As previously reported (Echelard et al., 1993), we did not observe expression of indian hedgehog or

desert hedgehog in the adult brain by in situ hybridization (data not shown). We did not observe expression of shh in the dorsal SVZ. However, shh mRNA was present in the medial septum, ventral forebrain, and in infrequent cells close to the ventral SVZ ( Figures S4A–S4C). This was confirmed by qRT-PCR on dissected ventral SVZ and septum ( Figure S4D) and is consistent learn more with previous reports of in situ hybridization in the rat brain ( Traiffort et al., 1999). We next used a genetic approach to label the cells producing Shh and visualize cell morphology. We first examined ShhCre; R26YFP animals ( Harfe et al., 2004), in which cells

http://www.selleckchem.com/products/abt-199.html expressing Shh at any point in development recombine the R26YFP reporter ( Figure S4E). In addition to labeling in the cerebellum and cortex, we also observed an accumulation of YFP+ cells in the ventral forebrain. By administering tamoxifen to ShhCreER; R26YFP animals, we induced YFP expression in cells producing Shh in the adult ( Figure 3; Harfe et al., 2004). YFP expression identified cells in the ventral forebrain, extending along the rostrocaudal axis adjacent and ventral to the SVZ. These cells primarily localized to the medial and ventral septum, the preoptic nuclei near the hypothalamus, and the bed nuclei of the stria terminalis. We also observed rare labeled cells in cortex ( Garcia et al., 2010). Within the septum, YFP+ cells localized to both the horizontal and vertical limbs of the diagonal band, approximately 0.25–1 mm ventromedial to the SVZ ( Figures 3A and 3B). A number of YFP+ cells in the bed nuclei of the stria terminalis were in close proximity to the ventral tip of the lateral ventricle (boxed area in Figures 3A and Etomidate 3D). We did not observe YFP-labeled cells

near the dorsal SVZ, the RMS, OB, or in the choroid plexus—other sites which have been suggested to produce this ligand ( Balordi and Fishell, 2007a and Angot et al., 2008). Labeled cells had the morphology of neurons, and all were colabeled by the neuronal marker protein NeuN ( Figure 3H). Most YFP-labeled cells were GABAergic (GABA-positive; Figure 3I), with a small number of cholinergic (ChAT-positive), YFP-positive cells ( Figure 3J). We did not observe YFP-positive dopaminergic (TH-positive) cells (data not shown). Labeled cells within the bed nucleus of the stria terminalis were of particular interest because of their close proximity to the ventral SVZ, and we examined these cells in greater detail. Computerized tracings of YFP-labeled cells highlighted thin processes that were located close to the basal side of ventral SVZ cells ( Figures 3E and 3F). In order to better characterize these cells, we used whole-mount preparations of ShhCreER; R26YFP animals, allowing en face visualization of the lateral and medial walls of the ventricles.

, 2003), epidermal hair tip and sensory bristle formation in Drosophila ( Cong et al., 2001 and Geng Selleck Apoptosis Compound Library et al., 2000), and dendritic morphogenesis and tiling in the worm and fly ( Emoto et al., 2004, Emoto et al., 2006, Gallegos and Bargmann, 2004 and Han et al., 2012). The

two Trc homologs mammalian (Stk38) and NDR2 (Stk38l; referred to as NDR1/2) are ∼86% identical. Their biochemical activation has been well characterized with no difference between NDR1 and NDR2 reported ( Hergovich et al., 2006). NDR1 and NDR2 are broadly expressed in the mouse brain ( Devroe et al., 2004, Stegert et al., 2004 and Stork et al., 2004). NDR1 knockout mice have increased susceptibility to tumor formation, implicating NDR1 as tumor suppressor ( Cornils et al., 2010). NDR2

levels are increased in NDR1 knockout mice and may compensate for the absence of NDR1 ( Cornils et al., 2010). The potential roles of NDR1/2 in regulating mammalian neuronal morphogenesis are unknown. Despite the importance of the NDR1/2 kinase pathway in regulating cellular morphogenesis in eukaryotes, the downstream phosphorylation targets of NDR1/2 remain largely unknown, except for two substrates for the PLX4032 NDR1/2 yeast homolog Cbk1: Sec2p (Kurischko et al., 2008) and Ssd1p, a nonconserved protein (Jansen et al., 2009), and a recently identified NDR1/2 substrate p21 (Cornils et al., 2011). To elucidate the mechanism of NDR1/2 kinase actions in neurons, it is important to identify the direct phosphorylation targets of NDR1/2 and their functions in the brain. In this study, we investigated NDR1/2 function in cultured rat hippocampal neurons and in mouse cortical neurons in vivo

by perturbing its function via the expression of dominant negative and constitutively active NDR1/2 and siRNA. We found that NDR1/2 kinases limit dendrite branching and length in cultures and in vivo, analogous to the roles of their fly homolog Trc. Additionally, NDR1/2 kinases were also required aminophylline for mushroom spine synapse formation as NDR1/2 loss of function led to more immature spines, both in cultures and in vivo, as well as a reduction in the frequency of miniature excitatory postsynaptic currents (mEPSCs) in neuronal cultures. To uncover the direct targets of NDR1/2, which control dendrite branching and mushroom spine formation, we used chemical genetics to create a mutant NDR1 capable of uniquely utilizing an ATP analog not recognized by endogeneous protein kinases (Blethrow et al., 2008 and Shah et al., 1997). An advantage of this method is that it identifies not only the substrates but also the phosphorylation sites. We identified five potential NDR1 substrates in the mouse brain and chose two for functional validation. We show that one NDR1 substrate is another kinase, AP-2 associated kinase-1 (AAK1), which regulates dendritic branching as a result of NDR1 phosphorylation.

, 2013, O’Reilly and McClelland, 1994 and Rolls and Kesner, 2006). Put into a computational perspective that has attracted considerable attention in recent years, the DG is critical for “pattern separation.” Pattern Ivacaftor clinical trial separation, as related to the DG, can be described as recoding cortical input information into a sparse, essentially orthogonal representation (McNaughton and Morris, 1987 and Treves and Rolls, 1992).

By manipulating the rate of adult neurogenesis, several groups of researchers have shown by ablating or overexpressing adult neurogenesis that newborn neurons are critical for making fine discriminations between neighboring spatial locations or highly similar environments in tests that reflect many of the computational characteristics of pattern separation (Clelland et al., 2009, Creer et al., 2010, Gu et al., 2012, Nakashiba et al., 2012 and Sahay et al., 2011). Together, these studies support the idea that a DG network dominated by young GCs is biased toward interpreting similar but not identical inputs as distinct, whereas older GCs are biased toward

interpreting similar inputs as equivalent. While adult neurogenesis in the DG is now generally accepted to occur in all adult mammals, there are many mechanistic details about how it takes place that will need to be determined before we have a more complete understanding of its functional contribution to hippocampus-mediated behaviors. That said, we need to know a lot more about PD184352 (CI-1040) Akt inhibitor how hippocampal circuits mediate

behaviors, and it is likely that understanding more about adult neurogenesis will contribute to a better understanding of hippocampal function. The field of stem cell biology changed forever when Takahashi and Yamanaka (Takahashi and Yamanaka, 2006) developed a simple and repeatable method to dedifferentiate mouse somatic cells (fibroblasts initially) to embryonic-like cells, termed induced pluripotent stem cells (iPSCs), that could give rise to every cell of the mouse body. The concept of reprogramming emerged from the early works of Briggs and King (Briggs and King, 1952) and Gurdon (Gurdon et al., 1958) but has become widely used as a technique since Takahashi and Yamanaka published their method and similar methods were shown to work for other species, including humans (Takahashi et al., 2007). A plethora of extensions and refinements followed, but the principle was established that essentially all cells in our body maintain an intrinsic plasticity for differentiating into a variety of cell types with completely different functions. The impact of this technology has been dramatic in all areas of biology but has been arguably most dramatic in the neurosciences. While much work remains to be done to improve and refine the technology, attempts to apply these techniques to the clinic are already ongoing.

The difference in heat responsiveness between Trpm3+/+ and Trpm3−/− mice was even more pronounced following injection of CFA. This inflammatory challenge caused a significant reduction in the response latencies of Trpm3+/+ mice, indicative

of heat hyperalgesia learn more but did not change the heat response latencies in Trpm3−/− mice. Taken together, these results establish TRPM3 as a chemo- and thermosensor in the somatosensory system, involved in the detection of noxious stimuli in healthy and inflamed tissue. Our analysis of the heat, capsaicin, and PS sensitivity of DRG and TG neurons from Trpm3+/+, Trpm3−/−, Trpv1+/+, and Trpv1−/− mice indicates the existence of at least four distinct subsets of heat-sensitive neurons. The largest subset encompasses heat-sensitive neurons that responded to both PS and capsaicin, suggesting coexpression of TRPV1 and TRPM3. In addition, we identified heat-sensitive neurons that responded to capsaicin but not to PS (TRPV1-expressing), or to PS but not to capsaicin (TRPM3-expressing). Finally, a fraction of heat-activated neurons was unresponsive to both PS and capsaicin, indicating the existence of a TRPM3- and TRPV1-independent heat-sensing mechanism. In line herewith, Selleck BMN-673 we observed

a substantial fraction of heat-sensitive cells after pharmacological inhibition of TRPV1 in DRG and TG preparations from Trpm3−/− mice. Moreover, Trpm3−/− mice treated with a selective TRPV1 antagonist still responded to noxious heat, albeit with increased latency. The molecular and cellular mechanisms underlying this residual thermosensitivity are currently unknown. How does the heat sensitivity of TRPM3 compare to that of TRPV1 and other thermosensitive TRP channels? From the temperature-induced change in inward TRPM3 current, we determined a maximal Q10 value of ∼7, which is comparable to the Q10 values between 6 and 25 that have been reported for other heat-activated TRP channels, including

TRPV1-TRPV4, TRPM2 and TRPM5 (Caterina et al., 1997, Caterina et al., 1999, Güler et al., 2002, Peier et al., 2002b, Smith et al., 2002, Talavera et al., 2005, Togashi et al., 2006 and Watanabe et al., 2002). Our analysis of the thermodynamic parameters associated with channel gating through further indicated that the temperature dependence of TRPM3 activation is shifted to higher temperature compared with TRPV1. It should be noted, however, that the thermal threshold for heat- or cold-induced action potential initiation in a sensory nerve will not only depend on the thermal sensitivity of the depolarizing thermosensitive (TRP) channels, but also on their expression levels at the sensory nerve endings and on the relative amplitude of other conductances, in particular voltage-gated Na+ channels and various K+ conductances (Basbaum et al., 2009, Madrid et al., 2009, Noël et al., 2009 and Viana et al., 2002). In addition, the thermal sensitivities of TRP channels are known to be modulated by various intra- and extracellular factors (Basbaum et al.

To verify that ePN activity vectors and their distances accurately reflect input to the LH, we expressed GCaMP3 (Tian et al., 2009 and Wang et al., 2003) under GH146-GAL4 control and imaged patterns of calcium selleck influx into ePN axonal branches in the LH ( Figure 1B). Distances between ePN activity vectors explained more than 50% of the observed variation in the structure

of these activity maps when responses to 21 odor pairs were compared across 13 individuals ( Figure 1C). Behavior was analyzed by tracking individual flies in narrow, 50 mm-long chambers (Claridge-Chang et al., 2009). The left and right halves of each chamber were perfused with independently controlled odor streams whose convergence at the midpoint

defined a ∼5 mm-wide choice zone. Each time a fly entered and left this choice zone, a decision was counted (Figure 2A). Choices in favor of either odor were tallied and combined into a single decision bias score. A bias of 100% indicates that a fly always chose one odor over the other; a bias of 0% signals unbiased or random choices. The measurement period was divided into two 2 min intervals, during which the left-right positions of the odorants were reversed (Figure 2A). We selected odors from the set characterized by Hallem et al. (2004) and Hallem and Carlson (2006) that would create odor pairs spanning the whole range of possible ePN distances (Table S1). Flies made an average of 19.9 ± 8.8 decisions per 4 min measurement see more period (mean ± SD, n = 10,102 experiments). When the same odor was delivered to both arms of the chamber, choices were unbiased (decision bias = 0.71% ± 3.30%;

mean ± SEM, n = 161 flies) (Figure 2); when different odors were presented, each odor combination elicited a characteristic bias (Table S1), which was expressed in a qualitatively similar fashion by all members of a population (Figures 2B and 2C). Therefore, the lack of a measurable bias in a population is not a consequence of averaging opposing individual preferences. Differences in behavioral bias can arise from two Resminostat sources: differences in odor discrimination and differences in odor preference. In our analysis, we conceptually separated the processes of odor discrimination and valuation. In this two-step model of odor choice, the animal must first distinguish the odors in a pair and then decide which (if any) it prefers. If it cannot distinguish the odors, it cannot express a preference. Thus, a measurable preference indicates successful discrimination. The converse is not true: a fly may be able to tell two odors apart but may choose randomly between them if it has no incentive to act on a perceived difference. In other words, our measurements cannot distinguish indiscrimination from indifference. Bearing in mind this limitation, we searched for predictors of behavioral bias across a data set of 51 odor pairs.

Q-PCR revealed that the expression levels of Bdnf, Vegf, and Igf1 mRNA were significantly increased by continuous IMI treatment, but were not affected by CUMS ( Figures S5B, S5D, and S5H). Interestingly, the mRNA levels of Gdnf and Nt-3 in the STR and HP, respectively, were significantly decreased by CUMS, and these effects were reversed by continuous IMI treatment ( Figures S5A and S5E). In addition, the mRNA expression level of Gdnf in stressed BALB mice

was significantly decreased in both the dorsal STR (dSTR) and the ventral STR (vSTR) ( Figure 1A). On the contrary, the mRNA expression level learn more of Gdnf in stressed B6 mice was significantly increased in the vSTR but not in the dSTR ( Figure 1B). These changes in GDNF expression were confirmed at the protein level using CH5424802 order an ELISA assay ( Figure 1C). These results suggest that the transcriptional regulation of Gdnf in the vSTR is differentially regulated in the two mouse

strains and may contribute to the observed behavioral responses to CUMS. We next investigated whether a correlation exists between Gdnf expression in the vSTR and behavioral performances in mice. We found that GDNF protein levels in the vSTR of nonstressed BALB and B6 mice were significantly correlated with social interaction time ( Figure 1D) and sucrose preferences ( Figure 1E), but not with immobility times in the forced only swim test ( Figure 1F) or the latency to feed in the novelty-suppressed feeding test ( Figure 1G). These data suggest an important role for GDNF in the vSTR for determining certain types of depression-like behaviors. To directly investigate the role of GDNF in depression-like behaviors, GDNF was overexpressed in the NAc of mice using the polyethylenimine (PEI) gene delivery system. The experimental design is shown in Figure S1B. The successful transduction of EGFP (Figure 1H) and GDNF (Figure 1I) into the NAc of mice using this system was confirmed. We first

assessed social interaction time and sucrose preference for nonstressed B6 mice 2 weeks after the injections of PEI/Gdnf or PEI/Egfp complexes. We found that GDNF overexpression increased the social interaction time ( Figure 1J), but not the sucrose preference ( Figure 1K). We next investigated the effect of GDNF overexpression in stressful conditions. BALB mice were subjected to 4 weeks of CUMS and injected bilaterally into the NAc with either PEI/Gdnf or PEI/Egfp complexes on day 14 of the CUMS session. After the CUMS session, we performed behavioral assays. We found that the social interaction time ( Figure 1J) and sucrose preference ( Figure 1K) of the stressed BALB mice that received PEI/Gdnf complexes were significantly greater than those of the mice receiving PEI/Egfp complexes. These results suggest a crucial role for GDNF in social interactions and sucrose preference.

This hybrid functional/anatomical analysis resulted in clusters of voxels that showed changes with task condition within an anatomical region of interest. In the main analysis, voxels within each functional/anatomical region of interest were then collapsed for the second level ROI analysis comparing control subjects and aMCI patients on placebo using a t test. A separate paired samples t test was used to compare the placebo condition to the levetiracetam condition in aMCI

patients for both the main and confirmatory analysis. We would like to thank Dr. Jason Brandt, Dr. Paul Dash, Dr. Argye Hillis-Trupe, find protocol Dr. Majid Fotuhi, and Dr. Peter Rabins for help with participant recruitment and the staff of the F.M. Kirby Center for Functional Brain Imaging and Alica Diehl, Benjamin Drapcho, and Christina Li for their assistance with data collection. This work was supported find more by NIH grant RC2AG036419 to M.G. M.G. is the founder of AgeneBio. She is an inventor on Johns Hopkins University intellectual property with patents pending and licensed to AgeneBio, and she consults for the company and owns company stock, which is subject to certain restrictions under University policy. M.G.’s

role in the current study was in compliance with the conflict of interest policies of the Johns Hopkins School of Medicine. G.L.K. is an investigator and received research support from UCB Pharma. “
“Recent studies in monkeys have shown that neurons in the lateral habenula (LHb) become active when an animal fails to receive an expected reward or if the animal receives a signal

indicating a negative outcome (Matsumoto and Hikosaka, 2007), i.e., these Vasopressin Receptor neurons encode “antireward” conditions and compute reward prediction errors—the difference between the amount of reward expected and the amount of reward received, a computation that is thought to drive reinforcement learning (Sutton and Barto, 1998). LHb neurons have also been shown to inhibit dopaminergic neurons in the ventral tegmental area (VTA) (Ji and Shepard, 2007), which encode “reward” conditions (Schultz, 1997; but see Matsumoto and Hikosaka, 2009). These findings are consistent with the view that an antireward (LHb) nucleus inhibits a reward (VTA) center and drives negative reward signals in dopamine neurons. However, the nature of the inputs that drive aversive responses in the LHb, as well as their possible modulation by other neurotransmitters, is poorly understood. The globus pallidus internus (GPi), an output region of the basal ganglia, and its nonprimate homolog, the entopeduncular (EP) nucleus, are major sources of input to the primate (Kim et al., 1976 and Parent et al., 2001) and rodent (Herkenham and Nauta, 1977) LHb, respectively, as well as the thalamus (Filion and Harnois, 1978, Harnois and Filion, 1982 and Parent et al., 2001).

Although cerebral endothelial cells exhibit low endocytosis activity, selective and tightly controlled trans-cellular transport mechanisms exist, via either nonspecific endocytosis or receptor-mediated endocytosis. Nonspecific endocytosis includes fluid-phase endocytosis (the capture of soluble molecules by endothelial membrane vesicles) and adsorptive endocytosis (binding of molecules by endothelial membrane proteins) ( Gloor et al., 2001). Receptor-mediated endocytosis involves endothelial transmembrane receptors, such as the transferrin receptor ( Zheng and Monnot, 2012), the insulin receptor ( Banks et al.,

2012), and the low-density lipoprotein (LDL) receptor-related proteins (LRPs), namely LRP-1 ( Deane et al., 2008). The family of ATP-binding cassette (ABC) transporters also plays a central role as efflux transporters for a wide range of lipophilic and amphipathic natural products, Ibrutinib in vivo among which are bacterial, herbal, and fungal toxins. They act as a detoxification system by protecting neurons selleckchem from toxic compounds present in their microenvironment ( ElAli and Hermann, 2011). The drug transporters ABCB1 and ABCG2 have been shown to be highly expressed at the luminal side of endothelial cells, acting as gatekeepers by impeding toxic compounds from CNS entry and accumulation ( Figure 1B). For decades, the immune privilege of the CNS was understood as

an absence of an immune system inside the CNS, and the BBB was considered only as a barrier isolating the CNS from the peripheral immune system, preventing the entry of infectious agents and immune cells into the CNS (Pachter et al., 2003). Extensive work in the last decade unravelled the presence of a specialized intrinsic innate immune system in the CNS (Rivest, 2009), which was accompanied by several observations showing that the BBB is not a neutral and passive barrier, from an immunological point of view, Thymidine kinase but rather contributes actively to the immune response of the CNS (Muldoon et al., 2013). More precisely, several data sets

showed that the peripheral immune cells can still cross an intact BBB (Carson et al., 2006), and the latter can modulate the function and control the fate of infiltrating cells (Ifergan et al., 2008), outlining a more active role of the BBB in the CNS intrinsic innate immunity. While there is limited infiltration of peripheral immune cells into the CNS in physiological conditions, neutrophils, eosinophils, T lymphocytes, monocytes, and others can be found in the CNS parenchyma after injuries to the CNS, including infections and chronic diseases such as multiple sclerosis (MS) (Wilson et al., 2010). However, the luminal side of the BBB is in constant contact with leukocytes patrolling the barrier. The advent of in vivo imaging techniques such as two-photon microscopy has allowed for the live imaging of cells constantly patrolling the brain vasculature (Coisne et al.

Behavioral context appears to even more strongly modulate pulvinar activity and, due to its connectivity, the pulvinar

is well positioned to influence feedforward and feedback information transmission between cortical areas. Because the TRN provides strong inhibitory input to both the LGN and pulvinar, the TRN may control and coordinate the information transmitted along both retino-cortical and cortico-cortical pathways. The visual thalamus serves as a useful model for the thalamus in general because of common cellular mechanisms and thalamo-cortical connectivity principles across different sensorimotor domains. Specifically, the LGN and pulvinar respectively serve as models for first- and Gemcitabine purchase higher-order thalamic nuclei, under

inhibitory control from associated sectors of the TRN. Because the pulvinar receives input from the SC to form an extra-geniculate pathway to cortex, the pulvinar also promises to be a useful model for higher-order thalamic nuclei that receive ascending sensory information from brainstem inputs—that is, nuclei exhibiting mixed first- and higher-order characteristics. As noted in our review, there are bold question marks regarding the exact role of the visual thalamus, particularly the pulvinar and TRN, in perception and cognition, and our account of these functional roles cannot be more than an approximation based on sparse experimental evidence at this time. While there has been much http://www.selleckchem.com/epigenetic-reader-domain.html study in in vitro and in anesthetized in vivo preparations of the cellular mechanisms involved in thalamo-cortical transmission, studies are missing that will link the mechanistic details to perceptual

and cognitive operations. For example, it is still not clear how firing modes or oscillatory activity in the thalamus relate to perceptual and cognitive processing. Basic electrophysiology studies of the thalamus in animals performing perceptual and cognitive tasks are much needed. Moreover, although selective attention has ADAMTS5 been shown to modulate neural activity in the LGN, pulvinar, and TRN, it is not clear how the TRN interacts with the LGN and pulvinar, nor how the thalamus interacts with cortex depending on behavioral context. These network properties will need investigation using simultaneous recordings from thalamic and cortical areas in awake, behaving primates. One reason for the scarcity of studies on the visual thalamus in awake, behaving animals may be the classical view that cognition is the exclusive domain of the cortex. An additional reason is presumably methodological, such as the difficulty in targeting thalamic regions. However, this problem has been greatly reduced since structural imaging of macaque brains has become routine. Moreover, combining electrophysiology with electrical stimulation (Berman and Wurtz, 2010) or diffusion tensor imaging (Saalmann, Y.B., Pinsk, M.A., Li, X., and Kastner, S.

Next, we examined whether calcium transients could also be detected in individual somata of Thy1-GCaMP mice. We first tested whether we could detect GCaMP responses in the soma of neurons in layer II/III of the motor cortex in anesthetized animals. In Thy1-GCaMP2.2c mice, we observed GCaMP-expressing neurons but could not detect activated

cells within an imaging window of 250 × 250 μm during a 10 min recording period. In contrast, in Thy1-GCaMP3 mice, we observed 9.3 ± 0.5 cells (n = 4 areas from 3 mice) using the same imaging conditions ( Figures 6A and 6B). Differences between Thy1-GCaMP2.2c and Thy1-GCaMP3 mice are likely due to the fact that significantly fewer layer II/III neurons are labeled in Thy1-GCaMP2.2c selleck products mice ( Figure 1; Figure S2). We also imaged neuronal activity in the motor cortex of awake mice using a fixed-head imaging design (Dombeck Alectinib et al., 2007, 2009). In awake, behaving animals,

we were able to detect activated neurons both in Thy1-GCaMP2.2c and Thy1-GCaMP3 mice ( Figures 6A and 6B; Figure S6; Movie S7). In Thy1-GCaMP3 mice, we detected many more activated neurons (34.4 ± 1.7 cells in a 250 × 250 μm imaging window, n = 5 areas from 3 mice) over a 10 min period ( Figures 6A and 6B) compared to anesthetized animals (see above). The population activity in the primary motor cortex of both Thy1-GCaMP mice was correlated with locomotor activity ( Figure 6C; Figure S6).

Repeated imaging of the same brain area at 15 days after the first imaging showed that most of the same neurons were active in both views ( Figure 6D). From these observations, we conclude that both Thy1-GCaMP2.2c and Thy1-GCaMP3 mice can be used to monitor neuronal activity over extended periods of time in the motor cortex of living animals. In recent studies, viral expression of the ratiometric GECIs (YC3.60 and D3cpV) and GCaMP3 in pyramidal neurons of the mouse somatosensory (barrel) cortex allowed the detection of neuronal activity induced by whisker stimulation (Lütcke et al., 2010; Mittmann et al., 2011; O’Connor et al., 2010; Wallace et al., 2008). To determine whether Ca2+ transients could be detected in the barrel cortex in response to sensory out stimulation in our Thy1-GCaMP mice, we performed a similar test. We used Thy1-GCaMP3 mice because in vivo two-photon imaging in the barrel cortex revealed sparse labeling of layer II/III pyramidal neurons in Thy1-GCaMP2.2c mice and dense labeling in Thy1-GCaMP3 mice (data not shown). To induce sensory stimulation, we deflected multiple mystacial vibrissae ten times using 500 ms air puffs with 10 s interpuff intervals. In Thy1-GCaMP3 transgenic mice, we routinely detected calcium transients associated with whisker stimulation in both cell somata and the adjacent layer II/III neuropil ( Figures 7A–7C and Movie S8).