Cholinergic inputs from mesopontine tegmentum activate midbrain dopamine (DA) neurons via M5 muscarinic receptors. The M5 receptor is important for mesopontine stimulation-induced accumbal or striatal DA efflux, brain stimulation reward or morphine-induced conditioned place preference (CPP). M5 receptor knockout (KO) mice show 40-50% less morphine-induced locomotion. Pedunculopontine tegmental nucleus (PPT) lesions in rodents block morphine CPP, but are ineffective after 18 hours food deprivation, opiate dependence, or intra-VTA BDNF. Based on these findings, we investigated whether acute food deprivation or intra-VTA BDNF alters morphine-induced locomotion (3 and 10 mg/kg, i.p.) in C57BL/6 M5 KO mice. Non-deprived M5 KOs showed reduced morphine-induced locomotion, suggesting M5 receptors partly mediate morphine-induced locomotion. Morphine-induced locomotion was reversed in food-deprived mice, suggesting the stimulant effects of morphine were altered to bypass the PPT. Unexpectedly, intra-VTA BDNF infusions were ineffective in altering morphine-induced locomotion. Additionally, M5 KOs receiving intra-VTA saline showed no deficits in morphine-induced locomotion.

Teneurin C-terminal associated peptide-1 (TCAP-1) is a recently characterized peptide that may act as one potential neuroprotective agent as it has been shown to regulate several stress-associated behaviours in rodents and possesses a number of protective actions on cells, however the mechanism remains unknown. Brain-derived neurotrophic factor (BDNF) is a neurotrophin recognized for mediating survival, differentiation, and proliferation. TCAP-1 may act, in part, via BDNF to provide neuroprotection via modulation of BDNF expression. The aim of this research was to further investigate the mechanism of TCAP’s neuroprotective actions. I show that TCAP-1 is neuroprotective and a potent enhancer of cell numbers under varying levels of oxygen. I also establish that TCAP-1 is able to influence neuronal behaviour by differentially regulating neurite growth. In addition, I indicate that TCAP-1 is able to regulate BDNF expression in immortalized mouse hypothalamic N38 cells, which suggests that TCAP-1’s neuroprotective mechanism may involve BDNF.

Teneurin C-terminal associated peptide-1 (TCAP-1) is a recently characterized peptide that may act as one potential neuroprotective agent as it has been shown to regulate several stress-associated behaviours in rodents and possesses a number of protective actions on cells, however the mechanism remains unknown. Brain-derived neurotrophic factor (BDNF) is a neurotrophin recognized for mediating survival, differentiation, and proliferation. TCAP-1 may act, in part, via BDNF to provide neuroprotection via modulation of BDNF expression. The aim of this research was to further investigate the mechanism of TCAP’s neuroprotective actions. I show that TCAP-1 is neuroprotective and a potent enhancer of cell numbers under varying levels of oxygen. I also establish that TCAP-1 is able to influence neuronal behaviour by differentially regulating neurite growth. In addition, I indicate that TCAP-1 is able to regulate BDNF expression in immortalized mouse hypothalamic N38 cells, which suggests that TCAP-1’s neuroprotective mechanism may involve BDNF.

Axonal degeneration is a major pathological feature of the central nervous system (CNS) inflammatory demyelinating disease multiple sclerosis (MS). This axonal degeneration has major consequences, as functional axonal regeneration in the CNS is largely absent. Cumulative axonal degeneration is the likely cause of the majority of progressive MS-related disability, and therefore, the need for novel neuroprotective therapies for MS exists. Experimental autoimmune encephalomyelitis (EAE), an animal model of MS pathology, also produces axonal injury. In particular, the optic nerve and spinal cord are key sites of neuroinflammation in mouse EAE. By utilizing this model, the short term and long term effects of the putative neuroprotective cytokine, leukaemia inhibitory factor (LIF), were investigated in the optic nerve and spinal cord utilising a number of outcome measures of axonal dysfunction. These included MRI measures of water diffusivity along (ADC ||) and across (ADC┴) the optic nerves, serum levels of phosphorylated neurofilament heavy chain subunit (pNF-H) and histological morphometric measures. LIF treatment reduced EAE grade and pNF-H plasma levels, decreased ADC┴, but had no effect on ADC ||, axon counts or inflammatory infiltration. / In contrast, genetic deletion of LIF and its sister cytokine ciliary neurotrophic factor (CNTF), not only increased EAE grade and pNF-H levels, but also decreased optic nerve ADC|| and optic nerve and spinal cord axon densities. After reviewing current literature, we hypothesize that the target cell for endogenously upregulated LIF in EAE may be the neuron or axon, whereas the target cell for exogenously administered therapeutic LIF may be another cell type, possibly infiltrating macrophages and activated microglial cells. LIF antagonist treatment did not have any affect on EAE grade, pNF-H levels or MRI parameters. This lack of effect may be due to the inability of the LIF antagonist to enter the CNS, supporting the hypothesis that endogenous LIF has a centrally acting mechanism.

Diss. (sammanfattning) Linköping : Univ., 2003. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Washington, 2005. / Vita. Includes bibliographical references (leaves 137-167).

Multiple sclerosis (MS) is a chronic neurological disease characterized by the destruction of central nervous system (CNS) myelin. Although the neurotrophin, brain derived neurotrophic factor (BDNF) has a beneficial role in re-myelination and/or myelin repair, these effects are hampered by the over-expression of a transcriptional repressor isoform of methyl CpG binding protein 2 (MeCP2) called MeCP2E1. We hypothesize that following experimental autoimmune encephalomyelitis (EAE) -induced myelin damage, the immune system induction of the pathogenic MeCP2E1 isoform hampers the re-myelination and/or myelin repair process by repressing BDNF expression. Our research identified the temporal gene and protein expression changes of MeCP2E1, MeCP2E2 and BDNF in an EAE mouse model of MS, and correlated them with the changes in the neurological disability scores (NDS). Our results indicated MeCP2E1 mRNA levels are elevated in EAE animals which is responsible for the repressed BDNF production in the spinal cord that prevents re-myelination and/or myelin repair. / February 2016

Contexto: As oscilações hormonais ao longo da vida estão associadas às variações de humor em mulheres normais. O estrógeno (E) parece estar associado aos níveis de fator neurotrófico derivado do cérebro (BDNF) em voluntárias saudáveis. No entanto, essa associação não foi investigada em pacientes com transtorno bipolar (TB). Sabe-se que os episódios de humor do TB estão associados a alterações dos níveis de BDNF; entretanto, não está claro o papel dos hormônios femininos. Objetivo: investigar a existência de uma possível associação entre os níveis de BDNF e os níveis de hormônios do eixo hipotálamo-hipófise-gonadal em mulheres com TB, incluindo pacientes durante o período reprodutivo e também na pós-menopausa. Métodos: Mulheres eutímicas (HAM-D e YMRS com escores menores que 8) com transtorno bipolar I(TB I), II (TB II) ou transtorno bipolar sem outra especificação (TB SOE) foram incluídas. As pacientes em idade reprodutiva tinham ciclos menstruais regulares (CMR) e não faziam uso de nenhum tipo de contracepção hormonal (CH); e as na pós-menopausa não estavam em uso de terapia de reposição hormonal (TRH). Condições endócrinas instáveis foram consideradas um fator de exclusão. Todas as pacientes estavam em tratamento farmacológico, associado ou não a intervenções psicossociais. Amostras de sangue foram retiradas para as medidas de BDNF, estrogênio (E), progesterona (P), LH e FSH, sendo coletadas nas fases folicular (FF) e lútea (FL) do ciclo menstrual, e uma única vez nas mulheres na pós-menopausa. Os diagnósticos foram confirmados através de entrevista clínica estruturada para o DSM-IV Transtornos do Eixo I (SCID-I), administrado por investigadores treinados. Resultados: Foram avaliadas 96 pacientes com TB. Destas, 64 não preenchiam critérios de inclusão ou apresentavam fatores de exclusão. Foram estudadas 32 mulheres com idades entre 22 e 69 anos (média = 52,78 anos). Considerando toda a amostra, o BDNF apresentou uma correlação positiva com os níveis de estradiol (r = 0,36, p = 0,043). Nas pacientes em período reprodutivo, na fase lútea(FL), houve uma correlação negativa entre o BDNF e o FSH (r = 0,831, p = 0,040). Um resultado semelhante foi encontrado com os níveis de LH (r = 0,908, p= 0,012) nessa mesma fase do ciclo menstrual. Conclusão: Os resultados encontrados na amostra de mulheres com TB foram semelhantes aos descritos na literatura em indivíduos saudáveis, que também apresentam correlação entre E e BDNF (Begliuomini et al., 2007). Estes achados indicam que o estímulo estrogênico pode ser importante na manutenção de s níveis fisiológicos de BDNF. A partir desses resultados novas vias devem ser incluídas na investigação na fisiopatologia das alterações de humor relacionadas a variações hormonais, bem como ao tratamento do TB. Além disso, ressalta a importância de incluir as variações hormonais femininas na equação diagnóstica e prognóstica do TB. / Background: Background: Hormonal oscilations across lifetime have been associated with mood variations in healthy women. Oestrogen (E) seems to be associated with Brain Derived Neurotrophic Factor (BDNF) levels in healthy volunteers. This assictaion was not studied in women with Bipolar Disorder (BD). Mood episodes of BD are associated with reductions in BDNF levels, although the role of feminine hormones in pathophysiology of BD hás not been completely studied. Objective: To investigate the association between BDNF levels and hormones involved in hypothalamus-pituitary-gonadal axis in women with BD, comparing a group during reproductive years with a menopausal group. In addition, differences across the two phases of menstrual cycle were also evaluated in the group during reproductive years. Methods: Women euthymic (HAM-D and YMRS scoring less than 8) with bipolar I, II or bipolar disorder not otherwise specified were included. Patients of reproductive age had regular menstrual cycles (RMC) and did not use any type of hormonal contraception (CH) and postmenopausal were not using hormone replacement therapy (HRT). Endocrine instable conditions were considered an exclusion factor. All patients were on pharmacotherapy, associed or not with psychosocial interventions. Blood samples withdrawn for measures of BDNF, oestrogen (E), progesterone (P), LH and FSH levels, being collected in the follicular phase (FP) and luteal (FL) of the menstrual cycle, menopausal women were held only one blood sample. Diagnoses were assessed using structured clinical interview for DSM-IV Axis I Disorders (SCID-I), administered by certified investigators. Results: Ninety six patients with BD were evaluated, of these, 64 did not meet inclusion criteria or met exclusion factors. The sample was constituted by 32 women with BD aged 22 to 69 years (mean = 52.78 years). Considering the whole sample, the BDNF was significantly correlated with estradiol levels (r = 0.36, p = 0.043). In patients in reproductive period, in the luteal phase, there was a negative correlation with FSH (r = 0.831, p = 0.040). A similar result was found with levels o LH (r = 0.908, p = 0.012) in the same menstrual cycle phase. Conclusion: The results found in the sample of women with TB were similar to those previously reported in healthy subjects, which also show a correlation between E and BDNF (Begliuomini et al., 2007). These findings indicate that estrogenic stimulation may be important in maintaining physiological BDNF levels. From these results, new avenues should be included in research on the pathophysiology of mood swings related to hormonal changes as well as the treatment of TB. Furthermore, the importance of including female hormonal changes in the equation of TB diagnostic and.

Fundamentação: O acidente vascular cerebral (AVC) é uma das principais causas de morte em todo o mundo e a maioria dos sobreviventes permanece com alguma sequela neurológica após o evento agudo. O presente estudo objetiva investigar a associação de alguns biomarcadores sanguíneos com as escalas de AVC, bem como avaliar a capacidade dos biomarcadores selecionados na predição de desfechos neurológicos durante o tempo de acompanhamento. Material e Métodos: Incluímos nesse estudo 60 pacientes com AVC agudo admitidos na unidade neurovascular da emergência ou na unidade de medicina intensiva do Hospital de Clínicas de Porto Alegre, nas primeiras 24 horas do início dos sintomas. Foram coletas amostras sanguíneas nas primeiras 24 horas, no terceiro e no quinto dias após o AVC para dosagem de enolase neurônio específica (ENS), proteína S100ß (S100ß), interleucina 6 (IL-6), proteína C reativa (PCR) e fator neurotrófico derivado do cérebro (BDNF). A gravidade do AVC e o grau de dependência funcional dos pacientes após o AVC foram mensurados através das escalas do National Institutes of Health Stroke Scale (NIHSS) e modified Rankin Scale (mRS) nos três momentos das coletas sanguíneas e na alta hospitalar. Resultados: Os níveis séricos de S100ß, IL-6 e PCR mostraram-se o melhor painel de biomarcadores após o AVC nesse estudo. Quando os pacientes foram subdivididos em dois grupos para a avaliação de desfechos neurológicos, usando as escalas do NIHSS (NIHSS ≤ 6 e NHISS > 6) e mRS (mRS ≤ 3 e mRS > 3), ambas as escalas apresentaram boa associação entre as concentrações de S100ß e de IL-6 em todas as medidas e as escalas de AVC para bom prognóstico (NIHSS ≤ 6 e mRS ≤ 3) na alta hospitalar. Dentre os biomarcadores selecionados para o estudo, foram os três citados acima que apresentaram as melhores correlações com as escalas de AVC e com o prognóstico pós AVC durante o tempo de acompanhamento. Conclusão: Os biomarcadores séricos podem ser úteis na avaliação da gravidade e do prognóstico após o AVC. A associação de S100ß, IL-6 e PCR parece acrescentar pouco às escalas validadas de AVC na capacidade de predizer desfechos após o evento agudo. / Background and Purpose: Stroke is an important cause of death worldwide, and the majority of stroke survivors suffer from some form of residual disability. This study aimed to investigate the association of blood biomarkers with stroke scales and their predictive value after acute stroke at the time of admission until hospital discharge. Design and Methods: We investigated 60 patients with acute stroke who were admitted within 24 h of event onset at the intensive care unit or neurovascular emergency unit of Clínicas Hospital. All patients provided venous blood samples for the measurement of neuron-specific enolase (NSE), S100ß protein (S100ß), interleukin-6 (IL-6), C-reactive protein (CRP) and brain-derived neurotrophic factor (BDNF) within 24 h of the acute event, on the third day and on the fifth day after the stroke. Neurological stroke severity and global disability were determined with the National Institutes of Health Stroke Scale (NIHSS) and modified Rankin Scale (mRS) at the same three times of blood collection and at the time of hospital discharge. Results: The serum levels of the S100ß protein, IL-6 and CRP seem to constitute the best panel of biomarkers after acute stroke in this study. When patients were subdivided into two groups according to the NIHSS (NIHSS ≤ 6 and NIHSS > 6) and mRS (mRS ≤ 3 and mRS > 3) scores, which were used as neurological outcome measures, both neurologic scores for good outcome (NIHSS ≤ 6 and mRS ≤ 3) at hospital discharge were significantly related to the S100ß protein and IL-6 levels at all of the measured time points. Among the analyzed blood markers, S100ß, IL-6 and PCR levels significanttly correlated with the stroke scales and prognostic value. Conclusion: Blood biomarkers may be useful in acute stroke either by suggesting stroke severity or providing a prognostic value. The addition of the S100ß protein, IL-6 and CRP to previously validated stroke scales slightly improves the ability of these scales to predict outcome.

During my thesis work I studied the expression and potential function of brain expressed microRNAs (miRNAs) in human prefrontal cortex (PFC). Initially, I used combinatorial computational analysis and microarray data to identify miRNAs that are predicted with high probability to target the human Brain Derived Neurotrophic Factor (BDNF) 3’ Untranslated Region (3’UTR) and are expressed in moderate to high levels in adult human prefrontal cortex. A subset of 10 miRNAs segregating into 5 different miRNA families (miR-30a-d, miR-103/107, miR-16/195, miR-191 and miR-495) met the above criteria. I then designed a protocol to detect these miRNAs with Locked Nucleic Acid (LNA) in situ hybridization in human prefrontal cortex and determine their layer and cellular expression patterns. LNA in situ revealed differential lamina and cellular enrichment of BDNF-related miRNAs. As an example, miR-30a-5p was found to be enriched in large pyramidal neurons of layer 3, which was verified using laser capture microdissection of layer 3 pyramidal neurons and quantitative Real Time Polymerase Chain Reaction (qRT-PCR) following dissection of upper and deeper layers of human PFC. Parallel to this, I used miRNA qRT-PCR to determine the developmental expression of miRNAs using postmortem PFC tissues ranging from embryonic age to old adulthood and compared miRNA to BDNF protein levels. My results revealed a robust inverse correlation between BDNF-related miRNAs and BDNF protein during late maturation and aging of human prefrontal cortex. In vitro luciferase assays and/or lentivirus mediated neuronal miRNA overexpression experiments validated that at least two miRNAs, miR-30a-5p and miR-195, target human BDNF 3’UTR and mediate its translational repression. In the second part of my thesis work I measured levels of miR-30a and miR-195 in the prefrontal cortex of patients with schizophrenia and compared them with levels of BDNF protein and BDNF-related GABAergic mRNAs. According to my results differences in miR-195 levels in a subset of subjects diagnosed with schizophrenia were found to be associated with disease related changes in BDNF protein levels and deficits in BDNF dependent GABAergic gene expression. In the last part of my work I focused on miR-30b, another member of the miR-30 family, which I found to be reduced in the prefrontal cortex of female but not male subjects with schizophrenia. More importantly, disease related changes in miR-30b levels were strongly associated with the age of onset of the disease. Additional experiments in mouse cortex and hippocampus revealed a gender dimorphic expression pattern of this miRNA with higher expression in female brain. Collectively, my results suggest that miRNAs could participate in novel molecular pathways that play an important role during cortical development and maturation and are potentially linked to the pathophysiology of neuropsychiatric disease.