Excess accumulation of manganese (Mn) in the brain results in a neurological syndrome with cognitive, psychiatric, and movement abnormalities. The highest concentrations of Mn in the brain are achieved in the basal ganglia, which may precipitate a form of parkinsonism with some clinical features that are similar and some that are different to those in Parkinson’s disease (PD). Recently, scientists have debated the possibility that Mn may have an etiological role in PD or that it may accelerate the expression of PD.The goal of this review was to examine whether chronic Mn exposure produces dopamine neuron degeneration and PD or whether it has a distinct neuropathology and clinical presentation.I reviewed available clinical, neuroimaging, and neuropathological studies in humans and nonhuman primates exposed to Mn or other human conditions that result in elevated brain Mn concentrations.Human and nonhuman primate literature was examined to compare clinical, neuroimaging, and neuropathological changes associated with Mn-induced parkinsonism.Clinical, neuroimaging, and neuropathological evidence was used to examine whether Mn-induced parkinsonism involves degeneration of the nigrostriatal dopaminergic system as is the case in PD.The overwhelming evidence shows that Mn-induced parkinsonism does not involve degeneration of midbrain dopamine neurons and that l-dopa is not an effective therapy. New evidence is presented on a putative mechanism by which Mn may produce movement abnormalities. Confirmation of this hypothesis in humans is essential to make rational decisions about treatment, devise effective therapeutic strategies, and set regulatory guidelines. Idiopathic Parkinson’s disease (PD) is a progressive neurodegenerative disorder with a slow onset, and compared with the familial forms of the disease, it is associated with advanced age (> 55 years of age). The four cardinal signs of idiopathic PD are tremor at rest, bradykinesia (hypokinesia and akinesia), rigidity, and postural instability . The diaBradykinesia is defined by slowness of movements or difficulty in initiating and executing movement, and it is one of the essential signs used to diagnose idiopathic PD . BradykiAlthough the clinical diagnosis of PD is based on a combination of the four cardinal motor signs, other parkinsonian disorders also express many of these signs , and a d18F]-fluoro-dopa PET , and action and postural tremor . Table 1Besides occupational exposures, other conditions have been shown to increase accumulation of Mn in the brain and have been valuable sources of information in understanding Mn-induced neurological dysfunction, including parkinsonism.Recently, there have been a number of reports, primarily originating from Eastern European countries and Russia, of young addicts afflicted with an atypical form of parkinsonism  Table 1Table 1. The parkinsonian signs in these young drug-abuse subjects are consistent with those in occupationally exposed Mn workers . An impoThere is evidence of parkinsonism associated with chronic liver disease. Patients with advanced cirrhosis have been documented with a form of parkinsonism with clinical symptoms similar to Mn-induced parkinsonism. This finding is likely because Mn is excreted in the bile, and in persons with chronic liver disease, the excretion of Mn is markedly impaired, with subsequent accumulation in the brain. The clinical symptoms associated with idiopathic PD, chronic liver disease, ephedrone abuse, and occupational exposures to Mn are described in With the advent of molecular imaging techniques in the 1980s, a number of neuroimaging modalities have been used to understand the structural, cellular, and molecular changes that occur in neurological and neurodegenerative diseases. In PD and conditions associated with Mn-induced parkinsonism, three principal neuroimaging modalities have been used: T1-weighted MRI, SPECT, and PET. Structural MRI and magnetic resonance spectroscopy have also been used, but to a lesser extent, and are not discussed in this review.In human conditions where Mn is the most likely etiological agent for the parkinsonism, a high percentage of the subjects exhibited a hyperintensive signal in T1-weighted MRI that is typically first observed in the globus pallidus and in other basal ganglia structures such as the substantia nigra, caudate, and putamen , Table 218F]-fluoro-dopa PET  and in a patient with liver disease (2005). These findings suggest the possibility of nigrostriatal dopamine neuron degeneration. However, other factors need to be considered. First, in the 2001 study, 15 career welders were studied, and [18F]-fluoro-dopa PET was performed in only two of them. Second, all of the welders were responsive to l-dopa therapy, and this response is not consistent with Mn-induced parkinsonism. Third, none of the welders tested with T1-weighted MRI scans (8 of 15) exhibited a hyperintensive signal in the basal ganglia despite being career welders. Further, one of the two welders that had reduced [18F]-fluoro-dopa uptake in the posterior putamen had a positive family history of PD, which suggests the possibility of a familial form of the disease. Taking all of the information together, the most parsimonious explanation is that the parkinsonism in these welders was not likely associated with the Mn in the welding fumes. The second study  in the basal ganglia of patients with liver disease .The early nonhuman primate studies of Mn exposure described decreased levels of dopamine in the caudate , striatu3 concentration of Mn for 5 days/week, 6 hr/day, for 2 years. They found decreased levels of dopamine in the caudate and globus pallidus but not in the putamen and substantia nigra. 2 at 20 mg/kg/day) for 18 months and reported significant reductions in dopamine concentrations in the striatum (−42%) and the midbrain (−15%). In this study, two observations were important. First, the brain was dissected, and the different regions were frozen at −20°C. For dopamine analysis, this temperature is not the most appropriate for storing fresh tissue, because dopamine autooxidizes readily. Second, the dopamine recovery for the method used was less than desirable and ranged from 72–84%. Although a comparison of control and treated dopamine tissue content is still possible, often in these early studies, the control and Mn-treated animals were euthanized at different times; thus potential differences in dopamine tissue content could result from differences in storage time and conditions. For example, in the study by In another study, The most consistent observation of these early studies was that Mn produced morphological changes in the globus pallidus, subthalamic nuclei, and substantia nigra pars reticulata, whereas the SNpc remained intact . More reIn vitro quantitative receptor autoradiography has been used to examine dopamine neuron markers in the brain of Mn-exposed nonhuman primates  was reduced by 75% in the caudate and putamen of Mn-exposed animals. However, there were technical problems that need to be discussed: a) The authors stated that the level of nonspecific [3H]-mazindol binding in the caudate and putamen was 50% and 60%, respectively. This level of nonspecific binding was very high, so the specific binding signal-to-noise (nonspecific binding) ratio was very low. Typically in this type of assay, one wants a low level of nonspecific binding, in the range of 10–15% of total binding. b) [3H]-Mazindol is known to bind to other monoaminergic uptake sites besides DAT (3H]-mazindol autoradiography results were a combination of DAT and other monoaminergic uptake sites. c) As pointed out, even if [3H]-mazindol was made selective for DAT by including antagonists for other monoaminergic transporters, a reduction in [3H]-mazindol binding to DAT would not necessarily reflect dopamine terminal loss, but it could represent a competitive inhibition of Mn with radioligand binding to DAT as shown by 3H]-SCH 23390 binding to D1R in the caudate and putamen with no change in [3H]-spiperone binding to D2R in the same regions. These results were opposite to what in vivo PET imaging and in vitro autoradiography—that is, decreased D2R and no effect on D1R.primates . The groprimates , 2008, hides DAT . To ensuin vivo and in vitro methods , [11C]-raclopride PET for D2R, and [11C]-l-dopa for l-dopa decarboxylase activity. They found a progressive decrease in [11C]-nomifensine binding as a function of cumulative dose in both animals. One of the animals had a transient decrease in [11C]-raclopride binding to D2R that returned to baseline by the end of the exposure period, and no effect of Mn treatment on [11C]-l-dopa PET was observed. These studies indicate that dopamine neurons are not degenerating, but there is a potential loss of dopamine terminals based on the [11C]-nomifensine PET. However, a firm conclusion that chronic Mn results in the loss of DAT and presumably dopamine neuron terminals in the striatum cannot be made because [11C]-nomifensine is not a selective DAT ligand and because Mn can directly inhibit ligand binding to DAT.PET imaging in nonhuman primates exposed to Mn was first performed by 18F]-fluoro-l-dopa PET, [11C]-raclopride PET, or [18F]-deoxyglucose PET despite the fact that the animals exhibited hypoactivity . They performed PET studies of presynaptic and postsynaptic “structural proteins” such as DAT ([11C]-methylphenidate as the PET radioligand) and D2R ([11C]-raclopride as the PET radioligand) to examine various markers of the dopaminergic synapse. A PET study to assess dopaminergic function was also included. Although [18F]-fluoro-dopa PET is a perfectly valid and useful functional marker of dopaminergic activity, this method interrogates the ability of dopamine neurons to synthesize dopamine, and it may indirectly measure the dopamine-releasing capacity of dopamine neurons. in vivo dopamine release using a bolus plus continuous infusion of [11C]-raclopride (D2R ligand) with amphetamine challenge  of in vivo dopamine release in the striatum measured by PET (in vivo dopamine release was observed in the absence of a change in DAT levels and with a small but significant decrease on D2R levels in the striatum (in vivo dopamine release measured by PET was not due to a reduction in tissue dopamine levels, because dopamine concentrations were not significantly different from all controls (in vivo dopamine release is associated with the ability of Mn to disrupt presynaptic release mechanisms.Since the study by hallenge . This tye to D2R . An animd by PET , 2008 (Td by PET . The effstriatum . Therefocontrols , 2008. IThe available evidence from human and nonhuman primate studies using behavioral, neuroimaging, neurochemical, and neuropathological end points provides strong support to the hypothesis that, although excess levels of Mn accumulation in the brain results in an atypical form of parkinsonism, this clinical outcome is not associated with the degeneration of nigrostriatal dopaminergic neurons as is the case in PD. In fact, the new evidence suggests that it may be the dysfunction of this system or the inability to release dopamine that produces the movement abnormalities documented in Mn-exposed subjects. This new finding is consistent with the fact that l-dopa therapy does not provide a benefit to the Mn-induced movement abnormalities as it does in PD  Table 1Table 1. Although the latest research clearly suggests that the effects of Mn on presynaptic aspects of the dopaminergic synapse need further exploration, delineating its effects on postsynaptic proteins of the dopaminergic synapse or other neuronal systems downstream and on the role of glial cells is essential in order to achieve a greater understanding of the neurotoxicity of Mn. The continued use of neuroimaging techniques that have already been used to study Mn neurotoxicity, as well as some others such as diffusion tensor imaging, are also likely to provide new information. Ultimately, the most significant studies in understanding Mn-induced parkinsonism are those in humans. From this perspective, persons who inject high doses of Mn in the ephedron preparations and workers who are occupationally exposed to moderate levels of Mn are cohorts in which comprehensive neurological assessments along with neuroimaging studies can provide longitudinal information about Mn neurotoxicity, its reversibility, and the usefulness of therapeutic strategies. Finally, although the preponderance of the behavioral, neuroimaging, and neuropathological evidence indicates that chronic Mn exposure does not cause degeneration of nigrostriatal dopamine neurons and PD, the parkinsonism and other neurological effects resulting from chronic Mn exposure are highly debilitating, and it should not be treated with any less importance.


Systems biology approaches offer novel insights into the development of chronic liver diseases. Current genomic databases supporting systems biology analyses are mostly based on microarray data. Although these data often cover genome wide expression, the validity of single microarray experiments remains questionable. However, for systems biology approaches addressing the interactions of molecular networks comprehensive but also highly validated data are necessary.http://www.medicalgenomics.org/databases/loma/news, currently holding approximately 1260 confirmed molecular associations for chronic liver diseases such as HCC, CCC, liver fibrosis, NASH/fatty liver disease, AIH, PBC, and PSC. We furthermore transformed these data into a powerful resource for molecular liver research by connecting them to multiple biomedical information resources.We have therefore generated the first comprehensive database for published molecular associations in human liver diseases. It is based on PubMed published abstracts and aimed to close the gap between genome wide coverage of low validity from microarray data and individual highly validated data from PubMed. After an initial text mining process, the extracted abstracts were all manually validated to confirm content and potential genetic associations and may therefore be highly trusted. All data were stored in a publicly available database, Library of Molecular Associations Together, this database is the first available database providing a comprehensive view and analysis options for published molecular associations on multiple liver diseases. The completely sequenced human genome has made it possible for modern medicine to step into an era rich in genetic information and high-throughput genomic analysis . Large gThese novel and readily available genetic resources and analytical tools may be the key to unravel the molecular basis of diverse chronic liver diseases as many of these must be regarded to be complex multigenic diseases. Moreover, since an efficient treatment for many of these conditions and diseases is lacking, further understanding of the genetic background of chronic liver disease will be crucial in order to develop new therapies aimed at selected targets -10.At present, large genetic association studies for liver diseases are mostly based on microarray data or SAGE ,12. SomeIn order to establish this database, the complete PubMed database, currently containing more than 17 million publications, has initially been searched by means of MeSH terms and text mining semi-automated searches  OR "hepatocellular carcinoma" OR "HCC" OR "hepatoma" OR "liver cancer" OR "primary liver cancer" OR "liver tumor" OR "liver carcinoma" OR "primary liver cancer" OR "hepatic tumor" for HCC2) "biliary tract cancer" OR "gallbladder cancer" OR "cholangiocellular carcinoma" OR cholangiocarcinoma for CCC3) "fibrosis" OR "fibroses" for liver fibrosis4) "NASH" OR "NAFLD" OR "nonalcoholic steatohepatitis" OR "non-alcoholic steatohepatitis" OR "nonalcoholic fatty liver disease" [MeSH] for NASH5) "AIH" OR "hepatitis, autoimmune" [MeSH] OR "autoimmune hepatitis" for AIH6) "PBC" OR "primary biliary cirrhosis" [MeSH] OR "biliary cirrhosis, primary"7) "PSC" OR "sclerosing cholangitis" OR "cholangitis, sclerosing" [MeSH] OR "primary sclerosing cholangitis" for PSChttp://www.hugo-international.org. Making use of the pattern matching capabilities of the Perl programming language http://www.perl.org, we used a pattern matching approach to identify gene names in the previously selected abstracts. E.g. if the gene to be searched was p53, the abstract was searched for any combinations of signs starting with the letter p followed by the numbers 5 and 3. This approach ensured a most flexible search strategy.The abstracts identified to be associated with the particular diseases were then searched for human, mouse, and rat gene names and alias gene names as provided by the Human Genome Organization  human studies.By this approach we gathered a total of 101026 abstracts, potentially holding information on genetic associations to chronic liver disease. In detail we identified 44548 abstracts suggesting genetic associations for HCC, 13710 for CCC, 917 for AIH, 37173 for liver fibrosis, 2022 for NASH, 1211 for PBC and 1445 for PSC.This strategy revealed all abstracts containing both the disease name and a gene name. However, also this semi-automated search provided a first approximation to genetic associations to liver diseases, as in multiple abstracts this genetic association could not be confirmed by reading the full abstract. E.g. the abstract may read that the gene XY is not related to disease Z, which would have also been detected by the described search strategy. Thus these automatically, by means of text mining identified abstracts, were then all individually read to confirm the suggested genetic association with the particular disease. We thereby obtained a large number of manually confirmed genetic associations to liver diseases.Thereby, we finally identified 574 molecular associations for HCC, 150 molecular associations for liver fibrosis, 310 molecular associations for CCC, and 82 molecular associations for NASH. Only a few genes were identified to be related to the development of autoimmune liver disease: 29 abstracts describing molecular associations were found to be related to AIH, 56 to PBC, and 60 to PSC. Overall, we were able to identify a total of 1260 molecular associations for major chronic liver diseases. As all these molecular associations were manually confirmed by reading the individual full published abstract, and thus these molecular associations can be trusted to be highly reliable.http://www.postgresql.org. Subsequently, this database was then made publicly accessible and searchable through a webinterface . Genetic mutations and a variable genetic background have been demonstrated to significantly influence the development and course of multiple diseases as well as the efficiency of treatment with diverse drugs.Over the past decades molecular mechanisms and individual factors have been shown to be involved in the development of liver diseases and it has become clear that most liver diseases such as liver cancer, cholangiocellular carcinoma, liver fibrosis, NASH or autoimmune liver diseases are complex systemic diseases. Thus they must not only be investigated focussing on individual, potentially key regulatory genes but also with respect to underlying genetic clusters and networks ,20-22. HTo overcome these obstacles, we designed a publicly available database for genetic associations with human (liver) diseases, Library of Molecular Associations (LOMA). Currently, this database holds 1260 molecular associations for a total of seven liver diseases, HCC, CCC, liver fibrosis, AIH, PBC and PSC. Most molecular associations were identified to be associated with HCC, 595, followed by CCC and liver fibrosis, 310 and 150 database entries, respectively. 82 entries were associated with the development of NASH. As expected and in concordance with a currently missing clear association of genetic networks with autoimmune liver diseases, only few genes were reported to be associated with AIH, PBC, or PSC. However, as some of these diseases, especially PBC, have been demonstrated an increased relative risk of the disease in twins and first grade relatives, a genetic basis of the disease must be suspected. Thus further research into the genetic basis of the disease is warranted to identify targets for therapeutic treatment of the disease.In contrast to other available genetic association databases such as the Genetic Association Database , our datThe LOMA database provides multiple search options to support complex genetic analyses. Firstly, LOMA offers the option to search for individual genes and their association with different liver diseases. This search may be performed by means of a search for individual gene names, NCBI Gene IDs , EnsemblAfter executing a search, the result page for these searches offers the genetic associations to individual diseases if present. Furthermore, the results page gives a summary on gene name, associated disease, NCBI Gene ID , EnsemblFor example, if one wants to know all molecular associations with the Wnt signaling pathway that have been published to play a role in HCC development, this is now easily possible with our database. On the search site under Explore molecular associations one would select "HCC" from the "Molecular Associations" column and "Wnt Signaling Pathway" from the KEGG column. The executed search will then return a number of Wnt signaling associated genes and target genes, APC, AXIN1, CTNNB1, MMP7, PRKCA, SMAD4, TP53.For these molecular associations further information is linked especially in the details section of each gene. With this information one could for example evaluate the enrichment of the Wnt signaling pathway among all CCC related molecular factors.A key issue in developing this database was to provide the hepatologic community with a powerful but simultaneously highly reliable and comprehensive database to perform systems biology based high-throughput searches and comparison of gene expression, our database was linked to multiple other sources of genomic or genetic information and gene expression information in particular. This rich embedding of our database into the current scenery of bioinformatics repositories provides valuable connections which may support advanced search and evaluation strategies.In detail, LOMA has been linked to the most commonly used bioinformatics databases, such as PubMed , the EurOur database has been evaluated against other public databases such as Genetic Association database, HuGENavigator, or OMIM. This evaluation was performed using the molecular associations to CCC development. Comparing our text mining strategy to a manually searched sample set of 1000 randomly selected CCC associated abstracts, we documented a sensitivity of our approach of 98% and a false negative rate for abstracts not selected by or text mining approach but containing molecular associations to CCC of 2%.For CCC development our database contained all associations also listed by other databases with two exceptions, MRP2/ABCC2 which was published only recently and the miRNA370 which was missed by our search strategy . In contThe Library of Molecular Associations (LOMA) was designed as a comprehensive database of highly reliable molecular associations conceived to close the gap between high-throughput molecular data for automated analysis and individual reliable experimental data by molecular biology. Currently this database supports information on molecular associations for several liver diseases, HCC, CCC, liver fibrosis, NASH/fatty liver disease, AIH, PBC and PSC. In addition, the database was extensively embedded into the currently available genomics repositories supporting advanced searches and cross analyses with other databases.Together, this database is the first available database providing a comprehensive view and analysis options for published molecular associations on multiple liver diseases.SB, JH, TM, FS, PRG, and AT: Abstract selection and database curation. MK, AW, KS, and AT: Database programming. All authors read and approved the final manuscript.


Water for Elephants . But Jacob is in "the old folks' home." Sitting down to mushy peas in the dining room, when he still has teeth and wants pot roast and corn on the cob, affronts his dignity. It only gets worse when he starts an argument with another resident at dinner and is sent to his room like a misbehaving child.Preventing Chronic Disease discusses the health concerns of these seniors. We thank Dr. Jaya Rao of the Division of Adult and Community Health, National Center for Chronic Disease Prevention and Health Promotion, Centers for Disease Control and Prevention, for serving as guest editor for this issue.Mr. Jankowski is a member of the population group called the oldest old. This issue of From 2000 to 2030, the number of people aged 80 years or older in the United States will double to 19.5 million (resilience. Lavretsky and Irwin's review of the literature on resilience defines it as the ability to maintain biologic and psychologic homeostasis under stress (However, quality of life in the last years depends on more than the items in this list; it requires r stress . Stress r stress ,5. A stur stress . The Gerr stress . A Swedir stress .Is resilience inborn, or does it develop over time? Can historical circumstances create resilience? Howe and Strauss popularized the concept that different U.S. generations hold different world views, depending on their life experiences . They reCan resilience be actively promoted? Will resilience vary between the GI and boomer generations as their histories vary? We know aging experiences will differ; for example, the baby boomer cohort will see more of its members survive to the oldest ages, and those seniors will be healthier. These differences could affect the health advice we offer to the oldest old. We will need to discover what characteristics remain constant across generations and what characteristics history may affect.resilient?Meanwhile, what of Jacob Jankowski? Throughout the story, we learn of his present and his past . He spenToday the circus is in town, just down the street from the old folks' home. One of Jacob's sons is scheduled to take him there. Jacob waits in the lobby in his wheelchair, and his son doesn't appear. One by one, the families of the other residents arrive, but still Jacob waits, impatient to see the Big Top one last time. Then he learns his son has forgotten their appointment, on today of all days. It's not fair, Jacob thinks. He's stuck in this lobby.Or is he?Carefully Jacob transitions from his wheelchair to his walker. Step by shuffling step, he moves out of the lobby, though the automatic doors, onto the sidewalk. Then he heads down the street, 10 inches at a time, making steady progress. Jacob Jankowski, age 90 or 93, one or the other, is running away to the circus.


Molecular Brain is a peer-reviewed, open-access online journal that aims at publishing high quality articles as rapidly as possible. The journal will cover a broad spectrum of neuroscience ranging from molecular/cellular to behavioral/cognitive neuroscience and from basic to clinical research. Molecular Brain will publish not only research articles, but also methodology articles, editorials, reviews, and short reports. It will be a premier platform for neuroscientists to exchange their ideas with researchers from around the world to help improve our understanding of the molecular mechanisms of the brain and mind.We are delighted to announce the arrival of a brand new journal dedicated to the ever-expanding field of neuroscience.  Molecular Brain.Neuroscience, the study of the nervous system, is not an emerging field . In factMolecular Brain is an open-access, peer-reviewed online journal [Molecular Brain will provide a valuable contribution to the rapidly expanding and diverse neuroscience field. As an online, open access journal, Molecular Brain will provide a home for high quality research on molecular, cellular and system levels of the physiological and pathological brain. journal . Open ac journal . The opeMolecular Brain publishes not only research articles, but also methodology articles, editorials, reviews on timely issues and short reports. It will provide a forum for neuroscientists to communicate news, thoughts, and ideas.The word "molecular" in the title of the journal is not meant to limit the journal's scope. The journal covers a broad spectrum of neuroscience and publishes studies of a wide range of topics in the field: from molecules to behavior, from basic science to clinical research, and from invertebrates to vertebrates, at the molecular, cellular, and systems levels. We especially encourage the submissions of works that contain studies at both molecular and behavioral levels. Molecular Brain aims to publish high quality articles as rapidly as possible, usually within 1 month. Our editors are committed to providing speedy reviews. We will constantly consider ways to improve the review process with one goal in mind: speed and quality. To ensure quality, Molecular Brain fully utilizes the journal's expert Editorial Board and reviewers from around the world in providing the most current perspective on a specific field of neuroscience. Molecular Brain operates a closed peer review process. Manuscripts will initially be evaluated by one of the Editors-in-Chief who will decide whether further peer review is warranted. An academic decision will be made based on the recommendations of at least two expert reviewers.Molecular Brain aims to provide a home for the latest research from neuroscientists all over the world. Members of the Editorial Board have a broad range of expertise representing diverse areas of neuroscience. Importantly they are committed to the success of Molecular Brain by serving as reviewers and publishing their own important work in the journal. Our goal is to make Molecular Brain a premier journal in neuroscience within five years. This will not be possible without your support. We welcome your contributions and invite you to join us in creating this new and high quality platform that is Molecular Brain.As a top-quality open-access online journal, Molecular Brain. The tools available for the exploration of the brain have now reached such a level of sophistication that, along with most neuroscientists, I believe this will be a golden age for research into the most complex structure in biology. The launch of Molecular Brain reflects this optimism, and with its strong editorial board and open-access policy it is well placed to play a leading role in the opening up of biology's last frontier." -Tim V. P. Bliss "It is a pleasure to extend a welcome to the first issue of


Involvement of the subcutis by Kaposi sarcoma (KS) occurs primarily when cutaneous KS lesions evolve into deep penetrating nodular tumors. Primary KS of the subcutaneous tissue is an exceptional manifestation of this low-grade vascular neoplasm.We present a unique case of acquired immune deficiency syndrome (AIDS)-associated KS manifesting primarily in the subcutaneous tissue of the anterior thigh in a 43-year-old male, which occurred without overlying visible skin changes or concomitant KS disease elsewhere. Radiological imaging and tissue biopsy confirmed the diagnosis of KS.This is the first documented case of primary subcutaneous KS occurring in the setting of AIDS. The differential diagnosis of an isolated subcutaneous lesion in an human immunodeficiency virus (HIV)-infected individual is broad, and requires both imaging and a histopathological diagnosis to guide appropriate therapy. Kaposi sarcoma (KS) is a low-grade vascular neoplasm associated with Human Herpesvirus-8 (HHV8) infection. There are four clinical-epidemiological types, including African (endemic) KS, AIDS-associated (epidemic) KS, classic KS, and transplant-associated (iatrogenic) KS. KS is a multifocal tumor that presents chiefly in mucocutaneous sites. AIDS-associated KS tends to be multicentric, often involving mucous membranes along the entire gastrointestinal tract and occurring in atypical locations. Patients with AIDS frequently manifest with skin lesions of the lower extremities, face, trunk, genitalia. In patients with AIDS, KS may also involve their lymph nodes and visceral organs. For patients with classic and transplant-associated KS, lesions are often limited to the skin, although visceral KS may occur. In African KS the legs are primarily involved, with more widespread KS involvement of the lymphoid system seen in children. Involvement of several unusual anatomical sites have been reported, such as KS of the musculoskeletal system, nervous system, heart, breast, major salivary glands, and endocrine organs .Involvement of the subcutaneous tissue (subcutis or hypodermis) by KS typically occurs when cutaneous KS lesions evolve from a plaque stage lesion into deep endophytic nodular tumors. Large KS tumors may even penetrate deep down to involve underlying contiguous bone . Hence, A 43-year-old homosexual man who was HIV positive for 18 years presented with a one-year history of a slowly enlarging mass in the proximal left anterior thigh. He described stabbing pain, often experiencing sharp shooting pains down the left thigh. He had been on and off antiretroviral medication, which he had stopped three years prior to this presentation. He had bilateral total hip replacements for avascular necrosis and osteoarthritis approximately three years prior to this visit. He reported no specific trauma or previous injection to his left thigh.3 and HIV viral load 72 copies/mL while off all antiretroviral medications.On physical examination, he appeared to be in good health. His gait was antalgic. He had no visible mucocutaneous KS lesions and he did not exhibit features of fat maldistribution. There was a firm 3 cm mass present deep in his left thigh that was tender to palpation. The mass was well away from the groin and inguinal region. In particular, there were no overlying skin changes or associated lymphedema. He had enlarged axillary lymph nodes. His complete blood count was unremarkable and his CD4 T-cell count was 249 cells/mmAn ultrasound test showed a 2.6 × 1.8 × 1.2 cm solid, vascular, heterogeneous lesion within the deep thigh soft tissue. A magnetic resonance image (MRI) showed a solid, vascular enhancing mass with spiculated margins , reactive/benign conditions (e.g. nodular fasciitis), benign neoplasms (e.g. lipoma), and malignant neoplasms . The anterior thigh compartment is an uncommon site for an enlarged lymph node to manifest. Although infection should always be excluded in the context of immunosuppression, other than tenderness to palpation there were no findings prior to biopsy in our case that were particularly indicative of infection. Soft tissue abscess due to mycobacterial infection has been noted in patients with AIDS . In our KS lesions develop as a result of the following combination of factors: HHV8, altered immunity (immunosuppression), and an inflammatory/angiogenic milieu . The etiIn our case, imaging studies revealed a solid vascular subcutaneous mass with features highly concerning for malignancy. In such a case a definitive tissue-based diagnosis is key to guiding appropriate KS therapy. KS needs to be high in the differential diagnosis in the setting of HIV infection, to avoid a major sarcoma surgical operation. KS disease was not identified elsewhere in or patient, confirming the unusual diagnosis of primary subcutaneous KS. For a soft tissue abscess MRI will show a well-demarcated fluid collection that is hypointense on T1-weighted images, hyperintense on T2-weighted images, surrounded by a low-signal-intensity pseudocapsule with all sequences, and will likely demonstrate peripheral rim enhancement after intravenous administration of gadolinium-based contrast material . For comWe present the first documented case of primary subcutaneous KS occurring in the setting of AIDS. The differential diagnosis of an isolated subcutaneous soft tissue tumor in an HIV-infected individual is broad, and requires imaging evaluation and a definitive pathological diagnosis in order to guide appropriate therapy. Awareness that KS can occur as an isolated deep soft tissue mass may avoid potential misdiagnosis.The authors declare that they have no competing interests.BJD, LP, and JM were involved in conception and design, in the drafting of the manuscript. All authors have read the final manuscript and approve of its submission.


Wilson and Schwarzman [Environ Health Perspect 117:A386 (2009)], the last sentence in the first paragraph was incorrect. The corrected sentence is as follows:In the letter by We would add that public policy that accurately reflects current science—and the needs of the chemicals market—is instrumental to the widespread adoption of green chemistry.EHP apologizes for the error.


In many scientific disciplines the use of a metaphor as an heuristic aid is not uncommon. A well known example in somatic medicine is the ‘defense army metaphor’ used to characterize the immune system. In fact, probably a large part of the everyday work of doctors consists of ‘translating’ scientific and clinical information  into information tailored to the needs and capacities of the individual patient. The ability to do so in an effective way is at least partly what makes a clinician a good communicator. Schizophrenia is a severe psychiatric disorder which affects approximately 1% of the population. Over the last two decades a large amount of molecular-biological, imaging and genetic data have been accumulated regarding the biological underpinnings of schizophrenia. However, it remains difficult to understand how the characteristic symptoms of schizophrenia such as hallucinations and delusions are related to disturbances on the molecular-biological level. In general, psychiatry seems to lack a conceptual framework with sufficient explanatory power to link the mental- and molecular-biological domains.Here, we present an essay-like study in which we propose to use visualized concepts stemming from the theory on dynamical complex systems as a ‘visual metaphor’ to bridge the mental- and molecular-biological domains in schizophrenia. We first describe a computer model of neural information processing; we show how the information processing in this model can be visualized, using concepts from the theory on complex systems. We then describe two computer models which have been used to investigate the primary theory on schizophrenia, the neurodevelopmental model, and show how disturbed information processing in these two computer models can be presented in terms of the visual metaphor previously described. Finally, we describe the effects of dopamine neuromodulation, of which disturbances have been frequently described in schizophrenia, in terms of the same visualized metaphor.The conceptual framework and metaphor described offers a heuristic tool to understand the relationship between the mental- and molecular-biological domains in an intuitive way. The concepts we present may serve to facilitate communication between researchers, clinicians and patients. Schizophrenia is a severe psychiatric disorder, characterized by the emergence at adolescence of psychotic phenomena: hallucinations, delusions and bizarre behavior. The neurodevelopmental hypothesis, which proposes a leading role for early aberrant brain development on which normal and/or abnormal brain maturation is superimposed has become the dominant paradigm for understanding the development of schizophrenia. The neurodevelopmental theory is usually underscored by a large amount of molecular-biological, imaging and genetic data which have been accumulated over the last two decades. Taken together, these findings point to reduced neuronal connectivity and synaptic stability. Psychotic symptoms are considered to be emergent properties on the psychological and behavioral level of the aberrantly developed neural system which start when during brain development a critical threshold is passed.However, it remains difficult to understand how psychotic symptoms are related to disturbances on the molecular-biological level. Kapur In somatic medicine the use of some kind of metaphor to bridge the biological and phenomenological domains is not uncommon. For instance, the immune system is characterized by the defense-army metaphor. Though the immune system is notoriously difficult on a molecular level, and the specific molecular phenomena which happen during, say, an HIV infection, may be only within the grasp of experts, the defense-army metaphor functions as a bridge between lay and professionals and facilitates understanding in an intuitive way.In this article we attempt to outline a heuristic framework that could provide a basis for uniting clinical phenomena and neurobiological theories. We will introduce what we have coined a ‘visual metaphor’ which is supposed to bridge the mental and neural domains. To this end we will combine two fields of research, namely the study of the behavior of complex systems and computer models of schizophrenia.During the last two decades a large body of literature has appeared concerning the study of complex systems and its associated theoretical framework ‘complexity theory’ The neurodevelopmental theory of schizophrenia has been supported by computer simulations of neural information processing. Hoffman and McGlashan reported a body of research on schizophrenia Both research efforts depend greatly on computer models and complicated mathematical ideas which are unfamiliar to most psychiatrists. However, the ideas developed in both fields have great heuristic value. In this paper we will try to visualize findings using concepts from the behavior of complex systems.This article will first briefly introduce some basic principles of computer models of mental processes, so-called Artificial Neural Networks (ANN). A conceptual framework which is related to the functioning of ANN is described and we show how this conceptual framework can be visualized. This is the visual metaphor we seek to describe. Subsequently, we describe how implementing developmental disturbances in ANN give rise to pathological phenomena, what their relationship is with ‘real-life’ pathology and how these phenomena can be understood in terms of the visual metaphor we introduced. Finally, we will show how psychotic symptoms originating from dopamine disturbances can be understood in terms of this visual metaphor.attractor neural networkThere is an extensive body of literature on the principles of ANN It is possible to present this ANN a certain pattern, for instance, a pattern that symbolizes the letter ‘A’  and ‘non-firing’ (‘0’). These two states can be graphically depicted as two points. Over time, the dynamical behavior of this one-neuron ‘system’ can be represented by a From a mathematical point of view, there is no difference with respect to the situation describing systems consisting of N neurons. They describe a trajectory through a N-dimensional space. Unfortunately, this situation cannot be visualized.We propose to simplify this high-dimensional situation to the depiction of a plane with the different states as points in the plane .basin of attraction, represents the influence of the attractor over its surroundings. In psychological terms, it represents how much of a stored memory has to be ‘fed into’ the neural system in order for the ANN to perform an associative recollection of the memory. Changing from one state to another can be conceptualized as the movement of a ball over the surface An attractor (a learned pattern) can be represented as a dent in this plane , center.Presenting a pattern to an ANN can be likened to moving a ball over the surface and letting it loose. The system will then seek some state guided by the attractors which dominate the systems' behavior and will settle in one of its attractors , center.phase transitions and appear when the system is near bifurcation points: unstable states in which the system chooses between one or another final state to evolve to. Such sudden changes in the state of the system can be imagined as popcorn, ‘popping’ from one attractor to another For a proper understanding, one has to imagine the dynamic aspects of the whole process. So, the state of the ANN is continuously moving through the space of its possible states guided by external influences and attractors with the ANN simultaneously creating new attractors which can be regarded as memory traces Efforts to simulate schizophrenic pathology using ANN assumed that schizophrenia arises from reductions in connectivity between brain regions as a result of developmental disturbances during synaptogenesis We can visualize this situation using the introduced metaphor see , bottom.Excessive pruning has created a so-called ‘parasitic’ attractor. Input will not bring the ANN into the regions in state space which correspond to a previously learned symbol but to a newly emerged attractor. The re-creation of the pattern can be interpreted as autonomous activation of the ANN. In terms of dynamical behavior the pathologically configured ANN is performing an inappropriate trajectory through its state space. .Later studies involved a more complex ANN (a back-propagation network with a recurrent layer) focusing on developmentally disturbed speech processing as a source of auditory hallucinations This ANN used featured a verbal working memory with linguistic expectations built up from prior exposure to a training set of grammatical correct sentences. The training set consisted of sentences like “Jane kiss girl” or “Cop chase man”. The ANN was programmed to process degraded input signals into identifiable words. The ANN was posited to ‘hallucinate’ when it recognized words during periods of input silence.Pruning of connections initially improved detection rates of phonetically degraded words. Higher levels of pruning however were associated with progressive impairment in word recognition and the emergence of hallucinations.Turning to the visual metaphor to describe what is happening see  we shoulPerceiving a word (like “Jane”) will change basins of attraction , and as a consequence the most likely trajectories that subsequently will be followed. Thus perceiving “Jane” will enlarge the basin of attraction of “kiss” and other words the ANN has learned to associate with “Jane” and at the same time shrink the basin of attraction of concepts associated with, for instance, “cop”. This situation is shown in As a result, after perceiving “Jane”, the most likely trajectories will lead to regions of state space associated with “Jane” (like “kiss” or whatever other words the ANN due to its prior expectations expects to follow “Jane”).The pruning process facilitates this process by improving the capacity to recognize degraded input, probably by enlarging basins of attraction and facilitating common trajectories (like “Jane” → “kiss” → “girl”). However, the drawback is that after a certain point recognizing becomes ‘too good’, and the system starts off to follow trajectories without any input: the system starts ‘hallucinating’.There is considerable evidence suggesting a role for dopamine involvement in schizophrenia, particularly motivated by the efficacy of antipsychotic medication, which derive their therapeutic effects from dopamine D2 antagonism An increasing body of research suggests that dopamine is a modulator of the signal-to-noise ratio of the neural system. One of the first to suggest this role were Servan-Schreiber et al Low levels of dopamine are associated with a low signal-to-noise ratio and in attractors terms this is equivalent to shallow attractors with large basins of attractions. In this situation frequent transitions from one attractor to another can take place see , bottom.High levels of dopamine are associated with a large signal-to-noise ratio and this comes down to the existence of a limited number of deep atractors with relatively large basins of attraction which dominate the information processing of the neural system , top. SuRecently, Kapur (1) presented an heuristic framework on the role of dopamine in delusions. In Kapur's model a central role of dopamine is to mediate the salience of environmental events. A  hyperdopaminergic state would lead, at a psychological level, to the aberrant assignment of salience to one's experience. Aberrant, because salience is assigned to experiences which (in fact) have none or only minor salience. Subsequently, delusions are the cognitive effort to make sense of aberrantly salient experiences.Kapur (1) provides a psychological description of the role of dopamine. Within the framework we presented (based on ‘information-processing with attractors’) an abormal hyperdopaminergic state is equivalent to the emergence of an attractor with a large basin of attraction which dominates the striatal information processing of the neural system. This dominance translates itself in ‘aberrant salience’. This is the situation depicted in Whereas hyperdopaminergic states in the striatum have been associated with delusions, reduced dopaminergic activity in the prefronal cortex has been associated with disorders of working memory Schizophrenia patients show several deficits in psychological tasks, specifically in tasks that place a demand on the active maintenance of internal representations of the context of the task. Cohen & Servan-Schreiber More recent insights have focused on the differential effects of D1- and D2- dopamine receptor stimulation in the prefrontal cortex The combined effects of D1- and D2-receptor stimulation cause dopamine to drive neural networks through a sequence of phases with opposing characteristics. In the case of prefrontal cortex this may consist of an initial phase where basins of attraction are flattened out  external influences or by limitations in the system itself. In most pathological circumstances, there will be a constant interaction between the system and the environment, resulting in a more or less pathological equilibrium.neurodynamicsIt has been suggested to name these aspects of neural functioning which focuses on the system properties of neural ensembles Our approach has been to simplify and visualize these processes, in order to produce an intuitive understanding.Ideally, medical thinking depends on the understanding of both physical reality and the availability of a metaconcept describing reality in a more abstract way. For instance, in cancer research, the metaconcept is ‘regulation of cell proliferation’, with ‘misregulation’ as its associated pathological state The line of reasoning introduced in this paper tries to approach this situation. The metaconcept defined is based on information processing theory  as instances of more general behavior (like the behavior of complex systems) and to construct a conceptual framework describing this general behavior.Finally, it should be borne in mind that theories and models need not be ‘explanations’ of observed phenomena but can also be useful as exploratory ‘heuristics’ This is an essay-like study aimed to contruct visualized concepts stemming from the theory on dynamical complex systems to be used as a ‘visual metaphor’ to bridge the mental- and molecular-biological domains in schizophrenia.The background for this study has been formed by a PubMed search with the terms  AND (‘schizophren*’ OR ‘psychosis’ OR ‘psychotic’ OR ‘psychiatr*’).The general concept of a  metaphor is inspired by the work of Globus & Arpaia To introduce the sought-after metaphor we first describe a computer model of neural information processing. We then show how the information processing in this model can be visualized, using concepts from the theory on complex systems. Finally, we use the visual metaphor to describe disturbed information processing and dopamine neuromodulation in schizophrenia.


This molecular chirality amplification in the LC provides a unique possibility for investigating the relationship between molecular structure, intermolecular interactions, and mesoscale organization. It is known that axially chiral or helical-shaped molecules with reduced conformational disorder are good candidates for high helical twisting power derivatives. In particular, biaryl derivatives are known to be efficient chiral inducers in biaryl nematic mesophases. In this paper, we focus on a new series of helicene-like molecules of known absolute configuration. We have integrated cholesteric pitch measurements with geometry optimization by DFT calculations and analysis of the twisting ability by the Surface Chirality model to shed light on the structural features responsible for the analogies and differences exhibited by these derivatives. The investigation of these dopants with well-defined geometry, by virtue of the low conformational freedom, and the substituents variously distributed around the core, allows us to extend our knowledge of the molecular origin of the chirality amplification in liquid crystals and to confirm the simple relationship “molecular P-helicity” → “cholesteric P-handedness” for helical-shaped helicene-like derivatives.The addition of a chiral non-racemic dopant to a nematic liquid crystal (LC) has the effect of transferring the molecular chirality to the phase organization and a  Nematic liquid crystals (LCs) are fluid phases formed by anisometric molecules which, though free to rotate as in ordinary liquids, are preferentially aligned along a common axis, called director. The addition of a chiral non-racemic dopant to a nematic liquid crystal has the effect of transferring the molecular chirality to the phase organization and a chiral nematic (or cholesteric) phase is formed, in which the director rotates perpendicularly to an axis in a helical way , which is defined as helicene quinones 6–8 has been described by Carreño, Urbano and coworkers [The enantioselective synthesis as well as the configuration assignment of enantiopure dihydro[5]helicene quinones or bisquinones oworkers –57.The twisting powers of the helicenes under investigation measured in the nematic solvent E7 are reported in P-helicity -7,8-Dihydro[5]helicene quinones or bisquinones 1–4 show moderate to high HTPs; despite the presence of the same tetracyclic core, derivative 5, with the tert-butyldimethylsilyl (TBDMS) blocking group, exhibits the lowest measured HTP. This is a confirmation of the fact that bulky substituents, though lacking centres of chirality (or other stereogenic elements), can strongly affect the molecule-to-phase chirality transfer. The data in 1–4, the [4]helicene quinones 6–8 have low twisting ability.We can see that all compounds possessing, as a common feature, homochiral lix axis ) follow elicenes . (P)-7,81–5, the naphthoquinone rings are approximately planar, with twist angles ranging between 42° and 43° for 1–4 and a slightly lower angle, around 40°, for 5. A larger twist angle of 50.1° is found in dihydro[4]helicene quinones 7 and 8, whereas a wider value, 81.7°, is allowed in tetrahydro[4]helicene quinone 6, because of the higher flexibility arising from the presence of an additional dihydro ring. In all derivatives, the core of the fused rings is conformationally constrained, and for most of them, even the introduced substituents do not provide high conformational freedom. The O–CAlkyl bond of alkoxy groups lies in the plane of the adjacent aromatic ring; for steric reasons, only the conformations shown in 2, 4, and 8, other structures, differing in the conformation of the ethoxy side chain, are also possible, but in view of their high energy, they have a negligible weight at room temperature. A different behavior is exhibited by derivative 5, which has bulkier TBDMS substituents; one of them is constrained by the proximity of the quinone ring, whereas the other is pointing towards the molecular periphery and is more free. Three structures with similar energy were obtained, differing essentially in the value of the torsional angle for the O–Si bond of the less hindered tert-butyl-silyl group. These structures are also displayed in The structure of the dopants under investigation was determined by DFT calculations at the B3LYP/6-31g** level ; the opt1–8 was analysed according to the SC method helicene quinone derivatives, which possess a wider extension of aromatic rings, capable of establishing stronger dispersion interactions with the host molecules. As to the effect of substituents, we can compare the results obtained for the pairs 1–2, 3–4, and 7–8, which differ by the replacement of a phenyl hydrogen by an ethoxy group. The measured HTPs are smaller for the derivatives with the ethoxy substituent, whereas the opposite change is predicted from the SC calculations; the discrepancy between calculations and experiments is especially evident in the case of derivative 8. These differences might indicate a role of electrostatic solute–solvent interactions, which are neglected in our model. It was already observed that these can be considered as a generally small, but non-negligible, correction to the underlying short-range interactions; their relative contribution can become relevant in the case of dopants with otherwise small twisting ability [According to our SC model results, the low HTP measured for Chirality is a peculiar molecular feature and its manifestations elude any trivial interpretation: different, often completely uncorrelated, responses depending on the experiment used are obtained in the attempt to quantify it.The helical twisting power cannot simply be correlated with a global stereochemical descriptor of the molecule, as demonstrated for homochiral series of propeller-like heptalenes and oligonaphthalenes for which the handedness of the induced cholesteric depends critically on the substituents attached to the chiral core –50.P-helicity” → “cholesteric P-handedness” exists for helicene-like compounds, in the absence of bulky and highly flexible substituents. Not surprisingly, taking into account molecular shape, the orientational behavior of 1–8 derivatives is analogous to that of bridged binaphthyls and also the helical twisting power can be interpreted in a similar way: the outcome is that pseudo-P dopants induce a right-handed (P)-cholesteric phase. The results obtained for the dopants investigated in the present work, with a clear molecular P-helicity and low conformational freedom, differing from each other in the presence of variously distributed substituents, confirm that short-range intermolecular interactions, parameterizable according to the molecular surface, are the main determinants of cholesteric induction in thermotropic liquid crystals. Other interactions, ascribable to the presence of electronegative groups, though present, are less relevant, and can have non-negligible effects in the case of dopants with low twisting ability.The investigation presented here confirms that, as already found for helicene derivatives , the simTi 60 °C). The standard error of the pitch determination is ca. 10%. The technique is described in detail in ref [Pitches and handedness of the cholesteric solutions in E7 have been obtained at room temperature using the lens version of the Grandjean-Cano method . E7 froml in ref .Q requires the molecular surface, which is generated on the basis of atomic coordinates. Since the chirality of the molecular surface strongly depends on the molecular geometry, accurate structures are needed to obtain reliable estimates of the twisting ability. The geometry of the molecules listed in Calculation of the chirality parameter S and the chirality parameter Q of dopants were calculated as explained in refs [−2 and the rolling sphere radius to 3 Å [rH = 1 Å, rC = 1.85 Å, rO = 1.5 Å, and rSi = 2.1 Å were used [−2 was assumed for the molecular surface.The Saupe ordering matrix  in refs –32. Once in refs ) was comere used . A densi


Vascular permeability factor/Vascular endothelial growth factor (VPF/VEGF), a multifunctional cytokine, is a potent inducer of vascular permeability, an important early step in angiogenesis. It is known that the neurotransmitter dopamine can inhibit VPF/VEGF mediated angiogenesis, in particular microvascular permeability, but the effectors of this action remain unclear.Here, we define the signaling pathway modulated by dopamine that inhibits VPF/VEGF induced vascular permeability in endothelial cells. Signals from VPF/VEGF lead to changes in the phosphorylation of tight junction protein zonula occludens (ZO-1) and adherens junction proteins like VE-cadherin and associated catenins, thus weakening endothelial cell-cell adhesion and increasing vascular permeability. We found VEGF receptor-2 (VEGFR-2) to be part of a multi-protein complex involving ZO-1, VE-cadherin and β-catenin. VPF/VEGF induced phosphorylations of VE-cadherin, β-catenin and ZO-1 were inhibited by dopamine treatment. Association of occludin with ZO-1 and ZO-1 with VE-cadherin were significantly inhibited by dopamine in VEGF treated cells. Furthermore, we identified Src as an important target for dopamine-mediated inhibition of VPF/VEGF induced permeability.Taken together, our results provide molecular insights of dopamine function in the vascular endothelium and suggest a central role of Src in regulating key molecules that control vascular permeability. Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is a multifunctional cytokine that plays a critical role in angiogenesis -3. It isVascular permeability, as induced by VPF/VEGF, is due to its action on the endothelium. Endothelial cell-to-cell junctions are complex structures formed by different adhesive molecules ,12. Endoin vivo  antibody were from Biosource International, Inc., CA; and AlexaFlour 546 secondary antibody was from Molecular Probes, OR. Aclar® fluoropolymer film was from Honeywell, NJ. Dopamine hydrochloride for in vitro, quinpirole and all other chemicals if not mentioned were from Sigma-Aldrich, MO. Dopamine hydrochloride for in vivo use was from Abbott Laboratories, IL. VPF/VEGF was from R &D Systems, MN. VEGF receptor 2 Kinase inhibitor IV was from Calbiochem (Cat # 676489).ZO-1 antibody (sc-10804), VE-cadherin antibody (sc-6458), β-catenin antibody (sc-7963), Occludin antibody (sc-8144), Src antibody (sc-5266) and VEGFR-2 (sc-504) antibody were from Santa Cruz Biotechnology, California, USA. Anti-phosphotyrosine monoclonal antibody (4G10) and phospho-Src [pTyrFive- to six-week-old NIHBNX-MMC mice were purchased from the Taconic Farms, Germantown, NY. The animals were caged in plastic tubs covered with stainless steel tops at a temperature between 23 and 25°C, humidity level of 50 ± 10%, and a 12:12-h light-dark cycle. The mice were given ad libitum water and food.3 HUVEC were seeded on collagen coated Lab-Tek chamber slides. After four days of culture when these cells were confluent, they were serum starved for 24 hours. Thereafter, these cells were treated with either dopamine (10 μM) or a specific dopamine D2 receptor agonist quinpirole (10 μM) [2 × 10 (10 μM) , followe (10 μM) ,38.HUVEC lysates in NP-40 buffer supplemented with protease inhibitor cocktail were immunoprecipitated with different antibodies (1:100 dilutions) and immunocomplexes were captured on protein A-agarose beads. Thereafter, samples were subjected to SDS-PAGE and then transferred to polyvinyl difluoride membranes and immunoblotted. Antibody-reactive bands were finally detected by enzyme-linked chemiluminescence (Amersham) and quantified by laser densitometry -23.5 HUVEC in 200 μL were seeded onto collagen coated inserts in 24 well plates. 700 μL complete EBM medium was added to the plates. The cells were allowed to grow for 4–5 days until a monolayer was formed. The medium was replaced with phenol free starving EBM medium containing 0.1% FBS for 12–14 hrs. The respective inserts containing HUVEC were then pre-treated with 10 μM Dopamine or Quinpirole for 15 min. Subsequently, FITC-Dextran containing medium (phenol free EBM) with or without VEGF at 10 ng/ml was added. Fluorescence was measured after 90 min on the Spectra Flour Plus using excitation and emission wavelengths of 485 nm/535 nm.The assay was performed according to established manufacturer's protocol, Catalog# ECM 640, Chemicon International Inc., CA. Briefly, 2 × 102 receptor agonist quinpirole (10 mg/kg ip) 10 min before VPF/VEGF injection. Briefly, normal mice previously treated with either dopamine or D2 receptor agonist was injected i.p. with 500 ng of recombinant human VPF/VEGF in 1 ml of HBSS. After 1 h, these animals were anesthetized with an injection of 0.4–0.75 mg/g ip Avertin and were euthanized by decapitation. The abdomen was then opened, loops of the small intestine were exteriorized, and mesentery windows including fatty vascular portions were dissected free and transferred to HBSS at 4°C. Four to six mesenteric windows, each roughly 1–3 cm2 in area, could be isolated from a single mouse.Animal protocols were approved by the Mayo Clinic Institutional Animal Care and Use Committee. Mesenteries were collected from normal 5–6 week-old NIHBNX-MMC mice and treated either with the anti-angiogenic dose of dopamine (50 mg/kg i.p) or specific dopamine D3 VO4, 2 mM EGTA, 1 mM phenylmethylsulfonyl fluoride, 10 ug/ml leupeptin, 0.5% aprotinin, and 2 mM pepstatin A for 10 s at 4°C. Homogenates were then centrifuged for 5 min at 4°C, and supernatants were immunoprecipitated with different antibodies (1:100 dilution), and immunocomplexes was captured on protein A-agarose beads. Finally, samples were subjected to SDS-PAGE and then transferred to polyvinyl difluoride membranes and immunoblotted. Antibody-reactive bands was detected by enzyme-linked chemiluminescence (Amersham) and quantified by laser densitometry [Thereafter, mesenteries were homogenized immediately upon collection with a polytron tissue homogenizer  in a precipitation assay buffer [50 mM Tris (pH 7. 5), 0.25% sodium deoxycholate, 1% NP40, 150 mM NaCl, 1 ml Naitometry . Since tt-test or Dunn's multiple-comparison test. Statistical significance was assigned when P < 0.05 [Differences among groups were evaluated by ANOVA and the unpaired Student's P < 0.05 -23. ErroVPF: Vascular permeability factor; VEGF: Vascular endothelial growth factor; VEGFR-2: Vascular endothelial growth factor receptor-2; HUVEC: Human umbilical vein endothelial cells; DA: Dopamine; D2DR: Dopamine D2 receptor; VE-cadherin: Vascular endothelial cadherin; ZO-1: Zonula occludens 1; TJ: Tight junctions; AJ: Adherens junctions.The authors declare that they have no competing interests.in vivo mouse models. All authors read and approved the manuscript.DM conceived the project. RB participated in the conception of the study, design and coordination and helped to draft the manuscript and performed some experiments with significant help from others as stated. SS performed majority of the experiments and helped to prepare the manuscript. Confocal microscopy was done by SPY. EW and SD performed most of the western blots. CP participated in cell culture and helped with


The WRAP53 protein regulates the formation and maintenance of Cajal bodies (nuclear sub-organelles), as well as directs the recruitment of nuclear factors to Cajal bodies. SMN1. This suggests that loss of WRAP53-mediated SMN trafficking contributes to spinal muscular atrophy.The WRAP53 gene gives rise to a p53 antisense transcript that regulates p53. This gene also encodes a protein that directs small Cajal body–specific RNAs to Cajal bodies. Cajal bodies are nuclear organelles involved in diverse functions such as processing ribonucleoproteins important for splicing. Here we identify the WRAP53 protein as an essential factor for Cajal body maintenance and for directing the survival of motor neuron (SMN) complex to Cajal bodies. By RNA interference and immunofluorescence we show that Cajal bodies collapse without WRAP53 and that new Cajal bodies cannot be formed. By immunoprecipitation we find that WRAP53 associates with the Cajal body marker coilin, the splicing regulatory protein SMN, and the nuclear import receptor importinβ, and that WRAP53 is essential for complex formation between SMN–coilin and SMN–importinβ. Furthermore, depletion of WRAP53 leads to accumulation of SMN in the cytoplasm and prevents the SMN complex from reaching Cajal bodies. Thus, WRAP53 mediates the interaction between SMN and associated proteins, which is important for nuclear targeting of SMN and the subsequent localization of the SMN complex to Cajal bodies. Moreover, we detect reduced WRAP53–SMN binding in patients with spinal muscular atrophy, which is the leading genetic cause of infant mortality worldwide, caused by mutations in  Cajal bodies, discovered more than 100 years ago by Santiago Ramón y Cajal, are sub-organelles found in the nucleus of proliferative cells and neurons. They have been implicated in a variety of nuclear functions including ribonucleoprotein maturation, spliceosome formation, histone mRNA processing, RNA polymerase assembly, telomerase biogenesis, and histone gene transcription. Concentrating relevant molecules within Cajal bodies may serve to increase the efficiency of specific nuclear functions. Here we identify the WRAP53 protein as an essential factor for Cajal body maintenance and for directing the splicing regulatory protein “survival of motor neuron” (SMN) complex to Cajal bodies. We show that WRAP53 is a constitutive component of Cajal bodies, and that knockdown of WRAP53 disrupts existing Cajal bodies and prevents formation of new Cajal bodies. Mechanistically, we find that WRAP53 recruits the SMN complex from the cytoplasm to Cajal bodies by mediating interactions between SMN, importinβ, and coilin. Finally, we report deficient WRAP53–SMN binding in patients with spinal muscular atrophy, suggesting a role in this pathology. This study not only reveals new functions of the WRAP53 protein, but also increases our understanding of the molecular mechanism behind Cajal body formation and recruitment of factors to Cajal bodies. WRAP53 as an antisense gene to the p53 tumor suppressor gene WRAP53 gives rise to a regulatory antisense transcript with a critical role for p53 function We previously discovered Cajal bodies are molecularly defined by the presence of the marker protein coilin. Coilin is essential for Cajal body integrity and function, and loss of coilin disrupts Cajal bodies. It has been proposed that coilin, upon oligomerization, provides a scaffold for the assembly of the different types of Cajal body components SMN1 gene cause the common neurodegenerative disorder spinal muscular atrophy (SMA), the leading genetic cause of infant mortality worldwide, which affects approximately one in 6,000 infants. A second copy of the SMN1 gene, SMN2, partially compensates for SMN1 loss. However, because of a single nucleotide change, most SMN2 transcripts lack exon 7, resulting in the production of the C-terminally truncated and unstable protein SMNΔC15 SMN2 and reduced number of nuclear structures containing the SMN protein (encoded by SMN2) The SMN protein is part of a large complex essential for the assembly of snRNPs in the cytoplasm n>300), clearly indicating that WRAP53 is a constitutive component of Cajal bodies  microscopy and Western blotting (WB). Two different small interfering RNA (siRNA) oligos targeting WRAP53 were used (siWRAP53#1 and siWRAP53#2), both knocking down WRAP53 mRNA with 90% efficiency . In contLoss of Cajal bodies was strictly associated with the degree of WRAP53 knockdown, where complete knockdown of WRAP53 led to the disappearance of all Cajal bodies, and cells still expressing low levels of nuclear WRAP53 showed Cajal body staining . We alsoWe next examined the effects on Cajal bodies in cells overexpressing WRAP53. Flag-tagged WRAP53 expressed at lower levels showed Cajal body accumulation . In contsnRNPs are known to be rate-limiting for Cajal body formation The finding that exogenous WRAP53 alters the localization of endogenous WRAP53, SMN, and coilin suggests a dominant negative effect of overexpressed WRAP53 that could be caused by WRAP53 self-interaction. Previous reports demonstrate such phenomena for coilin, where overexpressed coilin mislocalizes to nucleoli and disrupts Cajal bodies through dominant negative interference between exogenous and endogenous coilin IP of endogenous WRAP53 furthermore revealed that WRAP53 associates with coilin and SMN . ReciproTo investigate which region of WRAP53 mediates its localization to Cajal bodies, the panel of EGFP-WRAP53 deletion constructs was transiently transfected into U2OS cells, and protein localization was analyzed by IF. The cells were also stained for coilin to visualize Cajal bodies. Interestingly, only the WRAP53 constructs that bind coilin and SMN  accumulated in Cajal bodies . In contWe also observed that all WRAP53 constructs showed cytoplasmic localization  and thatPrevious studies failed to detect any WRAP53 protein in the cytoplasm To understand the organization of interaction between WRAP53, SMN, and coilin, we separately knocked down each of these proteins, and then performed IP analysis. This showed that WRAP53 co-precipitates coilin in SMN-depleted cells and co-precipitates SMN in coilin-depleted cells . Thus, WPrevious studies have described a direct binding between SMN and coilin We next analyzed whether knockdown of WRAP53 affects the cytoplasmic and nuclear distribution of SMN. Interestingly, a significant increase of SMN was found in the cytoplasm in WRAP53-depleted cells, which coincided with a decrease of SMN in the nucleus . This waInteraction with the nuclear import receptor importinβ is required for SMN nuclear import We next examined whether WRAP53 is required for directing other components of the SMN complex to Cajal bodies, and Gemin3 was chosen as a representative component. IP analysis showed that WRAP53 and Gemin3 indeed associate . DepletiWe also analyzed whether WRAP53 targets snRNPs to Cajal bodies. snRNPs consist of Sm proteins in complex with small nuclear RNAs and are carried by the SMN complex from the cytoplasm to Cajal bodies, where final maturation of the snRNPs takes place. In control cells the Sm proteins were enriched in Cajal bodies and distributed throughout the nucleus in speckles . In WRAPSMN2 allele is in fact weaker, we quantified the relative amount of SMN that interacts with WRAP53 in SMA patients and in normal cells. This revealed a lack of binding between SMN and WRAP53 in cells from SMA patients that cannot be explained by the lower amounts of SMN, and that the relative binding between WRAP53 and SMN in these cells was reduced by 83%  (data not shown). These observations, together with the fact that knockdown of WRAP53 or coilin disrupts all Cajal body structures, point to a more general function of WRAP53 in assisting coilin as a scaffold protein in Cajal body formation.Here we identify WRAP53 as an essential factor for Cajal body maintenance and for directing the SMN complex to Cajal bodies. We show that WRAP53 is a constitutive component of Cajal bodies that overlaps coilin in 100% of Cajal bodies in a variety of cell lines. Knockdown of WRAP53 disrupts Cajal bodies, prevents formation of new Cajal bodies, and relocates Cajal body proteins coilin and SMN from Cajal bodies to nucleoli. WRAP53 seems specifically important for Cajal body integrity, since depletion of WRAP53 does not affect gems or other nuclear structures, including nucleoli and PML bodies. We show that WRAP53 separately associates with coilin and SMN and is required for their complex formation. Previous studies demonstrated a direct interaction between SMN and coilin, suggesting that WRAP53 is not important for their binding but rather mediates interaction by bringing the proteins in close proximity. This may either occur within an already existing Cajal body or result in the formation of a new Cajal body. Importantly, WRAP53's role in Cajal body formation goes beyond bringing SMN and coilin together, since knockdown of SMN does not abolish all Cajal body structures. Residual Cajal bodies containing WRAP53 and coilin still remain. Moreover, in HeLa-PV cells, WRAP53 and coilin localize to all Cajal bodies and SMN to only 40% of them  and one heterozygous carrier  were obtained from Coriell Cell Repository and maintained in MEM supplemented with 10% fetal bovine serum. The GM03815 cells were initially described in the Coriell Cell Repository as derived from a heterozygous carrier . However, snRNP deficiency and genetic linkage analysis of these cells showed that they in fact are derived from a homozygous carrier with SMA type I  U2OS, H1299, HCT116, HEK293, MCF-7, HeLa-PV, HeLa-ATCC, and HDF cells were maintained in Dulbecco's modified medium supplemented with 10% fetal bovine serum (Invitrogen), 2 mM L-glutamine (Invitrogen), and 2.5 µg/ml Plasmocin (InvivoGen) at 37°C in 5% COTotal RNA was extracted from tumor cell lines using the Trizol reagent (Invitrogen). The RNA was reverse transcribed with First-Strand cDNA synthesis using Superscript II (Invitrogen). Quantitative real-time PCR was carried out in the Applied Biosystems 7500 Real-Time PCR using transcript-specific TaqMan Gene Expression Assays (Applied Biosystems). The following probes were used: Hs01126636_g1 for detection of WRAP53 and Hs00167441_m1 for detection of ALAS1 as endogenous control.3). Cover slips were subsequently incubated for 1 h in primary antibody and 40 min in secondary antibody diluted in blocking buffer. The cover slips were mounted with Vectorshield mounting medium with DAPI (Vector Laboratories). Images were acquired with a Zeiss Axioplan 2 microscope, equipped with an AxioCam HRm camera using 43× or 60× oil immersion lenses, and processed using Axiovision Release 4.7.For IF experiments, cells were grown on sterilized cover slips, fixed with 100% MeOH at −20°C, permeabilized with 0.1% Triton X-100 for 5 min, and then blocked in blocking buffer  for 15 min on ice, followed by sonication 2× for 10 s. Extracts were spun down at 6,000 rpm for 5 min at 4°C and then quantified by Bradford assay (Bio-Rad). Endogenous proteins were immunoprecipitated with 1 µg of affinity-purified antibody per 1 mg extract supplemented with 10 µl of Dynabeads Protein G (Invitrogen) overnight at 4°C. The beads were washed 4× for 15 min with 1 ml of NP40 buffer and prepared for WB.Cell extracts for WB analysis were prepared as previously described Cell fractionations were performed using a nuclear extract kit according to the manufacturer's instructions . For fractionation followed by IP, cells were lysed in hypotonic buffer  for 20 min on ice. Samples were spun down at 3,600 rpm for 10 min at 4°C, and the supernatant (cytoplasmic fraction) was converted into NP40 buffer. The remaining pellet was lysed in NP40 buffer. The IP was performed as described earlier. In Four different WRAP53 antibodies were used: rabbit α-WRAP53-C1 The following siRNAs from Qiagen were used: siWRAP53#1 (SI00388941), siWRAP53#2 (SI00388948), siSMN1#7 (SI03108084), siSMN1#11 (SI04950932), siSMN1#12 (SI04950939), siCoilin#3 (SI00350343), siCoilin#7 (SI04330830), and a control siRNA (1027280). siRNA (10–20 nM) was transfected into cells using either Oligofectamine (Invitrogen) or HiPerfect (Qiagen) transfection reagents, in accordance with the supplier's recommendations. Flag-WRAP53 was cloned as described in Mahmoudi et al. For the generation of cells with stable overexpression of WRAP53, Flag-tagged full-length WRAP53 cDNA was cloned into the pLenti6/V5-D-TOPO vector (Invitrogen). The pLenti6-Flag-WRAP53 plasmid along with the pMDLg/RRE, pCMV-VSVG, and pRSV-Rev plasmids required for viral production were transfected into HEK293FT cells using Lipofectamine 2000 (Invitrogen). U20S cells were infected with viruses containing either pLenti6-Flag-WRAP53 or empty pLenti6/V5-D-TOPO vector, and positive cells were selected 48 h after infection using 10 µg/ml Blasticidin (Invitrogen).t test was used to determine statistical significance.All analyses were performed using Microsoft Office Excel 2003. Two-tailed Student's 2 fragment (Jackson ImmunoResearch) was added in order to counterstain for WRAP53. Images were acquired using an epifluorescent microscope  equipped with a 100-W mercury lamp, a CCD camera , emission filters for visualization of DAPI, FITC, and Cy3.5, and a 63× objective (plan-neofluar). WRAP53 staining was used to select image position.In situ PLA experiments were performed as described previously Figure S1WRAP53 and coilin, but not SMN, are essential for Cajal body integrity. (A) Quantitative real-time PCR analysis of U2OS cells treated with the indicated siRNAs for 48 h. (B) WB analysis of WRAP53, coilin, and SMN levels in U2OS cells treated with the indicated siRNA oligos for 48 h. β-actin was used as loading control. (C) IF staining of SMN, coilin, and WRAP53 in HeLa cells treated with siControl and siWRAP53#1 oligos for 48 h. Nuclei were stained with DAPI in all IF experiments. (D) Immunostainings of U2OS cells treated with the indicated siRNA oligos for 48 h, followed by staining with fibrillarin, SMN, or PML. Arrows indicate nucleoli in the fibrillarin staining and a gem in the SMN staining. (E) IF staining of WRAP53, SMN, and coilin in U2OS and HeLa cells treated with siControl and siCoilin#7 oligos for 48 h. (F) IF staining of WRAP53, coilin, and SMN in U2OS and HeLa cells treated with siControl, siSMN1#7, or siSMN1#11 oligos for 48 h.(3.15 MB TIF)Click here for additional data file.Figure S2High exogenous expression of WRAP53 disrupts Cajal body structure. (A) Immunostaining of U2OS cells transiently transfected with Flag-WRAP53 for 16 h, followed by staining with Flag- and WRAP53-specific antibodies. (B) U2OS cells transiently transfected with EGFP-tagged full-length WRAP53 for 16 h and stained for coilin or SMN. (C) WB analysis of WRAP53, coilin, and SMN levels in U2OS cells overexpressing Flag-tagged WRAP53. β-actin was used as loading control. (D) IF staining of Flag and PML in U2OS cells transiently transfected with Flag-tagged WRAP53 for 16 h. Arrows indicate an untransfected cell.(1.84 MB TIF)Click here for additional data file.Figure S3WRAP53 is expressed in the cytoplasm and in Cajal bodies. (A) WB analysis of the four WRAP53 antibodies mentioned in the paper. Full-length filters are shown to demonstrate the specificity of the WRAP53 antibodies. The rabbit α-WRAP53  antibody was used in all IF stains shown in the main figures. This is also the same antibody employed by Tycowski et al. (1.30 MB TIF)Click here for additional data file.Figure S4Stable Flag-WRAP53 cells with high WRAP53 expression lack Cajal bodies. (A) IF analysis of U2OS cells stably transfected with either Flag–Empty vector (Mock cells) or Flag-WRAP53 (Flag-WRAP53 cells) stained for WRAP53 and coilin. (B) WRAP53 interacts with SMN both in the cytoplasm and in the nucleus. In situ PLA of WRAP53–SMN interaction in WRAP53-depleted U2OS cells as negative control. A clear reduction in in situ PLA signals was observed in siWRAP53 compared to siControl, confirming the specificity of detection in (1.34 MB TIF)Click here for additional data file.Figure S5Aberrant expression of WRAP53 leads to mislocalization of Gemin3 and Gemin2. (A) WB analysis of WRAP53 and SMN levels in fractionated U2OS cells treated with the indicated siRNA oligos for 48 h. Lamin A/C and Hsp90 were used as nuclear and cytoplasmic markers, respectively. (B) IP of endogenous WRAP53 from U2OS cells pretreated with the indicated siRNA oligos for 48 h. (C) IP of endogenous SMN from U2OS cells pretreated with the indicated siRNA oligos for 48 h. (D) IF of U2OS cells treated with the indicated siRNA oligos for 48 h, followed by staining with Gemin3-  and WRAP53-specific antibodies. Arrows indicate a gem. (E) IF staining of WRAP53 and Gemin3  in U2OS cells transiently transfected with Flag-tagged WRAP53 for 16 h. Arrows indicate an untransfected cell. (F) IF of U2OS cells treated with the indicated siRNA oligos for 48 h, followed by staining with SMN-, Gemin2-, Gemin3- , and WRAP53-specific antibodies.(3.60 MB TIF)Click here for additional data file.Figure S6WRAP53-depleted cells show altered localization of Sm but no change in intracellular distribution of Sm. (A) IF of U2OS cells treated with the indicated siRNA oligos for 48 h, followed by staining with Sm-, coilin-, and WRAP53-specific antibodies. (B) WB analysis of Sm levels in fractionated U2OS cells treated with the indicated siRNA oligos.(1.07 MB TIF)Click here for additional data file.Table S1PCR primers for cloning of EGFP-WRAP53 and Flag-SMN constructs.(0.71 MB TIF)Click here for additional data file.


Affiliation 1 is incorrect. It should read: Department of Molecular and Cellular Interactions, VIB, Brussels, Belgium


Sir,The paper ‘Comparison of norepinephrine and dopamine in the management of septic shock using impedance cardiography’ in the D


Aequidens pulcher undergoes a transformation of its optical properties every dawn and dusk as the eye adapts to changes in light conditions. During dusk the transformation result in an increase of the refractive power in the lens cortex, the outermost 40 percent. The change is thought to match the optical properties of the lens to the requirements of the retina. Using a short term in vitro lens culturing system together with optical measurements we here present data that confirm that the optical properties of the lens can change within hours and that dopamine influences the optical properties of the lens. Dopamine yields dose-dependent decrease of the refractive power in the lens cortex. The D1-agonist SKF-38393 induces a similar decrease of the refractive power in the cortex, while the D2-agonist quinpirole has no effect. The effect of dopamine can be blocked by using the D1-antagonist SCH 23390. Our results suggest that dopamine alone could be responsible for the light/dark adaptive optical changes in the lens, but the involvement of other signaling substances cannot be ruled out.The crystalline lens in the cichlid fish  Vertebrate eyes share a common ancestry and the general design appears to have been preserved over the past 500 million years. Similarities between vertebrate groups are present in both general morphology and development of the retina and lens as well as on a cellular level, for instance the photoreceptors and their opsins max. This creates a well-focused image on the retina composed of relevant wavelengths despite chromatic aberration Chromatic aberration arises from the same principle by which a prism divides white light into its spectral components: refraction is wavelength-dependent where longer wavelengths are refracted less than shorter. The defocusing effect of chromatic aberration is most severe in powerful lenses with large apertures, because such lenses have short depth of focus. Although multifocal optical systems occur throughout the vertebrate linage, it is best understood in teleosts (bony fishes) where the crystalline lens is the sole refractive element equidens pulcher, a South American cichlid, has its three spectral cone types in the focal plane during day that are replaced at dusk by one set of rods In most teleosts retinomotor movements result in a dramatic change in the outer layers of the retina during light/dark adaptation. The process is composed of two parts; a shift in position between the cone and rod inner/outer segments, and a migration of melanin granules in the retinal pigment epithelium. Circadian changes in the retina are controlled not only by light intensity variations, but also by endogenous signaling substances. Some changes are specifically activated by fluctuations in dopamine level, which acts as a signal for both light-induced and circadian endogenous light adaptation of the eye. Ocular dopamine is produced solely by one type of interplexiform cells in the retina from where it diffuses freely into the vitreous max changes. The optical properties of the lens change from multifocal during day towards monofocal during night, which is thought to compensate for the change occurring in the retina from three active photopigments to one. The change in the lens results in an increase of the refractive power in the outermost 40 percent of the lens radius. A similar, but stronger increase of the refractive power in this region occurs if the eye is depleted of dopamine, which suggests that dopamine is directly or indirectly involved in the regulation The lens optical properties change as the eye dark-adapts to compensate for the change occurring in the retina during dusk, where the animals' wavelength discrimination ability (color vision) disappears and λin vitro short-term culturing technique and treated the lenses with dopamine and dopamine analogs in order to determine whether dopamine is directly responsible for the changes occurring during light-dark adaptation and dopamine depletion. We found that dopamine influenced the lens directly and decreased the refractive power in the outermost 40 percent of the lens. Furthermore, we gained insight into what mechanism regulates the changes in refractive power by determining that the D1 receptor family is directly involved in the regulation while the addition of D2 family agonists had no effect.We used an i.e. the outermost 40 percent of the lens radius. Treating lenses in vitro with dopamine, at concentrations from 10−6 to 10−3 M, yielded a dose-dependent response   .−4 M of the D1-agonist SKF-38393 were similar to those where dopamine was used. In contrast, ΔBCD curves from experiments with 10−4 M of the D2-agonist quinpirole were similar to curves from the control group  .in vitro with dopamine leads to a decrease in refractive power in the outermost 40 percent of the lens cell layers. The reduction in refractive power increases with increased dopamine concentration in a dose-dependent manner within the concentration range tested.During light adaptation the refractive power of the lens cortex, the outermost 40 percent, is reduced. Dopamine levels are roughly three times higher in the vitreous during day than night and if dopamine is directly controlling the lens' refractive power, treatment with dopamine should induce a reduction of the refractive power in the same layers. The results obtained in this study are consistent with the hypothesized scenario; treatment of lenses A1, D1A2, D1B, and D1C receptors in teleosts 3, and increased activation of protein kinase C, PKC, through diacylglycerol. In mammalian lenses, PKC appears to have gap junction regulatory properties in the epithelial cells and could possibly therefore also affect the lens circulation 2+ by itself influences the water permeability of the aquaporin AQP0 in the lens cortex, which in turn might affect the lens circulation The D1-receptor family, consisting of the D1Our results are consistent with that dopamine alone may be responsible for the optical change in the lens occurring between night and day. If so, the lens would be in its natural state during night and the increased dopamine levels during day would decrease the refractive power in the lens periphery until dusk when the dopamine levels drop again. In the retina, however, several compounds work together and form a push-pull system that transforms the retina during dusk and dawn Dopamine directly induces a decrease in refractive power in the outermost 40 percent of the lens cell layers and acts through a D1 receptor family regulated pathway. The change is similar to that occurring between night and day.Malmö/Lunds djurförsöksetiska nämnd.All experiments involving animals were approved by the regional ethical committee for animal research, Aequidens pulcher, where obtained through a local distributor and kept in aquaria under 12h-12h light-dark cycles. The animals used in the experiments were isolated 24h prior to the experiments by translucent dividers in their aquaria to avoid damage to the eyes and effects caused by stress and aggression. All experiments were initiated 3 hours after subjective daybreak to exclude circadian influences.Fishes, 2, 1 mM MgCl2, 10 mM glucose and 3 mM HEPES. The pH was adjusted to 7.3 and osmolarity set to match that of the vitreous (330 mOsm) by using a slightly more concentrated solution.The animals were killed through pithing and the lenses were excised immediately. The lenses where cultured for four hours at room temperature, 20–22°C, in a modified H10 medium, 120 mM NaCl, 2.50 mM KCl, 0.80 mM CaClDuring culturing, both lenses remained untreated in the control group. In the experimental groups, one lens from each animal was treated, while the other lens remained untreated as internal control. Right and left lens was alternated as control. Lenses were treated with either dopamine hydrochloride, the D2 agonist (±)-quinpirole dihydrochloride, the D1-agonist (±)-SKF-38393 hydrochloride, or dopamine hydrochloride in combination with the D1-antagoinst (+)-SCH-23390 hydrochloride. All substances were obtained from Sigma Aldrich and dissolved in culturing medium. All substance concentrations were obtained from the literature. The concentration of the D2-antagonist was increased due to lack of response at the initial concentration The lenses' optical properties were determined with laser scanning that measures the back center distance (BCD) at each beam entrance position (BEP). BEP is the lateral distance from the optical axis to where the laser beam enters the lens and BCD is the longitudinal distance from the center of the lens to the where the beam deflected by the lens intercepts the optical axis. BCD as a function of BEP gives a longitudinal spherical aberration (LSA) curve representing the variation in refractive power over the lens radius. Since the lenses are spherically symmetric, the results from both halves of the lens diameter were averaged, which removes artifacts that occur because of small errors in the position of the optical axis. More in-depth descriptions have been published previously To reduce the impact of individual variation, the LSA curve from the untreated lens (control) was subtracted from the LSA curve from the treated lens in each animal. This yielded a ΔBCD curve that describes the difference in BCD between right and left lens. Untreated lenses from the same animal had very similar LSA curves and any deviation from 0 in the ΔBCD curve therefore indicated the effect of the treatment. For further comparisons the maximum deviation, max ΔBCD, was measured in each ΔBCD curve. The max ΔBCDs from the different treatments were used in statistical analyses. First, linear regression was used on the max ΔBCD from the dopamine concentration series and the statistical significance between the slope and 0 was tested with ANOVA (n = 34). Second, permutational ANOVA (Primer v6.0) was used to test for any statistical difference in max ΔBCD between agonists and control (n = 31). Finally, permutational ANOVA was also used to compare max ΔBCD from lenses treated either with dopamine and antagonist or only dopamine (n = 21).