A first aim of my studies was that of investigating the effect of intravenous administration of dipyridamole in a model of chronic ischemia obtained by bilateral common carotid artery occlusion (2VO). Chronic ischemia is a progressive, dynamic process, caused by cerebral hypoperfusion that may manifest with cognitive dysfunction as ischemic conditions persist, and ultimately leads to neuronal death. No pharmacological therapy able to improve cognitive performances of patients with chronic vascular pathologies is yet available. Dipyridamole was introduced into clinical medicine in the early 1960s as a coronary vasodilator. It is a potent inhibitor of platelet activation, reduces formation of thrombi in vivo and it is an antithrombotic agent used for secondary stroke prevention in combination with aspirin. Recent evidence indicates that dipyridamole has anti-inflammatory properties. Chronic cerebral hypoperfusion in the rat induced by 2VO is recognized as a valid model of chronic cerebral hypotension and it is assumed that it mimics, although with some limitations, a situation of chronic hypoperfusion-induced human dementia. Our results show that dipyridamole reverses the impairment of spatial working memory, evaluated by Y-Maze test, 90 days after 2VO. This protective effect might be in relation to dipyridamole’s anti-inflammatory properties. A subsequent goal of my studies was that of investigating on the source of extracellular adenosine release, under physiological and ischemic conditions, in the striatum of rat, using a selective inhibitor of the ATP degradation, POM-4 (100µM). Under physiological conditions, in the presence of POM4, the extracellular concentration of ATP increased significantly but the adenosine concentration was not altered. This result demonstrates that, under physiological conditions, adenosine is not a product of extracellular ATP. To assess if adenosine outflow occurred through the equilibrative nucleoside transporter (ENT), the adenosine transporter inhibitor, dipyridamole (100 μmol/L), was added to PV4. In the presence of PV4 plus dipyridamole, adenosine outflow progressively increased indicating that adenosine is transported in cells and it is not transported out of cells by ENT. By immunoelectron microscopy, we demonstrated the presence of the concentrative nucleoside transporter CNT2 on plasma and vesicle membranes isolated from the rat striatum. These results are in favor that adenosine is transported in vesicles and is released in an excitation-secretion manner under in vivo physiological conditions. Under ischemic conditions, induced by middle cerebral artery occlusion (MCAo), adenosine concentrations increased in vehicle-treated rats in the first 4 hours after MCAo. In the presence of POM-4, extracellular ATP concentrations were increased by about 10 times and adenosine concentrations significantly decreased. Results demonstrated that early after ischemia, extracellular ATP is hydrolyzed by ecto-nucleotidases and significantly contributes to the increase in extracellular adenosine. To establish the contribution of extracellular ATP to adenosine might constitute the basis for devising a correct putative purinergic strategy aimed at protection from ischemic damage. In the last part of my PhD studies, I investigated on the effect of the selective adenosine A2A receptor antagonist, SCH58261, at extended periods after focal ischemia induced by transient MCAo (1 hour) in the rat. SCH58261, chronically administered (i.p twice/day for 7 days), was protective from neurological deficit and motor deficiency 1 day after ischemia but it is not protective 7 days after tMCAo. Seven days after tMCAo, it did not protected against tissue infiltration. Our results indicate that SCH58261 is protective when administered in the first hours after ischemia by reducing cerebral excitotoxicity but it is not able to reduce infiltration and inflammatory cascade in the ischemic brain area at extended periods after ischemia.

Modalità di liberazione di adenosina ed effetti protettivi del dipiridamolo ed antagonista dei recettori A2A adenosinici in modelli di ischemia cerebrale cronica e focale / Corti F.. - STAMPA. - (2013).

Modalità di liberazione di adenosina ed effetti protettivi del dipiridamolo ed antagonista dei recettori A2A adenosinici in modelli di ischemia cerebrale cronica e focale

CORTI, FRANCESCA
2013

Abstract

A first aim of my studies was that of investigating the effect of intravenous administration of dipyridamole in a model of chronic ischemia obtained by bilateral common carotid artery occlusion (2VO). Chronic ischemia is a progressive, dynamic process, caused by cerebral hypoperfusion that may manifest with cognitive dysfunction as ischemic conditions persist, and ultimately leads to neuronal death. No pharmacological therapy able to improve cognitive performances of patients with chronic vascular pathologies is yet available. Dipyridamole was introduced into clinical medicine in the early 1960s as a coronary vasodilator. It is a potent inhibitor of platelet activation, reduces formation of thrombi in vivo and it is an antithrombotic agent used for secondary stroke prevention in combination with aspirin. Recent evidence indicates that dipyridamole has anti-inflammatory properties. Chronic cerebral hypoperfusion in the rat induced by 2VO is recognized as a valid model of chronic cerebral hypotension and it is assumed that it mimics, although with some limitations, a situation of chronic hypoperfusion-induced human dementia. Our results show that dipyridamole reverses the impairment of spatial working memory, evaluated by Y-Maze test, 90 days after 2VO. This protective effect might be in relation to dipyridamole’s anti-inflammatory properties. A subsequent goal of my studies was that of investigating on the source of extracellular adenosine release, under physiological and ischemic conditions, in the striatum of rat, using a selective inhibitor of the ATP degradation, POM-4 (100µM). Under physiological conditions, in the presence of POM4, the extracellular concentration of ATP increased significantly but the adenosine concentration was not altered. This result demonstrates that, under physiological conditions, adenosine is not a product of extracellular ATP. To assess if adenosine outflow occurred through the equilibrative nucleoside transporter (ENT), the adenosine transporter inhibitor, dipyridamole (100 μmol/L), was added to PV4. In the presence of PV4 plus dipyridamole, adenosine outflow progressively increased indicating that adenosine is transported in cells and it is not transported out of cells by ENT. By immunoelectron microscopy, we demonstrated the presence of the concentrative nucleoside transporter CNT2 on plasma and vesicle membranes isolated from the rat striatum. These results are in favor that adenosine is transported in vesicles and is released in an excitation-secretion manner under in vivo physiological conditions. Under ischemic conditions, induced by middle cerebral artery occlusion (MCAo), adenosine concentrations increased in vehicle-treated rats in the first 4 hours after MCAo. In the presence of POM-4, extracellular ATP concentrations were increased by about 10 times and adenosine concentrations significantly decreased. Results demonstrated that early after ischemia, extracellular ATP is hydrolyzed by ecto-nucleotidases and significantly contributes to the increase in extracellular adenosine. To establish the contribution of extracellular ATP to adenosine might constitute the basis for devising a correct putative purinergic strategy aimed at protection from ischemic damage. In the last part of my PhD studies, I investigated on the effect of the selective adenosine A2A receptor antagonist, SCH58261, at extended periods after focal ischemia induced by transient MCAo (1 hour) in the rat. SCH58261, chronically administered (i.p twice/day for 7 days), was protective from neurological deficit and motor deficiency 1 day after ischemia but it is not protective 7 days after tMCAo. Seven days after tMCAo, it did not protected against tissue infiltration. Our results indicate that SCH58261 is protective when administered in the first hours after ischemia by reducing cerebral excitotoxicity but it is not able to reduce infiltration and inflammatory cascade in the ischemic brain area at extended periods after ischemia.
2013
Felicita Pedata
Corti F.
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Utilizza questo identificatore per citare o creare un link a questa risorsa: https://hdl.handle.net/2158/797482
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