In this case, the cell does not monitor dendritic excitability, suggesting a sensor that is localized near the soma. A good candidate for the messenger would be Ca2+ influx during AP repolarization, which displays a relatively constant amplitude and duration in the soma compared with dendrites. Alternatively, because of their location distant from the nucleus, dendritic channels and receptors may simply PD173074 be untethered from strict homeostatic excitability mechanisms.

In any event, it is surprising that dendritic excitability is not more closely regulated. Dendritic voltage-gated ion channels regulate the processing and storage of incoming information in CA1 pyramidal neurons (Shah et al., 2010). Perhaps the dynamic nature of channel properties and expression during normal function in dendrites prohibits the establishment of a set point state of excitability. We should make the distinction that the applicable data come only

SB203580 order from recordings in CA1 primary apical dendrites. Oblique dendrites may well use mechanisms to homeostatically regulate their excitability. In CA1 neurons, AP back-propagation decreases with activity (Spruston et al., 1995) because of a combination of slow recovery from inactivation for dendritic Na+ channels and the activity of A- type K+ channels (Colbert et al., 1997 and Jung et al., 1997). We found DPP6 to be particularly important in the regulation of back-propagation at lower frequencies (Figures Ketanserin 5B and 5C). An explanation would be that normally a certain fraction of A-type K+ channels are able to recover from inactivation in between APs, but that without DPP6 the remaining A-type channels are too slow to recover from inactivation, allowing greater back-propagation. DPP6 therefore may be an important contributor to the cellular- and circuit-level mechanisms of theta rhythm (5–10 Hz) found in EEG recordings of the hippocampus during exploratory behavior and REM in the hippocampus. In addition to enhanced back-propagation, we observed that Ca2+

spikes were more readily generated in DPP6-KO dendrites. The activation of dendritic voltage-gated Ca2+ channels by back-propagating APs results at a “critical” frequency will induce a burst of mixed Ca2+ and Na+ action potentials in CA1 pyramidal neurons. Dendritic voltage-gated K+ channels modulate this change in AP firing mode from single to burst firing (Golding et al., 1999 and Magee and Carruth, 1999). We found that the critical frequency for Ca2+ electrogenesis in WT neurons of ∼130 Hz was dramatically lowered to only 85 Hz in DPP6-KO neurons. We have observed previously that this type of complex firing is critical for the induction of GluA1-independent LTP of synaptic inputs using a theta burst-pairing protocol (Hoffman et al., 2002). Using a similar protocol, it has been shown that Kv4.2-KO mice have a lower threshold for LTP induction than WT (Chen et al., 2006 and Zhao et al., 2010).

RT-PCR results demonstrate that after AAV1-VGLUT3 delivery, there is also more widespread VGLUT3 mRNA transcription than in just IHCs (Figure 1C). These results suggest that there is a posttranscriptional regulatory selleck chemicals mechanism acting on VGLUT3 mRNA, which leads to selective expression of the protein only within IHCs. Several types of posttranscriptional regulation have been described within the cochlea, and whether this specific mechanism involves microRNA inactivation (Elkan-Miller et al., 2011), transcription factor regulation (Masuda et al., 2011), or

another process remains to be determined. Such a mechanism, if appropriately elucidated and exploited, could theoretically allow the expression (or conversely suppression) of a number Trametinib of different proteins within the inner ear to alter function in pursuit of hearing preservation.

Another interesting finding was the variable success with the CO as compared to the RWM delivery technique. As noted, we initially started with an apical CO delivery method but abandoned it due to the low success rate of hearing restoration (17% of animals). Subsequently, we changed to an RWM delivery technique for several reasons; this would be the most likely method of delivery in any future human studies, and it was less likely to be traumatic, as evidenced by a number of recent human studies looking at hearing preservation with round window insertion of cochlear implants (von Ilberg et al., 2011). In fact, the change in technique resulted in hearing restoration in 100% of animals attempted. We believe the likely difference in success between the two techniques relates to the degree of trauma induced by each method. With a cochleostomy, a separate hole into the scala through bone must be created, which by its nature is traumatic, despite our best efforts

to minimize trauma. In contrast, an RWM injection simply involves piercing the membrane and sealing it with fascia after viral delivery. However, we were histologically unable to see any obvious differences between the ears of animals with and without hearing rescue in the cochleostomy group (data not shown) and there may be second other reasons for the variable success between the two techniques. Further, we noted that even earlier delivery via the RWM at P1–P3, as opposed to P10–P12, resulted in hearing recovery that was more consistently long lived, with all mice followed out through 9 months showing ongoing normal ABR thresholds (Figure 3D). Transgene expression with AAV1 should theoretically last for a year or longer (Henckaerts and Linden, 2010). However, it is not entirely clear why there is a variable loss of hearing after 7 weeks, regardless of delivery technique at the later P10–P12 delivery time point (Figure 3D).

For these reasons, recent alternative approaches emphasize either pure subjective reports, such as ratings of stimulus visibility (Sergent and Dehaene, 2004), or second-order commentaries such as postdecision wagering (e.g., would you bet that your response was correct?; Persaud et al., 2007). The wagering method and related confidence judgements provide a high motivation to respond truthfully and in an unbiased manner (Schurger Bortezomib chemical structure and Sher, 2008). Furthemore, they can be adapted to nonhuman subjects (Kiani and Shadlen, 2009 and Terrace and Son, 2009). However, they can sometimes exceed chance level

even when subjects deny seeing the stimulus (Kanai et al., 2010). Conversely, subjective report is arguably the primary data of interest selleck compound in consciousness research. Furthermore, reports of stimulus visibility can be finely quantified, leading to the discovery that conscious perception can be “all-or-none” in some paradigms (Del Cul et al., 2007, Del Cul et al., 2006 and Sergent and Dehaene, 2004). Subjective reports also present the advantage of assessing conscious access immediately and on every trial, thus permitting postexperiment sorting of conscious versus nonconscious trials

with identical stimuli (e.g., Del Cul et al., 2007, Lamy et al., 2009, Pins and Ffytche, 2003, Sergent et al., 2005 and Wyart and Tallon-Baudry, 2008). Although the debate about optimal measures of conscious perception continues, it is important to acknowledge that objective assessments, wagering indices and subjective reports are generally in excellent agreement (Del Cul et al., 2006, Del Cul et al., 2009 and Persaud et al., 2007). For instance, in visual masking, the conscious perception thresholds derived from objective and subjective data are essentially identical across subjects (r2 = 0.96, slope ≈ 1) (Del Cul et al., 2006). Those data suggest that

conscious access causes a major change in the global availability of information, Ketanserin whether queried by objective or by subjective means, whose mechanism is the focus of the present review. Conscious access must be distinguished from the related concept of attention. William James (1890) provided a well-known definition of attention as “the taking possession by the mind, in clear and vivid form, of one out of what seem several simultaneously possible objects or trains of thought.” The problem with this definition is that it conflates two processes that are now clearly separated in cognitive psychology and cognitive neuroscience (e.g., Huang, 2010 and Posner and Dehaene, 1994): selection and access.

The sections were then mounted in Vectashield (Vector Laboratories). For quantitative analysis, confocal image stacks from the MEC Cell Cycle inhibitor were obtained on a laser-scanning confocal microscope from four animals (5–10 histological sections each). Images were taken using a ×20 objective (NA 0.75) at 2 μm axial resolution and collected to cover the full extent of L2 and L3 of the MEC in the histological section. Image stacks obtained were registered and combined in Fiji (http://fiji.sc/wiki/index.php/Fiji) to form a montage of the sections. Additional higher resolution images

were taken using a ×60 objective (NA 1.3) at 0.5 μm axial resolution from selected areas. Immunostaining intensity for PV and VGAT over the neuropil of L2 and L3 was measured in 50 × 50 μm or 50 × 100 μm “regions of interests” positioned quasirandomly within 500 or 200 μm vertical wide strips of MEC in image planes from the top of the sections. Measured intensity values were normalized using the maximal intensity measured in each section. Putative GABAergic terminals and PV+ neuronal profiles were identified, and their numbers were determined in high-resolution image stacks of the immunolabeling for V-GAT by threshold-based segmentation using the “3D Objects Counter” plugin in Fiji. PF-01367338 solubility dmso Colocalization

of PV in V-GAT+ terminals was determined as the overlap in the segmented stacks. Cell densities were calculated using the optical dissector approach (West and Gundersen, 1990) from the full 50 μm thickness of the histological sections in 200 μm vertical strips of MEC from L2 and L3 separately. Regression analysis was performed on the two data sets plotted against the distance of the stripes along the DVA (measured from the dorsal end of the MEC to the midline of the stripes) using R software (http://www.r-project.org/). For graphical presentation, the data were rebinned at 1,000 μm, and mean ± SEM were plotted against the midposition of the pooled bins along the DVA. For studying gamma oscillations, slices were stored and recorded from in an interface-type recording chamber. The extracellular recording electrode was placed in

the superficial layers of MEC (LII), and baseline activity was recorded. Gamma oscillations were then induced by bath applying 300 nM kainate for up to 40 min. Both horizontal and sagittal slice Megestrol Acetate orientations that contained the MEC along with the rest of hippocampal formation were used. In sagittal preparations, two recording electrodes were used to record the gamma simultaneously from the dorsal and ventral MEC. In horizontal preparations, a dorsal and a ventral slice were recorded in parallel. In a subset of experiments, the GABAA receptor blocker gabazine (0.5 μM) was washed in to block inhibitory inputs. Male Wistar rats weighing 150–350 g were anesthetized with a combination of urethane and ketamine (Quilichini et al., 2010). The local field potential was recorded simultaneously using two tungsten electrodes (0.

Moreover, overexpression of ADAM10 in mouse primary neurons increased the production of sNLG1 ( Figure S2B). Lastly, to demonstrate the physiological significance of ADAM10 in NLG1 processing in vivo, we incubated the brain microsome from postnatal day (P) 18 neuron-specific conditional ADAM10 knockout

mice (Adam10flox/flox; Selleckchem Screening Library CamKII-Cre) ( Figure 3I). Notably, the levels of NLG1-FL were increased in the microsome fractions from brains of Adam10 conditional knockout mice, whereas sNLG1 production was significantly decreased ( Figures 3J and 3K). Taken together, we concluded that the major physiological sheddase of NLG1 in brain is ADAM10. We next examined the proteolytic processing of NLG1 at the ectodomain in more detail. According to the molecular Smad inhibitor weight of the CTF, the stalk region of NLG1 is predicted as the candidate cleavage site for shedding. To obtain further precise information on the location and characteristics of the cleavage site, we

analyzed mutant forms of NLG1 overexpressed in COS-1 cells (Figure 4A). Recombinant NLG1-FL was sequentially cleaved similarly to the endogenous NLG1-FL in primary neurons, whereas NLG1-ICD was not observed (Figures 4B and 4C). To identify the extracellular cleavage site of NLG1, we have systematically generated a series of alanine substitutions around the juxtamembrane stalk region, at sites consistent with the molecular weight of sNLG1 and NLG1-CTF: K674QDD/AAAA, P678KQQ/AAAA, P682SPF/AAAA, S686VDQ/AAAA, R690DYS/AAAA, and T694E/AA (Figure 4A). Among these mutants, generation of sNLG1 was significantly decreased in the PKQQ/AAAA mutant, whereas the level of

mutant FL protein was increased (Figures 4D and 4E). The observation that overexpressed PKQQ/AAAA mutant NLG1 accumulated at dendritic spines in rat primary neurons similarly to wild-type (WT) NLG1 and showed spinogenic effects supported the view that introduction of PKQQ/AAAA mutation did not affect the trafficking of NLG1 (see Figure 8C). In contrast, alanine substitutions at regions proximal to the membrane caused an augmentation of NLG1 shedding. Intriguingly, the levels of sNLG1 were significantly increased in the PSPF/AAAA and SVDQ/AAAA mutants, whereas the corresponding FL proteins were these decreased, suggesting that the region from Pro682 to Gln689 negatively regulates the NLG1 shedding. These data suggest that the region between Pro678 to Gln681 is critical for the sNLG1 production. We then overexpressed NLG1-ΔE, a recombinant polypeptide corresponding to NLG-CTF starting from residue Val682 fused with a signal peptide (Figure 4A). The levels of NLG1-ΔE were increased by DAPT treatment, indicating that NLG1-ΔE was processed by γ-secretase (Figure 4F). However, NLG1-ICD was again undetectable in lysates of transfected cells.

, 2010). To address whether Raf/MEK/ERK signaling in Schwann cells is involved in the recruitment of inflammatory cells, we compared P0-RafTR nerves to control nerve sections using a panel of inflammatory

cell markers (Figures 5 and S5). Remarkably, in nerves from injected P0-RafTR mice, we observed a large increase in the number of macrophages, GSK 3 inhibitor mast cells, neutrophils, and T cells, all of which have been shown to be recruited into nerves following injury (Figures 5A–5E and S5A–S5C). These results were confirmed by analysis of EM sections, where large numbers of macrophages and mast cells could be seen (Figures 5A and S5C). However, one cell type, fibroblasts, which are found in large numbers in damaged nerves were not recruited into P0-RafTR nerves following tamoxifen injection (Figure S5D), suggesting that Schwann cells may not be responsible for recruiting fibroblasts to damaged nerves. Interestingly, the influx of inflammatory cells mirrored the response seen following an injury, with neutrophils entering the nerve at day 3, followed by macrophages, mast cells and T cells at around day 5 with the numbers increasing over time (Hall, 2005 and Mueller et al., 2003). Importantly, at day 5, there was no observable myelin breakdown—suggesting that signals from dedifferentiated

Schwann cells, rather than www.selleckchem.com/products/AZD8055.html axons or myelin debris, are responsible for recruiting inflammatory cells. The simplest explanation for our findings is that substances secreted by Raf-activated Schwann cells were directly responsible for the inflammatory response. We therefore tested whether conditioned medium (CM) from cultures of tamoxifen-treated rat Schwann cells expressing and the RafTR (NSRafER) (Lloyd et al., 1997) was able to attract inflammatory cells in a similar fashion to that seen in vivo. We collected blood from adult rats, purified the white blood cell fraction and found that CM from tamoxifen-treated cells attracted significantly more monocytes, T cells, and granulocytes compared to vehicle-treated controls (Figures 6A and S6A). Moreover, this response was dependent

on signaling through the ERK pathway confirming the specificity of the response. In contrast, CM from tamoxifen-treated cells was unable to attract more fibroblasts suggesting that cytokines produced by dedifferentiated Schwann cells do not promote fibroblast attraction which is consistent with the lack of a fibroblast response in the RafTR nerves (Figure 6A). To determine the Schwann cell-derived molecules which may be involved in mediating the inflammatory response, we reexamined a microarray analysis performed on NSRafER cells (Parrinello et al., 2008) and found that a number of mRNAs encoding secreted factors were upregulated following Raf activation in dedifferentiated Schwann cells (Table 1). These included cytokines, some of which had been previously implicated in attracting inflammatory cells following nerve trauma such as the c-kit ligand and MCP-1 (Perrin et al.

He found that his depressed patients had a systematic negative bias. They almost invariably had unrealistically high expectations of themselves, put themselves down whenever possible, Dinaciclib and were pessimistic about their future. Beck addressed these distorted negative beliefs and found that his patients often improved with remarkable speed, feeling and functioning better after a few sessions. This led him to develop cognitive behavioral therapy, a systematic approach to therapy that focuses on the patient’s cognitive

style and distorted way of thinking (Beck, 1995). This systematic approach enabled Beck and others to study the outcomes of treatments for depression empirically. Their studies showed that cognitive behavioral therapy is as effective as, or more effective than, antidepressant medication in treating people with mild and moderate depression. It is less effective in severe depression, but it acts synergistically with antidepressants. Beck’s findings encouraged investigators to carry out

empirical outcome studies of psychoanalytically oriented insight therapy, and some progress has been made in this area selleck compound (Roose et al., 2008 and Shedler, 2010). In fact, a modest movement is now afoot to develop biological means of testing specific aspects of psychoanalytic theory and thus to link psychoanalysis to the biology of the mind. One reason we know so little about the biology of mental illness is that we know little about the neural circuits that are disturbed in psychiatric disorders; however, we are now beginning to discern a complex neural circuit that becomes disordered in depressive illnesses. Helen Mayberg, at Emory University, found and other scientists have used brain-scanning techniques to identify several components of this circuit,

two of which are particularly important. One is Area 25 (the subcallosal cingulate region), which mediates our autonomic and motor responses to emotional stress; the other is the right anterior insula, a region that becomes active during tasks that involve self-awareness as well as tasks that involve interpersonal experience. These two regions connect to other important regions of the brain, all of which can be disturbed in depressive illness. In a recent study of people with depression, Mayberg gave each person either cognitive behavioral therapy or an antidepressant medication (McGrath et al., 2013). She found that people who started with less than average activity in the right anterior insula responded well to cognitive behavioral therapy but not to the antidepressant. People with greater than average baseline activity responded to the antidepressant but not to cognitive behavioral therapy. Mayberg could actually predict a depressed person’s response to specific treatments from the baseline activity in their right anterior insula.

For the initial set of experiments we used extracellular recording in acutely prepared rat hippocampal slices and stimulated buy Antidiabetic Compound Library two independent inputs onto the same population of neurons. We decided to test the effects of the JAK inhibitor AG490 (10 μM), since this inhibitor has been shown to interfere with learning and memory (Chiba et al., 2009b). We found that AG490 had no effect on baseline transmission (100% ± 1% before and 101% ± 1% during AG490 application, n = 13). Next we tested the effects

of AG490 on NMDAR-LTP, since this is the most widely studied cellular correlate of learning and memory (Bliss and Collingridge, 1993). However, we found no difference between the level of LTP induced in the control

input, in which AG490 was applied immediately after the tetanus, or in the input tetanized in the presence of AG490 (Figure 1A). Thus, the level of LTP obtained 30 min following the tetanus, expressed as a percentage of baseline, was 135% ± 4% and 145% ± 3% (n = 4), respectively. These values were similar to the level of LTP induced in untreated inputs (140% ± 3% of baseline, n = 6; Figure 1C). Since more recent evidence has suggested that NMDAR-LTD is also involved in some forms of learning and memory (see Collingridge et al., 2010) we next tested AG490 on this form of synaptic plasticity. In all experiments, AG490 completely prevented the induction of NMDAR-LTD induced by low-frequency stimulation (LFS; Selleckchem Cabozantinib comprising of 900 stimuli delivered at 1 Hz), though usually a short-term depression remained (Figure 1B). In all cases, the block of NMDAR-LTD was fully reversible since a second, identical period of LFS induced LTD that was similar to that observed under control conditions. Thus, 60 min following the first LFS, delivered in presence of AG490, the responses were 99% ± 4% of baseline and 60 min following the second LFS, delivered after washout of AG490, they were 74% ± 11% of baseline (n = 6). In contrast to the dramatic effect on the induction of NMDAR-LTD, AG490 had no effect on the expression phase of this process. Farnesyltransferase Thus, LFS induced an LTD that was 71% ± 9% and 72% ± 9% of

baseline (n = 6), before and following the application of AG490, respectively. Since these experiments were all performed using two inputs, the ability of AG490 to selectively and reversibly block the induction of NMDAR-LTD without affecting baseline transmission or the expression of NMDAR-LTD were all internally controlled. Next, we explored whether the effects of AG490 were specific for de novo NMDAR-LTD or whether it blocked all forms of LTD. To do this we investigated depotentiation, the reversal of a previously potentiated input. For these experiments we compared, in the two inputs, the level of depotentiation before the application and in the presence of AG490. Under both sets of conditions, LFS reversed LTP to baseline conditions (Figure 1C).

We also observed increased bursting of complex spikes in the HCN1 knockout mice. Complex bursts are known to be important for information coding in hippocampus (Lisman, 1997) and bursts with shorter intervals are known to elicit LTP (Larson et al., 1986) and have an important role in synaptic plasticity (Thomas et al., 1998). We see a significant increase in complex bursting of CA1 place cells whereas CA3 place cells show

only a small, statistically insignificant increase in bursting, probably due to their low level of HCN1 expression. Complex bursts are thought to depend on the firing of dendritic Ca2+ spikes (Kamondi et al., 1998a). The increased CA1 bursting is consistent with the observation that the dendritic spikes are enhanced in CA1 neurons (Tsay et al., 2007). HCN1 channels are required for the large SB203580 Ih in the stellate neurons of layer II of EC (Garden et al., 2008) where they regulate low-frequency membrane potential oscillations (Giocomo and Hasselmo, 2009). Birinapant Therefore, one cannot rule out the possibility that the bursting properties of the hippocampal neurons could be driven by grid cell inputs. The results on the HCN1 knockout mice thus reveal

a series of phenotypic changes in learning and memory, on the one hand, and place cell properties on the other. Comparison of changes in CA1 and CA3 place cells indicate that these alterations are likely to reflect both changes in the entorhinal cortex grid cell inputs to these neurons as well as, in the case of CA1, a direct influence of HCN1 intrinsic to the place cell. Taken together with the results of Giocomo et al. (2011) on how HCN1 deletion alters grid cell properties, these results provide strong evidence that the firing properties of grid cells are important determinants of the properties of the downstream hippocampal place cells. Moreover, these properties are likely to contribute to the action of HCN1 to constrain spatial learning and memory. Forebrain restricted HCN1 KO mice (HCN1f/f,cre) and control littermates (HCN1f/f)

in a hybrid 50:50% C57BL/6J:129SVEV background were bred and raised in the New York State Psychiatric Institute animal care enough facilities as described (Nolan et al., 2004 and Nolan et al., 2003). Mice were studied between 3 and 6 months of age and weighed about 26–37 g at the time of electrode implantation surgery. The littermates were housed in groups of not more than five per cage. Following surgery, all mice were individually housed under 12 hr light/dark cycle and provided with food and water ad libitum. All breeding and housing procedures conformed to National Institute of Health (NIH) standards using protocols approved by the Institutional Animal Care and Use Committee (IACUC).

Strips were incubated overnight at 4 °C with sheep sera diluted 1:50 in PBS-TM 1% and then with biotinylated-rabbit

anti-sheep IgG diluted 1:500 followed by incubation with peroxidase-streptavidin (Sigma) diluted 1:1000 in PBS-TM at 1%. The reaction was developed by adding enzyme substrate (Fast™ 3,3′-diaminobenzidine tablet sets; Sigma). Samples were considered positive when showing seroreactivity of IgG antibodies to T. gondii SAG1 (p30) antigen ( Silva et al., 2002b) or at least two out of three clusters of immunodominant antigens (17, 29–32 and 35–37 kDa) of N. caninum, similarly to previous findings in several animal species, including cattle, sheep and goats ( Bjerkas et al., 1994, Schares et al., 1998 and Naguleswaran et al., 2004). Statistical analysis was performed using the GraphPad Prism 4.0 software (Graphpad Sofware Inc., San Diego, USA). Seropositivity percentages were compared by the Dinaciclib concentration Chi-square (χ2) test

or Fisher exact selleck chemicals test, when appropriate. The agreement between IFAT and ELISA for the detection of antibodies to T. gondii and N. caninum was analyzed by calculating the proportion of observed agreement (Po), the proportion of positive (Ppos) and negative (Pneg) agreeement, and the Kappa (κ) coefficient as previously reported ( Lasri et al., 2004 and Silva et al., 2007). Values of P < 0.05 were considered statistically significant. The distribution of IgG antibody titers or levels anti-T. gondii and anti-N. caninum determined by IFAT and ELISA are demonstrated in Table 1. From 155 analyzed serum samples, 72 (46.5%) were reagent for T. gondii (cutoff titer ≥ 64), with 80% of samples presenting titers between 512 and 2048 and the most frequent titer was 512 (30.5%). For N. caninum, 73 (47.1%) samples were reagent (cutoff titer ≥ 50)

with 78% of samples showing titers between 50 and 200, and the most frequent titer was 50 (36.9%). Seroreactivity by ELISA showed 75% of samples with higher EI values, between 2.0 and 3.0 for T. gondii (cutoff EI ≥ 1.3) and 54% presenting lower EI values, between 1.3 and 2.0 for N. caninum (cutoff EI ≥ 1.3). The comparison between IFAT and ELISA results for the detection of IgG antibodies to T. gondii ( Table 2) showed 84% of total agreement, with 85% and 82% of positive and negative agreement, respectively, resulting in a substantial Kappa coefficient (κ = 0.69). For N. caninum science ( Table 3), it was observed a lower total agreement (73%), with 63% and 78% of positive and negative agreement, respectively, resulting in a moderate Kappa coefficient (κ = 0.45). For T. gondii serology, 25 (16.1%) samples showed discordant results (ELISA+/IFAT−) and 22 (14.2%) were positive in immunoblot, by recognizing predominantly the p30 antigen from T. gondii. For N. caninum, 42 (27.1%) presented discordant results in both tests, with 37 (23.9%) samples showing ELISA−/IFAT+, and all of these samples were negative in immunoblot. Representative immunoblots for T. gondii and N.