K-edge imaging with the XPAD3 hybrid pixel detector, direct comparison of CdTe and Si sensors.

We investigate the improvement from the use of high-Z CdTe sensors for pre-clinical K-edge imaging with the hybrid pixel detectors XPAD3. We compare XPAD3 chips bump bonded to Si or CdTe sensors in identical experimental conditions. Image performance for narrow energy bin acquisitions and contrast-to-noise ratios of K-edge images are presented and compared. CdTe sensors achieve signal-to-noise ratios at least three times higher than Si sensors within narrow energy bins, thanks to their much higher detection efficiency. Nevertheless Si sensors provide better contrast-to-noise ratios in K-edge imaging when working at equivalent counting statistics, due to their better estimation of the attenuation coefficient of the contrast agent. Results are compared to simulated data in the case of the XPAD3/Si detector. Good agreement is observed when including charge sharing between pixels, which have a strong impact on contrast-to-noise ratios in K-edge images.

Fibronectin expression in glioblastomas promotes cell cohesion, collective invasion of basement membrane in vitro and orthotopic tumor growth in mice.

Glioblastoma multiforme (GBM) are highly invasive and angiogenic malignancies with a median survival time from diagnosis of <15 months. Previous work has revealed robust overexpression of fibronectin (FN) mRNA in GBM, although immunohistochemical staining of FN in these tumors is typically associated with the angiogenic vasculature. Here we sought to examine the expression of tumor cell FN and address its possible involvement in the invasive phenotype of GBM. We found that FN was expressed and assembled into fibrillar arrays in human tumors and in established GBM lines. Cultured cells spontaneously formed dense cellular networks and spheroid-like domes. Depletion of FN by targeted-short hairpin RNA expression disrupted matrix assembly and multicellular network organization by exerting profound effects on cell adhesion and motility. Although FN depletion enhanced persistent directional migration of single cells, it compromised collective invasion of spheroids through a laminin-rich matrix and sensitized cells to ionizing radiation. In orthotopic grafts, FN depletion significantly reduced tumor growth and angiogenesis. Together our results show that FN produced by the tumor cells has a role in GBM pathophysiology and they provide insights into the implications that targeting FN interactions may have for combating this dreaded disease.

Combined two-photon laser-scanning microscopy and spectral microCT X-ray imaging to characterize the cellular signature and evolution of microstroke foci.

Occlusive brain ischemia and micro-strokes are the most frequent brain pathologies, particularly in older patients and a major cause of dementia. Currently, we are missing appropriate methodology to study micro-strokes in experimental animals. In vivo two-photon laser-scanning microscopy (2P-LSM) and transgenic mouse models expressing cell type specific reporters have been used to examine ischemia-related insults, e.g. perturbations of neuronal process morphology and local blood flow in the MCAO - middle cerebral artery occlusion-model. Glia and pericytes can be visualized by selective fluorescent protein expression, e.g. astrocytes by their cyan-fluorescent ECFP, pericytes by red-fluorescent tdtomato and microglia by green fluorescent EGFP expression. In these mice, the breakdown of the blood brain barrier and the immediate as well as long-term cellular responses can be monitored. A new prototype of microCT incorporating a fast X-ray XPAD3 camera has been recently set up to allow cerebral angiography at high sampling rate. Preliminary data indicate that it is useful to monitor blood perfusion disturbance (i.e. lateralization) in the brain of tumor-bearing mice following retro-orbital injection of iodinated contrast agent. We expect this technology to be adequate to assess in real time the impact of acute stroke models on brain blood perfusion. By localizing perfusion anomalies, we will evaluate the extent of non-perfused areas and correlate these observations with subsequent behavioral deficits, and with local changes in myelin content in white matter tracks. The spectral properties of the XPAD3 detector moreover allow for the simultaneous identification and localization of several contrast agents opening the way to whole body multicolor imaging of vessels and inflammatory cells in the context of microstrokes.

Action potential propagation in dendrites of rat mitral cells in vivo.

Odors evoke beta-gamma frequency field potential oscillations in the olfactory systems of awake and anesthetized vertebrates. In the rat olfactory bulb, these oscillations reflect the synchronous discharges of mitral cells that result from both their intrinsic membrane properties and their dendrodendritic interactions with local inhibitory interneurons. Activation of dendrodendritic synapses is purportedly involved in odor memory and odor contrast enhancement. Here we investigate in vivo to what extent action potentials propagate to remote dendrodendritic sites in the entire dendritic tree and if this propagation is changed during discharges at 40 Hz. By combining intracellular recording and two-photon microscopy imaging of intracellular calcium ([Ca2+]i), we show that in remote branches of the apical tuft and basal dendrites, transient Ca2+ changes are triggered by single sodium action potentials. Neither the amplitude of these Ca2+ transients nor that of action potentials obtained from intradendritic recordings showed a significant attenuation as a function of the distance from the soma. Calcium channel density seemed homogeneous; however, propagating action potentials occasionally failed to trigger a Ca2+ transient at a site closer to the soma whereas it did farther. This suggests that measurements of calcium transients underestimate the occurrence of sodium action potentials. During 40 Hz bursts of action potentials, [Ca2+]i increases with the number of action potentials in all dendritic compartments. These results suggest that the presence of release sites in dendrites is accompanied by an "axonal-like behavior" of the entire dendritic tree of mitral cells, including their most distal dendritic branches.

Effect of bicuculline on thalamic activity: a direct blockade of IAHP in reticularis neurons.

The thalamic reticular nucleus (RTN) is the major source of inhibitory contacts in the thalamus and thus plays an important role in regulating the excitability of the thalamocortical network. Inhibition occurs through GABAergic synapses on relay cells as well as through GABAergic synapses between reticularis neurons themselves. Here we report that the role and mechanisms of this inhibition, which frequently have been studied using N-methyl derivatives of the gamma-aminobutyric acid-A (GABAA) receptor antagonist bicuculline, should be revisited. Using the whole cell patch-clamp technique in thalamic slices from young rats, we observed an enhancement by bicuculline methiodide, methobromide, and methochloride (collectively referred to as bicuculline-M; 5-60 microM) of the low-threshold calcium spike burst in RTN neurons that persisted in the presence of tetrodotoxin (1 microM) and was not reproduced in picrotoxin (100-300 microM). The effect did not involve activation of any GABA receptor subtype. Voltage-clamp recordings showed that bicuculline-M blocked the current underlying the low-threshold spike burst afterhyperpolarization (AHP), an effect that was mimicked by apamin (100 nM). Recordings from nucleated patches extracted from reticularis neurons demonstrated that this effect was not mediated by modulation of the release of an unidentified neurotransmitter but that bicuculline-M directly blocks small conductance (SK) channels. The AHP-blocking effect also was observed in other brain regions, demonstrating that although bicuculline-M is a potent GABAA receptor antagonist, it is of limited value in assessing GABAergic network interactions, which should be studied using picrotoxin or bicuculline-free base. However, bicuculline-M may provide a useful tool for developing nonpeptide antagonists of SK channels.