatory and inhibitory synapses, neuronal activity can trigger enduring increases or decreases in neurotransmitter release, thereby producing long-term potentiation (LTP) or long-term depression (LTD) of synaptic strength, respectively. In the last decade, many studies have revealed that these forms of plasticity are ubiquitously expressed in the mammalian brain, and accumulating evidence indicates that they may underlie behavioral adaptations occurring in vivo. These studies have also uncovered a wide range of induction mechanisms, which converge on the presynaptic terminal where an enduring modification in the neurotransmitter release process takes place. Interestingly, presynaptic forms of LTP/LTD can coexist with classical forms of postsynaptic plasticity. Such diversity expands the dynamic range and repertoire by which neurons modify their synaptic connections. This review discusses mechanistic aspects of presynaptic LTP and LTD at both excitatory and inhibitory synapses in the mammalian brain, with an emphasis on recent findings.     

Rab3-GAP controls the progression of synaptic homeostasis at a late stage of vesicle release

Homeostatic signaling systems stabilize neural function through the modulation of neurotransmitter receptor abundance, ion channel density, and presynaptic neurotransmitter release. Molecular mechanisms that drive these changes are being unveiled. In theory, molecular mechanisms may also exist to oppose the induction or expression of homeostatic plasticity, but these mechanisms have yet to be explored. In an ongoing electrophysiology-based genetic screen, we have tested 162 new mutations for genes involved in homeostatic signaling at the Drosophila NMJ. This screen identified a mutation in the rab3-GAP gene. We show that Rab3-GAP is necessary for the induction and expression of synaptic homeostasis. We then provide evidence that Rab3-GAP relieves an opposing influence on homeostasis that is catalyzed by Rab3 and which is independent of any change in NMJ anatomy. These data define roles for Rab3-GAP and Rab3 in synaptic homeostasis and uncover a mechanism, acting at a late stage of vesicle release, that opposes the progression of homeostatic plasticity.     

Differential regulation of spontaneous and evoked neurotransmitter release at central synapses

Recent studies have begun to scrutinize the presynaptic machinery and vesicle populations that give rise to action potential evoked and spontaneous forms of neurotransmitter release. In several cases this work produced unexpected results which lend support to the notion that regulation, mechanisms, postsynaptic targets and possibly presynaptic origins of evoked and spontaneous neurotransmitter release differ. Furthermore, the list of regulatory pathways that impact spontaneous and evoked release in a divergent manner is rapidly growing. These findings challenge our classical views on the relationship between evoked and spontaneous neurotransmission. In contrast to the well-characterized neuromodulatory pathways that equally suppress or augment all forms of neurotransmitter release, molecular substrates specifically controlling spontaneous release remain unclear. In this review, we outline possible mechanisms that may underlie the differential regulation of distinct forms of neurotransmission and help demultiplex complex neuronal signals and generate parallel signaling events at their postsynaptic targets.     

Signal regulatory proteins (SIRPS) are secreted presynaptic organizing molecules

Formation of chemical synapses requires exchange of organizing signals between the synaptic partners. Using synaptic vesicle aggregation in cultured neurons as a marker of presynaptic differentiation, we purified candidate presynaptic organizers from mouse brain. A major bioactive species was the extracellular domain of signal regulatory protein alpha (SIRP-alpha), a transmembrane immunoglobulin superfamily member concentrated at synapses. The extracellular domain of SIRP-alpha is cleaved and shed in a developmentally regulated manner. The presynaptic organizing activity of SIRP-alpha is mediated in part by CD47. SIRP-alpha homologues, SIRP-beta and -gamma also have synaptic vesicle clustering activity. The effects of SIRP-alpha are distinct from those of another presynaptic organizer, FGF22: the two proteins induced vesicle clusters of different sizes, differed in their ability to promote neurite branching, and acted through different receptors and signaling pathways. SIRP family proteins may act together with other organizing molecules to pattern synapses.     

Presynaptic N-type calcium channels regulate synaptic growth

Voltage-gated calcium channels couple changes in membrane potential to neuronal functions regulated by calcium, including neurotransmitter release. Here we report that presynaptic N-type calcium channels not only control neurotransmitter release but also regulate synaptic growth at Drosophila neuromuscular junctions. In a screen for behavioral mutants that disrupt synaptic transmission, an allele of the N-type calcium channel locus (Dmca1A) was identified that caused synaptic undergrowth. The underlying molecular defect was identified as a neutralization of a charged residue in the third S4 voltage sensor. RNA interference reduction of N-type calcium channel expression also reduced synaptic growth. Hypomorphic mutations in syntaxin-1A or n-synaptobrevin, which also disrupt neurotransmitter release, did not affect synapse proliferation at the neuromuscular junction, suggesting calcium entry through presynaptic N-type calcium channels, not neurotransmitter release per se, is important for synaptic growth. The reduced synapse proliferation in Dmca1A mutants is not due to increased synapse retraction but instead reflects a role for calcium influx in synaptic growth mechanisms. These results suggest N-type channels participate in synaptic growth through signaling pathways that are distinct from those that mediate neurotransmitter release. Linking presynaptic voltage-gated calcium entry to downstream calcium-sensitive synaptic growth regulators provides an efficient activity-dependent mechanism for modifying synaptic strength.     

Protein targeting and calcium signaling microdomains in neuronal cells

Over the last several years, a number of optical imaging, physiological, and molecular studies have clarified the mechanisms underlying differential calcium signaling in the postsynaptic neuron. These studies have revealed the existence of membrane-associated calcium microdomains, which are often specifically coupled to distinct protein signaling pathways. In this review, we discuss how these signaling microdomains are organized and regulated, emphasizing the structural and molecular features of synaptic protein complexes containing the metabotropic and N-methyl-D-aspartate (NMDA) glutamate receptors and the L-type voltage-dependent calcium channels (VDCCs). We conclude with a discussion of how these different signaling complexes may interact with one another, relationships which may be important in orchestrating the complex calcium signaling underlying developmental and activity-dependent changes in synaptic function.     

Calcium signaling in neurodegeneration

Calcium is a key signaling ion involved in many different intracellular and extracellular processes ranging from synaptic activity to cell-cell communication and adhesion. The exact definition at the molecular level of the versatility of this ion has made overwhelming progress in the past several years and has been extensively reviewed. In the brain, calcium is fundamental in the control of synaptic activity and memory formation, a process that leads to the activation of specific calcium-dependent signal transduction pathways and implicates key protein effectors, such as CaMKs, MAPK/ERKs, and CREB. Properly controlled homeostasis of calcium signaling not only supports normal brain physiology but also maintains neuronal integrity and long-term cell survival. Emerging knowledge indicates that calcium homeostasis is not only critical for cell physiology and health, but also, when deregulated, can lead to neurodegeneration via complex and diverse mechanisms involved in selective neuronal impairments and death. The identification of several modulators of calcium homeostasis, such as presenilins and CALHM1, as potential factors involved in the pathogenesis of Alzheimer's disease, provides strong support for a role of calcium in neurodegeneration. These observations represent an important step towards understanding the molecular mechanisms of calcium signaling disturbances observed in different brain diseases such as Alzheimer's, Parkinson's, and Huntington's diseases.     

Coactivation of pre- and postsynaptic signaling mechanisms determines cell-specific spike-timing-dependent plasticity

Synapses may undergo long-term increases or decreases in synaptic strength dependent on critical differences in the timing between pre-and postsynaptic activity. Such spike-timing-dependent plasticity (STDP) follows rules that govern how patterns of neural activity induce changes in synaptic strength. Synaptic plasticity in the dorsal cochlear nucleus (DCN) follows Hebbian and anti-Hebbian patterns in a cell-specific manner. Here we show that these opposing responses to synaptic activity result from differential expression of two signaling pathways. Ca2+/calmodulin-dependent protein kinase II (CaMKII) signaling underlies Hebbian postsynaptic LTP in principal cells. By contrast, in interneurons, a temporally precise anti-Hebbian synaptic spike-timing rule results from the combined effects of postsynaptic CaMKII-dependent LTP and endocannabinoid-dependent presynaptic LTD. Cell specificity in the circuit arises from selective targeting of presynaptic CB1 receptors in different axonal terminals. Hence, pre- and postsynaptic sites of expression determine both the sign and timing requirements of long-term plasticity in interneurons.     

Synapse development in health and disease

Recent insights into the genetic basis of neurological disease have led to the hypothesis that molecular pathways involved in synaptic growth, development, and stability are perturbed in a variety of mental disorders. Formation of a functional synapse is a complex process requiring stabilization of initial synaptic contacts by adhesive protein interactions, organization of presynaptic and postsynaptic specializations by scaffolding proteins, regulation of growth by intercellular signaling pathways, reorganization of the actin cytoskeleton, and proper endosomal trafficking of synaptic growth signaling complexes. Many neuropsychiatric disorders, including autism, schizophrenia, and intellectual disability, have been linked to inherited mutations which perturb these processes. Our understanding of the basic biology of synaptogenesis is therefore critical to unraveling the pathogenesis of neuropsychiatric disorders.     

Protein-protein interactions and protein modules in the control of neurotransmitter release

Information transfer among neurons is operated by neurotransmitters stored in synaptic vesicles and released to the extracellular space by an efficient process of regulated exocytosis. Synaptic vesicles are organized into two distinct functional pools, a large reserve pool in which vesicles are restrained by the actin-based cytoskeleton, and a quantitatively smaller releasable pool in which vesicles approach the presynaptic membrane and eventually fuse with it on stimulation. Both synaptic vesicle trafficking and neurotransmitter release depend on a precise sequence of events that include release from the reserve pool, targeting to the active zone, docking, priming, fusion and endocytotic retrieval of synaptic vesicles. These steps are mediated by a series of specific interactions among cytoskeletal, synaptic vesicle, presynaptic membrane and cytosolic proteins that, by acting in concert, promote the spatial and temporal regulation of the exocytotic machinery. The majority of these interactions are mediated by specific protein modules and domains that are found in many proteins and are involved in numerous intracellular processes. In this paper, the possible physiological role of these multiple protein-protein interactions is analysed, with ensuing updating and clarification of the present molecular model of the process of neurotransmitter release.     

Signaling by synaptogenic molecules

Multiple signaling pathways initiate and specify the formation of synapses in the central nervous system. General principles that organize nascent synapses have emerged from the studies in multiple model organisms. These include the synapse-organizing roles of dedicated synaptic adhesion molecules, synaptic signaling following receptor-ligand interactions, and the regulation of synapse formation by secreted molecules. Intracellularly, a range of effectors subsequently regulates signaling steps and cytoskeletal changes. Together, a blueprint of synapse formation is emerging into which these distinct signaling steps will need to be integrated temporally and spatially.     

Role of lipid microdomains in P/Q-type calcium channel (Cav2.1) clustering and function in presynaptic membranes

Lipid microdomains can selectively include or exclude proteins and may be important in a variety of functions such as protein sorting, cell signaling, and synaptic transmission. The present study demonstrates that two different voltage-gated calcium channels, which both interact with soluble N-ethyl-maleimide-sensitive fusion protein attachment protein receptor (SNARE) proteins but have distinct subcellular distributions and roles in synaptic transmission, are differently distributed in lipid microdomains; presynaptic P/Q (Cav2.1) but not Lc (Cav1.2) calcium channel subtypes are mainly accumulated in detergent-insoluble complexes. The immunoisolation of multiprotein complexes from detergent-insoluble or detergent-soluble fractions shows that the alpha1A subunits of Cav2.1 colocalize and interact with SNARE complexes in lipid microdomains. The altered organization of these microdomains caused by saponin and methyl-beta-cyclodextrin treatment largely impairs the buoyancy and distribution of Cav2.1 channels and SNAREs in flotation gradients. On the other hand, cholesterol reloading partially reverses the drug effects. Methyl-beta-cyclodextrin treatment alters the colocalization of Cav2.1 with the proteins of the exocytic machinery and also impairs calcium influx in nerve terminals. These results show that lipid microdomains in presynaptic terminals are important in organizing membrane sites specialized for synaptic vesicle exocytosis. The cholesterol-enriched microdomains contribute to optimizing the compartmentalization of exocytic machinery and the calcium influx that triggers synaptic vesicle exocytosis.     

Retrograde control of synaptic transmission by postsynaptic CaMKII at the Drosophila neuromuscular junction

Retrograde signaling plays an important role in synaptic homeostasis, growth, and plasticity. A retrograde signal at the neuromuscular junction (NMJ) of Drosophila controls the homeostasis of neurotransmitter release. Here, we show that this retrograde signal is regulated by the postsynaptic activity of Ca2+/calmodulin-dependent protein kinase II (CaMKII). Reducing CaMKII activity in muscles enhances the signal and increases neurotransmitter release, while constitutive activation of CaMKII in muscles inhibits the signal and decreases neurotransmitter release. Postsynaptic inhibition of CaMKII increases the number of presynaptic, vesicle-associated T bars at the active zones. Consistently, we show that glutamate receptor mutants also have a higher number of T bars; this increase is suppressed by postsynaptic activation of CaMKII. Furthermore, we demonstrate that presynaptic BMP receptor wishful thinking is required for the retrograde signal to function. Our results indicate that CaMKII plays a key role in the retrograde control of homeostasis of synaptic transmission at the NMJ of Drosophila.     

Postsynaptic regulation of synaptic plasticity by synaptotagmin 4 requires both C2 domains

Ca²⁺ influx into synaptic compartments during activity is a key mediator of neuronal plasticity. Although the role of presynaptic Ca²⁺ in triggering vesicle fusion though the Ca²⁺ sensor synaptotagmin 1 (Syt 1) is established, molecular mechanisms that underlie responses to postsynaptic Ca²⁺ influx remain unclear. In this study, we demonstrate that fusion-competent Syt 4 vesicles localize postsynaptically at both neuromuscular junctions (NMJs) and central nervous system synapses in Drosophila melanogaster. Syt 4 messenger RNA and protein expression are strongly regulated by neuronal activity, whereas altered levels of postsynaptic Syt 4 modify synaptic growth and presynaptic release properties. Syt 4 is required for known forms of activity-dependent structural plasticity at NMJs. Synaptic proliferation and retrograde signaling mediated by Syt 4 requires functional C2A and C2B Ca²⁺–binding sites, as well as serine 284, an evolutionarily conserved substitution for a key Ca²⁺-binding aspartic acid found in other synaptotagmins. These data suggest that Syt 4 regulates activity-dependent release of postsynaptic retrograde signals that promote synaptic plasticity, similar to the role of Syt 1 as a Ca²⁺ sensor for presynaptic vesicle fusion.