RESUMEN
Stroke is the second-leading cause of death worldwide, yet there are no drugs available to protect the brain from stroke-induced neuronal injury. Acid-sensing ion channel 1a (ASIC1a) is the primary acid sensor in mammalian brain and a key mediator of acidosis-induced neuronal damage following cerebral ischemia. Genetic ablation and selective pharmacologic inhibition of ASIC1a reduces neuronal death following ischemic stroke in rodents. Here, we demonstrate that Hi1a, a disulfide-rich spider venom peptide, is highly neuroprotective in a focal model of ischemic stroke. Nuclear magnetic resonance structural studies reveal that Hi1a comprises two homologous inhibitor cystine knot domains separated by a short, structurally well-defined linker. In contrast with known ASIC1a inhibitors, Hi1a incompletely inhibits ASIC1a activation in a pH-independent and slowly reversible manner. Whole-cell, macropatch, and single-channel electrophysiological recordings indicate that Hi1a binds to and stabilizes the closed state of the channel, thereby impeding the transition into a conducting state. Intracerebroventricular administration to rats of a single small dose of Hi1a (2 ng/kg) up to 8 h after stroke induction by occlusion of the middle cerebral artery markedly reduced infarct size, and this correlated with improved neurological and motor function, as well as with preservation of neuronal architecture. Thus, Hi1a is a powerful pharmacological tool for probing the role of ASIC1a in acid-mediated neuronal injury and various neurological disorders, and a promising lead for the development of therapeutics to protect the brain from ischemic injury.
Asunto(s)
Bloqueadores del Canal Iónico Sensible al Ácido/administración & dosificación , Canales Iónicos Sensibles al Ácido/metabolismo , Fármacos Neuroprotectores/administración & dosificación , Venenos de Araña/administración & dosificación , Accidente Cerebrovascular/tratamiento farmacológico , Bloqueadores del Canal Iónico Sensible al Ácido/química , Bloqueadores del Canal Iónico Sensible al Ácido/farmacología , Animales , Modelos Animales de Enfermedad , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Espectroscopía de Resonancia Magnética , Masculino , Fármacos Neuroprotectores/farmacología , Ratas , Venenos de Araña/química , Venenos de Araña/farmacología , Accidente Cerebrovascular/complicaciones , Accidente Cerebrovascular/metabolismoRESUMEN
It is quite well established that activation of the so-called protective arms of the renin-angiotensin system (RAS), involving both AT2 and Mas receptors, provides a counter-regulatory role to AT1 receptor overactivity that may drive pathological changes in the cardiovascular system. In this brief review, we will focus on recent evidence that identifies at least three different pathways that may be effective in the setting of stroke and may be complementary with AT1 receptor blockade. Such mechanisms include AT2 receptor stimulation, Mas receptor stimulation and insulin-regulated aminopeptidase blockade. This report highlights recent data demonstrating striking neuroprotective effects in preclinical models of stroke targeting each of these pathways, which may pave the way for translational opportunities in this field.
Asunto(s)
Antagonistas de Receptores de Angiotensina/uso terapéutico , Antihipertensivos/uso terapéutico , Hipertensión/tratamiento farmacológico , Sistema Renina-Angiotensina/efectos de los fármacos , Renina/metabolismo , Accidente Cerebrovascular/tratamiento farmacológico , Animales , Humanos , Sistema Renina-Angiotensina/fisiología , Accidente Cerebrovascular/metabolismoRESUMEN
Activation of acid-sensing ion channel 1a (ASIC1a) plays a major role in mediating acidosis-induced neuronal injury following a stroke. Therefore, the inhibition of ASIC1a is a potential therapeutic avenue for the treatment of stroke. Venom-peptide Hi1a, a selective and highly potent ASIC1a inhibitor, reduces the infarct size and functional deficits when injected into the brain after stroke in rodents. However, its efficacy when administered using a clinically relevant route of administration remains to be established. Therefore, the current investigation aims to examine the efficacy of systemically administered Hi1a, using two different models of stroke in different species. Mice were subjected to the filament model of middle cerebral artery occlusion (MCAO) and treated with Hi1a systemically using either a single- or multiple-dosing regimen. 24 h poststroke, mice underwent functional testing, and the brain infarct size was assessed. Rats were subjected to endothelin-1 (ET-1)-induced MCAO and treated with Hi1a intravenously 2 h poststroke. Rats underwent functional tests prior to and for 3 days poststroke, when infarct volume was assessed. Mice receiving Hi1a did not show any improvements in functional outcomes, despite a trend toward reduced infarct size. This trend for reduced infarct size in mice was consistent regardless of the dosing regimen. There was also a trend toward lower infarct size in rats treated with Hi1a. More specifically, Hi1a reduced the amount of damage occurring within the somatosensory cortex, which was associated with an improved sensorimotor function in Hi1a-treated rats. Thus, this study suggests that Hi1a or more brain-permeable ASIC1a inhibitors are a potential stroke treatment.
RESUMEN
It is quite well established that activation of the AT(2) receptor (AT(2)R) provides a counter-regulatory role to AT(1)R overactivity, particularly during pathological conditions. Indeed, a potential therapeutic role for the AT(2)R is currently being promulgated with the introduction of novel AT(2)R ligands such as compound 21 (C21). In this brief review, we will focus on recent evidence to suggest that AT(2)R exhibits promising organ protection in the context of the heart, kidney and brain, with inflammation and gender influencing outcome. However, this field is not without controversy since the 'flagship' ligand C21 has also come under scrutiny, although it is safe to say there is much evidence to support a potentially important role of AT(2)R in a number of cardiovascular diseases. This report updates recent data in this field.
Asunto(s)
Receptor de Angiotensina Tipo 2/fisiología , Angiotensina III/fisiología , Animales , Humanos , Riñón/fisiología , Fármacos Neuroprotectores , Oligopéptidos/metabolismo , Oligopéptidos/fisiología , Receptor de Angiotensina Tipo 2/agonistasRESUMEN
Microglia are the resident immune cells within the brain and retina, commonly known as the macrophages of the central nervous system (CNS). Microglia survey the surrounding milieu to eliminate invading microbes, clear cellular debris and enforce programmed cell death by removing apoptotic cells. Complementary to their 'house-keeping' role, microglia are capable of releasing brain-derived neurotrophic factor (BDNF), as well as various anti-inflammatory cytokines that sustain and support neuronal survival. Although microglia are essential for maintaining a healthy CNS, paradoxically they may undergo phenotypic changes to influence numerous neurodegenerative disorders, including Parkinson's disease and Alzheimer's disease. Understanding the underlying mechanisms that determine whether microglia are supportive or toxic could elucidate novel and more effective therapeutic targets to treat an array of neurological and retinal diseases. Although relatively little is known about the influences that evoke phenotypic changes in the microglial population, there is accumulating evidence illustrating an interaction with the renin-angiotensin system (RAS). The angiotensin AT1 and AT2 receptors may have differential roles in mediating the activity of microglia. Understanding the actions of these angiotensin receptors will be important in defining whether microglia are an important therapeutic target for RAS blockade in brain and ocular diseases.
Asunto(s)
Encéfalo/citología , Microglía/citología , Microglía/fisiología , Sistema Renina-Angiotensina/fisiología , Retina/citología , Animales , Regulación de la Expresión Génica/fisiologíaRESUMEN
The photothrombotic stroke model relies on the interaction between photosensitive-dye and light for clot formation. Interestingly, the relationship between the length of light exposure and stroke-outcome has never been examined. This model has yet to be established in the FVB/N strain, even though stroke-outcomes are strain-specific. Therefore, this study aimed to examine the effect of different lengths of light exposure in two strains of mice on photothrombotic stroke. Male FVB/N and C57Bl/6 mice were subjected to stroke using 15, 18, or 20-min light exposure. Mice underwent functional testing for up to 7 days. Infarct volume was assessed with thionin staining, and cellular responses to injury analysed via immunofluorescence at 7-days post-stroke. Blood brain barrier (BBB) breakdown was assessed using Evans blue dye at 4.5-h post-stroke. Increasing light exposure from 15 to 20-min increased infarct volume but not functional deficit. Interestingly, there were strain-specific differences in functional outcomes, with FVB/N mice having less deficit on the hanging wire test than C57BI/6 after 15-min of light exposure. The opposite was seen in the adhesive removal test. There was no difference in the number of neurons, astrocytes, microglia, macrophages, and T cells between the strains, despite FVB/N mice demonstrating greater BBB breakdown and an enlarged spleen post-stroke. Increasing light exposure systematically increases infarct volume but does not worsen functional outcomes. FVB/N and C57Bl/6 mice exhibit subtle differences in functional outcomes post stroke, which highlights the need to choose tests which are appropriate for the mouse strain being used.
Asunto(s)
Accidente Cerebrovascular , Animales , Barrera Hematoencefálica/metabolismo , Infarto , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones EndogámicosRESUMEN
BACKGROUND AND PURPOSE: The angiotensin II type 2 receptor (AT(2)R) is implicated to be neuroprotective in stroke, although this premise has not been directly tested. Therefore, we have examined the neuroprotective effect of AT(2)R stimulation after intracerebroventricular administration of AT(2)R agonist CGP42112 in a conscious rat model of stroke. METHODS: Spontaneously hypertensive rats were treated with either CGP42112 (0.1 to 10 ng/kg/min intracerebroventricularly) alone or in combination with the AT(2)R antagonist PD123319 (36 ng/kg/min intracerebroventricularly) beginning 5 days before stroke induction. A focal reperfusion model of stroke was induced in conscious spontaneously hypertensive rats by administering endothelin-1 to the middle cerebral artery through a surgically implanted cannula. Behavioral tests were used to assess the severity of neurological deficit as a result of the ischemic event. Cortical and striatal infarct volumes were measured 72 hours poststroke. RESULTS: Blood pressure was unaffected by treatments. CGP42112 dose-dependently reduced cortical infarct volume poststroke, and PD123319 abolished the neuroprotective effect of CGP42112. PD123319 had no effect on infarct volume alone. These results were consistent with the behavioral findings, indicating that CGP42112 reduced motor deficit on the ledged beam test at 72 hours poststroke and immunohistochemical analyses showing that CGP42112 increased neuronal survival and minimized the loss of AT(2)R expression in the infarcted region. CONCLUSIONS: Based on infarct, behavioral, and immunohistochemical data, these results indicate that centrally administered CGP42112 exhibits a neuroprotective effect, which was independent of blood pressure. Thus, for the first time, we have shown that central AT(2)R stimulation is neuroprotective in a conscious rat model of stroke.
Asunto(s)
Fármacos Neuroprotectores/farmacología , Oligopéptidos/farmacología , Receptor de Angiotensina Tipo 2/agonistas , Receptor de Angiotensina Tipo 2/metabolismo , Accidente Cerebrovascular/tratamiento farmacológico , Animales , Presión Sanguínea/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Infarto Cerebral/tratamiento farmacológico , Infarto Cerebral/metabolismo , Infarto Cerebral/patología , Estado de Conciencia , Modelos Animales de Enfermedad , Inmunohistoquímica , Inyecciones Intraventriculares , Masculino , Actividad Motora/efectos de los fármacos , Ratas , Ratas Endogámicas SHR , Accidente Cerebrovascular/metabolismo , Accidente Cerebrovascular/patología , Superóxidos/metabolismo , Vasodilatadores/farmacologíaRESUMEN
The renin angiotensin system (RAS) is intricately involved in normal cardiovascular homeostasis. Excessive stimulation by the octapeptide angiotensin II contributes to a range of cardiovascular pathologies and diseases via angiotensin type 1 receptor (AT1R) activation. On the other hand, tElsevier Inc.he angiotensin type 2 receptor (AT2R) is thought to counter-regulate AT1R function. In this review, we describe the enhanced expression and function of AT2R in various cardiovascular disease settings. In addition, we illustrate that the RAS consists of a family of angiotensin peptides that exert cardiovascular effects that are often distinct from those of Ang II. During cardiovascular disease, there is likely to be an increased functional importance of AT2R, stimulated by Ang II, or even shorter angiotensin peptide fragments, to limit AT1R-mediated overactivity and cardiovascular pathologies.
Asunto(s)
Enfermedades Cardiovasculares/metabolismo , Receptor de Angiotensina Tipo 2/fisiología , Sistema Renina-Angiotensina/fisiología , Envejecimiento , Angiotensinas/metabolismo , Angiotensinas/fisiología , Animales , Enfermedades Cardiovasculares/fisiopatología , Diabetes Mellitus/metabolismo , Humanos , Enfermedades Renales/metabolismo , Receptor de Angiotensina Tipo 1/metabolismoRESUMEN
Acid-sensing ion channel 1a (ASIC1a) is the primary acid sensor in mammalian brain and plays a major role in neuronal injury following cerebral ischemia. Evidence that inhibition of ASIC1a might be neuroprotective following stroke was previously obtained using "PcTx1 venom" from the tarantula Psalmopeous cambridgei. We show here that the ASIC1a-selective blocker PcTx1 is present at only 0.4% abundance in this venom, leading to uncertainty as to whether the observed neuroprotective effects were due to PcTx1 blockade of ASIC1a or inhibition of other ion channels and receptors by the hundreds of peptides and small molecules present in the venom. We therefore examined whether pure PcTx1 is neuroprotective in a conscious model of stroke via direct inhibition of ASIC1a. A focal reperfusion model of stroke was induced in conscious spontaneously hypertensive rats (SHR) by administering endothelin-1 to the middle cerebral artery via a surgically implanted cannula. Two hours later, SHR were treated with a single intracerebroventricular (i.c.v.) dose of PcTx1 (1 ng/kg), an ASIC1a-inactive mutant of PcTx1 (1 ng/kg), or saline, and ledged beam and neurological tests were used to assess the severity of symptomatic changes. PcTx1 markedly reduced cortical and striatal infarct volumes measured 72 h post-stroke, which correlated with improvements in neurological score, motor function and preservation of neuronal architecture. In contrast, the inactive PcTx1 analogue had no effect on stroke outcome. This is the first demonstration that selective pharmacological inhibition of ASIC1a is neuroprotective in conscious SHRs, thus validating inhibition of ASIC1a as a potential treatment for stroke.
Asunto(s)
Bloqueadores del Canal Iónico Sensible al Ácido/farmacología , Encéfalo/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Péptidos/farmacología , Venenos de Araña/farmacología , Accidente Cerebrovascular/tratamiento farmacológico , Canales Iónicos Sensibles al Ácido/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Encéfalo/metabolismo , Encéfalo/patología , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Endotelina-1 , Infarto de la Arteria Cerebral Media , Masculino , Actividad Motora/efectos de los fármacos , Mutación , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Péptidos/genética , Ratas Endogámicas SHR , Índice de Severidad de la Enfermedad , Venenos de Araña/genética , Arañas , Accidente Cerebrovascular/metabolismo , Accidente Cerebrovascular/patologíaRESUMEN
BACKGROUND: In this study, the neuroprotective effect of a novel nonpeptide AT2R agonist, C21, was examined in a conscious model of stroke to verify a class effect of AT2R agonists as neuroprotective agents. METHODS AND RESULTS: Spontaneously hypertensive rats (SHR) were pre-treated for 5 days prior to stroke with C21 alone or in combination with the AT2R antagonist PD123319. In a separate series of experiments C21 was administered in a series of 4 doses commencing 6 hours after stroke. A focal reperfusion model of ischemia was induced in conscious SHR by administering endothelin-1 to the middle cerebral artery (MCA). Motor coordination was assessed at 1 and 3 days after stroke and post mortem analyses of infarct volumes, microglia activation and neuronal survival were performed at 72 hours post MCA occlusion. When given prior to stroke, C21 dose dependently decreased infarct volume, which is consistent with the behavioural findings illustrating an improvement in motor deficit. During the pre-treatment protocol C21 was shown to enhance microglia activation, which are likely to be evoking protection by releasing brain derived neurotrophic factor. When drug administration was delayed until 6 hours after stroke, C21 still reduced brain injury. CONCLUSION: These results indicate that centrally administered C21 confers neuroprotection against stroke damage. This benefit is likely to involve various mechanisms, including microglial activation of endogenous repair and enhanced cerebroperfusion. Thus, we have confirmed the neuroprotective effect of AT2R stimulation using a nonpeptide compound which highlights the clinical potential of the AT2R agonists for future development.
Asunto(s)
Hipertensión/tratamiento farmacológico , Fármacos Neuroprotectores/uso terapéutico , Receptor de Angiotensina Tipo 2/agonistas , Receptor de Angiotensina Tipo 2/metabolismo , Animales , Endotelina-1/farmacología , Masculino , Ratas , Daño por Reperfusión/inducido químicamente , Daño por Reperfusión/tratamiento farmacológicoRESUMEN
We have demonstrated previously that pretreatment with an angiotensin II type 2 receptor (AT(2)R) agonist is neuroprotective against a subsequent stroke independent of any changes in blood pressure. Therefore, in the current study, we have examined the potential neuroprotective effect of AT(2)R stimulation initiated after stroke induction to mimic the clinical setting. Intracerebroventricular administration of the AT(2)R agonist CGP42112 was commenced 6 hours after an ischemic stroke had been induced in conscious spontaneously hypertensive rats. CGP42112 given over 4 doses in the same rats (3 µg/kg per dose centrally) at 6, 24, 48, and 72 hours after stroke induction reduced total infarct volume (32 ± 13 mm(3) versus vehicle, 170 ± 49 mm(3); P<0.05) and improved motor function. Furthermore, we have demonstrated that AT(2)R stimulation after stroke increased neuronal survival, decreased apoptosis, and caused an increase in the number of activated microglia in the core region of damage. The effects of CGP42112 were partially reversed with the coadministration of an AT(2)R antagonist, PD123319. Thus, the current study has shown for the first time that delayed central AT(2)R stimulation after a cerebral incident is neuroprotective in a conscious rat model of stroke.