These results suggest a possible functional role for the elaboration of both excitatory and inhibitory intracortical circuits, which are susceptible to changes in sensory experience in the period after eye opening (Ruthazer and

Stryker, 1996, Zufferey et al., 1999, White et al., 2001, Chattopadhyaya et al., 2004, Katagiri et al., 2007 and Ko et al., 2013). We propose that circuit connectivity is shaped by exposure to the statistical structure of the natural environment (e.g., extended contours or edges) after the onset of vision, which increases the effectiveness of surround modulation when viewing naturalistic stimuli to which animals are typically exposed. Our data suggest that Neratinib in vitro visual selleck products experience optimizes spiking output by refining the timing and magnitude of inhibition recruited by the surround.

In conclusion, our results support the idea that visual circuits mature in an experience-dependent manner to become sensitive to the statistical structure of natural stimuli extending beyond the boundaries of the RF. While the basic RF properties are established by the time of eye opening (Hubel and Wiesel, 1963, Blakemore and Van Sluyters, 1975, Chapman and Stryker, 1993, Krug et al., 2001, White et al., 2001, Rochefort et al., 2011 and Ko et al., 2013), efficient representations of natural stimulus features—in terms of selectivity, information Isotretinoin transfer, and energy consumption (Barlow, 1961, Simoncelli and Olshausen, 2001 and Laughlin, 2001)—are not inherent to sensory circuits but require visual experience to develop. All experimental procedures were licensed and performed in accordance with institutional and national animal welfare guidelines. Data were

obtained from C57BL/6 mice aged postnatal day (P) 14–19 (immature age group, n = 7) or P32–P40 (mature age group n = 10; dark-reared age group n = 8). For dark rearing, mice were kept in complete darkness from P13 until placed under anesthesia. Mice were initially anaesthetized with a mixture of fentanyl (0.05 mg/ml), midazolam (5.0 mg/kg), and medetomidin (0.5 mg/kg). Anesthesia was maintained with a low concentration of isoflurane (typically 0.5% mixed with O2) delivered by a small nose cone. Details of the surgery are given in Supplemental Experimental Procedures. The position and size of a neuron’s RF were determined in similar way as described before (Jones et al., 2001 and Jones et al., 2002). First, RF center position was mapped with pseudorandomized sparse noise stimulus sequence (white and black flashing patches on an isoluminant gray background). Then, the RF radius was estimated by determining a circular area of half-maximal spike responses to the same pseudorandomized sparse noise stimulus.

To further address this possibility, we turned to enhancer-supressor genetic assays dependent on Sema-1a/PlexA repulsive axon guidance. Ectopic expression of Sema-1a in muscles leads to reduced muscle innervation (Yu et al., 1998). These effects are due to the repulsive action of Sema-1a (Yu et al., 1998) and are suppressed by decreasing the levels

of the Sema-1a receptor, PlexA (Winberg et al., 1998b). In contrast, decreasing selleck products the levels of 14-3-3ε enhanced Sema-1a repulsion (Figures 3B–3D); suggesting that similar to PKA RII and Nervy (Figures 3C and 3D; Terman and Kolodkin, 2004), 14-3-3ε opposes Sema-1a repulsion. To further investigate these antagonistic interactions, we turned to genetic assays dependent on the repulsive effects of PlexA. Increasing the levels of neuronal PlexA generates abnormally defasciculated axons that result in discontinuous CNS longitudinal connectives and axons crossing the midline or projecting abnormally into the periphery ( Figures 4A and S3A; Ceritinib mw Winberg et al., 1998b and Ayoob et al.,

2004). Strikingly, decreasing the levels of 14-3-3ε significantly increased these PlexA-dependent guidance defects ( Figures 4A–4C), while increasing neuronal 14-3-3ε significantly decreased these PlexA-dependent guidance defects ( Figures 4B and 4C). Together, these results along with other in vivo Sema1a/PlexA-dependent CNS and motor axon guidance assays ( Figure S3) indicate that 14-3-3ε antagonizes Sema1a/PlexA-mediated repulsive axon guidance. To begin to investigate the mechanism aminophylline by which 14-3-3ε antagonizes Sema-1a/PlexA-mediated repulsive axon guidance, we sought to determine the site of interaction between PlexA and 14-3-3ε. We found that the portion of PlexA that was necessary and sufficient for the interaction with 14-3-3ε contains a consensus 14-3-3 binding sequence (Figures 5A and 5B).

In particular, 14-3-3 proteins typically bind to single phosphorylated serine or threonine residues on target proteins (Yaffe and Elia, 2001) and Drosophila PlexA contains a mode I 14-3-3 consensus binding motif, R/KSXpSXP, where p represents the phosphorylated serine (Ser1794) residue predicted to mediate the interaction with 14-3-3 proteins ( Figure 5B; Yaffe et al., 1997 and Rittinger et al., 1999). To test this possibility, we substituted alanine (Ala) for serine (Ser) and threonine (Thr) residues within this consensus 14-3-3ε binding motif. We found that the predicted Ser1794 residue was necessary for the observed PlexA interaction with 14-3-3ε ( Figures 5C and S4A). Next, we generated phospho-mimetic forms of Ser1794 (Ser1794 to Glu1794 or Asp1794), but found that as with other 14-3-3 interacting proteins adding one negative charge was not sufficient for the interaction between PlexA and 14-3-3ε ( Figures 5C and S4A).

These findings imply that Chx10off Shox2+ INs constitute part of the rhythm-generating network, providing key insights into the logic of iEIN diversity and motor rhythmicity. To identify distinct populations of iEINs, we performed a micro-array screen for genes preferentially enriched in ventral spinal cord at lumbar levels (Zagoraiou et al., 2009; Table

S1 available online). We found that the homeobox gene Shox2 was expressed at P0-P1 by a set of interneurons present along the entire rostrocaudal axis of the spinal cord. In the transverse plane, these neurons occupied an intermediate domain that extended mediolaterally from close to the central canal to the edge of the gray matter ( Figure 1A). To define the origin and distribution of Shox2 neurons in greater detail Bortezomib concentration we generated a Shox2::Cre mouse line ( Figure 1B) and performed lineage tracing with fluorescent protein (FP) conditional reporter mice (Rosa26-YFP/tdTomato and Z/EG lines). Comparison of FP and endogenous protein expression revealed isocitrate dehydrogenase targets that Shox2 expression begins around E11.5 and persists until postnatal stages, although expression is extinguished from many FP+ interneurons at later embryonic stages: ∼80% of FP+ neurons expressed Shox2 at E12.5, compared to ∼35% at P0-P1 ( Figures 1C and 1D). In our subsequent analyses, we define Shox2 interneurons (Shox2 INs) on

the basis of Shox2::Cre directed FP expression, independent of maintained Shox2 expression. To define the neurotransmitter phenotype of Shox2 INs, we monitored the status of vGluT2 expression in Shox2::Cre; Tau-GFP-nlsLacZ mice. We found that > 98%

of Shox2+ neurons expressed vGluT2 transcript (n = 3; Figure 1E), indicating that Shox2 INs are glutamatergic. We next addressed the extent of subtype diversity of Shox2 INs. The settling position of Shox2 INs overlapped that of V2a neurons, marked by expression of the transcription factor Chx10 (Jessell, 2000, Crone et al., 2008 and Lundfald et al., 2007). We therefore determined the extent of ADP ribosylation factor overlap of FP and Chx10 expression in lumbar spinal cord tissue derived from Shox2::Cre; FP reporter mice ( Figure 1F). At P0-1, we found that 77% of Shox2 INs coexpressed Chx10 and conversely that 60% of Chx10+ INs were marked by Shox2-directed FP expression ( Figure 1F). These studies reveal three distinct populations of ventrally positioned vGluT2+ excitatory interneurons: Shox2only INs, Chx10only INs, and Shox2/Chx10double INs. We next addressed the origin and diversity of the Shox2 IN class of EINs. Since Chx10+ INs derive from the p2 progenitor domain we considered whether Shox2only INs are p2 domain derived. p2 domain progenitors give rise to inhibitory GATA3-derived V2b and V2c INs as well as to excitatory Lhx3+/Chx10+ V2a INs (Peng et al., 2007 and Panayi et al., 2010). But our analysis of FP-marked neurons in Shox2::Cre; ROSA26-YFP reporter mice at E13.

, 2011 and von Philipsborn et al., 2011). Male courtship behavior is influenced by a range of sensory inputs (Krstic et al., 2009 and Koganezawa et al., 2010), especially the olfactory system (Billeter et al., 2009). Photoactivatable GFP

has been used to trace connectivity from the olfactory receptors that detect female flies through the antennal lobe to second order projections (Datta et al., 2008 and Ruta et al., 2010). Anatomical analysis suggests a compartment-level convergence of FruM neurons (Yu et al., 2010) and expression of dendritic buy GSK1349572 and synaptic reporters in candidate partners suggests connectivity (von Philipsborn et al., 2011). New understandings of the neural basis for courtship behavior have been reviewed (Manoli et al., 2006, Dickson, 2008 and Benton,

2011). The courtship circuit has several advantages: a single gene (or isoform) expressed in many neural components, sexually dimorphic anatomy and behavior, some known sensory inputs, and corroborative historical data from gynandromorphs and feminization screens (Hall, 1979, Ferveur et al., 1995 and Broughton et al., 2004), but the astute use of genetic tools to manipulate neurons has led to our current understanding of the neural circuit driving male courtship behavior. Recent work has demonstrated functional imaging of FruM neurons during a facsimile of courtship behavior (Kohatsu et al., 2011), which will allow interrogation of how neurons within the circuit respond to specific sensory stimuli and how their activity correlates with behavioral output. Everolimus clinical trial The same experimental setup could be used to deliver specific activity patterns with light-stimulated channelrhodopsin to determine how these neurons affect behavioral outcomes. Although

there is still much work to be done to connect the identified components of a courtship circuit and find the mafosfamide missing links, the potential to understand how a neural circuit actually works to drive this complex behavior is unmatched. Similar smart use of the powerful tools described here should enable mapping of circuits driving a range of different behaviors. This will permit circuit comparisons, identification of neurons that participate in several circuits, and the investigation of the way decisions between behavioral programs are made. To understand how the nervous system of an animal controls a particular behavior, one needs to identify the neurons involved, determine how their activity influences the behavior, and explore how they connect to other participating neurons. The abundance of tools for spatially and temporally targeting gene expression to specific neurons, manipulating or observing their activity, and assaying behavioral consequences makes Drosophila a premier system for exploring the principles guiding the development and function of neural circuits.

Morphed probe trials. After the acquisition of the discrimination problem, performance was evaluated by using feature-ambiguous probe trials. These probe trials increased the difficulty of the discrimination task by increasing the similarity of the S+ and S−. Probe trials were created by morphing the S+ and S− into one another in 14 steps (Morpheus Photo Animator; ACD Systems, Saanichton, Canada). Thus, one stimulus was gradually morphed into the other, physically changing each stimulus from one step to

the next ( Figure 2). This morphing procedure is similar to procedures used in previous work with monkeys ( Bussey et al., 2003) and humans ( Lee et al., http://www.selleckchem.com/products/azd9291.html 2005 and Shrager et al., 2006). Note that one stimulus was not blended into the other. Rather, the entire stimuli were gradually altered so that they became more alike. Probe level 1 consisted of the least amount of feature overlap (i.e., the two stimuli were quite distinct and most similar to the training stimuli). At level 14 the two stimuli contained substantial feature overlap and appeared quite similar ( Figure 2). During this phase of testing, mTOR inhibitor 80% of the trials were standard trials (training stimuli). The remaining 20% of the trials

were rewarded morphed probe trials. The order of the probe trials (levels 1–14) was pseudorandom with the constraint that each of the 14 difficulty levels had to be presented once before any one difficulty level could be repeated. This procedure ensured that data for probe trials accrued at the same rate for every difficulty level. This phase of testing continued until 150 probe trials were completed at each difficulty level. Thus, across this phase of testing each animal received 2,100

probe trials across the 14 different difficulty levels (150 × 14) and an additional 10,500 trials with the training stimuli. Surgery. Animals were assigned to a perirhinal lesion group or a normal control group based upon their trials-to-criterion score for the discrimination task (to create two equal groups). The intention was to CYTH4 remove the entire perirhinal cortex bilaterally. For surgery, the rat was placed in a Kopf stereotaxic instrument and the incisor bar was adjusted until bregma was level with lambda. Bilateral excitotoxic perirhinal lesions were produced by local microinjections of ibotenate acid (IBO; Biosearch Technologies, San Rafael, CA). IBO was dissolved in 0.1 M phosphate-buffered saline to provide a solution with a concentration of 10 mg/ml, pH 7.4. IBO was injected at a rate of 0.1 μl/min with a 10 μl Hamilton syringe mounted on a stereotaxic frame and held with a Kopf microinjector (model 5000). The syringe needle was lowered to the target coordinate and left in place for 1 min before beginning the injection. After the injection, the syringe needle was left in place for a further 5 min to reduce the spread of IBO up the needle tract. A total of 0.

With respect to both proliferation and fate determination, chromatin modification therefore represents an important mechanism for maintaining the adult VZ-SVZ stem cell pool during the lifetime of the organism. Determining how signaling by niche-provided factors ultimately drives transcriptional activity will help to develop a unified

understanding of how neurogenesis and stem cell persistence is maintained. Neurogenesis in the adult VZ-SVZ extends over a large area—approximately six square millimeters in mice. It was unclear why this large region evolved to support postnatal neuronal generation and why newly generated neurons had to migrate so far to integrate into the olfactory selleck circuitry. Clues to the biological heterogeneity of the adult VZ-SVZ came from examining the expression of transcription factors such as Pax6, which is present in specific subpopulations of migrating neuroblasts and olfactory bulb interneurons (Hack et al., 2005 and Kohwi et al., 2005). Subsequent experiments using viral targeting or genetic

lineage tracing in neonatal and adult mice revealed that specific subtypes of interneurons are made within specific adult VZ-SVZ locations (Figure 2; Kelsch et al., 2007, Kohwi et al., 2007, Merkle et al., 2007, Ventura and Goldman, 2007 and Young et al., 2007). While superficial Z-VAD-FMK nmr granule interneurons are largely generated by stem cells in the dorsal VZ-SVZ, deep granule interneurons are primarily derived

from the ventral VZ-SVZ. Distinct populations of periglomerular cells (PGCs) also arise from specific locations within the anterior and medial adult VZ-SVZ, and a population of glutamatergic olfactory bulb neurons is derived from the dorsal SVZ (Merkle et al., 2007 and Brill et al., 2009). Intriguingly, stem cells continued to generate specific types of progeny even after transplantation or multiple passages in culture, suggesting that the differentiation program for neuronal progeny is encoded at least in part by cell-intrinsic factors (Merkle et al., 2007). Although this patterning is present at birth, it is not yet known at what stage in embryonic development regional specification in the adult VZ-SVZ is organized, and whether there is a Bay 11-7085 window of time during development when the fate of stem cell progeny has not yet become restricted. Subregions of the adult VZ-SVZ express transcription factors that are involved in regional specification of the developing brain, suggesting that some of the same coding at play in development may continue to be active in the adult. However, the mechanisms by which this specification is generated and maintained are unknown. Additionally, the production of particular olfactory interneuron types appears to decline after birth, indicating that the repertoire of neuronal types derived from the VZ-SVZ may change over time (De Marchis et al., 2007 and Kohwi et al., 2007).

The segregation of Mib-GFP into the apical daughter was apparent at the time of birth (Figure 7F; Movie S3; ∼24 min). However, in the par-3 morphant, Mib-GFP selleck chemical was present in both the apical and basal daughter at the time of their birth ( Figure 7G; Movie S4; ∼18 min). Together, these results suggest that Mib is unequally segregated into the apical daughter upon asymmetric division in a Par-3-dependent manner, and such asymmetry is maintained in the daughter cells. In agreement with the disrupted Mib localization in the par-3 morphant,

we found that the asymmetry of both her4.1 ( Figures 8A–8C) and dla ( Figures 8D–8F) expression was lost in par-3-deficient embryos, demonstrating that Par-3 is essential for establishing Notch asymmetry in paired siblings. Mib mislocalization and lost asymmetry of Notch signaling components in par-3 morphants could result in either increased or diminished Notch activity in both daughter cells, which would in turn impact progenitor GW786034 supplier fate choice differently. To determine how Notch activity and cell fate might be affected in par-3-deficient embryos, we first analyzed the overall expression level of her4.1, dla, and the pan-neuronal marker Hu. These analyses showed that her4.1 expression ( Figures 8G and 8H; 88%, n = 16) was

increased, whereas neuronal numbers ( Figures 8I and 8J; 83%, n = 18) were Oxymatrine decreased in the par-3 morphant. In contrast the expression of dla was not changed significantly (

Figures 8K and 8L; 100%, n = 15). This is surprising, given the increase of her4.1 expression and the known negative feedback regulation of Notch ligands by hes/her genes. Quantitative reverse-transcription PCR analysis further confirmed the significant upregulation of her4.1 (and her6) mRNA expression, whereas the mRNA levels of dla, dld, notch1a, and notch1b were unchanged ( Figure 8M) ( Table S1). Thus, par-3 function is essential to restrict Notch activity, and is somehow also required for the feedback repression of Notch ligand expression. To understand the nature of these par-3 functions, we asked whether they are dependent on mib. In the mib−/− mutant, consistent with the disruption of Notch signaling, her4.1 expression was significantly reduced ( Figures S7J and S7K). The par-3 and mib double-deficient embryos also showed reduced her4.1 expression ( Figure S7M) that was indistinguishable from the mib−/− single mutant ( Figure S7K). This result indicates that Par-3 restricts Notch activity through Mib. Although the diminished Notch activation in the mib−/− mutant is expected to upregulate Notch ligand expression via the negative feedback loop, this was not what we observed. Instead, the dla mRNA level was significantly reduced in the mib−/− mutant ( Figures S7N and S7O) as well as in the par-3 and mib double-deficient embryos ( Figure S7Q).

Unlike the excitatory Selleckchem KRX-0401 channelrhodopsins, NpHR is a true pump and requires

constant light in order to move through its photocycle. Moreover, although optogenetic inhibition with NpHR was shown to operate well in freely moving worms and in mammalian brain slices ( Zhang et al., 2007) as well as cultured neurons ( Zhang et al., 2007 and Han and Boyden, 2007), several years passed before mammalian validation of any inhibitory optogenetic tool was obtained by successful application to behavioral studies in intact mammals ( Witten et al., 2010 and Tye et al., 2011), due to membrane trafficking problems that required additional engineering ( Gradinaru et al., 2008, Gradinaru et al., 2010 and Zhao et al., 2008). At high expression levels, NpHR-EYFP-expressing cells were found to show accumulations of intracellular fluorescence that colocalized with endoplasmic reticulum (Gradinaru et al., 2008). Addition of an ER export motif from the Kir2.1 potassium channel (ER2—identified PD0332991 cost after

a screen of many possible corrective motifs; Gradinaru et al., 2008) improved the surface membrane localization of NpHR and yielded eNpHR2.0 (Gradinaru et al., 2008 and Zhao et al., 2008), with higher currents suitable for use in intact rodent tissue (Sohal et al., 2009 and Tønnesen et al., 2009) as well as in human and nonhuman primate tissue (Busskamp et al., 2010 and Diester et al., 2011). Next, eNpHR3.0, which additionally contains a neurite trafficking sequence from the Kir2.1 potassium channel, showed further enhanced photocurrents (nanoampere scale at moderate light intensities, < 5 mW/mm2) that can be used to drive inhibition by yellow- or far-red-shifted wavelengths (up to 680 nm at the infrared Endonuclease border; Gradinaru et al., 2010). eNpHR3.0 ultimately enabled the loss-of-function

side of optogenetics for behavior in freely moving mammals (Witten et al., 2010 and Tye et al., 2011), complementing the engineered channelrhodopsins that had enabled gain-of-function in freely moving mammals (Adamantidis et al., 2007). eNpHR3.0 was first used along with bilateral optical fiber devices to inhibit the cholinergic neurons of the nucleus accumbens and elucidate a causal role for these rare cells in implementing cocaine conditioning in freely moving mice, which appears to operate via enhancing inhibition of inhibitory striatal medium spiny neurons (Witten et al., 2010). eNpHR3.0 was also used in a two-fiber approach to inhibit a specific intra-amygdala projection in freely moving mice, implicating a defined neural pathway in aspects of anxiety and anxiolysis (Tye et al., 2011).

These are robust during extended illumination and can be very sensitive to the external electric field. Zero-dimensional nanoparticles, i.e., quantum dots, could be directly used to measure

voltage in neurons. Other nanoparticles, such as nanodiamonds VEGFR inhibitor (Mochalin et al., 2012), may provide an even higher sensitivity to magnetic and electric fields. In addition, by acting as “antennas” for light, nanoparticles can greatly enhance optical signals emitted by more traditional voltage reporters. But regardless of the method chosen for imaging neuronal activity, to capture all spikes from all neurons, one needs to increase the number of imaged neurons and extend the depth of the imaged tissue. A variety of recent advancements in optical hardware and computational approaches could overcome these challenges (Yuste, 2011). Novel methods include powerful Selleckchem ATM/ATR inhibitor light sources for two-photon excitation of deep tissue, faster scanning strategies, scanless approaches using spatio-light-modulators to “bathe” the sample with light, high-numerical aperture objectives with large fields of view, engineered point spread functions and adaptive optics corrections of scattering distortions, light-field cameras to reconstruct signals emanating

in 3D, and, finally, advances in computational optics and smart algorithms that use prior information of the sample. A combination of many of these novel methods may allow simultaneous 3D imaging of neurons located in many different focal planes in an awake animal. In addition, GRIN fibers and endoscopes allow imaging deeper structures, such as the hippocampus, albeit with some invasiveness. Electrical recording of neuronal activity is now becoming possible on a massively parallel scale by harnessing novel developments in silicon-based nanoprobes (Figure 2). Silicon-based

neural probes with several dozen electrodes are already Liothyronine Sodium available commercially; it is now feasible to record from dozens of sites per silicon neural probe, densely, at a pitch of tens of μm (Du et al., 2009a). Stacking of two-dimensional multishank arrays into three-dimensional probe arrays would provide the potential for hundreds of thousands of recording sites. There are technical hurdles to be surmounted, but when the technology is perfected, recording from many thousands of neurons is conceivable with advanced spike-sorting algorithms. The “Holy Grail” will be to record from millions of electrodes, keeping the same bandwidth, reducing the electrode pitch down to distances of ∼15 μm, and increasing the probe length to cortical dimensions of several centimeters. This will require significant innovation in systems engineering. We also envision techniques for wireless, noninvasive readout of the activity of neuronal populations (Figure 2).

Moreover, the mGluR5 knockouts show a deficit in the developmental switch from Selleckchem Tariquidar NR2B to NR2A both at CA1 synapses and at inputs onto layer 2/3 pyramidal neurons in primary visual cortex. Finally, we show that the NR2B-NR2A switch driven by brief visual experience in layer 2/3 pyramidal neurons in dark-reared mice is absent in the mGluR5 knockout. These findings define the mechanism for the activity-dependent NR2B-NR2A switch and suggest a central role for this mechanism in the development- and experience-dependent regulation of cortical NMDAR NR2 subunit composition. Our results show that an LTP induction protocol increases

the relative amount of NR2A at CA1 synapses in an mGluR5 and NMDAR-dependent manner in the neonate. Moreover, mGluR5 function plays an important role in the rapid experience-driven switch in NR2 subunit composition in

pyramidal cells in layer 2/3 of the V1 cortex. In support of a requirement for mGluR5 and NMDARs in the activity-dependent change in the NR2 subunits, NMDARs are also required for this rapid experience-driven NR2B-NR2A switch in primary visual cortex (Quinlan et al., 1999). Together, these findings indicate that this mechanism may represent a ubiquitous process in the developing brain for the activity-dependent regulation of NMDAR function. This is in addition to the variety of other mechanisms described for the regulation Bortezomib of NMDAR function and trafficking in more mature brain (for reviews see Chen and Roche, 2007, Lau and Zukin, 2007 and Yashiro and Philpot, 2008). Whether the developmental regulation of NR2 subunit composition also involves some of the induction and expression mechanisms described in older animals is unclear and will be of interest to study in future work. High-frequency

stimulation can also have long-lasting potentiating effects on NMDAR-mediated synaptic transmission in adult CA1 hippocampus (Bashir et al., 1991). Interestingly, Idoxuridine this NMDAR LTP is also dependent on mGluR5 and NMDAR activation (O’Connor et al., 1994, Jia et al., 1998, Kotecha et al., 2003 and Rebola et al., 2008). Recent work shows that such NMDAR LTP also requires membrane fusion and causes a speeding in the kinetics of the NMDA EPSC (Peng et al., 2010). However, in the present study we did not observe significant changes in NMDAR peak amplitudes after the induction protocol, suggesting that in the neonate, NR2A-containing receptors replace NR2B-containing receptors as opposed to being added to the existing pool of synaptic NMDARs. Consistent with NMDAR replacement in our experiments, NR2B-containing receptors are more mobile and can diffuse to extrasynaptic sites at greater rates than NR2A-containing receptors (Groc et al., 2006 and Tovar and Westbrook, 2002), and NMDARs more rapidly internalize early in development (Washbourne et al., 2004 and Roche et al., 2001).