Targeting the αVß3/NgR2 pathway in neuroendocrine prostate cancer.

Highly aggressive, metastatic, neuroendocrine prostate cancer, which typically develops from prostate cancer cells acquiring resistance to androgen deprivation therapy, is associated with limited treatment options and hence poor prognosis. We have previously demonstrated that the αVß3 integrin is over-expressed in neuroendocrine prostate cancer. We now show that LM609, a monoclonal antibody that specifically targets the human αVß3 integrin, hinders the growth of neuroendocrine prostate cancer patient-derived xenografts in vivo. Our group has recently identified a novel αVß3 integrin binding partner, NgR2, responsible for regulating the expression of neuroendocrine markers and for inducing neuroendocrine differentiation in prostate cancer cells. Through in vitro functional assays, we here demonstrate that NgR2 is crucial in promoting cell adhesion to αVß3 ligands. Moreover, we describe for the first time co-fractionation of αVß3 integrin and NgR2 in small extracellular vesicles derived from metastatic prostate cancer patients' plasma. These prostate cancer patient-derived small extracellular vesicles have a functional impact on human monocytes, increasing their adhesion to fibronectin. The monocytes incubated with small extracellular vesicles do not show an associated change in conventional polarization marker expression and appear to be in an early stage that may be defined as "adhesion competent". Overall, these findings allow us to better understand integrin-directed signaling and cell-cell communication during cancer progression. Furthermore, our results pave the way for new diagnostic and therapeutic perspectives for patients affected by neuroendocrine prostate cancer.

Working in fours: generational communication in the emergency department.

BACKGROUND: This study examined the conflicts between different generations working in US emergency departments (ED). We sought to record generational differences involving communication preferences, perceived areas of conflict, work motivations, and attitudes regarding work-life balance. METHODS: We developed a survey to assess the physician perspective on generational conflict in the ED. The survey was distributed to members of the American College of Emergency Physicians, a professional organization comprising emergency medicine physicians in the USA. RESULTS: We received 696 completed responses. Men represented 60% of respondents and the largest proportion of respondents were emergency physicians working in community settings (53%); 11% were residents. Generation representation was smallest for Traditionalist (2%) and largest for Gen X (43%). Seventy percent reported observing conflict due to generational communication with the largest frequency being once a week (26%). In the associated open-ended questions, 247 (33%) provided 316 anecdotal descriptions of observed conflict. Responses clustered into seven themes (ordered by frequency): Work Ethic, Treatment Approach, Technology Application, Entitlement, Professionalism, Work Life/Balance, and Communication Style. Comparing Work Ethic responses, 52-70-year-olds reported that younger providers are less interested in "accomplishing anything" while 26-34-year-olds resented that attitude. Respondents completing the open-ended questions regarding preventing and responding to conflict provided some insight into helpful strategies including actions supportive of clear communication and standardized policies and expectations. Only 5% of respondents reported that they had discussed generational communication in department meetings with the odds of a woman reporting conflict being less than males (p = .01). CONCLUSION: Conflicts in the ED in the USA can be attributed to how an individual views the values of someone from another generation. Understanding the frequency and areas of generational conflict in the ED can help medical leaders find strategies to mitigate negative workplace interactions.

Antimicrobial Susceptibility Among Gram-Negative Isolates in Pediatric Patients in Latin America, Africa-Middle East, and Asia From 2016-2020 Compared to 2011-2015: Results From the ATLAS Surveillance Study.

BACKGROUND: Antimicrobial resistance (AMR) data in the pediatric population are limited, particularly in developing countries. This study assessed the AMR profile and key resistance phenotypes and genotypes for Gram-negative bacteria (GNB) isolates collected as part of the Antimicrobial Testing Leadership and Surveillance program from pediatric patients in Latin America, Africa-Middle East, and Asia in 2016-2020 versus 2011-2015. METHODS: Minimum inhibitory concentrations by broth microdilution methodology were interpreted per the Clinical and Laboratory Standards Institute. European Committee on Antimicrobial Susceptibility Testing breakpoints were used for interpreting colistin activity. ß-lactamase genes were screened by polymerase chain reaction and sequencing. RESULTS: For Acinetobacter baumannii, low susceptibility (<60.0%) was observed for all antimicrobials, except colistin (≥92.9%), across regions and year periods. Ceftazidime-avibactam, amikacin, colistin, and meropenem were mostly active (78.6%-100.0%) against Enterobacter cloacae, Escherichia coli, and Klebsiella pneumoniae. For Pseudomonas aeruginosa, susceptibility to ceftazidime-avibactam, amikacin, and colistin was ≥85.9%. Among resistance phenotypes, carbapenem-resistant (CR, ≥44.8%) and difficult-to-treat resistant (DTR, ≥37.1%) rates were the highest in A. baumannii. A consistent increase in CR and DTR K. pneumoniae was noted across regions over time. Extended-spectrum ß-lactamases (ESBL)-producing K. pneumoniae (32.6%-55.6%) were more frequent than ESBL-producing E. coli (25.3%-37.1%). CTX-M was the dominant ESBL among Enterobacterales. NDM-positive Enterobacterales species and VIM-positive P. aeruginosa were identified across regions. CONCLUSIONS: This study identified high susceptibility to few agents for key GNB in pediatric patients. Continued surveillance of resistance phenotypes and genotypes at regional levels may help to guide appropriate treatment decisions.

Optimizing intrauterine insemination and spontaneous conception in women with unilateral hydrosalpinx or tubal pathology: A systematic review and narrative synthesis.

BACKGROUND: Tubal surgery's role in infertile women with unilateral tubal pathology (e.g. hydrosalpinx, tubal occlusion) who desire spontaneous or intrauterine insemination (IUI) conception where in-vitro fertilisation is infeasible remains unclear. OBJECTIVE: To conduct a systematic review on fertility outcomes in women with unilateral tubal pathology desiring to conceive spontaneously or via IUI and to find guidance to support therapeutic tubal procedures to help these women conceive. SEARCH STRATEGY: Using a protocol registered on PROSPERO (ID CRD42021248720), we searched PubMed, EMBASE, CINAHL, and Cochrane Library from inception until June 2022. Bibliographies were reviewed to identify other relevant articles. DATA COLLECTION AND ANALYSIS: Two authors independently selected and extracted data. Disagreements were resolved by a third author. Studies presenting fertility outcome data in infertile women with unilateral tubal pathologies desiring spontaneous or IUI conception were included. Methodologic quality was assessed using a modified Newcastle Ottawa Scale for observational studies and the Institute of Health Economics Quality Appraisal Checklist for case series. Primary outcomes collated included cumulative pregnancy rate (CPR) and pregnancy rate per cycle (PR/cycle). Secondary outcomes such as ectopic pregnancy, birth outcomes, and pelvic inflammatory disease were collated. These were stratified by the types of unilateral tubal occlusion (UTO) i.e. hydrosalpinx, proximal tubal occlusion (PTO), or distal tubal occlusion (DTO) MAIN RESULTS: Two studies reported spontaneous or IUI pregnancies after treatment of unilateral hydrosalpinx with one reporting a pregnancy rate of 88% within 5.6 months on average. Thirteen studies compared IUI outcomes between women with UTO vs unexplained infertility and bilateral tubal patency (controls). Almost all were retrospective cohort studies and identified UTO by hysterosalpingography. In general, PTOs had no difference in PR/cycle and CPR compared to controls and significantly higher PR/cycle to DTOs. Women with DTOs had minimal incremental CPR benefit with each additional IUI cycle. CONCLUSIONS: Therapeutic salpingectomy or tubal occlusion improve IUI or spontaneous conception in women with hydrosalpinx, although more prospective studies are needed. While significant study heterogeneity hampered assessment of fertility outcomes, overall, infertile women with PTOs had similar IUI pregnancy outcomes to those with bilateral tubal patency while DTOs had inferior PR/cycle. This review highlights significant deficiencies in the evidence guiding management for this group of patients.

Emulation of deep-ultraviolet lithography using rapid-prototyping, electron-beam lithography for silicon photonics design.

We demonstrate a method to emulate the optical performance of silicon photonic devices fabricated using advanced deep-ultraviolet lithography (DUV) processes on a rapid-prototyping electron-beam lithography process. The method is enabled by a computational lithography predictive model generated by processing SEM image data of the DUV lithography process. We experimentally demonstrate the emulation method's accuracy on integrated silicon Bragg grating waveguides and grating-based, add-drop filter devices, two devices that are particularly susceptible to DUV lithography effects. The emulation method allows silicon photonic device and system designers to experimentally observe the effects of DUV lithography on device performance in a low-cost, rapid-prototyping, electron-beam lithography process to enable a first-time-right design flow.

Global Context Networks.

The non-local network (NLNet) presents a pioneering approach for capturing long-range dependencies within an image, via aggregating query-specific global context to each query position. However, through a rigorous empirical analysis, we have found that the global contexts modeled by the non-local network are almost the same for different query positions. In this paper, we take advantage of this finding to create a simplified network based on a query-independent formulation, which maintains the accuracy of NLNet but with significantly less computation. We further replace the one-layer transformation function of the non-local block by a two-layer bottleneck, which further reduces the parameter number considerably. The resulting network element, called the global context (GC) block, effectively models global context in a lightweight manner, allowing it to be applied at multiple layers of a backbone network to form a global context network (GCNet). Experiments show that GCNet generally outperforms NLNet on major benchmarks for various recognition tasks. The code and network configurations are available at https://github.com/xvjiarui/GCNet.

Simvastatin Attenuated Tumor Growth in Different Pancreatic Tumor Animal Models.

Newly diagnosed pancreatic cancer increases year by year, while the prognosis of pancreatic cancer has not been very good. Statin drugs were found to have protective effects against a variety of cancers, but their association with pancreatic cancer remains to be clarified. This study used different pancreatic cancer cell lines and in different animal models to confirm the relationship between simvastatin and pancreatic cancer. Flow cytometry and luciferase-based bioluminescent images were used to investigate the cell cycle and tumor growth changes under simvastatin treatment. Simvastatin decreased the MIA PaCa-2 cells, PANC-1 cells, and BxPC-3 cell viability significantly and may arrest the cell cycle in the G0 phase. During in vivo study, subcutaneously implanted simvastatin pre-treated pancreatic cancer cells and intraperitoneally treated simvastatin continuously demonstrated a slower tumor growth rate and decreased the tumor/body weight ratio significantly. In intravenous implant models, implanted simvastatin-pre-treated BxPC-3 cells and cells treated along with simvastatin significantly decreased the tumor growth curve. Implanting the simvastatin-pre-treated pancreatic cells in the subcutaneous model showed better growth inhibition than the intravenous model. These results suggest simvastatin treatment may relate to different signaling pathways in local growth and metastasis. Pancreatic cancer cells presented different growth patterns in different animal-induced models, which could be important for clinical reference when it comes to the relationship of long-term statin use and pancreatic cancer.

The NOGO receptor NgR2, a novel αVß3 integrin effector, induces neuroendocrine differentiation in prostate cancer.

Androgen deprivation therapies aimed to target prostate cancer (PrCa) are only partially successful given the occurrence of neuroendocrine PrCa (NEPrCa), a highly aggressive and highly metastatic form of PrCa, for which there is no effective therapeutic approach. Our group has demonstrated that while absent in prostate adenocarcinoma, the αVß3 integrin expression is increased during PrCa progression toward NEPrCa. Here, we show a novel pathway activated by αVß3 that promotes NE differentiation (NED). This novel pathway requires the expression of a GPI-linked surface molecule, NgR2, also known as Nogo-66 receptor homolog 1. We show here that NgR2 is upregulated by αVß3, to which it associates; we also show that it promotes NED and anchorage-independent growth, as well as a motile phenotype of PrCa cells. Given our observations that high levels of αVß3 and, as shown here, of NgR2 are detected in human and mouse NEPrCa, our findings appear to be highly relevant to this aggressive and metastatic subtype of PrCa. This study is novel because NgR2 role has only minimally been investigated in cancer and has instead predominantly been analyzed in neurons. These data thus pave new avenues toward a comprehensive mechanistic understanding of integrin-directed signaling during PrCa progression toward a NE phenotype.

The unique rectus extraocular muscles of cetaceans: Homologies and possible functions.

Each rectus extraocular muscle in cetaceans divides into two portions: a massive palpebral belly that inserts into the deep surface of the eyelids and a smaller scleral belly that inserts onto the eyeball. While the cetacean palpebral insertions have long been recognized, their homologies and functions remain unclear. To compare cetacean rectus EOM insertions with the global and orbital rectus EOM insertions of other mammals we dissected orbital contents of 20 odontocete species, 2 mysticete species and 18 non-cetacean species, both aquatic and terrestrial. Four cetacean species were also examined with magnetic resonance imaging (MRI). All four rectus muscles in cetaceans had well-developed palpebral bellies and insertions. Adjacent palpebral bellies showed varying degrees of fusion, from near independence to near complete fusion. Fusion was most complete towards palpebral insertions and less towards origins. A medial moiety of the superior rectus palpebral belly is likely the levator palpebrae superioris. Smaller but still robust scleral insertions were present on all recti, with the medial rectus (MR) being significantly more muscular than the others. All non-cetacean species examined had recti with distinct global and orbital insertions, the latter generally onto Tenon's capsule. Orbital insertions in pygmy hippopotamus and Florida manatee extended into the deep surfaces of the eyelids, hence qualifying as palpebral insertions. Our results suggest that rectus EOMs of mammals generally have both global and orbital insertions, and that palpebral bellies of cetaceans and other species are modified homologs of the orbital insertions. The presence of palpebral insertions in pygmy hippopotamus and absence in other cetartiodactyls suggests an intermediate condition between terrestrial cetartiodactyls and cetaceans. Palpebral insertions in Florida manatee and reports of their presence in some pinnipeds suggest parallel evolution in multiple aquatic lineages. Various functions of cetacean palpebral recti have been proposed, including eyelid dilators, protection during diving and thermogenesis for warming eye and brain. For further insight into their possible functions, we observed eye movements of captive bottlenose dolphins (Tursiops truncatus) at the U.S. National Aquarium. Our observations showed that in addition to rotation of the eyeball the entire surrounding palpebral region also moves during gaze changes. For example during upward gaze the globe not only rotates in supraduction but translates dorsally as well. It appears the rectus palpebral bellies are responsible for flexing the palpebral structures and thus also translating the globe, while the scleral insertions act directly for ocular rotation. Along with frequent non-conjugate eye movements, the oculomotor mechanics and repertoire of cetaceans are thus quite distinctive. Summarily, axial displacement within the orbit is a major 'eye movement' in cetaceans, with protrusion and retraction mediated by well-developed circular muscles and retractor bulbi respectively. Torsional eye movements driven by elaborate oblique EOMs are likewise significant. The roles of rectus EOMs for ocular rotation via their scleral insertions, especially the highly muscular MR, are for typical supra/infraductions and nasal/temporal ductions. The palpebral bellies accentuate these ductions by translating the globe and surrounding structures in the same direction.

Impact of Cardiopulmonary Bypass on Neurogenesis and Cortical Maturation.

OBJECTIVE: Neurodevelopmental delays and frontal lobe cortical dysmaturation are widespread among children with congenital heart disease (CHD). The subventricular zone (SVZ) is the largest pool of neural stem/progenitor cells in the postnatal brain. Our aim is to determine the effects of cardiopulmonary bypass (CPB) on neurogenesis and cortical maturation in piglets whose SVZ development is similar to human infants. METHODS: Three-week-old piglets (n = 29) were randomly assigned to control (no surgery), mild-CPB (34°C full flow for 60 minutes) and severe-CPB groups (25°C circulatory-arrest for 60 minutes). The SVZ and frontal lobe were analyzed with immunohistochemistry 3 days and 4 weeks postoperatively. MRI of the frontal lobe was used to assess cortical development. RESULTS: SVZ neurogenic activity was reduced up to 4 weeks after both mild and severe CPB-induced insults. CPB also induced decreased migration of young neurons to the frontal lobe, demonstrating that CPB impairs postnatal neurogenesis. MRI 4 weeks after CPB displayed a decrease in gyrification index and cortical volume of the frontal lobe. Cortical fractional anisotropy was increased after severe CPB injury, indicating a prolonged deleterious impact of CPB on cortical maturation. Both CPB-induced insults displayed a significant change in densities of three major inhibitory neurons, suggesting excitatory-inhibitory imbalance in the frontal cortex. In addition, different CPB insults altered different subpopulations of inhibitory neurons. INTERPRETATION: Our results provide novel insights into cellular mechanisms contributing to CHD-induced neurological impairments. Further refinement of CPB hardware and techniques is necessary to improve long-term frontal cortical dysmaturation observed in children with CHD. ANN NEUROL 2021;90:913-926.

ACP++: Action Co-Occurrence Priors for Human-Object Interaction Detection.

A common problem in the task of human-object interaction (HOI) detection is that numerous HOI classes have only a small number of labeled examples, resulting in training sets with a long-tailed distribution. The lack of positive labels can lead to low classification accuracy for these classes. Towards addressing this issue, we observe that there exist natural correlations and anti-correlations among human-object interactions. In this paper, we model the correlations as action co-occurrence matrices and present techniques to learn these priors and leverage them for more effective training, especially on rare classes. The efficacy of our approach is demonstrated experimentally, where the performance of our approach consistently improves over the state-of-the-art methods on both of the two leading HOI detection benchmark datasets, HICO-Det and V-COCO.

Broadband, silicon photonic, optical add-drop filters with 3 dB bandwidths up to 11 THz: publisher's note.

This publisher's note contains corrections to Opt. Lett.46, 2738 (2021)OPLEDP0146-959210.1364/OL.423745.

Broadband, silicon photonic, optical add-drop filters with 3 dB bandwidths up to 11 THz.

We present the designs, theory, and experimental demonstrations of ultra-broadband, optical add-drop filters on the silicon-on-insulator platform, realized using period-chirped contra-directional couplers. Our fabricated devices have ultra-broad 3 dB bandwidths of up to 11 THz (88.1 nm), with flat-top responses at their drop ports. All of our devices were fabricated using a commercial, CMOS-compatible, 193 nm deep-ultraviolet lithography process. By using lithography-prediction models, the measured bandwidths, insertion losses, central wavelengths, and extinction ratios of our devices are all in good agreement with our predicted, simulated results. Such filters are necessary for photonic integrated circuits to operate over multiple optical bands.

Evaluating the Risk of Digitalis Intoxication Associated With Concomitant Use of Dronedarone and Digoxin Using Real-World Data.

PURPOSE: Dronedarone may increase digoxin plasma levels through inhibition of P-glycoprotein. Using real-world data, we evaluated the risk of digitalis intoxication in concomitant users of dronedarone and digoxin compared digoxin-alone users. METHODS: We used the Clinformatics DataMart, a US claims database, to identify adult patients with atrial fibrillation (AF) or atrial flutter (AFL) who concomitantly used dronedarone and digoxin and those who used digoxin alone between July 2009 and March 2016. Digitalis intoxication during follow-up until March 2016 was ascertained using International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) and International Classification of Diseases, Tenth Revision, Clinical Modification (ICD-10-CM). Adjusted hazard ratios (HR) for digitalis intoxication in concomitant users versus users of digoxin alone were estimated, controlling for age, sex, cohort entry year, number of medical encounters for AF or AFL, history of congestive heart failure, diabetes, hypertension, stroke, myocardial infarction, renal failure, use of drugs interacting with digoxin, and digoxin dose. FINDINGS: Overall, 524 concomitant users and 32,459 users of digoxin alone were identified, among which 3 and 301 events of digitalis intoxication occurred during follow-up, respectively. Incidence rates were 17.25 and 9.17 cases per 1000 person-years, respectively. The adjusted HR for digitalis intoxication in concomitant users versus users of digoxin alone was 1.56 (95% CI, 0.50-4.88; P = 0.45). When digitalis intoxication was defined by ICD-9-CM and ICD-10-CM codes accompanied by laboratory testing for digoxin/digitoxin or hospitalization within 30 days, no events occurred in the concomitant users and 40 events occurred in the users of digoxin alone (incidence rate of 1.22 cases per 1000 person-years). IMPLICATIONS: Concomitant use of dronedarone and digoxin was uncommon in this study, and no significant increase in the risk of digitalis intoxication with concomitant use was found.

Dense Cross-Modal Correspondence Estimation With the Deep Self-Correlation Descriptor.

We present the deep self-correlation (DSC) descriptor for establishing dense correspondences between images taken under different imaging modalities, such as different spectral ranges or lighting conditions. We encode local self-similar structure in a pyramidal manner that yields both more precise localization ability and greater robustness to non-rigid image deformations. Specifically, DSC first computes multiple self-correlation surfaces with randomly sampled patches over a local support window, and then builds pyramidal self-correlation surfaces through average pooling on the surfaces. The feature responses on the self-correlation surfaces are then encoded through spatial pyramid pooling in a log-polar configuration. To better handle geometric variations such as scale and rotation, we additionally propose the geometry-invariant DSC (GI-DSC) that leverages multi-scale self-correlation computation and canonical orientation estimation. In contrast to descriptors based on deep convolutional neural networks (CNNs), DSC and GI-DSC are training-free (i.e., handcrafted descriptors), are robust to cross-modality, and generalize well to various modality variations. Extensive experiments demonstrate the state-of-the-art performance of DSC and GI-DSC on challenging cases of cross-modal image pairs having photometric and/or geometric variations.

Deep Depth from Uncalibrated Small Motion Clip.

We propose a novel approach to infer a high-quality depth map from a set of images with small viewpoint variations. In general, techniques for depth estimation from small motion consist of camera pose estimation and dense reconstruction. In contrast to prior approaches that recover scene geometry and camera motions using pre-calibrated cameras, we introduce in this paper a self-calibrating bundle adjustment method tailored for small motion which enables computation of camera poses without the need for camera calibration. For dense depth reconstruction, we present a convolutional neural network called DPSNet (Deep Plane Sweep Network) whose design is inspired by best practices of traditional geometry-based approaches. Rather than directly estimating depth or optical flow correspondence from image pairs as done in many previous deep learning methods, DPSNet takes a plane sweep approach that involves building a cost volume from deep features using the plane sweep algorithm, regularizing the cost volume, and regressing the depth map from the cost volume. The cost volume is constructed using a differentiable warping process that allows for end-to-end training of the network. Through the effective incorporation of conventional multiview stereo concepts within a deep learning framework, the proposed method achieves state-of-the-art results on a variety of challenging datasets.

Machine Boss: rapid prototyping of bioinformatic automata.

MOTIVATION: Many software libraries for using Hidden Markov Models in bioinformatics focus on inference tasks, such as likelihood calculation, parameter-fitting and alignment. However, construction of the state machines can be a laborious task, automation of which would be time-saving and less error-prone. RESULTS: We present Machine Boss, a software tool implementing not just inference and parameter-fitting algorithms, but also a set of operations for manipulating and combining automata. The aim is to make prototyping of bioinformatics HMMs as quick and easy as the construction of regular expressions, with one-line 'recipes' for many common applications. We report data from several illustrative examples involving protein-to-DNA alignment, DNA data storage and nanopore sequence analysis. AVAILABILITY AND IMPLEMENTATION: Machine Boss is released under the BSD-3 open source license and is available from http://machineboss.org/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Comparison of in vivo and in situ detection of hippocampal metabolites in mouse brain using 1 H-MRS.

The study of cerebral metabolites relies heavily on detection methods and sample preparation. Animal experiments in vivo require anesthetic agents that can alter brain metabolism, whereas ex vivo experiments demand appropriate fixation methods to preserve the tissue from rapid postmortem degradation. In this study, the metabolic profiles of mouse hippocampi using proton magnetic resonance spectroscopy (1 H-MRS) were compared in vivo and in situ with or without focused beam microwave irradiation (FBMI) fixation. Ten major brain metabolites, including lactate (Lac), N-acetylaspartate (NAA), total choline (tCho), myo-inositol (mIns), glutamine (Gln), glutamate (Glu), aminobutyric acid (GABA), glutathione (GSH), total creatine (tCr) and taurine (Tau), were analyzed using LCModel. After FBMI fixation, the concentrations of Lac, tCho and mIns were comparable with those obtained in vivo under isoflurane, whereas other metabolites were significantly lower. Except for a decrease in NAA and an increase in Tau, all the other metabolites remained stable over 41 hours in FBMI-fixed brains. Without FBMI, the concentrations of mIns (before 2 hours), tCho and GABA were close to those measured in vivo. However, higher Lac (P < .01) and lower NAA, Gln, Glu, GSH, tCr and Tau were observed (P < .01). NAA, Gln, Glu, GSH, tCr and Tau exhibited good temporal stability for at least 20 hours in the unfixed brain, whereas a linear increase of tCho, mIns and GABA was observed. Possible mechanisms of postmortem degradation are discussed. Our results indicate that a proper fixation method is required for in situ detection depending on the targeted metabolites of specific interests in the brain.

The oblique extraocular muscles in cetaceans: Overall architecture and accessory insertions.

The oblique extraocular muscles (EOMs) were dissected in 19 cetacean species and 10 non-cetacean mammalian species. Both superior oblique (SO) and inferior oblique (IO) muscles in cetaceans are well developed in comparison to out-groups and have unique anatomical features likely related to cetacean orbital configurations, swimming mechanics, and visual behaviors. Cetacean oblique muscles originate at skeletal locations typical for mammals: SO, from a common tendinous cone surrounding the optic nerve and from the medially adjacent bone surface at the orbital apex; IO, from the maxilla adjacent to lacrimal and frontal bones. However, because of the unusual orbital geometry in cetaceans, the paths and relations of SO and IO running toward their insertions onto the temporal ocular sclera are more elaborate than in humans and most other mammals. The proximal part of the SO extends from its origin at the apex along the dorsomedial aspect of the orbital contents to a strong fascial connection proximal to the preorbital process of the frontal bone, likely the cetacean homolog of the typical mammalian trochlea. However, the SO does not turn at this connection but continues onward, still a fleshy cylinder, until turning sharply as it passes through the external circular muscle (ECM) and parts of the palpebral belly of the superior rectus muscle. Upon departing this "functional trochlea" the SO forms a primary scleral insertion and multiple accessory insertions (AIs) onto adjacent EOM tendons and fascial structures. The primary SO scleral insertions are broad and muscular in most cetacean species examined, while in the mysticete minke whale (Balaenoptera acutorostrata) and fin whale (Balaenoptera physalus) the muscular SO bellies transition into broad fibrous tendons of insertion. The IO in cetaceans originates from an elongated fleshy attachment oriented laterally on the maxilla and continues laterally as a tubular belly before turning caudally at a sharp bend where it is constrained by the ECM and parts of the inferior rectus which form a functional trochlea as with the SO. The IO continues to a fleshy primary insertion on the temporal sclera but, as with SO, also has multiple AIs onto adjacent rectus tendons and connective tissue. The multiple IO insertions were particularly well developed in pygmy sperm whale (Kogia breviceps), minke whale and fin whale. AIs of both SO and IO muscles onto multiple structures as seen in cetaceans have been described in humans and domesticated mammals. The AIs of oblique EOMs seen in all these groups, as well as the unique "functional trochleae" of cetacean SO and IO seem likely to function in constraining the lines of action at the primary scleral insertions of the oblique muscles. The gimble-like sling formed by SO and IO in cetaceans suggest that the "primary" actions of the cetacean oblique EOMs are not only to produce ocular counter-rotations during up-down pitch movements of the head during swimming but also to rotate the plane containing the functional origins of the rectus muscles during other gaze changes.