Camin Dean

Neurexin mediates the assembly of presynaptic terminals

Neurexins are a large family of proteins that act as neuronal cell-surface receptors. The function and localization of the various neurexins, however, have not yet been clarified. Beta-neurexins are candidate receptors for neuroligin-1, a postsynaptic membrane protein that can trigger synapse formation at axon contacts. Here we report that neurexins are concentrated at synapses and that purified neuroligin is sufficient to cluster neurexin and to induce presynaptic differentiation. Oligomerization of neuroligin is required for its function, and we find that beta-neurexin clustering is sufficient to trigger the recruitment of synaptic vesicles through interactions that require the cytoplasmic domain of neurexin. We propose a two-step model in which postsynaptic neuroligin multimers initially cluster axonal neurexins. In response to this clustering, neurexins nucleate the assembly of a cytoplasmic scaffold to which the exocytotic apparatus is recruited.

Neuroligins and neurexins: linking cell adhesion, synapse formation and cognitive function.

Cell adhesion represents the most direct way of coordinating synaptic connectivity in the brain. Recent evidence highlights the importance of a trans-synaptic interaction between postsynaptic neuroligins and presynaptic neurexins. These transmembrane molecules bind each other extracellularly to promote adhesion between dendrites and axons. This signals the recruitment of presynaptic and postsynaptic molecules to form a functional synapse. Remarkably, neuroligins alone can induce the formation of fully functional presynaptic terminals in contacting axons. Conversely, neurexins alone can induce postsynaptic differentiation and clustering of receptors in dendrites. Therefore, the neuroligin-neurexin interaction has the unique ability to act as a bi-directional trigger of synapse formation. Here, we review several recent studies that offer clues as to how these proteins form synapses and how they might function in the brain to establish and modify neuronal network properties and cognition.

Synaptotagmin-IV modulates synaptic function and long-term potentiation by regulating BDNF release

Synaptotagmin-IV (syt-IV) is a membrane trafficking protein that influences learning and memory, but its localization and role in synaptic function remain unclear. We found that syt-IV localized to brain-derived neurotrophic factor (BDNF)-containing vesicles in hippocampal neurons. Syt-IV/BDNF–harboring vesicles underwent exocytosis in both axons and dendrites, and syt-IV inhibited BDNF release at both sites. Knockout of syt-IV increased, and overexpression decreased, the rate of synaptic vesicle exocytosis from presynaptic terminals indirectly via changes in postsynaptic release of BDNF. Thus, postsynaptic syt-IV regulates the trans-synaptic action of BDNF to control presynaptic vesicle dynamics. Furthermore, selective loss of presynaptic syt-IV increased spontaneous quantal release, whereas a loss of postsynaptic syt-IV increased quantal amplitude. Finally, syt-IV knockout mice showed enhanced long-term potentiation (LTP), which depended entirely on disinhibition of BDNF release. Thus, regulation of BDNF secretion by syt-IV emerges as a mechanism for maintaining synaptic strength in a useful range during LTP.

Neuropeptide Exocytosis Involving Synaptotagmin-4 and Oxytocin in Hypothalamic Programming of Body Weight and Energy Balance

Hypothalamic neuropeptides play essential roles in regulating energy and body weight balance. Energy imbalance and obesity have been linked to hypothalamic signaling defects in regulating neuropeptide genes; however, it is unknown whether dysregulation of neuropeptide exocytosis could be critically involved. This study discovered that synaptotagmin-4, an atypical modulator of synaptic exocytosis, is expressed most abundantly in oxytocin neurons of the hypothalamus. Synaptotagmin-4 negatively regulates oxytocin exocytosis, and dietary obesity is associated with increased vesicle binding of synaptotagmin-4 and thus enhanced negative regulation of oxytocin release. Overexpressing synaptotagmin-4 in hypothalamic oxytocin neurons and centrally antagonizing oxytocin in mice are similarly obesogenic. Synaptotagmin-4 inhibition prevents against dietary obesity by normalizing oxytocin release and energy balance under chronic nutritional excess. In conclusion, the negative regulation of synaptotagmin-4 on oxytocin release represents a hypothalamic basis of neuropeptide exocytosis in controlling obesity and related diseases.

Autapses and Networks of Hippocampal Neurons Exhibit Distinct Synaptic Transmission Phenotypes in the Absence of Synaptotagmin I

Synaptotagmin-I (syt-I) is required for rapid neurotransmitter release in mouse hippocampal neurons. However, contradictory results have been reported regarding evoked and spontaneous secretion from syt-I knock-out (KO) neurons. Here, we compared synaptic transmission in two different hippocampal neuron preparations: autaptic cultures in which a single isolated cell innervates itself, and dissociated mass cultures in which individual cells are innervated by neighboring cells. In autaptic cultures, the total extent of evoked release, size of readily releasable pool of synaptic vesicles, and release probability were unchanged in syt-I KO neurons. In contrast, in cultures containing multiple interconnected neurons, total evoked release, the number of docked vesicles, and release probability, were significantly reduced in syt-I KO neurons. Using a micronetwork system in which we varied the number of cells on an island, we found that the frequency of spontaneous synaptic vesicle fusion events (minis) was unchanged in syt-I KO neurons when two or fewer cells were present on an island. However, in micronetworks composed of three or more neurons, mini frequency was increased threefold to fivefold in syt-I KO neurons compared with wild type. Moreover, interneuronal synapses exhibited higher rates of spontaneous release than autaptic synapses. This higher rate was attributable to an increase in release probability because excitatory hippocampal neurons in micronetworks formed a set number of synapses per cell regardless of the number of connected neurons. Thus, aspects of synaptic transmission differ between autaptic and dissociated cultures, and the synaptic transmission phenotype, resulting from loss of syt-I, is dictated by the connectivity of neurons.

HDAC inhibitor-dependent transcriptome and memory reinstatement in cognitive decline models

Aging and increased amyloid burden are major risk factors for cognitive diseases such as Alzheimers disease (AD). Effective therapies for these diseases are lacking. Here, we evaluated mouse models of age-associated memory impairment and amyloid deposition to study transcriptome and cell type-specific epigenome plasticity in the brain and peripheral organs. We determined that aging and amyloid pathology are associated with inflammation and impaired synaptic function in the hippocampal CA1 region as the result of epigenetic-dependent alterations in gene expression. In both amyloid and aging models, inflammation was associated with increased gene expression linked to a subset of transcription factors, while plasticity gene deregulation was differentially mediated. Amyloid pathology impaired histone acetylation and decreased expression of plasticity genes, while aging altered H4K12 acetylation-linked differential splicing at the intron-exon junction in neurons, but not nonneuronal cells. Furthermore, oral administration of the clinically approved histone deacetylase inhibitor vorinostat not only restored spatial memory, but also exerted antiinflammatory action and reinstated epigenetic balance and transcriptional homeostasis at the level of gene expression and exon usage. This study provides a systems-level investigation of transcriptome plasticity in the hippocampal CA1 region in aging and AD models and suggests that histone deacetylase inhibitors should be further explored as a cost-effective therapeutic strategy against age-associated cognitive decline.

Synaptotagmin IV: a multifunctional regulator of peptidergic nerve terminals

Many members of the synaptotagmin (Syt) protein family bind Ca2+ and trigger exocytosis, but some Syt proteins appear to have no Ca2+-dependent actions and their biological functions remain obscure. Syt IV is an activity-induced brain protein with no known Ca2+-dependent interactions and its subcellular localization and biological functions have sparked considerable controversy. We found Syt IV on both micro- and dense-core vesicles in posterior pituitary nerve terminals in mice. In terminals from Syt IV knockout mice compared with those from wild types, low Ca2+ entry triggered more exocytosis, high Ca2+ entry triggered less exocytosis and endocytosis was accelerated. In Syt IV knockouts, dense-core and microvesicle fusion was enhanced in cell-attached patches and dense-core vesicle fusion pores had conductances that were half as large as those in wild types. Given the neuroendocrine functions of the posterior pituitary, changes in Syt IV levels could be involved in endocrine transitions involving alterations in the release of the neuropeptides oxytocin and vasopressin.

Distinct Subsets of Syt-IV/BDNF Vesicles Are Sorted to Axons versus Dendrites and Recruited to Synapses by Activity

BDNF plays a critical role in the regulation of synaptic strength and is essential for long-term potentiation, a phenomenon that underlies learning and memory. However, whether BDNF acts in a diffuse manner or is targeted to specific neuronal subcompartments or synaptic sites to affect circuit function remains unknown. Here, using photoactivation of BDNF or syt-IV (a regulator of exocytosis present on BDNF-containing vesicles) in transfected rat hippocampal neurons, we discovered that distinct subsets of BDNF vesicles are targeted to axons versus dendrites and are not shared between these compartments. Moreover, syt-IV- and BDNF-harboring vesicles are recruited to both presynaptic and postsynaptic sites in response to increased neuronal activity. Finally, using syt-IV knockout mouse neurons, we found that syt-IV is necessary for both presynaptic and postsynaptic scaling of synaptic strength in response to changes in network activity. These findings demonstrate that BDNF-containing vesicles can be targeted to specific sites in neurons and suggest that syt-IV-regulated BDNF secretion is subject to spatial control to regulate synaptic function in a site-specific manner.

Alternative Splicing of Neuroligin Regulates the Rate of Presynaptic Differentiation

Neuroligins (NLGs) and Neurexins (NRXs) are important adhesion molecules that promote synapse formation. Multiple splice variants of NLG and NRX exist, but their specific functions are unclear. Here we report that a surrogate postsynaptic cell expressing full-length NLG-1 triggers slow presynaptic differentiation in a contacting axon. In contrast, a version of NLG-1, which lacks insert B (NLG-1ΔB), induces rapid presynaptic differentiation, reaching the rate seen at native neuronal synapses. We show that this acceleration is attributed to the removal of the N-linked glycosylation site within insert B. NLG-1ΔB also increases synaptic density at neuro-neuronal synapses more than does full-length NLG-1. Other postsynaptic adhesion proteins, such as N-cadherin, EphB2, and SynCAM-1, alone or in combination with full-length NLG-1, do not trigger fast differentiation, suggesting that rapid presynaptic differentiation depends on a unique interaction of NLG-1ΔB with axonal proteins. Indeed, we find that NLG-1ΔB recruits more axonal α-NRX. Our results suggest that the engagement of α-NRX is a key to rapid induction of synapses at new sites of axo-dendritic contact.

Axonal and dendritic synaptotagmin isoforms revealed by a pHluorin-syt functional screen

Synaptotagmin-1 regulates synaptic vesicle fusion, but little is known about the remaining syt isoforms. Syt-pHluorin reporters are used to show that only two syts are on synaptic vesicles, whereas other isoforms are selectively targeted to dendrites, axons, or both axons and dendrites, where they undergo exocytosis and endocytosis with distinct kinetics.