Xiao Bo Liu

Disrupted-In-Schizophrenia 1 Regulates Integration of Newly Generated Neurons in the Adult Brain

Adult neurogenesis occurs throughout life in discrete regions of the adult mammalian brain. Little is known about the mechanism governing the sequential developmental process that leads to integration of new neurons from adult neural stem cells into the existing circuitry. Here, we investigated roles of Disrupted-In-Schizophrenia 1 (DISC1), a schizophrenia susceptibility gene, in adult hippocampal neurogenesis. Unexpectedly, downregulation of DISC1 leads to accelerated neuronal integration, resulting in aberrant morphological development and mispositioning of new dentate granule cells in a cell-autonomous fashion. Functionally, newborn neurons with DISC1 knockdown exhibit enhanced excitability and accelerated dendritic development and synapse formation. Furthermore, DISC1 cooperates with its binding partner NDEL1 in regulating adult neurogenesis. Taken together, our study identifies DISC1 as a key regulator that orchestrates the tempo of functional neuronal integration in the adult brain and demonstrates essential roles of a susceptibility gene for major mental illness in neuronal development, including adult neurogenesis.

Switching of NMDA Receptor 2A and 2B Subunits at Thalamic and Cortical Synapses during Early Postnatal Development

Switching of the NMDA receptor 2A (NR2A) and NR2B subunits at NMDA receptors is thought to underlie the functional changes that occur in NMDA receptor properties during the developmental epoch when neural plasticity is most pronounced. The cellular expression of NR2A and NR2B and the NR2 synaptic binding protein postsynaptic density-95 (PSD-95) was examined in the mouse somatosensory cortex and thalamus from postnatal day 2 (P2) to P15 using reverse transcription-PCR, in situ hybridization histochemistry, and immunocytochemistry. The localization of NR2A and NR2B subunits and PSD-95 was then studied at synapses in layer IV of somatosensory cortex and in the ventral posterior nucleus of the thalamus using high-resolution immunoelectron microscopy. At both cortical and thalamic synapses, a quantitative switch in the dominant synaptic subunit from NR2B to NR2A was accompanied by a similar change in the cellular expression of NR2A but not of NR2B. Synaptic PSD-95 developed independently, although both NR2A and NR2B colocalized with PSD-95. Displacement of NR2B subunits from synapses was not accompanied by an increase in an extrasynaptic pool of this subunit. Thus, the switch in synaptic NR2 subunit predominance does not occur by changes in expression or displacement from synapses and may reflect the formation of new synapses from which NR2B is lacking.

Temporal and regional expression of NMDA receptor subunit NR3A in the mammalian brain

NR3A is a developmentally regulated N‐methyl‐D‐aspartate receptor (NMDAR) subunit that was previously known as NMDAR‐L or χ‐1. Unlike other NMDAR subunits, NR3A inhibits the NMDAR‐associated ion channel in a novel manner, and a role in synaptogenesis has been suggested for this subunit. Here, we report a comprehensive study to delineate the temporal and anatomic expression of NR3A protein in the mammalian brain by using a monoclonal anti‐NR3A antibody. NR3A protein was found to peak at postnatal day (P) 8, and to decrease gradually from P12 to adulthood in the rat central nervous system. Moreover, NR3A protein was heavily expressed in all areas of the isocortex, portions of the amygdaloid nuclei, and selective cell layers and nuclei of the hippocampus, thalamus, hypothalamus, brainstem, and spinal cord. NR3A protein was also expressed in the cerebellar cortex, whereas only weak signal was detected in the previous in situ studies by using riboprobes. At an ultrastructural level, NR3A was associated specifically with asymmetrical synapses and localized to postsynaptic membranes. This information will facilitate future research on NMDARs by providing clues to possible inclusion of the NR3A subunit in NMDARs in many brain regions. J. Comp. Neurol. 450:303–317, 2002.

Development of hippocampal mossy fiber synaptic outputs by new neurons in the adult brain

New neurons are continuously generated in restricted regions of the adult mammalian brain. Although these adult-born neurons have been shown to receive synaptic inputs, little is known about their synaptic outputs. Using retrovirus-mediated birth-dating and labeling in combination with serial section electron microscopic reconstruction, we report that mossy fiber en passant boutons of adult-born dentate granule cells form initial synaptic contacts with CA3 pyramidal cells within 2 weeks after their birth and reach morphologic maturity within 8 weeks in the adult hippocampus. Knockdown of Disrupted-in-Schizophrenia-1 (DISC1) in newborn granule cells leads to defects in axonal targeting and development of synaptic outputs in the adult brain. Together with previous reports of synaptic inputs, these results demonstrate that adult-born neurons are fully integrated into the existing neuronal circuitry. Our results also indicate a role for DISC1 in presynaptic development and may have implications for the etiology of schizophrenia and related mental disorders.

Cycling of NMDA Receptors during Trafficking in Neurons before Synapse Formation

The trafficking of glutamate receptors in neurons is of the utmost importance for synapse formation and synaptic plasticity. Recently, we demonstrated that both NMDA and AMPA receptors reside in mobile transport packets that are recruited rapidly and independently to nascent synapses. Here, we show that a large proportion of the glutamate receptor clusters in young cortical neurons are present on the surface of dendrites before synapses are formed and these surface-exposed transport packets are mobile. Exocytosis of glutamate receptors to the dendritic surface occurs via a SNARE [soluble n-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor]-dependent SNAP-23-mediated mechanism. Endocytosis occurs rapidly after surface exposure; >50% of surface-labeled NMDA receptors (NMDARs) are endocytosed within 5 min. NMDARs are transported along microtubules on large tubulovesicular organelles, as indicated by immunoelectron microscopy, and are associated with EEA1 (early endosomal antigen 1) and SAP102 (synapse-associated protein 102), as indicated by immunocytochemistry. Most surprisingly, a large proportion of these transport packets cycle through the dendritic plasma membrane before synapse formation. These results suggest a novel model in which NMDARs cycle with the plasma membrane during pauses of movement along microtubules while trafficking.

Endocytosis and synaptic removal of NR3A-containing NMDA receptors by PACSIN1/syndapin1

A key step in glutamatergic synapse maturation is the replacement of developmentally expressed N-methyl-D-aspartate receptors (NMDARs) with mature forms that differ in subunit composition, electrophysiological properties and propensity to elicit synaptic plasticity. However, the mechanisms underlying the removal and replacement of synaptic NMDARs are poorly understood. Here we demonstrate that NMDARs containing the developmentally regulated NR3A subunit undergo rapid endocytosis from the dendritic plasma membrane in cultured rat hippocampal neurons. This endocytic removal is regulated by PACSIN1/syndapin1, which directly and selectively binds the carboxy-terminal domain of NR3A through its NPF motifs and assembles a complex of proteins including dynamin and clathrin. Endocytosis of NR3A by PACSIN1 is activity dependent, and disruption of PACSIN1 function causes NR3A accumulation at synaptic sites. Our results reveal a new activity-dependent mechanism involved in the regulation of NMDAR expression at synapses during development, and identify a brain-specific endocytic adaptor that confers spatiotemporal and subunit specificity to NMDAR endocytosis.

MHCI negatively regulates synapse density during the establishment of cortical connections

Major histocompatibility complex class I (MHCI) molecules modulate activity-dependent refinement and plasticity. We found that MHCI also negatively regulates the density and function of cortical synapses during their initial establishment both in vitro and in vivo. MHCI molecules are expressed on cortical neurons before and during synaptogenesis. In vitro, decreasing surface MHCI (sMHCI) on neurons increased glutamatergic and GABAergic synapse density, whereas overexpression decreased it. In vivo, synapse density was higher throughout development in β2m−/− mice. MHCI also negatively regulated the strength of excitatory, but not inhibitory, synapses and controlled the balance of excitation and inhibition onto cortical neurons. sMHCI levels were modulated by activity and were necessary for activity to negatively regulate glutamatergic synapse density. Finally, acute changes in sMHCI and activity altered synapse density exclusively during early postnatal development. These results identify a previously unknown function for immune proteins in the negative regulation of the initial establishment and function of cortical connections.

Vesicular glutamate transporters define two sets of glutamatergic afferents to the somatosensory thalamus and two thalamocortical projections in the mouse

The ventral posterior nucleus of the thalamus (VP) receives two major sets of excitatory inputs, one from the ascending somatosensory pathways originating in the dorsal horn, dorsal column nuclei, and trigeminal nuclei, and the other originating from the cerebral cortex. Both systems use glutamate as neurotransmitter, as do the thalamocortical axons relaying somatosensory information from the VP to the primary somatosensory cortex (SI). The synapses formed by these projection systems differ anatomically, physiologically, and in their capacity for short‐term synaptic plasticity. Glutamate uptake into synaptic vesicles and its release at central synapses depend on two isoforms of vesicular glutamate transporters, VGluT1 and VGluT2. Despite ample evidence of their complementary distribution, some instances exist of co‐localization in the same brain areas or at the same synapses. In the thalamus, the two transcripts coexist in cells of the VP and other nuclei but not in the posterior or intralaminar nuclei. We show that the two isoforms are completely segregated at VP synapses, despite their widespread expression throughout the dorsal and ventral thalamus. We present immunocytochemical, ultrastructural, gene expression, and connectional evidence that VGluT1 in the VP is only found at corticothalamic synapses, whereas VGluT2 is only found at terminals made by axons originating in the spinal cord and brainstem. By contrast, the two VGluT isoforms are co‐localized in thalamocortical axon terminals targeting layer IV, but not in those targeting layer I, suggesting the presence of two distinct projection systems related to the core/matrix pattern of organization of thalamocortical connectivity described in other mammals. J. Comp. Neurol. 507:1258–1276, 2008.

Stereotyped axon pruning via plexin signaling is associated with synaptic complex elimination in the hippocampus

Plexin signaling is required for stereotyped pruning of long axon collaterals in the vertebrate CNS; however, a cellular basis for plexins on stereotyped pruning has not been determined. Using quantitative electron microscopy and immunocytochemistry, we found that infrapyramidal mossy fiber axon collaterals form transient synaptic complexes with basal dendrites of CA3 pyramidal cells in the early postnatal mouse hippocampus. At later postnatal ages, these synaptic complexes stop maturing and are removed before stereotyped pruning by a mechanism that does not involve axon degeneration and glial cell engulfment. In knock-out mice that lack plexin-A3 signaling, the synaptic complexes continue to mature, and, as a result, the collaterals are not pruned. Thus, our results suggest that intact plexin-A3 signaling contributes to synaptic complex elimination, which is associated with stereotyped axon pruning.

Plexin signaling selectively regulates the stereotyped pruning of corticospinal axons from visual cortex

Neurons in the developing CNS tend to send out long axon collaterals to multiple target areas. For these neurons to attain specific connections, some of their axon collaterals are subsequently pruned—a process called stereotyped axon pruning. One of the most striking examples of stereotyped pruning in the CNS is the pruning of corticospinal tract (CST) axons. The long CST collaterals from layer V neurons of the visual and motor cortices are differentially pruned during development. Here we demonstrate that select plexins and neuropilins, which serve as coreceptors for semaphorins, are expressed in visual cortical neurons at the time when CST axon collaterals are stereotypically pruned. By analyzing mutant mice, we find that the pruning of visual, but not motor, CST axon collaterals depends on plexin-A3, plexin-A4, and neuropilin-2. Expression pattern study suggests that Sema3F is a candidate local cue for the pruning of visual CST axons. Using electron microscopic analysis, we also show that visual CST axon collaterals form synaptic contacts in the spinal cord before pruning and that the unpruned collaterals in adult mutant mice are unmyelinated and maintain their synaptic contacts. Our results indicate that the stereotyped pruning of the visual and motor CST axon collaterals is differentially regulated and that this specificity arises from the differential expression of plexin receptors in the cortex.