Attention-deficit hyperactivity disorder (ADHD) is a common neuropsychiatric disorder with onset at preschool age Approximately 5-10% of school-aged children worldwide have ADHD. Psychostimulants are the most common treatments for ADHD, although the precise etiology and pathological mechanisms underlying ADHD are poorly understood. Animal models could help to elucidate and further the understanding of this disorder. Among the major rodent models of ADHD of the genetic and neurotoxin-exposed animal models, the spontaneously hypertensive rats (SHR) are more extensively studied. Nevertheless, the mechanism of ADHD is complex and the evidence of SHR model for ADHD has been conflicting. Objective. In this work, we combined behavioral, neurochemical, neuroimaging, pharmacological and molecular studies to examine SHR as an animal model of ADHD. At the same time, the results of our studies could help us to explore the potential mechanism of ADHD. Material and methods. We compared the locomotor activity, attention, inhibition, learning and memory of juvenile male SHR with those of age- and gender-matched genetic control Wistar-Kyoto rats (WKY) by using the open field test, Morris water maze and prepulse inhibition test. We employed magnetic resonance imaging (MRI) to measure potential morphological differences between different brain areas of SHR and WKY, and the functional MRI (fMRI) for functional differences in these brain areas. We also measured dopamine concentration and dopamine related genes expression in the different dopamine pathways by using enzyme-linked immunosorbent assay (ELISA) to measure the dopamine concentration and by using real time PCR to assay genes expression. We examined SHR responses to D-amphetamine (D-AMP), which is psychostimulant. These included locomotor activity and inhibition ability during D-AMP treatment, expression of dopamine related genes after D-AMP treatment measured by real time PCR and c-fos protein after repeated treatment of D-AMP by the Western Blotting. Results . Hyperactivity, impulsivity and attention deficit were observed in SHR. Decreased brain volume in caudate-putamen and vermis cerebelli in SHR were demarcated using MRI. Functional MRI (fMRI) and altered c-fos expression indicated plasticity changes of the prefrontal cortex (PFC) in SHR. Dopamine content was found to decrease in mesocortical and mesolimbic dopamine pathways, but increased in the striatum. Dopamine D4 receptors gene and protein expression were decreased in the PFC in SHR. We also found that the expression of the synaptosomal-associated protein 25 (SNAP-25) gene was initially lower in the PFC but higher in the striatum in SHR. However, this disparity of SNAP-25 in the PFC vanished after repeated treatment of D-AMP between SHR and WKY. Conclusions. In the present study, we demonstrated that SHR could be established as an ADHD model by completing complex assessments of face validity, construct validity and prediction validity. We suggested that the "synaptogenesis hypotheses" might contribute to the abnormal release of dopamine and dysfunction of PFC and the striatum in SEER. In conclusion, our results have provided further new information relevant to the understanding of ADHD in human via the analysis of the SHR model. / Li, Qi. / Adviser: David Yen. / Source: Dissertation Abstracts International, Volume: 69-02, Section: B, page: 1375. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 108-125). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / School code: 1307.

Made available in DSpace on 2015-07-22T20:49:41Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-06-29. Added 1 bitstream(s) on 2015-08-11T03:26:12Z : No. of bitstreams: 1 Publico-12851a.pdf: 1993449 bytes, checksum: 6fad4a58fc1b395d08412bb6e3b076ea (MD5). Added 1 bitstream(s) on 2015-08-11T03:26:12Z : No. of bitstreams: 2 Publico-12851a.pdf: 1993449 bytes, checksum: 6fad4a58fc1b395d08412bb6e3b076ea (MD5) Publico-12851b.pdf: 1501009 bytes, checksum: ab8946f01511cdc67f2383a0a9fabaac (MD5). Added 1 bitstream(s) on 2015-08-11T03:26:12Z : No. of bitstreams: 3 Publico-12851a.pdf: 1993449 bytes, checksum: 6fad4a58fc1b395d08412bb6e3b076ea (MD5) Publico-12851b.pdf: 1501009 bytes, checksum: ab8946f01511cdc67f2383a0a9fabaac (MD5) Publico-12851c.pdf: 1843176 bytes, checksum: 32033d7bc48d53d7cb1127e1ae58221f (MD5). Added 1 bitstream(s) on 2015-08-11T03:26:13Z : No. of bitstreams: 4 Publico-12851a.pdf: 1993449 bytes, checksum: 6fad4a58fc1b395d08412bb6e3b076ea (MD5) Publico-12851b.pdf: 1501009 bytes, checksum: ab8946f01511cdc67f2383a0a9fabaac (MD5) Publico-12851c.pdf: 1843176 bytes, checksum: 32033d7bc48d53d7cb1127e1ae58221f (MD5) Publico-12851d.pdf: 1909278 bytes, checksum: 7f3b132a4365c4aeab410fd8f4041429 (MD5) / Objetivos: avaliar os efeitos da fumaça lateral de cigarro (FLC) sobre o controle neural do sistema cardiovascular em ratos normotensos e ratos espontaneamente hipertensos (SHR). Método: ratos Wistar, Wistar-Kyoto (WKY) e SHR foram expostos à FLC durante três semanas, cinco dias por semana, 180 minutos por dia numa concentração de monóxido de carbono entre 100 e 300 ppm. Barorreflexo foi estimulado por uma dose vasodepressora de nitroprussiato de sódio (NPNa, 50Og/kg, i.v.) e uma dose pressora de fenilefrina (PE, 8Og/kg , i.v.). Para avaliar os efeitos da inibição da catalase no quarto ventrículo cerebral (4ºV) sobre as respostas cardiovasculares, foi injetado o inibidor de catalase 3-amino-1,2,4-triazole (ATZ, 0,01Og/100OL). Resultados: nos ratos Wistar expostos à FLC, foi observado que a inibição da catalase causou respostas mais intensas quanto a FC basal e ao pico bradicárdico. A inibição da catalase afetou de maneira mais intensa a FC basal e o pico bradicárdico, nos ratos WKY expostos à FLC. Por outro lado, nos animais SHR a exposição à FLC afetou o pico taquicárdico após inibição central de catalase de maneira mais intensa. Conclusão: a exposição à FLC altera os componentes simpáticos do barorreflexo em ratos WKY e SHR, além de causar respostas cardiovasculares mais intensas perante a inibição de catalase no 4ºV em ratos Wistar e WKY. / Objectives: To evaluate the effects of sidestream cigarette smoke (SSCS) on neural control of cardiovascular system in normotensive and spontaneously hypertensive rats (SHR). Method: Wistar, Wistar-Kyoto rats (WKY) and SHR were exposed to SSCS for three weeks, five days per week, 180 minutes per day at a concentration of carbon monoxide between 100 and 300 ppm. Baroreflex was stimulated with a vasodepressor dose of sodium nitroprusside (NPNa, 50ìg/kg, iv) and with a pressor dose of phenylephrine (PE, 8ìg/kg, iv). In order to evaluate the effects of catalase inhibition into the fourth cerebral ventricle (4th V) on cardiovascular responses, we injected the catalase inhibitor 3-amino-1,2,4-triazole (ATZ, ìg/100ìL 0.01). Results: It was observed in Wistar rats exposed to SSCS that catalase inhibition caused more intense responses on basal HR and bradycardic peak. Central catalase inhibition affected in a higher intensity baseline HR and bradycardic peak WKY rats exposed to SSCS. On the other hand, in SHR SSCS exposure affected the tachycardic peak after central inhibition of catalase in a higher intensuty. Conclusion: Exposure to SSCS alters the sympathetic component of the baroreflex in WKY and SHR and caused more severe cardiovascular responses to catalase inhibition into the 4th V in Wistar and WKY rats. / TEDE / BV UNIFESP: Teses e dissertações

Made available in DSpace on 2014-06-11T19:27:47Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-08-20Bitstream added on 2014-06-13T19:15:14Z : No. of bitstreams: 1 cursino_nm_me_araca.pdf: 2792202 bytes, checksum: 7bc02b29243b06952aa21b86edc0566a (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / A hipertensao arterial representa um fator de risco sistemico e condicao desfavoravel para tratamentos dentarios, especialmente aqueles que necessitam de reparacao ossea. O objetivo deste estudo foi avaliar o reparo alveolar em ratos espontaneamente hipertensos (SHR) e o efeito do atenolol sobre este processo. Wistar e SHR tratados ou nao com 100mg/kg/dia (atenolol), foram submetidos a extracao do dente incisivo superior direito e sacrificados aos 7, 14, 21, 28 e 42 dias apos a cirurgia. As hemi-maxilas foram removidas e as imagens radiograficas foram realizadas. A analise radiografica foi obtida por meio do sistema digital Digora. Analises histologicas, histomorfometricas e reacoes imunoistoquimicas foram feitas em cortes histologicos de 5ƒÊm de espessura, os quais foram corados com hematoxilina-eosina ou submetidos a imunomarcacao para RANK, RANKL, OPG e proteinas MMP-9. A analise histologica foi realizada por microscopia optica e a analise histomorfometrica pelo software RGB / Leica Qwin Color. Os resultados densitometricos e histomorfometricos foram analisados pela Anova two-way. Na analise imunoistoquimica, utilizando um microscopio optico, foram atribuidos scores as imagens. Os resultados foram analisados pelos testes estatisticos Kruskal-Wallis e Mann Whitney. As diferencas entre os resultados foram consideradas significativas quando p<0,05. Reducao da densidade mineral ossea (DMO), menor porcentagem de osso e menor espessura do trabeculado osseo foram observadas nos periodos finais do reparo alveolar em SHR. Aumento da imunomarcacao para RANKL, RANK e MMP-9 foi observado em 28 dias apos a cirurgia no alveolo em SHR. Consistente efeito do atenolol foi observado no reparo alveolar de ratos hipertensos. O atenolol aumentou a DMO observada na maioria dos periodos analisados e aumentou a espessura do trabeculado... / Hypertension represents a systemic risk factor and unfavorable condition for dental treatments, especially treatments that require bone healing. The purpose of this study was to evaluate the alveolar wound healing in spontaneously hypertensive rats (SHR) and the atenolol effect on this process. Normotensive Wistar rats and SHR, untreated or treated with atenolol (100mg/kg/day), were submitted to the extraction of the upper right incisive tooth and sacrificed at 7, 14, 21, 28 and 42 days after surgery. The hemi-jaws were extracted and the radiographic images were obtained. Radiographic analysis was performed by using the digital system Digora. Histological, histomorphometric and immunohistochemical reactions were done in histological sections, 5 μm thick, stained with hematoxylin and eosin or subjected to immunolabeling to RANK, RANKL, OPG and MMP-9 proteins. Histological analysis was performed by light microscopy and histomorphometric analysis by Leica Qwin Color/RGB software. The densitometric and histomorphometric results were also analyzed by two-way ANOVA. In immunohistochemical analysis, using an optical microscopy, scores were assigned to the images. Results were analyzed by Kruskal-Wallis and Mann Whitney statistical tests. Differences between results were considered significant when p <0.05. Reduced bone mineral density (BMD), lower bone percentage and less thickness of trabecular bone was observed in the final periods of alveolar bone healing in SHR. Increased RANKL, RANK and MMP-9 immunolabeling were observed at 28 days after surgery in SHR alveolus. Consistent atenolol effect was observed on alveolar bone healing of hypertensive rats. Atenolol increased the BMD observed in most of the periods analyzed and increased trabecular bone thickness at 28 and 42 days in SHR alveolus. Increased OPG immunolabeling... (Complete abstract click electronic access below)

Para avaliar as repercussões da insuflação prolongada da cavidade peritoneal com gás carbônico sobre a hipertensão arterial essencial, utilizou-se ratos machos espontaneamente hipertensos (SHR) e como normotensos ratos machos Wistar-Kioto (WKY). No total foram utilizados 34 animais, sendo 22 SHRs e 12 WKYs, onde os ratos SHR foram distribuídos aleatoriamente aos grupos G1 e G3. O primeiro grupo (G1) com 12 animais SHRs e o segundo (G2) com 12 animais WKYs foram expostos a pneumoperitônio com dióxido de carbono por 120 minutos, enquanto que o terceiro grupo (G3) com 10 animais SHRs, passou por insuflação da cavidade peritoneal, seguida de punção com trocarte e esvaziamento do pneumoperitônio. Os animais deste grupo permaneceram anestesiados e com o abdome puncionado por 2 horas. Previamente a confecção do pneumoperitônio, a artéria e veia femorais direita foram dissecadas e canuladas. A artéria foi conectada ao transdutor de pressão para o registro contínuo da pressão arterial (PA), após a coleta inicial de 0,2 ml para dosagem da gasometria basal e 0,8 ml para as dosagens de uréia (U) e creatinina (Cr) basais. A veia femoral foi uttilizada para a expansão volêmica lenta com 10 ml de solução fisiológica após a coleta inicial de 1,0 ml de sangue arterial. Feito isto, procedeu-se a insuflação e punção abdominal mantendo ou não o pnemoperitônio, conforme o grupo. Foram feitas medidas da pressão arterial a cada 15 minutos e 5 minutos após o esvaziamento do abdome. Após a última aferição, foi colhido aproximandamente 3 ml de sangue arterial e 1 ml para a gasometria mais dosagem da U e Cr. A análise multivariada para medidas repetidas ao longo do tempo permitiu concluir que: nos cinco minutos após a desinsuflação, houve diferença estatística significante (p<0,0001) nas pressões arteriais sistólica, diastólica e média no G1 com uma curva ascendente em relação ao G2 e G3; O pH diminuiu (p<0,0001) de maneira similar nos três grupos de intervenção, enquanto a pCO2 aumentou (p<0,0001) de maneira similar nos três grupos de intervenção; não houve mudanças significativas na creatinina (p=0,3232); a uréia apresentou um efeito de momento com significância estatística (p<0,0001) e a atividade da renina plasmática foi significativamente maior no G2 em relação aos outros dois grupos / To assess the effects of prolonged peritoneal cavity insufflation with carbon dioxide on the essential hypertension, a experimental study was designed using male spontaneously hypertensive rats (SHR) and male normotensive Wistar-Kyoto (WKY). Thirty-four animals were used, 22 SHRs and 12 WKYs, where SHR rats were randomly assigned to groups G1 and G3. The first group (G1) with 12 animals SHRs and second group (G2) with 12 animals WKYs were exposed to pneumoperitoneum with carbon dioxide for 120 minutes, while the third group (G3) with 10 animals SHRs, had the peritoneal cavity insufflated, followed by puncture with trocarte and released the pneumoperitoneum. The animals of this group remained anesthetized and the abdomen punctured by 2 hours. Before making the pneumoperitoneum, right femoral artery and vein were dissected and cannulated. The artery was connected to the transducer pressure for the continuous recording of blood pressure (BP), after the initials blood samples: 0.2 ml for blood gases measurement and 0.8 ml for urea (U) and creatinine (Cr ). The femoral vein was used to volume expansion with 10 ml of saline solution after the initial sample of 1.0 ml arterial blood. Afterwards, a pnemoperitoneum insufflation and maintaining is done or not, depending on group. Blood pressure was recorded every 15 minutes and 5 minutes after pnemoperitoneum released. After last blood pressure record, a 3.0 ml blood sample was collected to measure plasma renin activity (PRA), and 1.0 ml for blood gases measurement, urea (U) and creatinine (Cr). The multivariate analysis for repeated measurements over time has concluded that: five minutes after pnemoperitoneum released, systolic, diastolic and mean blood pressure has significant statistic differences (p <0.0001) in G1 with an upward curve in relation to G2 and G3; The pH decreased (p <0.0001) in a similar way in the three groups of intervention, while pCO2 increased (p <0.0001) in a similar way in the three groups of intervention, with no significant changes in creatinine (p = 0.3232), but the urea had a moment effect with statistical significance (p <0.0001) and the plasma renin activity (PRA) was significantly higher in G2 compared with the other two groups

INTRODUÇÂO: Durante o desenvolvimento da hipertensão arterial sistêmica (HAS) ocorre a hiperatividade simpática, que está relacionada ao comprometimento dos sistemas baro e quimiorreflexo arteriais e disfunção ventricular esquerda (VE). Entretanto, a função ventricular direita (VD) tem sido pouco avaliada no contexto da HAS associada à desnervação sino-aórtica (DSA). OBJETIVO: Avaliar a função biventricular de forma não-invasiva e invasiva, a capacidade funcional, a sensibilidade barorreflexa e o controle autonômico cardiovascular em ratos Wistar (W) e ratos espontaneamente hipertensos (SHR) submetidos ou não à DSA. MÉTODOS: Após 10 semanas de DSA, a função cardíaca foi avaliada pelo teste de esforço (TE), ecocardiograma transtorácico e transesofágico, e a pressão diastólica final biventricular; as funções hemodinâmica e autonômica foram avaliadas pelo registro da pressão arterial (PA) e da freqüência cardíaca (FC), variabilidade da PA e da FC e sensibilidade barorreflexa. Os ratos (n = 32) foram divididos em 4 grupos: 16 W com (n = 8) e sem DSA (n = 8), 16 SHR com (n = 8) ou sem DSA (n = 8). RESULTADOS: A PA e a FC não apresentaram alterações entre os grupos DSA e não-DSA, entretanto, os SHR apresentaram níveis mais elevados da PA comparado com W. O TE mostrou que os SHR apresentaram melhor capacidade funcional em relação ao DSA e SHRDSA (W: 1,16±0,3m/s, DSA: 0,9±0,15m/s, *SHR: 1,46±0,29m/s, SHR-DSA: 1,02±0,31, *p< 0,05 vs. DSA e SHRDSA). Os SHRs apresentaram aumento da variabilidade da PA comparados aos W. Após a DSA houve aumento da variabilidade PA em todos os grupos comparados ao W (W: 15±29 mmHg2, *DSA: 49±27 mmHg2, *SHR: 60±29 mmHg2, *SHR-DSA: 137±76 mmHg2, *p<0,05 vs. W). Foi observado hipertrofia concêntrica do VE; disfunção sistólica segmentar e diastólica global do VE; disfunção sistólica global e segmentar, e diastólica global do VD; sinais indiretos de hipertensão arterial pulmonar pela ecocardiografia, mas evidentes no grupo SHRDSA. A pressão diastólica final do VD mostrou aumento em todos os grupos comparados com W (W: 3±0.39mmHg, *DSA:4,7±0,52mmHg, *SHR: 6;6±1.1mmHg, *SHRDSA: 7,8±0.87mmHg, *p< 0,05 vs. W), enquanto a pressão diastólica final do VE mostrou aumento dos grupos SHR e SHRDSA em relação ao W, e dos SHRDSA em relação aos DSA (W: 5,83±0,19 mmHg, DSA: 8,98±1,2 mmHg, *SHR: 12,51±4,73 mmHg, *#SHRDSA: 14,57±2.52 mmHg, *p< 0,05 vs. W, #p< 0,05 vs. DSA). Houve relação entre medidas não- invasivas e invasivas do VD, mostrando uma boa acurácia das medidas ecocardiográficas. CONCLUSÕES: Nossos resultados sugerem que a disfunção baroreflexa compromete a função biventricular. Além disso, os achados observados nos índices ecocardiográficos do VD indicam que a DAS pode induzir a elevação da pressão arterial pulmonar, reforçando o papel da disfunção barorreflexa na patogênese da doença cardíaca hipertensiva / INTRODUCTION: During the development of hypertension, sympathetic hyperactivity commonly seems to be related to the left ventricular (LV) dysfunction and baro and chemoreflexes impairment. However, right ventricle (RV) function has not been evaluated specially regarding the association of hypertension and baroreflex dysfunction. OBJECTIVE: To evaluate noninvasively and invasively the biventricular myocardial function, the functional capacity, the baroreflex sensitivity and the cardiovascular autonomic control in Wistar (W) rats and spontaneously hypertensive rats (SHR) submitted or not to sinoaortic denervation (SAD). METHODS: Ten weeks after DSA, cardiac function was evaluated by the maximal exercise test (MET), by transthoracic (TT) and transesophageal echocardiography (TEE) and the biventricular end diastolic pressures (EDP). Additionally, hemodynamic and autonomic functions were evaluated by the blood pressure (BP) and heart rate (HR) records, BP and HR variability and baroreflex sensitivity. The rats (n=32) were divided in 4 groups: 16 Wistar (W) with (n=8) or without SAD (n=8) and 16 SHR, with (n=8) or without SAD (n=8). RESULTS: Blood pressure and HR did not show any change between the groups SAD and without SAD, although, SHR showed higher BP levels in comparison to W. MET results showed that SHR had better functional capacity compared to SAD and SHRSAD (W: 1,16±0,3m/s, DSA: 0,9±0,15m/s, *SHR: 1,46±0,29m/s, SHR-DSA: 1,02±0,31, *p< 0.05 vs. SAD and SHRSAD). BP variability was increased in SHR groups compared to W. After SAD, BP variability increased in all groups compared to W (W: 15±29 mmHg2, *DSA: 49±27 mmHg2, *SHR: 60±29 mmHg2, *SHR-DSA: 137±76 mmHg2, *p<0.05 vs. W). Left ventricular concentric hypertrophy; segmental systolic dysfunction and global diastolic LV dysfunction; segmental and global systolic dysfunction, and global diastolic RV dysfunction; indirect signals of pulmonary arterial hypertension were shown by echocardiography, mostly evident in SHRSAD. The RV-EDP increased in all groups compared to W (W: 3±0.39mmHg, *SAD:4.7±0.52mmHg, *SHR: 6.6±1.1mmHg, *SHRSAD: 7.8±0.87mmHg, *p<0.05 vs. W), and the LV-EDP increased in SHR and SHRSAD groups compared to W, and in SHRSAD compared to SAD (W: 5,83±0,19 mmHg, SAD: 8.98±1.2 mmHg, *SHR: 12.51±4.73 mmHg, *#SHRSAD: 14.57±2.52 mmHg, *p<0.05 vs. W, #p<0.05 vs. DSA). There was a relation between invasive or noninvasive measurements of the RV showing good accuracy of echocardiographic measurements. CONCLUSIONS: Our results suggest that baroreflex dysfunction impaired biventricular function. Moreover, the findings of RV echocardiographic indices indicate that SAD may lead to increased pulmonary artery pressure, supporting a role for baroreflex dysfunction in the pathogenesis of the hypertensive cardiac disease

O emprego de \"Total Genome Scan\" em modelos genéticos de doenças complexas tem sido fundamental para seleção de regiões cromossômicas envolvidas com traços complexos. Em nosso laboratório, identificamos cinco regiões cromossômicas associadas ao traço quantitativo pressão arterial (BP-QTL) que explicam 43% da variação da pressão arterial numa progênie obtida a partir de animais espontaneamente hipertensos (SHR) e \"Brown Norway\" (BN). Os BP-QTLs foram, então, validados por desenvolvimento de linhagens congênicas, incluindo uma para o cromossomo 4 (SHR.BN4) cuja substituição das sequências SHR pelo do animal BN levou a redução da pressão arterial sistólica basal (~14 mm Hg). O objetivo deste trabalho foi identificar as variantes genéticas candidatas neste intervalo cromossômico com base em diferenças no padrão de expressão gênica e na presença de alterações genéticas não sinônimas \"missense\" ou em regiões regulatórias conservadas que possam estar envolvidas na gênese da hipertensão. Identificamos 533 genes com expressão renal, dentre os 682 do intervalo, sendo que 28 apresentaram padrão de expressão diferente entre amostras de animais adultos (congênico vs. SHR) e seis apresentaram alterações não sinônimas \"missense\". É importante salientar que dos genes diferentemente expressos, encontramos alterações estruturais em regiões conservadas com potencial de participar na regulação em 11. Em conjunto, utilizamos uma plataforma integrada para selecionar 34 genes candidatos no cromossomo 4, dos quais 17 genes serão priorizados, para ser investigados quanto sua contribuição na hipertensão arterial do SHR e na hipertensão primária humana / Total genome scan in genetic models of complex diseases have been instrumental to select candidate genes underlying complex traits. We previously mapped 5 blood pressure related quantitative trait loci (BP-QTLs) that explain about 43% of the BP variance in a progeny derived from Spontaneous Hypertensive Rat (SHR) and Brown Norway (BN) rats. The BP-QTLs were then validated by derivation of congenic strains, including one for chromosome 4 (SHR.BN4) in which a segment from BN replaced the SHR sequences reducing basal systolic BP (~14 mm Hg). The aim of this project is to identify the candidate genetic variants within the chromosome interval based on differences in renal gene expression patterns and structural changes in both non-synonymous missense or within adjacent regulatory sequences that may contribute to hypertension. We identified 533 genes with renal expression, out of 682 in the interval, in which 28 presented differences in expression pattern in adult samples (congenic vs. SHR) and six presented non-synonymous missense alterations. In addition, 11 out of 28 differentially expressed genes showed structural alterations in adjacent conserved regions that potentially contribute to gene regulation. Taken together, using the proposed combination of strategies, we selected 34 hypertensive candidate genes in chromosome 4, in which 17 will be prioritized, to be further explored to assess their contribution to hypertension in the SHR and to essential hypertension in humans

A hipertensão arterial essencial é caracterizada pela elevação crônica da pressão arterial e representa o principal fator de risco para doenças cardiovasculares e renais. O rim participa do controle da pressão arterial e alterações intrínsecas no manuseio renal de sódio desempenham papel importante na patogênese da hipertensão essencial. Os túbulos proximais renais são responsáveis pela reabsorção da maior parte do sódio filtrado nos glomérulos e a maior parte da reabsorção de sódio neste segmento faz-se através da troca de Na+ por H+ em membrana apical, mediada pela isoforma 3 do trocador Na+/H+ (NHE3). Entretanto, os dados existentes referentes à modulação renal do NHE3 em modelos de hipertensão são ainda conflitantes. Este estudo teve como objetivo avaliar as possíveis alterações funcionais do trocador Na+/H+ NHE3 em túbulo proximal renal na linhagem SHR no estágio de préhipertensão (5 semanas) e de hipertensão (14 semanas) e investigar se estas alterações são acompanhadas de alterações na atividade e na expressão da proteína cinase A (PKA) e de proteínas fosfatase-1 (PP1). Por meio de microperfusão estacionária in vivo mediu-se a atividade do NHE3 em túbulo proximal e verificou-se que a reabsorção de bicarbonato foi reduzida em 62 ± 6 % (P < 0,001) na transição do J-SHR para o A-SHR enquanto foi aumentada em 113 ± 10 % (P < 0,001) na transição entre o J-WKY e o A-WKY. A atividade estimulada do NHE3 em J-SHR é decorrente da redistribuição do NHE3 do domínio intermicrovilar (IMV) para o domínio das microvilosidades (MMV) e do baixo nível de fosforilação da serina 552, sítio consenso para a PKA. Por outro lado, durante a fase de hipertensão, a atividade diminuída do NHE3 deve-se à sua redistribuição para o IMV e ao aumento da fosforilação na serina 552. Para testar a hipótese de que os níveis de fosforilação do NHE3 estariam aumentados em túbulo proximal de SHR adulto devido ao aumento da atividade da PKA e/ou à diminuição na atividade da PP1, foram avaliados tanto os níveis de fosforilação quanto a atividade do NHE3 em SHR jovens e adultos em resposta ao 6MB-cAMP (análogo ao cAMP que ativa especificamente a PKA). O JSHR apresentou um aumento tanto nos níveis de fosforilação da serina 552 (179 ± 14 %, P < 0,001) quanto nos de inibição da atividade (65 ± 10 %, P < 0,001) do NHE3 em relação ao J-SHR em resposta ao 6MB-cAMP. Já no A-SHR, a fosforilação da serina 552 aumentou moderadamente (36 ± 4 %, P < 0,01), assim como inibiu moderadamente (23 ± 9 %, P < 0,05) a atividade do NHE3 em resposta ao 6MBcAMP. Adicionalmente, verificou-se que não houve alteração da atividade da PKA entre os animais nem ao longo da idade e nem entre as linhagens. Por sua vez, o JSHR apresentou maior atividade da PP1 que o A-SHR (1640 ± 107 vs. 940 ± 119 pM/?g, P < 0,01). Além disso, houve uma diminuição na expressão da PP1? no ASHR (32 ± 8 %, P < 0,01) quando comparado ao J-SHR. Os dados sugerem que o NHE3 é diferencialmente regulado antes e após o desenvolvimento da hipertensão em SHR por mecanismos que envolvem modificações pós-transcricionais e distribuição subcelular. Além do mais, a regulação diferencial dos níveis de fosforilação do NHE3 tubular proximal antes e após o desenvolvimento da hipertensão em SHR é devida, provavelmente, a alterações na atividade e na expressão da PP1 / Essential hypertension is characterized by chronic elevation of blood pressure and represents the major risk factor for cardiovascular and renal diseases. The kidney participates in the blood pressure control and intrinsic changes in renal sodium handling play an important role in the pathogenesis of essential hypertension. The renal proximal tubule is responsible for reabsorption of the great majority of sodium that is filtered by the glomerulus and the principal apical membrane mechanism for sodium reabsorption in this nephron is Na+/H+ exchanger isoform 3 (NHE3)- mediated Na+/H+ exchange. However, conflicting data have been reported with regard to NHE3 modulation in experimental models of hypertension. This study aimed to evaluate the possible functional changes of the Na+/H+ exchanger NHE3 in the renal proximal tubule of SHR both at the pre-hypertensive (5 weeks) and at hypertensive (14 weeks) stages and to investigate whether these changes were accompanied by changes in the activity and/or expression of protein kinase A (PKA) and protein phosphatase 1 (PP1). Proximal tubule NHE3 activity was measured by means of stationary microperfusion. Bicarbonate reabsorption was found to be decreased by 62 ± 6 % (P < 0.001) in the transition from youth to adulthood in SHR (Y-SHR to A-SHR), whereas in the transition from Y-WKY to A-WKY it increased by 113 ± 10 % (P < 0.001). Stimulated NHE3 activity in Y-SHR was due to redistribution of NHE3 from intermicrovilar domain (IMV) to microvilar domain (MMV) and to a lower level of serine 552 phosphorylation, a consensus site for PKA. Conversely, during the hypertensive stage, decreased NHE3 activity was due to increased redistribution of NHE3 to the IMV domain and increased phosphorylation at serine 552. To test the hypothesis that the increased levels of NHE3 phosphorylation in the proximal tubule of adult SHR were due to increased PKA activity and/or decreased PP1 activity, it was evaluated both phosphorylation levels and activity of NHE3 in young and adult SHR in response to 6MB-cAMP (an cAMP analog that specifically activates PKA). Y-SHR showed an increase both in the phosphorylation levels at serine 552 (179 ± 14 %, P < 0.001) and in the inhibition of NHE3 transport activity (65 ± 10 %, P < 0.001) compared to Y-SHR in response to 6MB-cAMP. With respect to A-SHR, the phosphorylation of serine 552 was slightly increased (36 ± 4 %, P < 0.01) and NHE3 activity was mildly inhibited (23 ± 9 %, P < 0.05) in response to 6MB-cAMP. Additionally, PKA activity remained unchanged with both age and strain. Nevertheless, Y-SHR exhibited higher PP1 activity than A-SHR (1640 ± 107 vs. 940 ± 119 pM/?g, P < 0.01). Furthermore, PP1? expression was decreased in the renal cortex of A-SHR (32 ± 8 %, P < 0.01) compared to Y-SHR. Taken together, these data suggest that NHE3 is differentially regulated before and after development of hypertension in SHR by mechanisms involving post-translational modifications and subcellular distribution. Moreover, the differential regulation of proximal tubule NHE3 phosphorylation levels before and after development of hypertension in SHR is most likely due to changes on the activity and expression of PP1

Brain stroke is considered as one of the three diseases that threaten human health all over the world. Hypertension and cerebral arteriosclerosis are thought to be the most dangerous risk factors of brain stroke, and they frequently occur together, leading to ischemia of brain tissue. Unfortunately, it is not clear whether the pathological changes resulting from hypertension are related to those resulting from cerebral arteriosclerosis. There have been no ideal animal models mimicking the pathological changes in such a combined condition. In this thesis, an animal model of hypertension combined with cerebral arteriosclerosis in rats was established by occlusion of both the left and right common carotid arteries in spontaneous hypertension rats. Pien Tze Huang (PTH), a reputed traditional Chinese medicinal complex, contains Radix notoginseng, snake bile, calculus bovis, and musk and some other components that are known to protect vessels and cells from injuries. Since different tissue injuries share many common cellular mechanisms, the protection by PTH to in nerves and the circulation systems may also be benefical to cerebrovascular conditions as well. In present experiments, PTH was used to treat hypertension rats that also developed chronic brain ischemia as a result of the bilateral carotid occlusion, and its protective role for neurons and blood vessels was investiaged. / From the data above, more severe damage could be caused by hypertension combined with chronic ischemia. The model of SHR with bilaterally occluded common carotid artery can be used to study pathological changes resulted from hypertension combined with chronic ischemia. PTH was able to protect neurons in stroke. / In the initial part of the work, patients from clinics in two cities in South and North China were compared and analysed; they had been suffering from brain ischemic stroke. About two thirds of the stroke patients were found to have hypertension before the onset of stroke. Their prognosis was significantly worse than those stroke patients without hypertension. In the hypertensive rats with occluded arteries, mean of functional magnetic resonance imaging (fMRI) examination showed that brain blood flow was very weak or even transiently became undetectable at the beginning of the acute stage of brain ischemia, but was restored one hour after the occlusion surgery. In addition, pathological changes in brains of hypertensive rats with induced brain ischemia (carotid occlusion) were examined by Nissl staining, TUNEL staining, cell death ELISA and anti-oxidation enzymes. At day 15 after ischemia, a large number of pyramid cells in the hippocampus of SHR were lost and a great deal of apoptotic cells were found in the CA1 of the hippocampus, while activities of some enzyme including superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) were increased. At day 30 and 60, some degenerative changes appeared to have subsided and the cells appeared morphologically normal. The activities of the above enzymes were also decreased at day 60. In WKY control rats with normal blood pressure, neurons in the CA1 were found less damaged after the bilateral carotid occlusion. It was found that apoptotic and dead cells were significantly reduced in rats with hypertension combined with chronic brain ischemia if they had been pre-treated with PTH. Moreover, brain stroke damage was less severe in this pretreated rats. / Zhang, Lihong. / "March 2010." / Adviser: WH Kwong. / Source: Dissertation Abstracts International, Volume: 72-01, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 116-134). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. Ann Arbor, MI : ProQuest Information and Learning Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

O emprego de \"Total Genome Scan\" em modelos genéticos de doenças complexas tem sido fundamental para seleção de regiões cromossômicas envolvidas com traços complexos. Em nosso laboratório, identificamos cinco regiões cromossômicas associadas ao traço quantitativo pressão arterial (BP-QTL) que explicam 43% da variação da pressão arterial numa progênie obtida a partir de animais espontaneamente hipertensos (SHR) e \"Brown Norway\" (BN). Os BP-QTLs foram, então, validados por desenvolvimento de linhagens congênicas, incluindo uma para o cromossomo 4 (SHR.BN4) cuja substituição das sequências SHR pelo do animal BN levou a redução da pressão arterial sistólica basal (~14 mm Hg). O objetivo deste trabalho foi identificar as variantes genéticas candidatas neste intervalo cromossômico com base em diferenças no padrão de expressão gênica e na presença de alterações genéticas não sinônimas \"missense\" ou em regiões regulatórias conservadas que possam estar envolvidas na gênese da hipertensão. Identificamos 533 genes com expressão renal, dentre os 682 do intervalo, sendo que 28 apresentaram padrão de expressão diferente entre amostras de animais adultos (congênico vs. SHR) e seis apresentaram alterações não sinônimas \"missense\". É importante salientar que dos genes diferentemente expressos, encontramos alterações estruturais em regiões conservadas com potencial de participar na regulação em 11. Em conjunto, utilizamos uma plataforma integrada para selecionar 34 genes candidatos no cromossomo 4, dos quais 17 genes serão priorizados, para ser investigados quanto sua contribuição na hipertensão arterial do SHR e na hipertensão primária humana / Total genome scan in genetic models of complex diseases have been instrumental to select candidate genes underlying complex traits. We previously mapped 5 blood pressure related quantitative trait loci (BP-QTLs) that explain about 43% of the BP variance in a progeny derived from Spontaneous Hypertensive Rat (SHR) and Brown Norway (BN) rats. The BP-QTLs were then validated by derivation of congenic strains, including one for chromosome 4 (SHR.BN4) in which a segment from BN replaced the SHR sequences reducing basal systolic BP (~14 mm Hg). The aim of this project is to identify the candidate genetic variants within the chromosome interval based on differences in renal gene expression patterns and structural changes in both non-synonymous missense or within adjacent regulatory sequences that may contribute to hypertension. We identified 533 genes with renal expression, out of 682 in the interval, in which 28 presented differences in expression pattern in adult samples (congenic vs. SHR) and six presented non-synonymous missense alterations. In addition, 11 out of 28 differentially expressed genes showed structural alterations in adjacent conserved regions that potentially contribute to gene regulation. Taken together, using the proposed combination of strategies, we selected 34 hypertensive candidate genes in chromosome 4, in which 17 will be prioritized, to be further explored to assess their contribution to hypertension in the SHR and to essential hypertension in humans

A hipertensão arterial essencial é caracterizada pela elevação crônica da pressão arterial e representa o principal fator de risco para doenças cardiovasculares e renais. O rim participa do controle da pressão arterial e alterações intrínsecas no manuseio renal de sódio desempenham papel importante na patogênese da hipertensão essencial. Os túbulos proximais renais são responsáveis pela reabsorção da maior parte do sódio filtrado nos glomérulos e a maior parte da reabsorção de sódio neste segmento faz-se através da troca de Na+ por H+ em membrana apical, mediada pela isoforma 3 do trocador Na+/H+ (NHE3). Entretanto, os dados existentes referentes à modulação renal do NHE3 em modelos de hipertensão são ainda conflitantes. Este estudo teve como objetivo avaliar as possíveis alterações funcionais do trocador Na+/H+ NHE3 em túbulo proximal renal na linhagem SHR no estágio de préhipertensão (5 semanas) e de hipertensão (14 semanas) e investigar se estas alterações são acompanhadas de alterações na atividade e na expressão da proteína cinase A (PKA) e de proteínas fosfatase-1 (PP1). Por meio de microperfusão estacionária in vivo mediu-se a atividade do NHE3 em túbulo proximal e verificou-se que a reabsorção de bicarbonato foi reduzida em 62 ± 6 % (P < 0,001) na transição do J-SHR para o A-SHR enquanto foi aumentada em 113 ± 10 % (P < 0,001) na transição entre o J-WKY e o A-WKY. A atividade estimulada do NHE3 em J-SHR é decorrente da redistribuição do NHE3 do domínio intermicrovilar (IMV) para o domínio das microvilosidades (MMV) e do baixo nível de fosforilação da serina 552, sítio consenso para a PKA. Por outro lado, durante a fase de hipertensão, a atividade diminuída do NHE3 deve-se à sua redistribuição para o IMV e ao aumento da fosforilação na serina 552. Para testar a hipótese de que os níveis de fosforilação do NHE3 estariam aumentados em túbulo proximal de SHR adulto devido ao aumento da atividade da PKA e/ou à diminuição na atividade da PP1, foram avaliados tanto os níveis de fosforilação quanto a atividade do NHE3 em SHR jovens e adultos em resposta ao 6MB-cAMP (análogo ao cAMP que ativa especificamente a PKA). O JSHR apresentou um aumento tanto nos níveis de fosforilação da serina 552 (179 ± 14 %, P < 0,001) quanto nos de inibição da atividade (65 ± 10 %, P < 0,001) do NHE3 em relação ao J-SHR em resposta ao 6MB-cAMP. Já no A-SHR, a fosforilação da serina 552 aumentou moderadamente (36 ± 4 %, P < 0,01), assim como inibiu moderadamente (23 ± 9 %, P < 0,05) a atividade do NHE3 em resposta ao 6MBcAMP. Adicionalmente, verificou-se que não houve alteração da atividade da PKA entre os animais nem ao longo da idade e nem entre as linhagens. Por sua vez, o JSHR apresentou maior atividade da PP1 que o A-SHR (1640 ± 107 vs. 940 ± 119 pM/?g, P < 0,01). Além disso, houve uma diminuição na expressão da PP1? no ASHR (32 ± 8 %, P < 0,01) quando comparado ao J-SHR. Os dados sugerem que o NHE3 é diferencialmente regulado antes e após o desenvolvimento da hipertensão em SHR por mecanismos que envolvem modificações pós-transcricionais e distribuição subcelular. Além do mais, a regulação diferencial dos níveis de fosforilação do NHE3 tubular proximal antes e após o desenvolvimento da hipertensão em SHR é devida, provavelmente, a alterações na atividade e na expressão da PP1 / Essential hypertension is characterized by chronic elevation of blood pressure and represents the major risk factor for cardiovascular and renal diseases. The kidney participates in the blood pressure control and intrinsic changes in renal sodium handling play an important role in the pathogenesis of essential hypertension. The renal proximal tubule is responsible for reabsorption of the great majority of sodium that is filtered by the glomerulus and the principal apical membrane mechanism for sodium reabsorption in this nephron is Na+/H+ exchanger isoform 3 (NHE3)- mediated Na+/H+ exchange. However, conflicting data have been reported with regard to NHE3 modulation in experimental models of hypertension. This study aimed to evaluate the possible functional changes of the Na+/H+ exchanger NHE3 in the renal proximal tubule of SHR both at the pre-hypertensive (5 weeks) and at hypertensive (14 weeks) stages and to investigate whether these changes were accompanied by changes in the activity and/or expression of protein kinase A (PKA) and protein phosphatase 1 (PP1). Proximal tubule NHE3 activity was measured by means of stationary microperfusion. Bicarbonate reabsorption was found to be decreased by 62 ± 6 % (P < 0.001) in the transition from youth to adulthood in SHR (Y-SHR to A-SHR), whereas in the transition from Y-WKY to A-WKY it increased by 113 ± 10 % (P < 0.001). Stimulated NHE3 activity in Y-SHR was due to redistribution of NHE3 from intermicrovilar domain (IMV) to microvilar domain (MMV) and to a lower level of serine 552 phosphorylation, a consensus site for PKA. Conversely, during the hypertensive stage, decreased NHE3 activity was due to increased redistribution of NHE3 to the IMV domain and increased phosphorylation at serine 552. To test the hypothesis that the increased levels of NHE3 phosphorylation in the proximal tubule of adult SHR were due to increased PKA activity and/or decreased PP1 activity, it was evaluated both phosphorylation levels and activity of NHE3 in young and adult SHR in response to 6MB-cAMP (an cAMP analog that specifically activates PKA). Y-SHR showed an increase both in the phosphorylation levels at serine 552 (179 ± 14 %, P < 0.001) and in the inhibition of NHE3 transport activity (65 ± 10 %, P < 0.001) compared to Y-SHR in response to 6MB-cAMP. With respect to A-SHR, the phosphorylation of serine 552 was slightly increased (36 ± 4 %, P < 0.01) and NHE3 activity was mildly inhibited (23 ± 9 %, P < 0.05) in response to 6MB-cAMP. Additionally, PKA activity remained unchanged with both age and strain. Nevertheless, Y-SHR exhibited higher PP1 activity than A-SHR (1640 ± 107 vs. 940 ± 119 pM/?g, P < 0.01). Furthermore, PP1? expression was decreased in the renal cortex of A-SHR (32 ± 8 %, P < 0.01) compared to Y-SHR. Taken together, these data suggest that NHE3 is differentially regulated before and after development of hypertension in SHR by mechanisms involving post-translational modifications and subcellular distribution. Moreover, the differential regulation of proximal tubule NHE3 phosphorylation levels before and after development of hypertension in SHR is most likely due to changes on the activity and expression of PP1