A detailed spatial analysis of these clusters with respect to the cell’s main axis reveals patterns of microcircuit design that, to our knowledge, have not been described for other cortical areas. The size of these input clusters depends on the cell type of the target cell; the spatial spread of inputs from deep to superficial L2Ps and L3Ps is two times larger when compared to L2Ss. The deep input clusters projecting to L3Ps display a medial asymmetric

offset to their main axis when compared to L2Ps and L2Ss. A microcircuit has been defined as the “minimal number of interacting neurons that can collectively produce a functional output” (Grillner et al., 2005 and Silberberg et al., 2005). Cells in the superficial find more layers of the MEC integrate position, direction, and speed signals to compute a grid-like matrix of external space, information that is then relayed

to the hippocampus proper (Sargolini et al., 2006). The organization of superficial MEC microcircuitry described here is likely to be instrumental for this integrative computational task, which has already been speculated to be organized in spatially confined integrative units (Sargolini et al., 2006). The observed input clusters defined by the deep to superficial microcircuitry could constitute these integrative units at the microcircuit level. Future work will have to relate Crenolanib the specific patterns of microcircuit design to the systems and behavioral before level function of integrative functional units in the MEC superficial layers. Acute cortical slices were prepared from Wistar rats (age = postnatal day 15–25). Animals were anesthetized and decapitated. The brains were quickly removed and placed in ice-cold ACSF (pH 7.4) containing (in mM) 87 NaCl, 26 NaHCO3, 25 Glucose, 2.4 KCl, 7 MgCl2, 1.25 NaH2PO4, 0.5 CaCl2, and 75 Sucrose. Tissue blocks containing the brain region of interest were mounted on a vibratome (Leica VT 1200, Leica Microsystems, Wetzlar, Germany), cut at 300 μm thickness, and incubated at 35°C for 30 min. The slices were then transferred to ACSF containing (in mM): 119 NaCl, 26 NaHCO3, 10 Glucose, 2.5 KCl, 2.5 CaCl2,

1.3 MgSO4, and 1.25 NaH2PO4. The slices were stored at room temperature in a submerged chamber for 1–5 hr before being transferred to the recording chamber. Whole-cell voltage- and current-clamp recordings were performed with an Axopatch 700B Amplifier (Molecular Devices, Sunny Vale, CA, USA). Data were digitized (National Instruments BNC-2090, Austin, TX, USA) at 5 kHz, low-pass filtered at 2 kHz and recorded in a stimulation-point-specific manner with custom-made software. For calibration experiments, patch electrodes (with electrode resistances ranging from 3–6 MΩ) were filled with (in mM): 135 K-gluconate, 20 KCl, 2 MgATP, 10 HEPES, 0.2 EGTA, and 5 phosphocreatine (final solution pH 7.3). For mapping experiments, the intracellular solution consisted of (in mM): 150 K-gluconate, 0.

We also found no effect of learning on song coding or auditory scene processing in the higher-level AC, in contrast with previous reports that used the European Starling (e.g., Gentner and Margoliash, 2003 and Meliza and Margoliash, 2012), which may suggest differences in cortical plasticity between selleck chemical species with open-ended (European Starling) and close-ended (zebra finch) learning periods.

We propose and model a cortical circuit based on feedforward inhibition that recapitulates salient aspects of the neural coding transformations observed between the primary and higher-level AC. Although the results of the simulation are in close agreement with our physiologic and pharmacologic findings, the model makes assumptions regarding the identity and connectivity of excitatory and inhibitory neurons, and the relative timing of excitatory and inhibitory inputs. The model also assumes that excitatory and inhibitory inputs to BS neurons are perfectly cotuned in frequency,

because in the model excitation is directly supplied and inhibition is indirectly supplied by the same neuron in the primary AC. Although we do not explicitly verify these assumptions, they are supported by previous studies showing that the higher-level AC receives direct XAV-939 datasheet synaptic input from the primary AC and is richly interconnected by local interneurons (Vates et al., 1996), and that neurons

in the songbird (Mooney and Prather, 2005) and mammalian (Atencio and Schreiner, 2008) cortex can be segregated based on action potential width into excitatory (broad) and inhibitory (narrow) populations. Our data show that primary AC and NS neurons in the higher-level AC have similar spike train patterns, firing rates, selectivity, and STRFs, in support of NS neurons receiving direct excitatory first input from the primary AC. Spectrally cotuned but temporally offset excitation and inhibition have been demonstrated in the mammalian auditory cortex (Wehr and Zador, 2003). Our proposed model captures our experimental findings and makes testable hypotheses about how the auditory cortex is organized to transform behaviorally relevant information. Across organisms and sensory modalities, examples of sparse coding (Crochet et al., 2011, DeWeese et al., 2003, Stopfer et al., 2003 and Weliky et al., 2003), contextual sparsification (Haider et al., 2010 and Vinje and Gallant, 2000), and feedforward inhibition (Tiesinga et al., 2008, Vogels et al., 2011 and Wehr and Zador, 2003) are common.

, 2002); from the OPN, Sox14-positive cells extend laterally in the thin layer of cells that make up the nucleus of the optic tract (NOT) ( Figures 2C and 2D). In a more ventral location, Sox14-positive cells cluster at the thalamus-prethalamus border to form the IGL (labeled by Npy expression) with scattered cells in the vLGN ( Figures 2C, 2D, and S1). As at E12.5, all Sox14-positive clusters coexpress the GABAergic marker Gad1 ( Figure S1). GFP-positive axons of Sox14-expressing nuclei extend into the hypothalamus to reach and surround the SCN ( Figure 2C). GFP-positive axons also extend between

the IGL and the PLi and between the PLi and the OPN and CPA ( Figures 2C and 2D). Based on their anatomical location and on check details their cross-connections, we define the pretectal and thalamic domains of Sox14-expressing cells as being part of the SVS. To show that Sox14-expressing cells selleck are part of the non-image-forming circuit originating with ipRGCs, we followed the retrograde transsynaptic spread of the Bartha strain of the pseudorabies virus (PRV152tdTomato). Upon injection in the eye chamber, PRV152 spreads through the parasympathetic circuit

controlling the PLR, eventually reaching ipRGCs in the contralateral eye 72 hr after infection ( Figure 2F) ( Pickard et al., 2002; Viney et al., 2007). We have found that at P3, pups are old enough to survive the procedure and expression from the Sox14 locus is still detectable, albeit at reduced levels and in fewer cells than at P2. Colabeling of GFP and tdTomato highlighted several Sox14-positive cells that contained viral particles within the OPN, CPA, and IGL ( Figure 2E). In contrast, hypothalamic nuclei that are

also part of the PLR circuit only contained viral particles but no GFP-expressing cells (SCN and paraventricular nucleus [PVN]) ( Figure 2E). We also noticed very few and isolated viral particles in the LHa, sometimes coexpressed with the Sox14-expressing cells in the region ( Figure 2E). Examination of the Sox14gfp/+ diencephalon at E12.5 did not show GFP-expressing cells at the thalamus-pretectum border alsactide or next to the habenula ( Figure 3A). By contrast, at E14.5, GFP-positive cells are visible at the future PLi and extend toward the LHa ( Figure 3A). Given that no progenitor domain other than the ones we described at E12.5 arises at this location, we supposed that GFP-positive cells move to the LHa and PLi by tangential migration. To test this hypothesis, we performed live time-lapse imaging on Sox14gfp/+ diencephalic explants in culture. GFP-positive cells are first seen migrating tangentially from the r-Th toward the pretectum at E12.5 ( Figures 3B and 3C; Movie S1). Migration starts in the ventralmost part of the thalamus and moves dorsally, eventually concerning only the dorsalmost tip of the GFP-positive r-Th at E14.5 ( Figures 3B and 3C; Movie S3). By E15.

Furthermore, our data indicate that LEPRs on non-AgRP GABAergic neurons are predominantly responsible for this effect. The following three findings support this view: (1) leptin-mediated reduction of IPSC frequency is minimally affected when LEPRs are deleted from AgRP neurons (Agrp-ires-Cre, Leprlox/lox mice), but is totally abrogated when LEPRs are deleted from all GABAergic neurons (Vgat-ires-Cre, Leprlox/lox mice, Figure 5B); (2) this response is unimpaired in mice that cannot release GABA from AgRP neurons (Agrp-ires-Cre, Vgatlox/lox mice,

Figure 5B); and (3) deletion of LEPRs from GABAergic neurons (Vgat-ires-Cre, Leprlox/lox mice) markedly increases IPSC frequency and amplitude in POMC neurons while, in contrast, no effect is seen when LEPRs are deleted from AgRP neurons Navitoclax solubility dmso (Agrp-ires-Cre, Leprlox/lox mice, Figure 6A). EGFR inhibitor These results clearly attest to the important role played by non-AgRP neurons in leptin-mediated disinhibition of POMC neurons and, of interest, are congruent with the presence of massive obesity versus minimal obesity, respectively, in Vgat-ires-Cre, Leprlox/lox mice ( Figure 2) versus Agrp-ires-Cre, Leprlox/lox mice ( van de Wall et al., 2008). One notable caveat on the above analysis is the possibility of compensation as was observed after

diphtheria toxin-mediated ablation of AGRP neurons in neonates ( Luquet et al., 2005). If such compensation were to occur after genetic deletion of LEPRs in AgRP neurons, then our approach could underestimate the contribution of AgRP GABAergic neurons. However, given that toxin ablation kills neurons while LEPR deletion, on the other hand, leaves neurons largely intact, it is unclear whether similar degrees or forms of compensation should be expected. To summarize, our results and those of others ( Cowley et al., 2001) demonstrate that leptin reduces inhibitory tone to POMC neurons. This effect

is mediated by LEPRs on presynaptic GABAergic neurons, some of which may express AgRP and many of which probably do not. It has previously been established that leptin’s antiobesity effects require Tyr1138 of the LEPR, which allows for phosphorylation-dependent docking and activation (via subsequent phosphorylation) of the latent Vasopressin Receptor transcription factor STAT3 (Bates et al., 2003). Of note, marked obesity, similar in magnitude to that observed in mice totally lacking LEPRs, occurs in mice homozygous for the LeprS1138 allele. This requirement for Tyr1138 strongly implicates STAT3-mediated gene expression in leptin’s antiobesity effects. The relevant downstream transcriptional targets, however, are not yet known but are of great interest. Prior studies have focused on the Pomc gene ( Münzberg et al., 2003). However, given the important role of leptin-responsive GABAergic neurons in regulating body weight, most of which do not express AgRP and none of which appear to express POMC ( Figure 3; Ovesjö et al., 2001 and Yee et al.

, 2011 and Jossin and Cooper, 2011), most likely by sequestering cytoplasmic binding partners find more of endogenous cadherins. Deletion of the binding site for p120ctn within DN-Cdh (Figure 6B) released the dominant-negative effect (Figures 6E and 6F), likely because p120ctn was no longer sequestered, indicating that p120ctn binding to Cdh2 is important for glia-independent somal translocation. The nectin/afadin complex does not bind p120ctn directly, but does so via the small guanosine triphosphatase (GTPase) Rap1, which binds to both afadin and p120ctn (Figure 6A) (Hoshino et al., 2005 and Sato et al., 2006). We hypothesized

that Rap1 might be the crucial link between nectin3 and afadin and Cdh2 and p120ctn pairs. Several lines of evidence support this model. First, Rap1 is required for glia-independent somal translocation, and overexpression

of Cdh2 can rescue the migration defect caused by Rap1 loss of function, demonstrating that Cdh2 acts downstream of Rap1 in this process (Franco et al., 2011). In addition, we now show that a constitutively active form of Rap1 rescued the migration defect caused by nectin3 knockdown (Figures 6C and 6D). Finally, an afadin construct lacking the Rap1 binding site (Figure 6B) acted as a dominant negative and disrupted radial migration selleck chemicals (Figures 6G and 6H). Taken together, our data suggest that nectin3 in migrating neurons recruits an afadin/Rap1 complex that regulates Cdh2 function via p120ctn, thereby promoting leading-process attachment in the MZ and glia-independent somal translocation. At adherens

junctions, cadherins are recruited between neighboring cells through nectin and afadin to form stable adhesions. We therefore reasoned that CR cells might also express Cdh2 that acts in concert with nectin1 to mediate interactions with neurons. Indeed, Cdh2 was expressed in CR cells (Figures 7A and 7B). For functional tests, we electroporated the cortical hem at E11.5 with Dcx-iGFP or Dcx-DN-Cdh-iGFP Fluocinolone acetonide then electroporated the neocortical VZ of the same embryos at E13.5 with Dcx-mCherry to label migrating neurons. By E17.5, GFP+ CR cells had migrated into the neocortical MZ (Figure 7C), while mCherry+ radially migrating neurons populated the emerging CP (Figure 7D). Expression of DN-Cdh did not inhibit the migration of CR cells within the MZ (Figure 7C), but the positions of radially migrating neurons were significantly altered (Figures 7D and 7E). Neurons in controls migrated into the upper CP, whereas large numbers of neurons remained in the lower CP following expression of DN-Cdh in CR cells (Figures 7D and 7E). In addition, neurons in controls had leading processes that branched extensively in the MZ, but branch density was decreased following expression of DN-Cdh in CR cells (Figures 7F and 7G).

1 ± 0.12, n = 15) compared to Ex[DD::UNC-104] cyy-1(+) animals (average intensity of dorsal GFP::RAB-3, normalized; 1.0 ± 0.06, n = 26). We found that during DD remodeling, fluorescently tagged CDK-5 and UNC-104 both exhibit punctate localization patterns and colocalize in the V+D ( Figure S6C) as well as the commissures ( Figure S6D). Furthermore, when we overexpressed CDK-5 in the unc-104(e1265) background, all GFP::RAB-3 was located in the ventral processes and cell bodies ( Figure S6A, A4–A6), just as in the unc-104(e1265) single mutant ( Figure S6A, A1–A3). Again,

these data support the model that UNC-104 is an essential motor protein needed for the transport of synaptic material to the dorsal processes and that CDK-5 facilitates the UNC-104-mediated transport of synaptic selleck compound components to the dorsal Afatinib nmr sites of new synapse formation. When we closely examined the localization of GFP::RAB-3 in the dorsal process, we uncovered an intriguing interplay between the anterograde and retrograde motors during this remodeling event. We found that in 27% of the wild-type worms, GFP::RAB-3 fluorescence first accumulated at the most distal ends of the DD neurons at around

the 20–22 hr time points (Figure 8A, upper image; Figure 8E). Subsequently, fluorescence became evenly redistributed along the dorsal processes at around the 26 hr time point (Figure 8A, lower image), suggestive of a two-step trafficking process during DD remodeling: (1) early anterograde trafficking of synaptic vesicles from the ventral process all the way to the anterior and posterior ends of the dorsal process, and (2) late retrograde movement that results in the even distribution pattern of synaptic vesicles along the dorsal process. P-type ATPase Interestingly, we found that overexpression of UNC-104 led to dramatic, fully penetrant accumulation of GFP::RAB-3 at the distal ends of the dorsal process, even at the 26 hr time

point (Figures 8B, 8E, and 8F), while loss-of-function mutants of UNC-104 showed a complete block of dorsal delivery of GFP::RAB-3 (Figure 8C). As shown in Figure 8A, the accumulation of GFP::RAB-3 at both ends is transient, and fluorescence was then redistributed along the entire dorsal axon in a punctate pattern, similar to the pattern observed in adult animals. Because an anterograde motor UNC-104 directs GFP::RAB-3 to both ends of the dorsal processes, we hypothesized that a retrograde motor Dynein might be responsible for the redistribution of GFP::RAB-3 by delivering the end-accumulated GFP::RAB-3 toward the opposite direction to UNC-104. To test this hypothesis, we disrupted the function of Dynein using a loss-of-function allele of dynein, dhc-1(js319), which has been shown to result in an accumulation of SNB-1/synaptobrevin at the tips of mechanosensory neuronal processes ( Koushika et al., 2004).

, 2011). The authors first demonstrate that mInsc is expressed in the neocortex during mid-neurogenesis and is enriched in the spindle midzone in anaphase progenitor cells. To assess whether or not mInsc is a functional homolog of Drosophila Insc, the authors took an elegant approach and generated transgenic flies expressing mInsc, observing similar localization of mInsc in the Drosophila neuroblast. The authors next investigated the function of mInsc by generating conditional loss-of-function and gain-of-function mice. mInsc mediates the orientation of retina precursor

division (Zigman et al., 2005), but whether this is also true in RG cells has not been clear. Through careful find more measurements of spindle orientation and the angle of division in RG cells, the authors showed that 63% of the mitotic spindles in control embryos were at angles between 0 and 30 (horizontal) while 33% were between 30 and 60 (oblique). Vertically orientated spindles (between 60 and 90) were rare, representing less than 3% of all the mitotic cells. The authors then evaluated mInsc conditional knockout mice (NesCre/+;mInscfl/fl) and found that the majority of mitotic spindles (95%) were between 0 and 30, with oblique and vertical spindles strongly reduced. Overexpression of mInsc in the conditional knock-in

mouse (NesCre/+;R26ki/ki) yielded the opposite phenotype, where oblique and vertical spindles were significantly increased (63%). Therefore, loss of mInsc results in the enrichment of horizontal divisions, whereas overexpression learn more of mInsc randomizes the cleavage plane. What then are the consequences of changing the mitotic spindle angle of RG cells? Analysis of conditional mInsc knockout mice revealed a decrease in cortical thickness, while conditional mInsc overexpression led to an increase in cortical thickness. These phenotypes were attributed to major changes in the number of neurons, as histological

analysis using layer-specific neuronal markers demonstrated a uniform decrease in neurons with mInsc deletion and an increase with mInsc overexpression Plasmin across all cortical layers (Postiglione et al., 2011). To link the alterations of neuron production to the progenitor cell subtypes responsible, the authors examined the M phase index and the cell cycle exit index (Q fraction). Surprisingly, the average cell cycle length and exit rates of neural progenitors did not change in the NesCre/+;mInscfl/fl or the NesCre/+;R26ki/ki mice, indicating that mInsc has little to no general role in regulating the cell cycle. Finally, the authors carefully examined the composition of progenitor cells in the mutants that would lead to the observed changes in neuron number.

, 2008 and Huberman et al., 2003) or whether it can act in an instructive way to guide neural circuit formation through specific spatiotemporal NVP-BGJ398 patterns of neural activity (Feller, 2009 and Huberman et al., 2008). These

issues have been investigated in some detail in the mammalian visual system, where retinal ganglion cell (RGC) projections to the dorsal lateral geniculate nucleus (dLGN) and superior colliculus (SC) form two sensory maps, one reflecting eye of origin and the other retinotopic location (Huberman et al., 2008). Molecular factors are clearly involved in forming these neural circuits, directing RGC axons whether to cross at the optic chiasm (Petros et al., 2008) and where to branch in the dLGN and SC (Huberman et al., 2008 and McLaughlin and O’Leary, 2005). Evidence concerning the role of neuronal activity in early Adriamycin visual map development is more equivocal, failing to distinguish whether neuronal activity acts in a passive way to promote cell survival and neurite outgrowth, or in an instructive way to guide neural circuit formation through specific spatiotemporal patterns of neural activity (Crair, 1999, Stellwagen and Shatz, 2002 and Huberman

et al., 2003). This fundamental question has been difficult to answer because manipulations that change the spatiotemporal pattern of ongoing spontaneous neuronal activity typically also alter the activity of individual neurons (their overall spike rate, or burst frequency, etc.). This completely confounds changes in interneuronal activity patterns with changes in single-neuron activity levels, making it impossible to distinguish between a passive and active role for neuronal activity in visual map development (Chalupa, 2009 and Feller, 2009). As in many parts of the developing brain and spinal cord (Meister et al., 1991, Bekoff et al., 1975 and Feller, 1999), coordinated waves

of spontaneous neuronal Heterotrimeric G protein activity are found in the retina of all mammalian species examined (Wong, 1999 and Warland et al., 2006), well before the onset of sensory experience. Maps for eye of origin and retinotopy emerge in neonatal mice in the first week after birth, a period in which spontaneous retinal activity is mediated by nicotinic acetylcholine receptors containing the β2 subunit (β2-nAChRs; Feller et al., 1996 and Bansal et al., 2000). Genetic and pharmacologic manipulations that impair β2-nAChR-mediated retinal waves cause deficits in visual system development, including defects in retinotopy and eye segregation (Stellwagen and Shatz, 2002, Chandrasekaran et al., 2005, Mrsic-Flogel et al., 2005, Rossi et al., 2001, Grubb et al., 2003, McLaughlin et al., 2003, Penn et al., 1998, Pfeiffenberger et al.

An increasing body of evidence indicates that there is a specific

population of peripheral osmosensory neurons, which represent the afferent arm of a complex reflex response triggered by water intake (Boschmann et al., 2003, Boschmann et al., Trichostatin A 2007, Jordan et al., 1999, Jordan et al., 2000, McHugh et al., 2010, Scott et al., 2000, Scott et al., 2001 and Tank et al., 2003). We postulated that osmosensory neurons detect very small hypo-osmotic shifts in blood osmolality in the hepatic circulation following water intake. Using an activity marker, we could show that hepatic afferent fibers are activated by the small osmolality changes induced by physiological water intake in the mouse. The magnitude of the stimulus was comparable to that shown in healthy humans to rapidly activate a sympathetic reflex that can elevate blood pressure and increase metabolic rate (Boschmann et al., 2007, Jordan et al., 1999, Jordan et al., 2000, Lipp et al.,

2005, Scott et al., 2000 and Scott et al., 2001). By analogy with our animal model, the water-evoked reflexes anti-PD-1 antibody inhibitor observed in humans are also probably mediated by hepatic osmoreceptors capable of detecting a decrease of just 8% in blood osmolality. A key feature of the osmoreceptors described here, is that they can signal changes in blood osmolality well before water intake impacts systemic blood osmolality. Systemic osmolality changes following water intake will be even smaller than what we observed in the hepatic portal vein and would follow the stimulus with some delay (Adachi et al., 1976, Baertschi and Vallet, 1981 and Choi-Kwon and Baertschi, 1991). We used our animal model first to identify the cellular nature of the hepatic osmoreceptor and second to characterize the physiological and molecular nature of osmosensitive transduction in these neurons. The liver is innervated by both vagal and thoracic sensory Heterotrimeric G protein afferents (Carobi

and Magni, 1985, Choi-Kwon and Baertschi, 1991, Magni and Carobi, 1983 and Vallet and Baertschi, 1982). We show that virtually all identified hepatic sensory neurons in the thoracic ganglia possess an osmosensitive current whereas nodose sensory neurons innervating the liver do not (Figure 6B). Using a transgenic animal model in which EGFP is expressed by thoracic ganglion neurons innervating the liver, we have shown that the peripheral endings of hepatic neurons are activated by physiological changes in blood osmolality. Although almost all hepatic sensory neurons could be shown to be osmosensing, it is likely that many nonhepatic thoracic sensory neurons are also osmosensitive. Thus, normally only very few thoracic ganglion neurons are labeled by injection of fluorescent tracers into the liver (<5% of the ganglia).

, 2010). To elucidate the role of Plk2 phosphorylation in AMPAR surface expression, click here we stimulated neuronal activity while blocking Plk2 kinase activity (with BI2536) or Plk2 expression (with Plk2-RNAi). PTX treatment

markedly decreased surface GluA1 (sGluA1) expression only in proximal dendrites, with no change in distal dendrites, and this decrease was abolished by either BI2536 or Plk2 RNAi (Figures 7A and 7C). In contrast, PTX reduced sGluA2 in both proximal and distal dendrites (Figures 7B and 7D), consistent with previous findings (Evers et al., 2010). Interestingly, coincubation of BI2536 with PTX rescued sGluA2 expression only in proximal dendrites, but not distal dendrites, while Plk2 RNAi increased basal sGluA2 expression in both proximal and distal dendrites and abolished PTX-induced removal of sGluA2 in either region (Figures 7B and 7D). No changes in total GluA1/A2 were observed under any conditions (data not shown and Evers et al., GSK1120212 mw 2010). Thus, sGluA1/A2 on proximal dendrites were regulated by a Plk2 kinase-dependent mechanism, whereas the kinase-independent mechanism specifically affected sGluA2 in distal dendrites. We next examined the role of Ras/Rap regulators in overactivity-induced reduction of AMPARs. Cultured neurons were transfected

with shRNA against RasGRF1 or SPAR in the absence of synaptic stimulation to test whether inactivation of Ras or activation of Rap is sufficient to cause loss of surface AMPARs. As expected, knockdown of SPAR reduced sGluA1/A2 expression in proximal dendrites (Figures 7E–7H). Silencing of RasGRF1 also decreased sGluA1 but

only showed a nonsignificant trend for sGluA2 removal (Figures 7E–7H, p = 0.10). We then transfected neurons with shRNA constructs for SynGAP or PDZGEF1 and stimulated with PTX to induce endogenous Histidine ammonia-lyase Plk2. PTX-mediated loss of sGluA1/A2 was completely abolished by silencing SynGAP or PDZGEF1 (Figures 7E–7H). These results demonstrate that tuning down of Ras or tuning up of Rap is necessary and sufficient for PTX-induced reduction of AMPARs in proximal dendrites. Finally, we investigated whether Plk2 phosphorylation of Ras/Rap regulators is important for the PTX effects on surface AMPARs. As before, PTX stimulation reduced sGluA1/A2 levels in proximal dendrites (Figures 7I–7L). Overexpression of RasGRF1 WT or its phosphomutant (S71A) significantly increased sGluA1 expression, and the sGluA1 loss by PTX was partially blocked in neurons expressing S71A (Figures 7I and 7K). In contrast, RasGRF1 expression did not increase sGluA2 levels or prevent PTX-mediated removal of sGluA2 (Figures 7J and 7L), confirming the above result that silencing of RasGRF1 did not greatly reduce sGluA2 (Figures 7G and 7H). Expression of SynGAP WT or PDZGEF1 WT reduced sGluA1/A2, and there was further reduction of sGluA1/A2 after PTX stimulation (Figures 7I–7L).