EPSCs during and after train stimulation.

<p><b>A</b>, Upper trace: original recording with a 5 s train of stimuli (10 Hz,). Lower traces are details of Pre, 10 Hz Stimulation, Post and Recovery conditions of same recording. <b>B</b>, Group data of EPSCs. EPSC are increased during stimulation until at least 5 s post stimulation (* <i>p</i><0.05 compared to Control 5 s prior). Oxytocin antagonist OTA reduces the EPSC frequency during and 5 s post stimulation (#, 5 s 10 Hz stim Control vs. OTA, <i>p</i> = 0.015; 5 s post Control vs OTA, <i>p</i> = 0.012).</p

PVN monosynaptic glutamatergic projections to CVNs.

<p><b>A</b>, Schematic of approach, left. Right, postsynaptic responses were completely blocked by antagonists for NMDA (AP-5; 50 µM) and AMPA/kainate receptors (CNQX; 50 µM). <b>B</b>, Paired-pulse facilitation (10 Hz) of glutamatergic neurotransmission from PVN to CVNs (first trace) abolished by oxytocin antagonist OTA (10 µM; middle trace) and partly restored upon wash out (right trace) <b>C</b>, Average data of OTA abolished paired-pulse facilitation (n = 7). * <i>p</i> = 0.021. <b>D</b>, Five consecutive stimulations (10 Hz;) with increasing amplitudes (black trace). OTA reduced the amplitude (red trace). Traces are normalized to amplitude of first response. <b>E</b>, Average data of the five stimulation paradigm (* <i>p</i> = 0.014)).</p

Imaging oxytocin and oxytocin receptor activation.

<p>(<b>A</b>) Co-localization of ChR2-EYFP and oxytocin in brainstem fibers within the DMNV is shown (scale bar represents 28 microns). 55.7+3.7% of ChR2-EYFP PVN fibers in the DMNV are positive for oxytocin. (<b>B</b>) Both the oxytocin and vasopressin dose-response relationships of sniffer CHO cells expressing an oxytocin receptor and a R-GECO Ca²⁺ indicator were characterized. These oxytocin receptor expressing CHO cells are considerably more sensitive and responsive to oxytocin than vasopressin with a half maximal response (EC-50) for oxytocin of 1.5 nM, and an EC-50 for vasopressin of 12.1 nM. Responses to oxytocin were considerably more robust than that for vasopressin; at the concentration (1 µM) at which oxytocin maximally activates these cells, vasopressin evoked a blunted response of only 24+5% of the oxytocin response. Sniffer CHO cells deposited on slices with ChR2 PVN fibers (green) in dorsal motor nucleus of the vagus (DMNV) (<b>C</b>) detect optogenetic oxytocin receptor activation in brainstem DMNV tissue in close apposition to PVN fibers, (3-D reconstruction top down view (left) and side view (right)). <b>D</b>, Repeated stimulations (5 min apart) of ChR2 axons in DMNV increased Ca²⁺. Representative traces of one sniffer CHO cell, top, and averages of repeated stimulations in 9 CHO cells, * p<0.0001, bottom. <b>E</b>, Oxytocin antagonist OTA blocks Ca²⁺ response, representative trace of one cell, top, and average control increase in 7 cells (; * p<0.0001), bottom.</p

Expression profiles of septins among developmental stages of <i>Schistosoma mansoni</i>.

<p>Expression analysis by qPCR of each of four septins in seven developmental stages of the blood fluke. Panels A to D represent expression levels of <i>SmSept5</i>, <i>SmSept10</i>, <i>SmSept7.1</i> and <i>SmSept7.2</i>, respectively. Absolute quantification was used to evaluate the expression levels and is presented as copy number per ng of total RNA. Copy number of each transcript was estimated by interpolation of the sample signal from a standard curve for each gene; bars represent standard deviation of the mean of three technical replicates. Mir, miracidia; Spor, sporocyst; Cerc, cercariae; 2 d S, 2 days old schistosomula; 14 d S, 14 day old schistosomula; Fem, adult female; Mal, adult male. (A biological replicate revealed the same trend among stages; <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002602#pntd.0002602.s002" target="_blank">Figure S2</a>.)</p

Septin fibers co-localize with actin fibers at the surface of the schistosomulum.

<p>Panel A: Cross-section at the surface of a two day old schistosomulum showing DAPI stained nuclei. B: F-actin structure stained with phalloidin. C: Septin labeled with anti-<i>Sm</i>SEPT5. D: Merged channels. Scale bars, 20 µm.</p

Septin in germ cells of miracidia and sporocysts of schistosomes.

<p>Confocal optical sections of a miracidium (panel A) and a two day old sporocyst (B); nuclei stained with DAPI (blue) and actin filaments stained with phalloidin conjugated with Alexa Fluor 568 (green). Probing with anti-<i>Sm</i>SEPT10 immunoglobulin (red) revealed the prevalence of septin in germ cells of both miracidia and sporocysts. The insets of A and B highlight germ cell rich regions in these developmental stages. Scale bar, 20 µm.</p

Protonephridial ducts of the schistosome cercaria are septin rich structures.

<p>Optical section of cercariae labeled with phalloidin (green) and anti-<i>Sm</i>SEPT10 (red). The arrows indicate the flame cells at anterior termini of protonephridial canals. Scale bar, 20 µm.</p

Evolutionary relationship among septins of humans and <i>Schistosoma mansoni</i>.

<p>Phylogenetic tree (based on Bayesian inference) generated from a multiple alignment of the conserved GTPase domains of septins from <i>S. mansoni</i> and two other informative protostomes and three deuterostomes. The numbers on the tree nodes are posterior probabilities calculated by MrBayes. Branches with the four discrete groups of septins are enclosed by the dotted lines. Species are identified by the small circles of different shapes and colors as indicated in the lower panel.</p

Superficial structures of miracidia and sporocysts of <i>Schistosoma mansoni</i>.

<p>Panels A and C represent a superficial cross section of miracidia and B and D depict a superficial layer of two-day-old sporocysts. The green panels A and B show F-actin stained with phalloidin conjugated with Alexa Fluor 568. The red panels C and D reveal septin structures labeled with anti-<i>Sm</i>SEPT10 immunoglobulin. The upper left inset in panel C highlights an epidermal plate of a miracidium, rich in septins, and in panel A the same region revealed a muscular structure stained with phalloidin. Scale bar, 20 µm.</p

Septins are ubiquitous in tissues of the schistosomulum.

<p>Cross section at an inner intersection of a schistosomulum cultured for 14 days. Panel A: Nuclei stained with DAPI. B: F-actin structure stained with phalloidin. C: Septin labeled with anti-<i>Sm</i>SEPT5. D: Merged channels. Scale bar, 20 µm.</p