<p>Mutations in the gene SHANK2, which encodes a synaptic scaffolding protein, have been shown to cause a spectrum of neuropsychiatric disorders including: intellectual disability, autism spectrum disorders (ASDs), bipolar disorder (BD), and schizophrenia. However, many aspects of SHANK2 including the array of isoforms expressed, the expression pattern of the protein, biochemical and regulatory mechanisms, and in vivo protein function remain elusive. This body of work aims to uncover the function of the SHANK2 gene and its role in neuropsychiatric disorders using in vitro and in vivo experimental systems.</p> <p>Using a molecular genetics approach, I revealed the transcript architecture of the mouse Shank2 gene including characterization of promoters, isoforms and protein domains. I then outlined the temporal and spatial pattern of the Shank2 isoform expression throughout development. To further explore the protein’s function, we sought to identify novel SHANK2 interacting proteins using a yeast-2-hybrid screen and characterized the interacting proteins. Lastly, in order to understand how Shank2 deficiencies alter brain function we generated and characterized both Shank2 conventional (∆e24) and conditional mutant mice (e24floxed) by deleting or floxing exon 24 that encodes the Homer binding site and has nonsense mutations in human patients with neuropsychiatric disorders.</p> <p>Collectively, these studies 1) provide insight into the transcriptional regulation of Shank2 during brain development; 2) support the value of using Shank2 to further dissect the pathophysiology and circuitry mechanism underlying manic and autism like behaviors; 3) offers a novel mechanistic link between ubiquitination-mediated protein modification and SHANK2 function that may elucidate the molecular basis underlying SHANK2-related neuropsychiatric disorders. Ultimately, these findings may lead to the development of new therapeutic interventions for SHANK2-related neuropsychiatric disorders.</p> / Dissertation

The basic helix-loop-helix PAS domain transcription factor gene NPAS3 is a risk factor for psychiatric disorders. A knockout mouse model also exhibits behavioural and adult neurogenesis deficits consistent with human illness. To define the location and mechanism of NPAS3 aetiopathology immunofluorescent and transcriptomic approaches were used. Npas3 was co-localised with Dcx, but not other neurogenesis markers, in the hippocampal subgranular zone - the site of adult neurogenesis. This implied that NPAS3 might be involved in maturing, rather than proliferating, neuronal precursor cells. Microarray analysis revealed that the transcriptional activities of NPAS3 and its truncated form (C-terminal deletion) in the HEK293 cell line are sensitive to circadian rhythm context. The most highly up-regulated NPAS3 target gene, VGF, encodes secretory peptides with established roles in neurogenesis, depression and schizophrenia. VGF was one of many NPAS3 target genes also shown to be regulated by the SOX family of transcription factors, suggesting an overlap in neurodevelopmental pathways. The transcriptional repression of multiple glycolytic genes indicated that NPAS3 has a second role in metabolic regulation. This finding was also confirmed by collaboration with a metabolomics research group at the University of Strathclyde. SOX11, a transcription factor known to play a role in neuronal and glial cell differentiation, was shown to be down-regulated by NPAS3. The set of genes targeted by SOX11 and their ontologies were deduced by a microarray analysis in a SOX11 overexpressing HEK293 cell line. Regulated genes include a previously established SOX11 target, known markers of neurogenesis as well as genes implicated in neuropsychiatric disorders. Multiple histone and zinc finger genes are regulated by SOX11, many of which were located in two clusters on chromosomes 6 and 19. The chromosome 6 cluster lies within a region of the genome showing the strongest genetic association with schizophrenia. SOX11 may alter localised expression competence and its targets induce a complex programme of chromatin remodelling and downstream gene expression changes to achieve the mature neuronal phenotype. This thesis details how transcription factors are involved in biological processes linked to psychiatric illness. The dual neurodevelopmental and metabolic aspects of NPAS3 activity described here increase our understanding of aspects of neurogenesis relevant to mental illness and may explain the innate and medication-induced susceptibility to diabetes reported in psychiatric patients.

This study explores the effectiveness of “In-Sight”, a holistic recovery-based group training for people with bipolar disorder. Drawn from professional therapies and lived experience of the user-researcher who is diagnosed with having the illness, the training was delivered over 12 weekly sessions. Within a case study approach, an experimental design incorporated pilot (eight participants), main study (five) and control groups (six). Participants with a wide range of DSM-IV bipolar disorders were sought. Three non-bipolar participants experiencing severe mood swings were maintained in the pilot. One bipolar pilot participant later trained as the main study group co-facilitator in delivering the training. Self-report scales measured mood, coping, empowerment and quality of life pre-, post- and six months post-training. Semi-structured interviews noted individual change within the same time frame. Mental health professional interviews, medical note analysis and user-researcher observations also informed the study. Findings from self-report questionnaires indicated promising evidence that participants experienced improved mood stability, symptom severity, coping and quality of life and greater empowerment. A model has emerged to illustrate the various course components that appeared to benefit participants (user-led approach, self-help iii group format, illness management techniques, interpersonal skills development, healthy lifestyle, structure and planning, and intellectual change of perspective). The model illustrates how these components appeared to impact upon coping strategies, in turn leading to greater mood stability, maintaining wellness and personal development resulting in greater empowerment, improved outlook on self and the future and improved quality of life over time. The three non-bipolar pilot participants also appeared to benefit from the training compared to controls albeit to a lesser degree than the bipolar participants. The co-facilitator showed evidence of the greatest improvement. Four controls indicated evidence of continued use of poor coping and risk of further deterioration in illness, of whom two experienced major episodes with psychosis. Two controls indicated slight improvements. The “In-Sight” training is proposed as a step towards a more comprehensive approach for recovery from bipolar disorder informing the development of a new Expert Patients Programme in the UK.

Background: The Behavioural Activation System dysregulation theory of Bipolar Disorder (BD) proposes that (hypo)manic episodes represent prolonged periods of elevated approach-motivation and high-activation positive affect (PA). Excessive goal-setting behaviour and increased engagement in stimulating activities have been found amongst people with BD and may interact with elevated approach-motivation, contributing to an “upward spiral”. Hypotheses were: both i) approach-related behaviours in response to PA; and ii) high-activation PA, will be more common in individuals with BD than those without; iii) individuals with BD will be more likely to respond to high-activation PA than to low-activation PA with approach-related behaviours, compared to individuals without BD. Method: Individuals with BD and a non-clinical control group were tested. Participants completed measures of current hypomanic/depressive symptoms, trait PA, and two versions of two measures of response to PA, asking about low-activation and high-activation PA. Results: The BD group used more approach-related responses to PA, particularly within high-activation mood states, compared to controls. The groups differed in their experience of different types of PA, due to the control group experiencing more low-activation PA. Limitations: It is unclear whether the findings are specific to BD or affective disorders in general, due to a lack of a clinical control group. The groups may also differ in their expectations of PA due to medication use and previous experience of mania. Conclusions: It may be therapeutically beneficial to help individuals with BD use non-approach-related strategies in response to high-activation PA. Further research would identify which strategies are most useful.

The purpose of this study proposal is to examine the potential relationship between linguistic creativity and mood disorders, specifically depression and bipolar disorder. Participants will be approximately 67 adults who have either bipolar disorder, major depressive disorder, or serve as a healthy control group. Participants will complete prompts in order to measure linguistic creativity and then fill out several questionnaires relating to depressed mood, mania, general creativity, and rumination levels. It is predicted that bipolar disorder will have higher levels of certain types of linguistic creativity, such as lexical and semantic creativity, whereas depression will have more syntactic creativity. Furthermore, it is anticipated that higher rumination levels in the depressed group will be associated with higher levels of linguistic creativity, as opposed to participants in the depressed group with lower levels of rumination. Lastly, it is predicted that the type of writing prompt will influence the amount of creativity exhibited by each participant group. The proposed study has implications for therapeutic benefits, the emergence of a new area of research in two separate fields, and a new way of analyzing shifts in speech patterns of those with mood disorders.

Bipolar disorder and borderline personality disorder are common psychiatric diagnoses. One is a mood disorder with a strong genetic basis while the other is a disorder of personality commonly related to abusive experiences in childhood. Despite contrasting aetiologies they can be difficult to differentiate because of overlapping clinical presentations and symptoms. Diagnostic accuracy is important because of their polarised treatment approaches: long term treatment with mood stabilizers for bipolar disorder and psychotherapy for borderline personality disorder. A qualitative study of psychiatrists revealed comprehensive knowledge of the diagnostic criteria however, many expressed the view that diagnostic criteria did not assist diagnostic differentiation. These findings were validated in a large electronic survey of UK psychiatrists. A detailed study of actual diagnostic processes revealed that this scepticism appeared to influence actual practice. Clinicians largely ignored diagnostic criteria but continued to give diagnoses. Age and IQ matched women with bipolar disorder, borderline personality disorder and a healthy control group were compared in a series of cognitive tasks. Borderline personality disorder was associated with a failure to establish and maintain reciprocal cooperation in a game theoretic measure of social exchange. This behavioural change was not seen in euthymic bipolar disorder. Borderline personality disorder was also associated with an insensitivity to reward and losses in a risky decision-making task. Using a simple two-choice reaction task post error slowing was significantly amplified in the borderline group despite overall reaction times and error rates being similar in all three groups. Clinical diagnostic practice as revealed in this study is not adequate to reliably differentiate between bipolar disorder and borderline personality disorder. Laboratory measures of social exchange, decision making and post-error slowing highlight fundamental difficulties in borderline personality disorder not seen in euthymic bipolar disorder. These findings support the differentiation of bipolar disorder from borderline personality disorder and offer translational models for developing and evaluating new treatments for borderline personality disorder.

O Transtorno Afetivo Bipolar (TAB) é um transtorno mental crônico e recorrente. Está situado entre as principais doenças que causam incapacitação e morbidade em todo mundo. A substituição do modelo de atendimento ao doente mental centrado no hospital, por uma rede de serviços prestados na comunidade, tem como conseqüência o aumento do número de pacientes vivendo em família. Estas famílias são solicitadas a oferecer apoio emocional, tolerar estigma, lidar com perdas financeiras e laboral, interrupções na rotina do lar, entre outras. Tais implicações correspondem à sobrecarga que são as conseqüências que o familiar enfrenta ao conviver com pessoa portadora de transtorno mental. O objetivo deste estudo foi identificar e descrever a sobrecarga do TAB nas famílias que convivem com algum portador deste transtorno. Foram entrevistados 15 familiares, sendo três filhos, quatro irmãs, duas cunhadas, uma ex- cunhada, três cônjuges, uma tia e uma mãe. Foram utilizados para a coleta de dados um questionário com informações sobre dados clínicos do paciente com TAB, Critério Padrão de Classificação Econômica Brasil 2008 e roteiro de entrevista semi-estruturado adaptado por Koga (1997) que engloba a sobrecarga familiar. As respostas foram analisadas utilizando-se a técnica de análise de conteúdo de Bardin. O impacto do TAB sobre a família foi descrito nos seguintes aspectos: trabalho, gastos, lazer, relacionamento interpessoal, futuro, recursos e adoecimento do familiar como expressão de sobrecarga. Observou-se que a família se sente mais sobrecarregada nos períodos de crise da doença. Nos períodos estáveis somente os aspectos trabalho e relacionamento interpessoal permanecem como ponto de preocupação e sofrimento familiar. O fato de não se conhecer as causas e tampouco a cura, faz com que apareçam insegurança e temor quanto ao futuro. Este estudo contribuiu para compreender que TAB também sobrecarrega a família que necessita e solicita apoio dos profissionais de saúde para obter melhora na qualidade de vida. A principal limitação do presente estudo é o número pequeno de familiares entrevistados e o fato de todos terem sido recrutados em um único serviço de saúde. Pesquisas semelhantes deverão ser realizadas a fim de verificar se outras famílias manifestam sobrecarga semelhante as apresentadas pelos familiares deste estudo. / The bipolar disorder (BD) is a chronic and recurring mental disorder. Nowadays, it is one of most important disease that is responsible for incapacity and morbidity worldwide. The replacement treatment model of mental illness that was focus at hospital for services accessible to community is responsible for increase of number of patients living in family contexts. These families are responsible to offer emotional support, sometimes tolerate stigma, deal with financial and work loses, disruption at home routine and other ones. These implications are the result of relationship with a person with mental illness. The aim of this study was to identify and describe the burden of bipolar disorder in the family that lives with a person that has this disorder. Questioners were applied for fifteen relatives: tree children, four sisters, tree sistersin- law, tree spouse, 1 aunt and 1 mother. The data were collected using questionnaires with information about clinic data of BD patients, Standard Economic Classification Brazil 2008 and semi structured interview adapted by Koga. The results were analyzed by Bardins content technique. The impact in the family resulting of the relationship with BD patients was described following these aspects: work, costs with daily life, leisure, interpersonal relationship, future, resource and falling ill like expression of burden. As a result BD patients family reported that they were burdensome in the severe periods of disease. In the stable periods, only work and interpersonal aspects are continuous factors of worry and familiar suffer. Ignore the causes and the cure emerge insecurity and fear about the future. This study contributed to comprehension of that BD also burdensome the family of the patients. In addition, this people require support of health professionals to provide quality of life. The limits of this study were the short number of relatives submitted to questioners and they were come from the one source. More studies will be necessary to analyze other families and the pattern of BD illness burdensome.

O Transtorno Bipolar (TB) é uma doença psiquiátrica grave, altamente incapacitante que está associada com diversas comorbidades médicas e altas taxas de suicídio. Embora sua fisiopatologia não esteja completamente elucidada, inúmeros estudos têm mostrado alterações no sistema imune de indivíduos com TB. A resposta crônica destes indivíduos ao estresse parece gerar um aumento da inflamação sistêmica bem como da neuroinflamação. A micróglia ativada devido aos estímulos inflamatórios contínuos deve ocasionar diferentes prejuízos tanto bioquímicos quanto funcionas. Os macrófagos, primeira linha de defesa, são células de característica plástica de extrema importância do sistema imune e podem ser estimulados a polarizar para diferentes formas com liberação de fatores pró e antiinflamatórios, estimulando ou mantendo a homeostase no ambiente agredido de alguma forma. Desta forma, nosso trabalho buscou investigar a resposta fenotípica dos macrófagos contra o meio ambiente pró-inflamatório sistêmico observado no plasma de pacientes bipolares eutímicos, maníacos e depressivos em comparação aos controles. A amostra incluiu 5 controles saudáveis, 8 pacientes bipolares remetidos, 5 pacientes maníacos e 5 pacientes depressivos. As citocinas e quimiocinas de RNAm em células U937 tratadas com plasma mostraram um padrão de expressão diferente relativo entre controles saudáveis e pacientes com TB. As citoquinas inflamatórias tais como IL-1β e TNF-α, em pacientes bipolares maníacos e depressivos demonstram maiores quantidades de IL-1β mRNA do que os pacientes eutímicos e pacientes depressivos induziram maiores quantidades de RNAm de TNF-α do que os pacientes eutímicos em células U937. Já a expressão das quimiocinas CXCL9 e CXCL10 no plasma de pacientes com TB depressivos, demostraram ser de menor expressão significativa no grupo de pacientes maníacos quando comparados a controles e pacientes bipolares eutímicos. Nossos resultados sugerem que as citocinas periféticas devem modular a polarização M1 ou M2 de macrófagos no TB. / Bipolar Disorder (BD) is a severe and highly incapacitating psychiatric disorder which is associated with the presence of medical comorbidities. The progression of BD is related to an important cognitive deficit and also to biological and clinical manifestations that lead to treatment resistance and worse prognosis. Immune disturbances have been widely observed and investigated in BD patients. Chronic inflammatory responses induce neuroinflammation, mainly by pro-inflammatory microglial activation, and result in biochemical and functional impairment. Macrophages are the first line of defense of the immune system and exhibit cell plasticity. As well, microglia represents the resident macrophage of the central nervous system been responsible for its protection. Both cells can be stimulated to polarize into two different phenotypes, mainly pro- and anti-inflammatory, maintaining the homeostasis under physiologic and pathologic conditions. Therefore, we aimed to investigate macrophages phenotypical response when submitted to BD patients plasma in different episodes, which is considered a pro-inflammatory environment, and healthy controls plasma. Subjects included healthy controls (n=5), remitted BD patients (n=8), manic patients (n=5) and depressive patients (n=5). The mRNA expression of chemokynes and cytokines from U937 cells treated with BD patients plasma were different from those submitted to healthy controls plasma. Higher mRNA expression of IL-1β was observed in those cells submitted to manic and depressive BD patients plasma when compared to euthymic patients. Also, depressive BD patients plasma induced higher expression of TNF-α compared to euthymic patients. However, chemokynes expression, such as CXCL9 and CXCL10, were reduced in depressive BD patients. However, chemokynes expression, such as CXCL9 and CXCL10, were reduced in depressive BD patients. Inflammatory cytokines such as IL-1β and TNF-α in bipolar manic and depressive patients demonstrate higher amounts of IL-1β mRNA that euthymic patients and depressive patients induced higher amounts of TNF-α mRNA levels than the patients in euthymic U937. Since the expression of CXCL9 and CXCL10 chemokines in plasma from patients with depressive TB, proved less significant expression in the group of manic patients when compared to controls and euthymic bipolar patients.

A disfunção do sistema imune inato e a neuroinflamação tem sido cada vez mais reconhecidas como elementos importantes na fisiopatologia do transtorno bipolar (TB). Como componentes essenciais da imunidade inata, os macrófagos tem múltiplas funções tanto na inibição como na promoção da proliferação celular e na reparação tecidual, sendo a diversidade e a plasticidade características marcantes deste tipo celular. A polarização M1 clássica e a polarização alternativa M2 de macrófagos representam dois extremos de um estado dinâmico na mudança da ativação dos mesmos. Os macrófagos do tipo M1 sintetizam citocinas próinflamatórias que inibem a proliferação de células circundantes e danificam tecidos, enquanto os macrófagos do fenótipo M2 liberam citocinas antiinflamatórias que podem promover reparo tecidual. Um desequilíbrio da polarização M1-M2 dos macrófagos é frequentemente associado a várias doenças ou condições inflamatórias. O objetivo desta tese foi, além de revisar a importância da inflamação sistêmica na modulação da resposta inflamatória da microglia/macrófagos e consequentemente seu potencial envolvimento na fisiopatologia do TB, avaliar o perfil de polarização M1/M2 em cultura de macrófagos de sujeitos com TB comparados a indivíduos saudáveis. Monócitos foram isolados a partir de sangue periférico de dez sujeitos com TB e dez indivíduos saudáveis e diferenciados em macrófagos através da adição de fator estimulante de colônia de macrófagos (MCSF) ao meio de cultura. Para induzir a polarização M1 ou M2, as culturas foram incubadas com IFN-y e LPS ou IL-4 respectivamente. Após a incubação, recolheram-se os sobrenadantes e mediram-se as citocinas (IL-1β, IL-6, IL-10 e TNF-α) por ensaio multiplex. A secreção das citocinas IL-1β, TNF-α e IL-6 características do protótipo M1 e citocinas IL-10 do protótipo M2 foram semelhantes entre os pacientes e os controles. Utilizou-se a razão TNF-α / IL-10 do fenótipo M1 para refletir o estado inflamatório dos participantes. Não foi observada diferença entre os grupos (p=0,627). Duas hipóteses diferentes poderiam explicar esses resultados: todos os pacientes incluídos neste estudo representam um estágio inicial da doença como evidenciado pela pontuação FAST total inferior a 11. De acordo com o modelo de estadiamento em TB, as alterações biológicas (incluindo a inflamação) parecem estar relacionadas com os episódios de humor e progressão da doença. Juntamente com estudos anteriores, os nossos dados sugerem que os pacientes nos estágios iniciais ainda preservam a função do sistema imunológico sem apresentar um desequilíbrio a favor do perfil de macrófagos M1 como tem sido observado em pacientes no estágio tardio, destacando a relevância da intervenção precoce no TB. Ainda, estes pacientes estavam em tratamento com estabilizadores de humor e é plausível especular que esses fármacos exerçam efeitos sobre a polarização de macrófagos. Estudos futuros em pacientes drug-free são essenciais para avaliar esta questão. Em conclusão, nossos achados sugerem que os pacientes TB não apresentam desequilíbrio na polarização dos macrófagos em favor do fenótipo pró-inflamatório M1. O fato de todos estes pacientes estarem em estágios iniciais da doença reforça os efeitos protetores da intervenção precoce no TB na prevenção de alterações do sistema imune e, consequentemente, na progressão da doença. / Innate immune system dysfunction and neuroinflammation have been recognized as important elements in the pathophysiology of bipolar disorder (BD). As essential players of innate immunity, macrophages have multiple roles in inhibition and promotion of cell proliferation and tissue repair. The classical M1 polarization and the M2 alternative polarization of macrophages represent two extremes of a dynamic state in their change of activation. M1 macrophages synthesize proinflammatory cytokines that inhibit the proliferation of surrounding cells and damage tissues, whereas macrophages of the M2 phenotype release antiinflammatory cytokines that may promote tissue repair. An imbalance of the M1-M2 polarization of macrophages is often associated with various diseases or inflammatory conditions. The aim of this thesis was to review the importance of systemic inflammation in modulating the inflammatory response of microglia/ macrophages and consequently their potential involvement in the pathophysiology of BD, and also evaluate the M1/M2 polarization profile in macrophages of patients with BD compared to healthy individuals. Blood monocytes were obtained from ten BD patients and ten healthy controls. These cells were activated/polarized into the M1 (IFNγ + LPS) or M2(IL-4) phenotype. Supernatants were collected and the cytokines (IL-1β, IL-6, IL-10 and TNF-α) were measured by multiplex assay. Secretion of the IL- 1β, TNF-α, IL-6 and IL-10 were similar between patients and controls. The TNF-α/IL- 10 ratio of the M1 phenotype was used to reflect the inflammatory state of the participants. There was no difference between groups (p = 0.627). Two hypotheses could explain these results: all patients included in this study represent an early stage of disease as evidenced by the FAST score below 11. According to the BD staging model, biological changes (including inflammation) appear to be related to mood episodes and disease progression. Together with previous studies, our data suggest that patients in early stages of BD still preserve immune system function without presenting an imbalance in favor of M1 macrophages as has been observed in latestage patients, highlighting the relevance of early intervention. Moreover, these patients were under treatment with mood stabilizers and it is plausible to speculate that these drugs have effects on macrophage polarization. Future studies in drug-free patients are essential to assess this issue. In conclusion, our findings suggest that BD patients do not present imbalance in macrophage polarization in favor of the M1 proinflammatory phenotype. The fact that all these patients are in the early stages of the disease reinforces the protective effects of early intervention in BD to prevent changes in the immune system and, consequently, prevent the progression of the disease.

Background: Suicide prevention strategies recognise the need to address suicide in high-risk groups, such as people with psychiatric illness. People with bipolar disorder are known to be at particularly high risk of suicide and self-harm, with around half of people diagnosed with bipolar disorder making at least one suicide attempt during their lifetime. It is important that clinicians can identify who is most at risk among people with bipolar disorder so that interventions that meet the needs of this high risk group can be implemented. Method: A multiple-methods approach was used to explore suicidal behaviour in bipolar disorder. Descriptive analysis, case-control methods, and survival analysis were used on data held by The National Confidential Inquiry into Suicide and Homicide by People with Mental Illness (NCI), and the Manchester Self-Harm (MaSH) Project, to identify characteristics and risk factors associated with suicide in bipolar disorder. Semi-structure interviews were carried out with people who had a range of experiences of suicidal behaviour in bipolar disorder, and these data were analysed using Thematic Analysis to add context and depth to the quantitative results. Results: Suicidal behaviours were common in people with bipolar disorder, accounting for around 10% of all psychiatric suicide deaths in England; this rate was fairly stable over time. Characteristics associated with suicidal behaviour in bipolar disorder included; being aged 45 to 64 years old, experiencing negative life events, comorbid alcohol use, multiple inpatient admissions; there was a particularly strong association with a history of self-harm. It is clinically important that people with bipolar disorder were often seen by services in the 24 hours before they died. This both emphasises the weaknesses in current risk assessment, and highlights the potential for successful intervention if risk can be determined more accurately. Key issues identified in the interview study included being able to access care rapidly during time periods when risk was elevated, the importance of obtaining a correct diagnosis of bipolar disorder, and the potential benefits of including family in the care of people with bipolar disorder. Conclusion: Suicidal behaviours are common in people with bipolar disorder. People with bipolar disorder who die by suicide tend to have several markers that may indicate a more severe (e.g. multiple inpatient admission, history of self-harm) and complex course of illness (e.g. comorbid alcohol use, personality disorder). Diagnosis-specific risk assessment is needed to better identify risk of suicide in an illness that is often characterised by fluctuating mood states. Family involvement in care may aid detection of increased suicide risk.