Plug-and-play adaptive optics for commercial laser scanning fluorescence microscopes based on an adaptive lens.

In this Letter, we present a solution for simple implementation of adaptive optics in any existing laser scanning fluorescence microscope. Adaptive optics are implemented by the introduction of a multiactuator adaptive lens between the microscope body and the objective lens. Correction is performed with a sensorless method by optimizing the quality of the images presented on screen by the microscope software. We present the results acquired on both a commercial linear excitation confocal microscope and a custom-made multiphoton excitation microscope.

Modal-based phase retrieval using Gaussian radial basis functions.

In this paper, we propose the use of Gaussian radial basis functions (GRBFs) to model the generalized pupil function for phase retrieval. The selection of the GRBF hyper-parameters is analyzed to achieve an increased accuracy of approximation. The performance of the GRBF-based method is compared in a simulation study with another modal-based approach considering extended Nijboer-Zernike (ENZ) polynomials. The almost local character of the GRBFs makes them a much more flexible basis with respect to the pupil geometry. It has been shown that for aberrations containing higher spatial frequencies, the GRBFs outperform ENZ polynomials significantly, even on a circular pupil. Moreover, the flexibility has been demonstrated by considering the phase retrieval problem on an annular pupil.

Nonlinear spline wavefront reconstruction through moment-based Shack-Hartmann sensor measurements.

We propose a spline-based aberration reconstruction method through moment measurements (SABRE-M). The method uses first and second moment information from the focal spots of the SH sensor to reconstruct the wavefront with bivariate simplex B-spline basis functions. The proposed method, since it provides higher order local wavefront estimates with quadratic and cubic basis functions can provide the same accuracy for SH arrays with a reduced number of subapertures and, correspondingly, larger lenses which can be beneficial for application in low light conditions. In numerical experiments the performance of SABRE-M is compared to that of the first moment method SABRE for aberrations of different spatial orders and for different sizes of the SH array. The results show that SABRE-M is superior to SABRE, in particular for the higher order aberrations and that SABRE-M can give equal performance as SABRE on a SH grid of halved sampling.

High speed wavefront sensorless aberration correction in digital micromirror based confocal microscopy.

The quality of fluorescence microscopy images is often impaired by the presence of sample induced optical aberrations. Adaptive optical elements such as deformable mirrors or spatial light modulators can be used to correct aberrations. However, previously reported techniques either require special sample preparation, or time consuming optimization procedures for the correction of static aberrations. This paper reports a technique for optical sectioning fluorescence microscopy capable of correcting dynamic aberrations in any fluorescent sample during the acquisition. This is achieved by implementing adaptive optics in a non conventional confocal microscopy setup, with multiple programmable confocal apertures, in which out of focus light can be separately detected, and used to optimize the correction performance with a sampling frequency an order of magnitude faster than the imaging rate of the system. The paper reports results comparing the correction performances to traditional image optimization algorithms, and demonstrates how the system can compensate for dynamic changes in the aberrations, such as those introduced during a focal stack acquisition though a thick sample.

Preoperative arterial embolization facilitates multivisceral transplantation for portomesenteric thrombosis.

Multivisceral transplantation (MvTx) for diffuse venous portomesenteric thrombosis is a surgically and anesthesiologically challenging procedure, partly because of the risk of massive bleeding during visceral exenteration. Preoperative visceral artery embolization might reduce this risk. In three consecutive MvTx, the celiac trunk (CT) and superior mesenteric artery (SMA) were embolized immediately pretransplant. We analyzed demographics, serum D-lactate, pH, base excess, hemoglobin, blood pressure, transfused packed cell (PC) units, intervention time and outcome. Results are reported as median (range). All recipients were male (43, 22, 47 years old). Portomesenteric thrombosis followed antiphospholipid syndrome, neuroendocrine tumor and liver cirrhosis. A peritransplant D-lactate peak of 6.1 (5.1-7.6) mmol/L, lowest pH of 7.24 (7.18-7.36) and lowest base excess level of -9.5 (-7.6 to -11.5) were observed. Values normalized within 3 h posttransplant. Embolization and exenteration times were 80 (70-90) min and 140 (130-165) min, respectively, during which blood pressure remained stable, lowest hemoglobin was 6.1 (6.1-7.6) g/dL and three (2-4) PC were administered. All procedures were uneventful. Follow-up was 7 (4-9) months. The first patient died 4 months post-MvTx after an intracranial bleeding; the other patients are doing well. Our experience suggests that preoperative embolization of CT and SMA facilitates native organ resection in MvTx.

Snapshot coherence-gated direct wavefront sensing for multi-photon microscopy.

Deep imaging in turbid media such as biological tissue is challenging due to scattering and optical aberrations. Adaptive optics has the potential to compensate the tissue aberrations. We present a wavefront sensing scheme for multi-photon scanning microscopes using the pulsed, near-infrared light reflected back from the sample utilising coherence gating and a confocal pinhole to isolate the light from a layer of interest. By interfering the back-reflected light with a tilted reference beam, we create a fringe pattern with a known spatial carrier frequency in an image of the back-aperture plane of the microscope objective. The wavefront aberrations distort this fringe pattern and thereby imprint themselves at the carrier frequency, which allows us to separate the aberrations in the Fourier domain from low spatial frequency noise. A Fourier analysis of the modulated fringes combined with a virtual Shack-Hartmann sensor for smoothing yields a modal representation of the wavefront suitable for correction. We show results with this method correcting both DM-induced and sample-induced aberrations in rat tail collagen fibres as well as a Hoechst-stained MCF-7 spheroid of cancer cells.

Predictive control of thermally induced wavefront aberrations.

In this paper we experimentally demonstrate the proof of concept for predictive control of thermally induced wavefront aberrations in optical systems. On the basis of the model of thermally induced wavefront aberrations and using only past wavefront measurements, the proposed adaptive optics controller is able to predict and to compensate the future aberrations. Furthermore, the proposed controller is able to correct wavefront aberrations even when some parameters of the prediction model are unknown. The proposed control strategy can be used in high power optical systems, such as optical lithography machines, where the predictive correction of thermally induced wavefront aberrations is a crucial issue.

High precision wavelength estimation method for integrated optics.

A novel and simple approach to optical wavelength measurement is presented in this paper. The working principle is demonstrated using a tunable waveguide micro ring resonator and single photodiode. The initial calibration is done with a set of known wavelengths and resonator tunings. The combined spectral sensitivity function of the resonator and photodiode at each tuning voltage was modeled by a neural network. For determining the unknown wavelengths, the resonator was tuned with a set of heating voltages and the corresponding photodiode signals were collected. The unknown wavelength was estimated, based on the collected photodiode signals, the calibrated neural networks, and an optimization algorithm. The wavelength estimate method provides a high spectral precision of about 8 pm (5 · 10(-6) at 1550 nm) in the wavelength range between 1549 nm to 1553 nm. A higher precision of 5 pm (3 · 10(-6)) is achieved in the range between 1550.3 nm to 1550.8 nm, which is a factor of five improved compared to a simple lookup of data. The importance of our approach is that it strongly simplifies the optical system and enables optical integration. The approach is also of general importance, because it may be applicable to all wavelength monitoring devices which show an adjustable wavelength response.

Iterative linear focal-plane wavefront correction.

We propose an efficient approximation to the nonlinear phase diversity (PD) method for wavefront reconstruction and correction from intensity measurements with potential of being used in real-time applications. The new iterative linear phase diversity (ILPD) method assumes that the residual phase aberration is small and makes use of a first-order Taylor expansion of the point spread function (PSF), which allows for arbitrary (large) diversities in order to optimize the phase retrieval. For static disturbances, at each step, the residual phase aberration is estimated based on one defocused image by solving a linear least squares problem, and compensated for with a deformable mirror. Due to the fact that the linear approximation does not have to be updated with each correction step, the computational complexity of the method is reduced to that of a matrix-vector multiplication. The convergence of the ILPD correction steps has been investigated and numerically verified. The comparative study that we make demonstrates the improved performance in computational time with no decrease in accuracy with respect to existing methods that also linearize the PSF.

An innovative and efficient method to control the shape of push-pull membrane deformable mirror.

We carry out performance characterisation of a commercial push and pull deformable mirror with 48 actuators (Adaptica Srl). We present a detailed description of the system as well as a statistical approach on the identification of the mirror influence function. A new efficient control algorithm to induce the desired wavefront shape is also developed and comparison with other control algorithms present in literature has been made to prove the efficiency of the new approach.

Model-based aberration correction in a closed-loop wavefront-sensor-less adaptive optics system.

In many scientific and medical applications, such as laser systems and microscopes, wavefront-sensor-less (WFSless) adaptive optics (AO) systems are used to improve the laser beam quality or the image resolution by correcting the wavefront aberration in the optical path. The lack of direct wavefront measurement in WFSless AO systems imposes a challenge to achieve efficient aberration correction. This paper presents an aberration correction approach for WFSlss AO systems based on the model of the WFSless AO system and a small number of intensity measurements, where the model is identified from the input-output data of the WFSless AO system by black-box identification. This approach is validated in an experimental setup with 20 static aberrations having Kolmogorov spatial distributions. By correcting N=9 Zernike modes (N is the number of aberration modes), an intensity improvement from 49% of the maximum value to 89% has been achieved in average based on N+5=14 intensity measurements. With the worst initial intensity, an improvement from 17% of the maximum value to 86% has been achieved based on N+4=13 intensity measurements.

Therapeutic window for melanoma treatment provided by selective effects of the proteasome on Bcl-2 proteins.

Melanoma cells depend on sustained proteasomal function for survival. However, bortezomib, the first proteasome inhibitor in clinical use, is not sufficient to improve the poor prognosis of metastatic melanoma patients. Since the proteasome is also expressed in all normal cell compartments, it is unclear how to enhance the efficacy of bortezomib without exacerbating secondary toxicities. Here, we present pharmacological and genetic analyses of mechanisms of resistance to proteasome inhibition. We focused on Bcl-2, Bcl-x(L) and Mcl-1 as main antiapoptotic factors associated with melanoma progression. Despite an efficient blockage of the proteasome, bortezomib could not counteract the intrinsically high levels of Bcl-2 and Bcl-x(L) in melanoma cells. Moreover, Mcl-1 was only downregulated at late time points after treatment. Based on these results, a combination treatment including (-)-gossypol, an inhibitor of Mcl-1/Bcl-2/Bcl-x(L), was designed and proven effective in vivo. Using a specific RNA interference approach, the survival of bortezomib-treated melanoma cells was found to rely primarily on Mcl-1, and to a lesser extent on Bcl-x(L) (but not on Bcl-2). Importantly, neither Mcl-1 nor Bcl-x(L) inactivation affected the viability of normal melanocytes. This hierarchical requirement of Bcl-2 family members for the maintenance of normal and malignant cells offers a therapeutic window to overcome melanoma chemoresistance in a tumor cell-selective manner.

Abdominal vagus nerve stimulation as a new therapeutic approach to prevent postoperative ileus.

BACKGROUND: Electrical stimulation of the cervical vagus nerve (VNS) prevents postoperative ileus (POI) in mice. As this approach requires an additional cervical procedure, we explored the possibility of peroperative abdominal VNS in mice and human. METHODS: The effect of cervical and abdominal VNS was studied in a murine model of POI and lipopolysaccharide (LPS)-induced sepsis. Postoperative ileus was quantified by assessment of intestinal transit of fluorescent dextran expressed as geometric center (GC). Next, the effect of cervical and abdominal VNS on heart rate was determined in eight Landrace pigs to select the optimal electrode for VNS in human. Finally, the effect of sham or abdominal VNS on LPS-induced cytokine production of whole blood was studied in patients undergoing colorectal surgery. KEY RESULTS: Similar to cervical VNS, abdominal VNS significantly decreased LPS-induced serum tumor necrosis factor-α (TNFα) levels (abdominal VNS: 366±33 pg/mL vs sham: 822±105 pg/mL; P<.01). In line, in a murine model of POI, abdominal VNS significantly improved intestinal transit (GC: sham 5.1±0.2 vs abdominal VNS: 7.8±0.6; P<.01) and reduced intestinal inflammation (abdominal VNS: 35±7 vs sham: 80±8 myeloperoxidase positive cells/field; P<.05). In pigs, heart rate was reduced by cervical VNS but not by abdominal VNS. In humans, abdominal VNS significantly reduced LPS-induced IL8 and IL6 production by whole blood. CONCLUSIONS & INFERENCES: Abdominal VNS is feasible and safe in humans and has anti-inflammatory properties. As abdominal VNS improves POI similar to cervical VNS in mice, our data indicate that peroperative abdominal VNS may represent a novel approach to shorten POI in man.

Characterization of protein phosphokinase activities in horse thyroid nuclei.

The distribution of protein phosphokinase (EC 2.7.1.37) activities has been established in horse thyroid nuclei. The presence of several enzyme activities has been demonstrated, two of which are clearly distinct. The first one acts on histone as substrate and is activated by cyclic AMP. Physico-chemical properties of this nuclear cyclic AMP-dependent histone kinase and of the cytosol histone kinase are different, demonstrating the absence of a contamination from the cytosol. The second enzyme acts on casein as substrate and is not stimulated by cyclic AMP POR CYCLIC GMP. The findings are consistent with the observation of thyrotropin stimulation of histone phosphorylation in thyroid nuclei.

Regulation of lipogenesis in adipocytes. Independent effects of thyroid hormones, cyclic AMP and insulin on the uptake of deoxy-D-glucose.

Thyroidectomy is known to enhance fat cell phosphodiesterase activity; as a result, the response to lipolytic hormones is markedly reduced. Thyroidectomy also stimulates overall lipogenesis and the uptake of glucose: the present experiments investigated whether there was a correlation between cyclic AMP and glucose uptake. The parameter measured was the transport and phosphorylation (uptake) of deoxy-D-glucose in the presence of two modifiers of the cyclic AMP pool: phosphodiesterase inhibitors and the analogue, dibutyryl cyclic AMP. The inhibition by methylxanthines and dibutyryl cyclic AMP of deoxy-D-glucose uptake observed, was the same in fat cells from normal and thyroidectomized rats: the latter nonetheless still maintained their enhanced glucose uptake. It was therefore concluded that thyroid hormones and cyclic AMP control this step by different, separate pathways. Insulin, well known for its lipogenic effect, enhanced deoxy-D-glucose uptake in fat cells from both normal and thyroidectomized rats to the same extent (about 40%). An additive effect of thyroidectomy and insulin on glucose uptake was thus demonstrated. These results imply that glucose uptake in the adipocyte is controlled by at least three factors: thyroid hormones, cyclic AMP and insulin, each of which can act independently. Maximum glucose uptake is achieved in the presence of a combination of low concentrations of cyclic AMP, of insulin, and in the absence of thyroid hormones.

The role of electrode size on the incidence of spreading depression and on cortical cerebral blood flow as measured by H2 clearance.

Cerebral blood flow was measured by the H2 clearance method 30 and 60 min after the implantation of 300, 250, 125, or 50 microns diameter platinum-iridium electrodes 2 mm deep into the right parietal cortex of normothermic, normocarbic halothane-anesthetized rats. Another group of animals had 50 microns electrodes inserted 1 mm. In all animals, the presence or absence of a wave of spreading depression (SD) was noted at the time of implantation, with recordings made with glass micropipettes. H2 flow values were compared with those measured in gray matter from the same anatomical region (but from different rats), using [3H]nicotine. The incidence of SD ranged from 60% following insertion of 300 microns electrodes to 0% with 50 microns electrodes. H2 clearance flows also varied with electrode size, from 77 +/- 21 ml 100 g-1 min-1 (mean +/- standard deviation) with 300 microns electrodes to 110 +/- 31 and 111 +/- 16 ml 100 g-1 min-1 with 125 and 50 microns electrodes, respectively (insertion depth of 2 mm). A CBF value of 155 +/- 60 ml 100 g-1 min-1 was obtained with 50 micron electrodes inserted only 1 mm. Cortical gray matter blood flow measured with [3H]nicotine was 154 +/- 35 ml 100 g-1 min-1. When the role of SD in subsequent flow measurements was examined, there was a gradual increase in CBF between 30 and 60 min after electrode insertion in those animals with SD, while no such change was seen in rats without SD.(ABSTRACT TRUNCATED AT 250 WORDS)

Cyclic AMP and lipogenesis in fat cells from thyroidectomized rats.

Thyroid hormones regulate lipid metabolism by affecting lipogenesis as well as lipolysis. The present paper discusses the way thyroidectomy induced an enhancement in lipogenesis in rat fat cells. The doubling in the conversion of glucose to CO2 and fatty acids seen after thyroidectomy was found to be due to a modification in the actual pathway of glucose metabolism: there was a preferential stimulation of the conversion of glucose to CO2 by the pentose cycle (utilisation of [1-14C]glucose) while the production of fatty acids and glyceride-glycerol proceeded, respectively, much more, or only slightly more, via the pathway of [6-14C]glucose metabolism. Studies employing the phosphodiesterase inhibitor MIX, or the cyclic AMP analogue, DBcAMP showed that the lipogenic process depends on cyclic AMP. As the stimulatory effect of thyroidectomy was not abolished, however, lipogenesis must be under the independent control of both cyclic AMP and absence of thyroid hormones. Insulin, a further mediator of lipogenesis was found to further enhance the already preexisting high conversion of glucose to CO2 in fat cells from thyroidectomized rats. It is concluded that at least three factors modify lipogenesis: thyroidectomy, cyclic AMP and insulin; each achieving its effect in an independent manner.

The influence of different concentrations of volatile anesthetics on the threshold for cortical spreading depression in rats.

We studied the influence of halothane and isoflurane on the electrical threshold for triggering spreading depression (SD) in rats. While the N-methyl-D-aspartate (NMDA) antagonist ketamine dramatically increased the triggering threshold (and blocked propagation), neither volatile agent had any dose-related effect. High doses of both agents slightly slowed SD propagation rate. These results indicate that volatile agents in typical anesthetic doses can be used in studies of SD in rats, and also suggest that these agents have little effect on NMDA receptor-mediated neurotransmission.

Effects of intra-ischemic blood pressure on outcome from 2-vessel occlusion forebrain ischemia in the rat.

Halothane anesthetized Sprague-Dawley rats underwent 10 min of bilateral carotid artery occlusion with mean arterial pressure (MAP) held at 30, 50 or 60 mmHg. Sham rats did not undergo ischemia. A 7-day recovery interval was allowed. Intra-ischemic electroencephalographic (EEG) changes, behavioral function (Days 5-7), and histologic injury (Day 7) were evaluated. Under similar conditions, cerebral blood flow was determined after 10 min ischemia by the [3H]nicotine indicator fractionation technique. EEG isoelectricity was observed in 11 of 11, 5 of 10, and 2 of 11 rats in the 30 mmHg, 50 mmHg, and 60 mmHg groups respectively. Neither passive avoidance cross-over latencies nor general motor scores were affected by intra-ischemic MAP and no differences from sham performance were observed. The per cent of CA1 neurons counted as dead (left and right hemispheres combined) was significantly affected by intra-ischemic MAP (72, 46 and 28% in the 30 mmHg, 50 mmHg, and 60 mmHg groups, respectively; P less than 0.001). A greater than 50% CA1 neuronal mortality rate was present only in those rats exhibiting EEG isoelectricity. However, the number of rats demonstrating greater than a 25% interhemispheric difference in CA1 neuronal loss was greatest in the 50 mmHg group (P less than 0.02). Hippocampal blood flow decreased in association with severity of hypotension (8 +/- 1, 35 +/- 8, and 48 +/- 2 ml/100 g/min (mean +/- S.E.M.) for 30, 50, and 60 mmHg, respectively; P less than 0.01). Again, however, the greatest variability in blood flow was observed at MAP = 50 mmHg.(ABSTRACT TRUNCATED AT 250 WORDS)

Aquatic ape theory, speech origins, and brain differences with apes and monkeys.

Humans and apes show clear differences in brain anatomy. In the human cerebral cortex, for instance, the areas that control the fine movements of the hand, the areas that control the breathing and speech musculature, and the association areas have strongly expanded. It will be argued that these differences are best explained by the aquatic ape theory of human evolution (AAT) and originated in our semi-aquatic past, notably in the adaptations necessary for diving and shellfish collection at sea coasts.