O córtex pré-límbico (PL) é uma importante área límbica envolvida em vários processos funcionais correlatos ao estresse, tais como respostas cardiovasculares, hormonais e comportamentais. O modelo de estresse de restrição (ER) foi padronizado na literatura como uma situação aversiva capaz de promover aumento da pressão arterial e frequência cardíaca, queda da temperatura cutânea e estimulação do eixo hipotálamo-pituitária-adrenal (HPA). Trabalhos da literatura evidenciaram que ratos submetidos ao ER apresentavam aumento da atividade neuronial no PL, sugerindo que essa estrutura module respostas ao ER. Assim, a inibição temporária de sinapses no PL potencializou a resposta taquicárdica induzida pelo ER, sem alterar a resposta pressora. Além do controle cardiovascular, outros trabalhos demonstraram que o PL também participa do controle hormonal durante o ER. O ER agudo também está envolvido com a produção de espécies reativas de oxigênio (EROs), fator que pode estar envolvido nas alterações a longo prazo observadas após exposição a uma situação aversiva. O sistema renina angiotensina (SRA) central modula respostas cardiovasculares, inclusive aquelas induzidas por situações aversivas, além de ter um papel reconhecido na produção de EROs. Além disso, foi demonstrado que o PL possui SRA funcional com presença dos peptídeos a ele relacionados. Baseado nos fatos mencionados acima, a hipótese do presente projeto é que a neurotransmissão angiotensinérgica do PL está envolvida na modulação de respostas autonômicas (aumento de pressão arterial e frequência cardíaca, e queda da temperatura cutânea) e hormonal (aumento plasmático de corticosterona) evocadas pelo ER, e que essa via envolveria a formação de EROs. A microinjeção do inibidor da enzima conversora de angiotensina (ECA) lisinopril no PL, nas doses de 0,5 e 1nmol/100nL, reduziu a resposta pressora, sendo a dose de 1nmol/100nL de lisinopril também capaz de reduzir a resposta taquicárdica induzida pelo ER; porém nenhuma dose utilizada ocasionou mudanças na queda da temperatura cutânea evocada pelo ER. O pré-tratamento do PL com o antagonista de receptores do subtipo AT1 candesartan reduziu o efeito pressor induzido pelo ER, porém não alterou a resposta taquicárdica e queda da temperatura cutânea associadas ao ER. Por sua vez, o pré-tratamento com o antagonista de receptores do subtipo AT2, PD123177, reduziu a resposta taquicárdica sem alterar a resposta pressora e a queda da temperatura cutânea evocadas pelo ER. Em adição, o estresse de restrição agudo e os pré-tratamentos realizados não foram capazes de alterar a atividade da enzima NADPH oxidase no PL. Em conclusão, os presentes resultados sugerem a participação do SRA na modulação da resposta cardiovascular ao ER, através da ativação de receptores AT1, e AT2 do PL, afetando respectivamente, o componente vascular e o cardíaco da resposta autonômica causada pelo ER. Além disso, os resultados da atividade da enzima NADPH oxidase no PL sugerem que o ER agudo, os receptores AT1, AT2 e a ECA não modulam o status oxidativo local. / The prelimbic cortex is an important limbic structure involved in several stressrelated functional processes, such as cardiovascular, hormonal and behavior responses. Restraint stress (RS) was standardized in literature as an aversive situation able to promote blood pressure and heart rate increases, reduction in tail temperature and stimulation of the hypothalamic-pituitary-adrenal axis (HPA). Previous studies demonstrated that rats submitted to RS exhibited increased neuronal activity in the PL, suggesting that this structure modulates RS-evoked responses. Temporary, synaptic temporary inhibition in the PL markedly increased the RS-evoked tachycardiac response, without affecting the pressor one. Beyond cardiovascular control, other studies demonstrated that PL also participates in hormonal control during RS. Acute RS is also involved in the production of reactive oxygen species (ROS), which could be involved in long- term changes observed after exposure to an aversive situation. The central renin-angiotensin system (RAS) modulates cardiovascular responses, including those induced by aversive situations. In addition, this system has a well-known role in ROS production. Furthermore, the presence of angiotensinergic peptides in PL has also been demonstrated, suggesting the existence of a functional RAS in this structure. Based on the facts mentioned above, the hypothesis of the present study was that the angiotensinergic neurotransmission in PL is involved in the modulation of autonomic responses (blood pressure and heart rate increase, and reduction in tail temperature) evoked by RS, and this pathway would involve ROS formation. Microinjection of lisinopril (0.5 and 1nmol/100nL), an inhibitor of angiotensinconverting enzyme (ACE), into PL reduced the pressor response, and the dose 1nmol/nL was also able to reduce the tachycardiac response induced by RS; however, none of doses changed the reduction in tail temperature evoked by RS. PL treatment with candesartan, an AT1 receptors antagonist, reduced the RS-evoked pressor response, but did not affect the RS-evoked tachycardiac response and reduction in tail temperature. In addition, pretreatment with PD123177, an AT2 receptors antagonist, reduced the RS-evoked tachycardiac response, without affecting the pressor response or the RS-evoked reduction in tail temperature. In addition, neither acute RS or local treatments affected NADPH oxidase activity in the PL. In conclusion, the present results suggests the involvement of the central RAS in the modulation of the cardiovascular responses caused by RS, through the activation of both AT1 and AT2 receptors in the PL. The PL AT1 receptors modulating the vascular, and the AT2 modulating the cardiac component of RS-evoked autonomic response. Furthermore, our study suggests that neither acute RS or local AT1, AT2 and ACE affect oxidative status in the PL.

O córtex pré-límbico (PL) é uma importante área límbica envolvida em vários processos funcionais correlatos ao estresse, tais como respostas cardiovasculares, hormonais e comportamentais. O modelo de estresse de restrição (ER) foi padronizado na literatura como uma situação aversiva capaz de promover aumento da pressão arterial e frequência cardíaca, queda da temperatura cutânea e estimulação do eixo hipotálamo-pituitária-adrenal (HPA). Trabalhos da literatura evidenciaram que ratos submetidos ao ER apresentavam aumento da atividade neuronial no PL, sugerindo que essa estrutura module respostas ao ER. Assim, a inibição temporária de sinapses no PL potencializou a resposta taquicárdica induzida pelo ER, sem alterar a resposta pressora. Além do controle cardiovascular, outros trabalhos demonstraram que o PL também participa do controle hormonal durante o ER. O ER agudo também está envolvido com a produção de espécies reativas de oxigênio (EROs), fator que pode estar envolvido nas alterações a longo prazo observadas após exposição a uma situação aversiva. O sistema renina angiotensina (SRA) central modula respostas cardiovasculares, inclusive aquelas induzidas por situações aversivas, além de ter um papel reconhecido na produção de EROs. Além disso, foi demonstrado que o PL possui SRA funcional com presença dos peptídeos a ele relacionados. Baseado nos fatos mencionados acima, a hipótese do presente projeto é que a neurotransmissão angiotensinérgica do PL está envolvida na modulação de respostas autonômicas (aumento de pressão arterial e frequência cardíaca, e queda da temperatura cutânea) e hormonal (aumento plasmático de corticosterona) evocadas pelo ER, e que essa via envolveria a formação de EROs. A microinjeção do inibidor da enzima conversora de angiotensina (ECA) lisinopril no PL, nas doses de 0,5 e 1nmol/100nL, reduziu a resposta pressora, sendo a dose de 1nmol/100nL de lisinopril também capaz de reduzir a resposta taquicárdica induzida pelo ER; porém nenhuma dose utilizada ocasionou mudanças na queda da temperatura cutânea evocada pelo ER. O pré-tratamento do PL com o antagonista de receptores do subtipo AT1 candesartan reduziu o efeito pressor induzido pelo ER, porém não alterou a resposta taquicárdica e queda da temperatura cutânea associadas ao ER. Por sua vez, o pré-tratamento com o antagonista de receptores do subtipo AT2, PD123177, reduziu a resposta taquicárdica sem alterar a resposta pressora e a queda da temperatura cutânea evocadas pelo ER. Em adição, o estresse de restrição agudo e os pré-tratamentos realizados não foram capazes de alterar a atividade da enzima NADPH oxidase no PL. Em conclusão, os presentes resultados sugerem a participação do SRA na modulação da resposta cardiovascular ao ER, através da ativação de receptores AT1, e AT2 do PL, afetando respectivamente, o componente vascular e o cardíaco da resposta autonômica causada pelo ER. Além disso, os resultados da atividade da enzima NADPH oxidase no PL sugerem que o ER agudo, os receptores AT1, AT2 e a ECA não modulam o status oxidativo local. / The prelimbic cortex is an important limbic structure involved in several stressrelated functional processes, such as cardiovascular, hormonal and behavior responses. Restraint stress (RS) was standardized in literature as an aversive situation able to promote blood pressure and heart rate increases, reduction in tail temperature and stimulation of the hypothalamic-pituitary-adrenal axis (HPA). Previous studies demonstrated that rats submitted to RS exhibited increased neuronal activity in the PL, suggesting that this structure modulates RS-evoked responses. Temporary, synaptic temporary inhibition in the PL markedly increased the RS-evoked tachycardiac response, without affecting the pressor one. Beyond cardiovascular control, other studies demonstrated that PL also participates in hormonal control during RS. Acute RS is also involved in the production of reactive oxygen species (ROS), which could be involved in long- term changes observed after exposure to an aversive situation. The central renin-angiotensin system (RAS) modulates cardiovascular responses, including those induced by aversive situations. In addition, this system has a well-known role in ROS production. Furthermore, the presence of angiotensinergic peptides in PL has also been demonstrated, suggesting the existence of a functional RAS in this structure. Based on the facts mentioned above, the hypothesis of the present study was that the angiotensinergic neurotransmission in PL is involved in the modulation of autonomic responses (blood pressure and heart rate increase, and reduction in tail temperature) evoked by RS, and this pathway would involve ROS formation. Microinjection of lisinopril (0.5 and 1nmol/100nL), an inhibitor of angiotensinconverting enzyme (ACE), into PL reduced the pressor response, and the dose 1nmol/nL was also able to reduce the tachycardiac response induced by RS; however, none of doses changed the reduction in tail temperature evoked by RS. PL treatment with candesartan, an AT1 receptors antagonist, reduced the RS-evoked pressor response, but did not affect the RS-evoked tachycardiac response and reduction in tail temperature. In addition, pretreatment with PD123177, an AT2 receptors antagonist, reduced the RS-evoked tachycardiac response, without affecting the pressor response or the RS-evoked reduction in tail temperature. In addition, neither acute RS or local treatments affected NADPH oxidase activity in the PL. In conclusion, the present results suggests the involvement of the central RAS in the modulation of the cardiovascular responses caused by RS, through the activation of both AT1 and AT2 receptors in the PL. The PL AT1 receptors modulating the vascular, and the AT2 modulating the cardiac component of RS-evoked autonomic response. Furthermore, our study suggests that neither acute RS or local AT1, AT2 and ACE affect oxidative status in the PL.

Zugl.: Dresden, Techn. Univ., Habil.-Schr., 2008

Non invasive in vivo imaging will lead to better understanding of Angiotensin II Type 1 Receptor’s (AT1R) role in disease progression and may guide therapy in cardiovascular patients. Two models were used in this project: 5/6 nephrectomy and transient left anterior descending (LAD) ligation. Rats were scanned with [13N]ammonia and [11C]methyl-candesartan, both of which are Positron Emission Tomography (PET) tracers, at 8 weeks (nephrectomy) and 2 weeks (LAD ligation) after surgery. Western blot analysis was used to corroborate PET data. Nephrectomy: Renal AT1R image analysis displayed a 40% decrease in kidney AT1R in nephrectomized animals compared to sham (p<0.05) which was confirmed with Western blot and biodistribution. LAD ligation: Left Ventricle AT1R Western blot analysis exhibited a 60% increase in 20min ligation (p<0.05) with maintained myocardial blood flow. In conclusion, changes in renal AT1R were successfully imaged using [11C]methyl-candesartan in nephrectomized animals, and 20min LAD ligation/reperfusion is an appropriate model to image an increase in cardiac AT1R following ischemic injury.

Non invasive in vivo imaging will lead to better understanding of Angiotensin II Type 1 Receptor’s (AT1R) role in disease progression and may guide therapy in cardiovascular patients. Two models were used in this project: 5/6 nephrectomy and transient left anterior descending (LAD) ligation. Rats were scanned with [13N]ammonia and [11C]methyl-candesartan, both of which are Positron Emission Tomography (PET) tracers, at 8 weeks (nephrectomy) and 2 weeks (LAD ligation) after surgery. Western blot analysis was used to corroborate PET data. Nephrectomy: Renal AT1R image analysis displayed a 40% decrease in kidney AT1R in nephrectomized animals compared to sham (p<0.05) which was confirmed with Western blot and biodistribution. LAD ligation: Left Ventricle AT1R Western blot analysis exhibited a 60% increase in 20min ligation (p<0.05) with maintained myocardial blood flow. In conclusion, changes in renal AT1R were successfully imaged using [11C]methyl-candesartan in nephrectomized animals, and 20min LAD ligation/reperfusion is an appropriate model to image an increase in cardiac AT1R following ischemic injury.

Non invasive in vivo imaging will lead to better understanding of Angiotensin II Type 1 Receptor’s (AT1R) role in disease progression and may guide therapy in cardiovascular patients. Two models were used in this project: 5/6 nephrectomy and transient left anterior descending (LAD) ligation. Rats were scanned with [13N]ammonia and [11C]methyl-candesartan, both of which are Positron Emission Tomography (PET) tracers, at 8 weeks (nephrectomy) and 2 weeks (LAD ligation) after surgery. Western blot analysis was used to corroborate PET data. Nephrectomy: Renal AT1R image analysis displayed a 40% decrease in kidney AT1R in nephrectomized animals compared to sham (p<0.05) which was confirmed with Western blot and biodistribution. LAD ligation: Left Ventricle AT1R Western blot analysis exhibited a 60% increase in 20min ligation (p<0.05) with maintained myocardial blood flow. In conclusion, changes in renal AT1R were successfully imaged using [11C]methyl-candesartan in nephrectomized animals, and 20min LAD ligation/reperfusion is an appropriate model to image an increase in cardiac AT1R following ischemic injury.

Non invasive in vivo imaging will lead to better understanding of Angiotensin II Type 1 Receptor’s (AT1R) role in disease progression and may guide therapy in cardiovascular patients. Two models were used in this project: 5/6 nephrectomy and transient left anterior descending (LAD) ligation. Rats were scanned with [13N]ammonia and [11C]methyl-candesartan, both of which are Positron Emission Tomography (PET) tracers, at 8 weeks (nephrectomy) and 2 weeks (LAD ligation) after surgery. Western blot analysis was used to corroborate PET data. Nephrectomy: Renal AT1R image analysis displayed a 40% decrease in kidney AT1R in nephrectomized animals compared to sham (p<0.05) which was confirmed with Western blot and biodistribution. LAD ligation: Left Ventricle AT1R Western blot analysis exhibited a 60% increase in 20min ligation (p<0.05) with maintained myocardial blood flow. In conclusion, changes in renal AT1R were successfully imaged using [11C]methyl-candesartan in nephrectomized animals, and 20min LAD ligation/reperfusion is an appropriate model to image an increase in cardiac AT1R following ischemic injury.

Les douleurs neuropathiques se caractérisent par l’apparition de symptômes positifs tels qu’une allodynie et de symptômes négatifs tel qu’une hypoalgésie. Les douleurs neuropathiques ont un retentissement important sur la qualité de vie et il n’existe à ce jour aucune thérapie efficace pour leur prise en charge préventive. Récemment, un système rénine-angiotensine tissulaire a été mis en évidence au sein du système nerveux périphérique sensitif et il a été démontré que sa modulation pharmacologique modifie la perception douloureuse chez l’animal. Dans ce travail, nous nous sommes intéressés à la physiopathologie et à la prévention thérapeutique des neuropathies sensitives par des modulateurs du SRA. Pour cela, deux modèles murins de neuropathie sensitive à l’origine de douleurs neuropathiques ont été utilisés ; un modèle de neuropathie induite par la résinifératoxine (RTX), toxine naturelle spécifique des petites fibres nociceptives et un modèle de neuropathie induite par la vincristine (VCR), un agent anticancéreux particulièrement neurotoxique. Une étude pharmacologique menée sur la neuropathie induite par la RTX nous a permis de mettre en évidence que seul le candésartan prévenait le développement de la neuropathie et que son effet était AT2R-dépendant. L’efficacité du candésartan a été confirmée dans le modèle de neuropathie chimio-induite, développé et caractérisé au cours de ce travail. Ce modèle de neuropathie induite par la VCR a permis de révéler, pour la première fois, le potentiel neuroprotecteur du C21 (agoniste direct du récepteur AT2R, Vicore Pharma) dans un contexte de neuropathie périphérique. L’ensemble de ces résultats confirme l’intérêt de la stimulation du récepteur AT2R dans le traitement des douleurs neuropathiques associées à une chimiothérapie, et plus largement d’origine toxique. / Neuropathic pain was characterized by positive symptoms as allodynia and negative symptoms as hypoalgesia. Neuropathic pain has a major impact on patient’s quality of life and there is, currently, no specific treatment for its preventive management. Recently, a specific renin angiotensin system in sensory peripheral nervous system has been showed and it has been demonstrated that its pharmacological modulation could modify pain perception in animals. In this work, we studied the neuroprotective potential of RAS modulators in two animal models of sensory neuropathy leading to neuropathic pain; a model of neuropathy induced by resiniferatoxin (RTX), a specific natural toxin of nociceptive nerve fibers, and a model of neuropathy induced by vincristine (VCR), a neurotoxic anticancer agent. Pharmacological study on mice with RTX-induced neuropathy allowed to conclude that only candesartan was neuroprotective and that its effect was AT2R-dependent.The effective neuroprotective effect of candesartan was confirmed on the model of VCR-induced neuropathy which was previously developed and characterized. This VCR-induced neuropathy mouse model allowed to demonstrate, for the first time, that C21 (a direct AT2R receptor agonist, Vicore Pharma) was neuroprotective against a peripheral neuropathy. All these results confirm the interest of stimulation of the AT2R receptor in the treatment of neuropathic pain associated with chemotherapy and more generally of toxic origin.

<p>Live donor nephrectomy (LDN) is a unique surgical challenge where surgery is performed on healthy individuals. It is of great importance to keep the morbidity of donors as low as possible, as well as harvesting a kidney in optimal condition. Lowering morbidity is the motive for introducing the endoscopic technique in LDN. Oliguria and impaired kidney function can, however, be seen during pneumoperitoneum and endoscopic LDN have been criticized for not yet being proven safe enough.</p><p>The aims of this study were to investigate the changes in renal function during elevated intraabdominal pressure (IAP) in donors and rats and to evaluate donor morbidity and safety of the new endoscopic techniques compared to the open LDN.</p><p>In two studies, a rat model was used. It was found that elevation of IAP diminished glomerular filtration rate (GFR). Cardiac output (CO) and renal blood flow decreased as well. Elevation of IAP activates the renin system and aldosterone was increased. Acute angiotensin II receptor 1 blockade (candesartan) treatment lowered blood pressure significantly and impaired renal function during elevated IAP. Volume expansion prior to, and during, pneumoperitoneum reduces the deleterious effects on renal function.</p><p>Three studies on kidney live donors show that traditional laparoscopic surgery (TLS) takes longer time to perform than open LDN. Hand-assistance facilitates the operation and increases the safety margin as well as shortens the operation by 27% compared to TLS. Evaluation of a hand-assisted retroperitoneoscopy (HARS), performed for the first time ever in Uppsala 2001, show that the operation is short and safe, the donors experience little pain and the renal function is favourable compared to open surgery, TLS and hand-assisted transperitoneal laparoscopic approaches.</p><p>In conclusion, the results indicate that elevated IAP decreases GFR due to decreased CO and activation of the RAAS, which can be avoided with adequate hydration. Endoscopy can be facilitated if hand-assistance is applied and in particular hand-assisted retroperitoneoscopic nephrectomy shows advantages for the donor.</p>

Live donor nephrectomy (LDN) is a unique surgical challenge where surgery is performed on healthy individuals. It is of great importance to keep the morbidity of donors as low as possible, as well as harvesting a kidney in optimal condition. Lowering morbidity is the motive for introducing the endoscopic technique in LDN. Oliguria and impaired kidney function can, however, be seen during pneumoperitoneum and endoscopic LDN have been criticized for not yet being proven safe enough. The aims of this study were to investigate the changes in renal function during elevated intraabdominal pressure (IAP) in donors and rats and to evaluate donor morbidity and safety of the new endoscopic techniques compared to the open LDN. In two studies, a rat model was used. It was found that elevation of IAP diminished glomerular filtration rate (GFR). Cardiac output (CO) and renal blood flow decreased as well. Elevation of IAP activates the renin system and aldosterone was increased. Acute angiotensin II receptor 1 blockade (candesartan) treatment lowered blood pressure significantly and impaired renal function during elevated IAP. Volume expansion prior to, and during, pneumoperitoneum reduces the deleterious effects on renal function. Three studies on kidney live donors show that traditional laparoscopic surgery (TLS) takes longer time to perform than open LDN. Hand-assistance facilitates the operation and increases the safety margin as well as shortens the operation by 27% compared to TLS. Evaluation of a hand-assisted retroperitoneoscopy (HARS), performed for the first time ever in Uppsala 2001, show that the operation is short and safe, the donors experience little pain and the renal function is favourable compared to open surgery, TLS and hand-assisted transperitoneal laparoscopic approaches. In conclusion, the results indicate that elevated IAP decreases GFR due to decreased CO and activation of the RAAS, which can be avoided with adequate hydration. Endoscopy can be facilitated if hand-assistance is applied and in particular hand-assisted retroperitoneoscopic nephrectomy shows advantages for the donor.