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1.
Int J Mol Sci ; 24(18)2023 Sep 19.
Article in English | MEDLINE | ID: mdl-37762606

ABSTRACT

Despite the overwhelming advances in the understanding of the pathogenesis of stroke, a devastating disease affecting millions of people worldwide, currently there are only a limited number of effective treatments available. Preclinical and clinical studies show that stroke is a sexually dimorphic disorder, affecting males and females differently. Strong experimental evidence indicates that estrogen may play a role in this difference and that exogenous 17ß-estradiol (E2) is neuroprotective against stroke in both male and female rodents. However, the molecular mechanisms by which E2 intervenes in ischemia-induced cell death, revealing these sex differences, remain unclear. The present study was aimed to determine, in female rats, the molecular mechanisms of two well-known pro-survival signaling pathways, MAPK/ERK1/2 and PI3K/Akt, that mediate E2 neuroprotection in response to acute ischemic stroke. E2 pretreatment reduced brain damage and attenuated apoptotic cell death in ovariectomized female rats after an ischemic insult. Moreover, E2 decreased phosphorylation of ERK1/2 and prevented ischemia/reperfusion-induced dephosphorylation of both Akt and the pro-apoptotic protein, BAD. However, MAPK/ERK1/2 inhibitor PD98059, but not the PI3K inhibitor LY294002, attenuated E2 neuroprotection. Thus, these results suggested that E2 pretreatment in ovariectomized female rats modulates MAPK/ERK1/2 and activates Akt independently of PI3K to promote cerebroprotection in ischemic stroke. A better understanding of the mechanisms and the influence of E2 in the female sex paves the way for the design of future successful hormone replacement therapies.

2.
STAR Protoc ; 4(4): 102630, 2023 Dec 15.
Article in English | MEDLINE | ID: mdl-38264871

ABSTRACT

Global cerebral ischemia occurs when blood flow to the entire brain is transiently blocked, which results in delayed neurologic deficits. Here, we present a protocol for performing the four-vessel occlusion rat model to study the neurodegeneration and cognitive deficits associated with global ischemia. We describe steps for carrying out the vertebral and common carotid artery occlusion which enables sufficient blockage of cerebral blood flow. We then detail expected outcomes using histology assays and behavioral tests. For complete details on the use and execution of this protocol, please refer to Chung et al. (2022).1.


Subject(s)
Cognition Disorders , Ischemia , Animals , Rats , Cerebrovascular Circulation , Biological Assay , Brain
3.
Front Cell Neurosci ; 15: 633610, 2021.
Article in English | MEDLINE | ID: mdl-34040505

ABSTRACT

Cerebral ischemia is a devastating disease that affects many people worldwide every year. The neurodegenerative damage as a consequence of oxygen and energy deprivation, to date, has no known effective treatment. The ischemic insult is followed by an inflammatory response that involves a complex interaction between inflammatory cells and molecules which play a role in the progression towards cell death. However, there is presently a matter of controversy over whether inflammation could either be involved in brain damage or be a necessary part of brain repair. The inflammatory response is triggered by inflammasomes, key multiprotein complexes that promote secretion of pro-inflammatory cytokines. An early event in post-ischemic brain tissue is the release of certain molecules and reactive oxygen species (ROS) from injured neurons which induce the expression of the nuclear factor-kappaB (NF-κB), a transcription factor involved in the activation of the inflammasome. There are conflicting observations related to the role of NF-κB. While some observe that NF-κB plays a damaging role, others suggest it to be neuroprotective in the context of cerebral ischemia, indicating the need for additional investigation. Here we discuss the dual role of the major inflammatory signaling pathways and provide a review of the latest research aiming to clarify the relationship between NF-κB mediated inflammation and neuronal death in cerebral ischemia.

4.
J Neuroendocrinol ; 31(8): e12751, 2019 08.
Article in English | MEDLINE | ID: mdl-31127971

ABSTRACT

Because neuroprotection in stroke should be revisited in the era of recanalisation, the present study analysed the potential neuroprotective effect of the selective oestrogen receptor modulator, bazedoxifene acetate (BZA), in an animal model of diabetic ischaemic stroke that mimics thrombectomy combined with adjuvant administration of a putative neuroprotectant. Four weeks after induction of diabetes (40 mg kg-1 streptozotocin, i.p.), male Wistar rats were subjected to transient middle cerebral artery occlusion (intraluminal thread technique, 60 minutes) and assigned to one of three groups treated with either: vehicle, BZA (3 mg kg-1  day-1 , i.p.) or 17ß-oestradiol (E2 ) (100 µg kg-1  day-1 , i.p.). At 24 hours post-ischaemia-reperfusion, brain damage (neurofunctional score, infarct size and apoptosis), expression of oestrogen receptors (ER)α, ERß and G protein-coupled oestrogen receptor), and activity of the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK)1/2 and phosphoinositide 3-kinase/Akt pathways were analysed. At 24 hours after the ischaemic insult, both BZA- and E2 -treated animals showed lower brain damage in terms of improved neurofunctional condition, decreased infarct size and decreased apoptotic cell death. Ischaemia-reperfusion induced a significant decrease in ERα and ERß expression without affecting that of G protein-coupled oestrogen receptor, whereas BZA and E2 reversed such a decrease. The ischaemic insult up-regulated the activity of both the MAPK/ERK1/2 and phosphoinositide 3-kinase/Akt pathways; BZA and E2 attenuated the increased activity of the ERK1/2 pathway, without affecting that of the Akt pathway. The results of the present study lend further support to the consideration of BZA as an effective and safer alternative overcoming the drawbacks of E2 with respect to improving diabetic ischaemic stroke outcome after successful reperfusion.


Subject(s)
Brain Ischemia/prevention & control , Diabetic Angiopathies/prevention & control , Estradiol/pharmacology , Indoles/pharmacology , Receptors, Estrogen/genetics , Stroke/prevention & control , Animals , Brain Ischemia/genetics , Brain Ischemia/metabolism , Brain Ischemia/pathology , Diabetes Mellitus, Experimental/complications , Diabetes Mellitus, Experimental/drug therapy , Diabetes Mellitus, Experimental/genetics , Diabetes Mellitus, Experimental/pathology , Diabetic Angiopathies/genetics , Diabetic Angiopathies/metabolism , Diabetic Angiopathies/pathology , MAP Kinase Signaling System/drug effects , MAP Kinase Signaling System/physiology , Male , Neuroprotective Agents/pharmacology , Rats , Rats, Wistar , Receptors, Estrogen/metabolism , Selective Estrogen Receptor Modulators/pharmacology , Streptozocin , Stroke/genetics , Stroke/metabolism , Stroke/pathology
5.
Int Neurourol J ; 23(Suppl 1): S11-21, 2019 Feb.
Article in English | MEDLINE | ID: mdl-30832463

ABSTRACT

PURPOSE: Transient global ischemia arising in human due to cardiac arrest causes selective, delayed neuronal death in hippocampal CA1 and cognitive impairment. Growth arrest and DNA-damage-inducible protein 45 beta (Gadd45b) is a wellknown molecule in both DNA damage-related pathogenesis and therapies. Emerging evidence suggests that Gadd45b is an anti-apoptotic factor in nonneuronal cells and is an intrinsic neuroprotective molecule in neurons. However, the mechanism of Gadd45b pathway is not fully examined in neurodegeneration associated with global ischemia. METHODS: Rats were subjected to transient global ischemia by the 4-vessel occlusion or sham operation. The animals were sacrificed at 24 hours, 48 hours, and 7 days after ischemia. The hippocampal CA1 was microdissected and processed to examine mRNA and protein level. To assess neuronal death, tissue sections were cut and processed for Fluoro-Jade and Nissl staining. RESULTS: Here we show that ischemic insults increase abundance of Gadd45b and brain-derived neurotrophic factor, a known target of Gadd45 mediated demethylation, in selectively-vulnerable hippocampal CA1 neurons. We further show that knockdown of Gadd45b increases abundance of a pro-apoptotic Bcl-2 family member Bax while decreasing the antiapoptotic protein Bcl-2, which together promote neuronal death. CONCLUSION: These findings document a protective role of Gadd45b against neuronal insults associated with global ischemia and identify Gadd45b as a potential therapeutic target for the amelioration of hippocampal neurodegeneration.

6.
Eur J Pharmacol ; 853: 33-40, 2019 Jun 15.
Article in English | MEDLINE | ID: mdl-30876977

ABSTRACT

Hydrogen sulfide (H2S) is a potential endothelium-derived hyperpolarizing factor (EDHF) and adventitium- or adipocyte-derived relaxing factor (ADRF) which vasorelaxant action is mediated by potassium channels. H2S could also play an important role in the pathophysiology of diabetic cardiovascular complications. The present study has investigated the influence of alloxan-induced diabetes on the role of potassium channels mediating the relaxant response of the rabbit carotid artery to NaHS, a donor of H2S. NaHS (10-8-3 × 10-5 M) relaxed phenylephrine-precontracted carotid arteries, with higher potency in diabetic than in control rabbits. The selective blockers of potassium channels charybdotoxin, 4-amynopiridine and glibenclamide significantly inhibited the relaxant action of NaHS in diabetic rabbits, but not in control rabbits. When compared to control rabbits, carotid arteries from diabetic rabbits showed significantly reduced expression of big conductance Ca+2-activated potassium channels (BKCa), significantly enhanced expression of intermediate conductance Ca+2-activated potassium channels (IKCa) and not significant different expression of voltage-sensitive potassium channels (KV) and ATP-sensitive potassium channels (KATP). These results suggest that an enhanced role of IKCa, KV and KATP potassium channels could be involved in the increased sensitivity of the rabbit carotid artery to H2S in diabetes.


Subject(s)
Carotid Arteries/drug effects , Carotid Arteries/physiopathology , Diabetes Mellitus, Experimental/metabolism , Hydrogen Sulfide/pharmacology , Potassium Channels/metabolism , Vasodilation/drug effects , Animals , Carotid Arteries/metabolism , Diabetes Mellitus, Experimental/physiopathology , Dose-Response Relationship, Drug , Male , Rabbits
7.
Neuroscience ; 388: 263-273, 2018 09 15.
Article in English | MEDLINE | ID: mdl-30077000

ABSTRACT

Preclinical and clinical studies support a promising, albeit not definitive, neuroprotective effect of emergent uric acid (UA) administration in ischemic stroke. We assessed the effects of UA in an ischemic stroke model relevant to the current treatment paradigm of mechanical thrombectomy within the STAIR/RIGOR recommendations. A cohort of male and female Wistar rats was subjected to ischemic stroke with mechanical recanalization under physiological monitoring. The effects of transient middle cerebral artery occlusion (tMCAO) with adjunctive UA (IV, 16 mg/kg) or vehicle treatment were assessed at 24 h and 7 days. Outcomes included neurofunctional impairment, brain infarct (TTC staining, MRI imaging and cresyl violet staining) and edema. At 24 h after tMCAO, neurofunctional scores and brain infarct were significantly reduced in rats subjected to UA treatment compared to vehicle, with a selective effect of UA on cortical infarct. No differential effect of UA between male and female rats was evidenced, as no significant interaction of sex with stroke outcomes was found. Rats achieving higher reperfusion levels after tMCAO showed superior reduction of neurofunctional impairment, cortical infarct and edema by UA. After a 7-day follow-up, male rats subjected to UA treatment still showed reductions in neurofunctional impairment and infarct size, compared to vehicle treatment. In conclusion, UA treatment immediately after transient ischemia results in a sex-independent, maintained reduction of brain damage and neurological impairment, better manifested in hyperperfusion conditions. This synergistic effect of UA with mechanical recanalization supports additional clinical testing of UA as an adjunctive treatment to mechanical thrombectomy.


Subject(s)
Brain Ischemia/therapy , Mechanical Thrombolysis , Neuroprotective Agents/pharmacology , Stroke/therapy , Uric Acid/pharmacology , Animals , Brain/drug effects , Brain/pathology , Brain Ischemia/pathology , Combined Modality Therapy , Disease Models, Animal , Female , Male , Random Allocation , Rats, Wistar , Recovery of Function , Stroke/pathology
8.
Mol Cell Endocrinol ; 472: 1-9, 2018 09 05.
Article in English | MEDLINE | ID: mdl-29842904

ABSTRACT

Along with its role in regulating blood pressure and fluid homeostasis, the natriuretic peptide system could be also part of an endogenous protective mechanism against brain damage. We aimed to assess the possibility that exogenous atrial natriuretic peptide (ANP) could protect against acute ischemic stroke, as well as the molecular mechanisms involved. Three groups of rats subjected to transient middle cerebral artery occlusion (tMCAO, intraluminal filament technique, 60 min) received intracerebroventricular vehicle, low-dose ANP (0.5 nmol) or high-dose ANP (2.5 nmol), at 30 min reperfusion. Neurofunctional condition, and brain infarct and edema volumes were measured at 24 h after tMCAO. Apoptotic cell death and expression of natriuretic peptide receptors (NPR-A and NPR-C), K+ channels (KATP, KV and BKCa), and PI3K/Akt and MAPK/ERK1/2 signaling pathways were analyzed. Significant improvement in neurofunctional status, associated to reduction in infarct and edema volumes, was shown in the high-dose ANP group. As to the molecular mechanisms analyzed, high-dose ANP: 1) reduced caspase-3-mediated apoptosis; 2) did not modify the expression of NPR-A and NPR-C, which had been downregulated by the ischemic insult; 3) induced a significant reversion of ischemia-downregulated KATP channel expression; and 4) induced a significant reversion of ischemia-upregulated pERK2/ERK2 expression ratio. In conclusion, ANP exerts a significant protective role in terms of both improvement of neurofunctional status and reduction in infarct volume. Modulation of ANP on some molecular mechanisms involved in ischemia-induced apoptotic cell death (KATP channels and MAPK/ERK1/2 signaling pathway) could account, at least in part, for its beneficial effect. Therefore, ANP should be considered as a potential adjunctive neuroprotective agent improving stroke outcome after successful reperfusion interventions.


Subject(s)
Atrial Natriuretic Factor/therapeutic use , Brain Ischemia/drug therapy , Neuroprotective Agents/therapeutic use , Stroke/drug therapy , Animals , Apoptosis/drug effects , Atrial Natriuretic Factor/pharmacology , Brain/drug effects , Brain/pathology , Brain Ischemia/complications , Caspase 3/metabolism , DNA Cleavage/drug effects , Down-Regulation , Infarction, Middle Cerebral Artery/complications , Infarction, Middle Cerebral Artery/pathology , Injections, Intraventricular , MAP Kinase Signaling System/drug effects , Male , Neuroprotective Agents/pharmacology , Phosphatidylinositol 3-Kinases/metabolism , Potassium Channels/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Rats, Wistar , Receptors, Atrial Natriuretic Factor/metabolism , Reperfusion Injury/complications , Reperfusion Injury/pathology , Stroke/complications
9.
Naunyn Schmiedebergs Arch Pharmacol ; 391(5): 501-511, 2018 05.
Article in English | MEDLINE | ID: mdl-29464270

ABSTRACT

Diabetic nephropathy is associated with increased risk of cardiovascular disease. B-type natriuretic peptide (BNP) plays an important role in cardiovascular pathophysiology and therapeutics. The aim of the present study was to investigate the influence of experimental diabetes on the mechanisms that regulate the relaxant response of the rabbit renal artery to BNP. Arterial relaxations to BNP were enhanced in diabetic rabbits. Indomethacin enhanced BNP-induced relaxation in control rabbits but showed no effect in diabetic rabbits. BNP-induced release of thromboxane A2 or prostacyclin was not different in both groups of animals. Iberiotoxin had no effect on relaxations to BNP in both groups of animals. Charybdotoxin displaced to the right the concentration-response curve to BNP in both group of animals, and inhibited BNP-induced relaxation only in diabetic rabbits. Glibenclamide did not modify the BNP-induced relaxations in control rabbits, but inhibited it in diabetic rabbits. These results suggest that diabetes induces hypereactivity of the rabbit renal artery to BNP by mechanisms that at least include (1) a reduced vasoconstrictor influence of arachidonic acid metabolites via cyclooxygenase 2, which is not related with changes in thromboxane A2 and prostacyclin release from the arterial wall and (2) a selectively increased modulatory activity of KATP and endothelial IKCa channels.


Subject(s)
Diabetes Mellitus, Experimental/physiopathology , Natriuretic Peptide, Brain/physiology , Potassium Channels/physiology , Prostaglandins/physiology , Renal Artery/physiology , Animals , Male , Rabbits , Vasodilation
10.
Sci Rep ; 8(1): 1610, 2018 01 25.
Article in English | MEDLINE | ID: mdl-29371613

ABSTRACT

Brain preconditioning (PC) refers to a state of transient tolerance against a lethal insult that can be evoked by a prior mild event. It is thought that PC may induce different pathways responsible for neuroprotection, which may involve the attenuation of cell damage pathways, including the apoptotic cell death. In this context, p53 is a stress sensor that accumulates during brain ischemia leading to neuronal death. The murine double minute 2 gene (MDM2), a p53-specific E3 ubiquitin ligase, is the main cellular antagonist of p53, mediating its degradation by the proteasome. Here, we study the role of MDM2-p53 pathway on PC-induced neuroprotection both in cultured neurons (in vitro) and rat brain (in vivo). Our results show that PC increased neuronal MDM2 protein levels, which prevented ischemia-induced p53 stabilization and neuronal death. Indeed, PC attenuated ischemia-induced activation of the p53/PUMA/caspase-3 signaling pathway. Pharmacological inhibition of MDM2-p53 interaction in neurons abrogated PC-induced neuroprotection against ischemia. Finally, the relevance of the MDM2-p53 pathway was confirmed in rat brain using a PC model in vivo. These findings demonstrate the key role of the MDM2-p53 pathway in PC-induced neuroprotection against a subsequent ischemic insult and poses MDM2 as an essential target in ischemic tolerance.


Subject(s)
Brain/pathology , Ischemia/pathology , Ischemic Preconditioning , Neurons/pathology , Proto-Oncogene Proteins c-mdm2/metabolism , Signal Transduction , Tumor Suppressor Protein p53/metabolism , Animals , Cell Survival , Cells, Cultured , Disease Models, Animal , Mice , Rats
11.
Eur J Pharmacol ; 815: 64-72, 2017 Nov 15.
Article in English | MEDLINE | ID: mdl-29024691

ABSTRACT

Atrial natriuretic peptide (ANP) is a vasodilator with significant regional differences and controversial effects in the cerebral circulation, a vascular bed particularly prone to diabetes-induced complications. The present study has investigated how alloxan-induced diabetes modifies the mechanisms involved in the response of the rabbit basilar artery to ANP. ANP (10-12-10-7M) relaxed precontracted basilar arteries, with higher potency in diabetic than in control rabbits. In arteries from both groups of animals, endothelium removal reduced ANP-induced relaxations. Inhibition of NO-synthesis attenuated ANP-induced relaxation but this attenuation was lower in diabetic than in control rabbits. In control rabbits, indomethacin displaced to the left the concentration-response curve to ANP, without significantly modifying the Emax value. In diabetic rabbits, indomethacin significantly enhanced arterial relaxations to ANP. In KCl-depolarised arteries, relaxation to ANP was almost abolished both in control and in diabetic rabbits. Iberiotoxin inhibited relaxations to ANP in both groups of rabbits. Glibenclamide and 4-aminopyridine inhibited the ANP-induced relaxations more in diabetic than in control rabbits. Basilar arteries from diabetic rabbits showed decreased natriuretic peptide receptor C expression and no changes in natriuretic peptide receptor A, large conductance calcium-activated K+ channels (BKCa), ATP-sensitive K+ channels (KATP) and voltage-sensitive K+ channels (KV) expression. These results suggest that diabetes enhances the sensitivity of the rabbit basilar artery to ANP by mechanisms that at least include reduced expression of natriuretic peptide receptor C, and enhanced activity of KATP and KV channels. Furthermore, diabetes reduces endothelial NO and prostacyclin which mediate arterial relaxation to ANP.


Subject(s)
Atrial Natriuretic Factor/pharmacology , Basilar Artery/drug effects , Basilar Artery/metabolism , Diabetes Mellitus, Experimental/metabolism , Animals , Dose-Response Relationship, Drug , Male , Nitric Oxide/metabolism , Prostaglandins/metabolism , Rabbits , Receptors, Atrial Natriuretic Factor/metabolism
12.
J Steroid Biochem Mol Biol ; 171: 296-304, 2017 07.
Article in English | MEDLINE | ID: mdl-28479229

ABSTRACT

As the knowledge on the estrogenic system in the brain grows, the possibilities to modulate it in order to afford further neuroprotection in brain damaging disorders so do it. We have previously demonstrated the ability of the selective estrogen receptor modulator, bazedoxifene (BZA), to reduce experimental ischemic brain damage. The present study has been designed to gain insight into the molecular mechanisms involved in such a neuroprotective action by investigating: 1) stroke-induced apoptotic cell death; 2) expression of estrogen receptors (ER) ERα, ERß and the G-protein coupled estrogen receptor (GPER); and 3) modulation of MAPK/ERK1/2 and PI3K/Akt signaling pathways. For comparison, a parallel study was done with 17ß-estradiol (E2)-treated animals. Male Wistar rats subject to transient right middle cerebral artery occlusion (tMCAO, intraluminal thread technique, 60min), were distributed in vehicle-, BZA- (20.7±2.1ng/mL in plasma) and E2- (45.6±7.8pg/mL in plasma) treated groups. At 24h from the onset of tMCAO, RT-PCR, Western blot and histochemical analysis were performed on brain tissue samples. Ischemia-reperfusion per se increased apoptosis as assessed by both caspase-3 activity and TUNEL-positive cell counts, which were reversed by both BZA and E2. ERα and ERß expression, but not that of GPER, was reduced by the ischemic insult. BZA and E2 had different effects: while BZA increased both ERα and ERß expression, E2 increased ERα expression but did not change that of ERß. Both MAPK/ERK1/2 and PI3K/Akt pathways were stimulated under ischemic conditions. While BZA strongly reduced the increased p-ERK1/2 levels, E2 did not. Neither BZA nor E2 modified ischemia-induced increase in p-Akt levels. These results show that modulation of ERα and ERß expression, as well as of the ERK1/2 signaling pathway accounts, at least in part, for the inhibitory effect of BZA on the stroke-induced apoptotic cell death. This lends mechanistic support to the consideration of BZA as a potential neuroprotective drug in acute ischemic stroke treatment.


Subject(s)
Brain Ischemia/drug therapy , Indoles/therapeutic use , Neurons/drug effects , Neuroprotective Agents/therapeutic use , Reperfusion Injury/prevention & control , Selective Estrogen Receptor Modulators/therapeutic use , Stroke/drug therapy , Animals , Apoptosis/drug effects , Brain Ischemia/metabolism , Brain Ischemia/pathology , Cerebral Cortex/drug effects , Cerebral Cortex/metabolism , Cerebral Cortex/pathology , Estradiol/therapeutic use , Estrogen Receptor alpha/agonists , Estrogen Receptor alpha/genetics , Estrogen Receptor alpha/metabolism , Estrogen Receptor beta/agonists , Estrogen Receptor beta/genetics , Estrogen Receptor beta/metabolism , Estrogens/therapeutic use , MAP Kinase Signaling System/drug effects , Male , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Neurons/metabolism , Neurons/pathology , Phosphatidylinositol Phosphates/agonists , Phosphatidylinositol Phosphates/metabolism , Rats, Wistar , Receptors, G-Protein-Coupled/genetics , Receptors, G-Protein-Coupled/metabolism , Second Messenger Systems/drug effects , Stroke/metabolism , Stroke/pathology
13.
J Cardiovasc Pharmacol ; 68(4): 313-321, 2016 Oct.
Article in English | MEDLINE | ID: mdl-27389095

ABSTRACT

We have previously shown that the selective estrogen receptor modulator, bazedoxifene, improves the consequences of ischemic stroke. Now we aimed to characterize the effects and mechanisms of action of bazedoxifene in cerebral arteries. Male rabbit isolated basilar arteries were used for isometric tension recording and quantitative polymerase chain reaction. Bazedoxifene relaxed cerebral arteries, as 17-ß-estradiol, 4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol [estrogen receptor (ER) α agonist], and G1 [G protein-coupled ER (GPER) agonist] did it (4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol > bazedoxifene = G1 > 17-ß-estradiol). 2,3-Bis(4-hydroxyphenyl)-propionitrile (ERß agonist) had no effect. Expression profile of genes encoding for ERα (ESR1), ERß (ESR2), and GPER was GPER > ESR1 > ESR2. As to the endothelial mechanisms, endothelium removal, N-nitro-L-arginine methyl ester, and indomethacin, did not modify the relaxant responses to bazedoxifene. As to the K channels, both a high-K medium and the Kv blocker, 4-aminopyridine, inhibited the bazedoxifene-induced relaxations, whereas tetraethylammonium (nonselective K channel blocker), glibenclamide (selective KATP blocker) or iberiotoxin (selective KCa blocker) were without effect. Bazedoxifene also inhibited both Ca- and Bay K8644-elicited contractions. Therefore, bazedoxifene induces endothelium-independent relaxations of cerebral arteries through (1) activation of GPER and ERα receptors; (2) increase of K conductance through Kv channels; and (3) inhibition of Ca entry through L-type Ca channels. Such a profile is compatible with the beneficial effects of estrogenic compounds (eg, SERMs) on vascular function and, specifically, that concerning the brain. Therefore, bazedoxifene could be useful in the treatment of cerebral disorders in which the cerebrovascular function is compromised (eg, stroke).


Subject(s)
Basilar Artery/drug effects , Estrogens/pharmacology , Indoles/pharmacology , Selective Estrogen Receptor Modulators/pharmacology , Vasodilation/drug effects , Animals , Basilar Artery/physiology , Dose-Response Relationship, Drug , Male , Organ Culture Techniques , Rabbits , Vasodilation/physiology
14.
Eur J Pharmacol ; 701(1-3): 159-67, 2013 Feb 15.
Article in English | MEDLINE | ID: mdl-23340222

ABSTRACT

The relation between diabetes and stroke is bidirectional: diabetes is an important risk factor for ischemic stroke, and acute stroke frequently induces hyperglycemia. On the other hand, plasma B-type natriuretic peptide (BNP) levels are raised in diabetes and stroke. The purpose was to study how alloxan-induced diabetes might modify the effects of BNP in rabbit carotid arteries and the mechanisms involved in such actions. To do this, isometric tension in isolated rabbit carotid artery was recorded and prostanoids release and plasma NT-proBNP were measured by enzyme immunoassay. BNP induced a relaxation of phenylephrine-precontracted carotid arteries, and this relaxation was lower in diabetic than in control rabbits. Endothelium removal did not modify the relaxation to BNP in control rabbits but increased this relaxation in diabetic rabbits. In control rabbits, indomethacin inhibited the BNP-induced relaxation in the presence and in the absence of endothelium. In diabetic rabbits, indomethacin did not modify the BNP-induced relaxation in arteries with endothelium and inhibited it in arteries without endothelium. In the presence of BNP the carotid artery released thromboxane A2 and prostacyclin, and the release of endothelial prostacyclin was inhibited in diabetic rabbits. Glibenclamide and 4-aminopyridine inhibited the relaxation to BNP, and these inhibitions were lower in diabetic than in control rabbits. In conclusion, our results provide a new understanding concerning the mechanisms of the diabetes-induced hyporeactivity of the carotid artery to BNP, that at least include the loss of endothelial prostacyclin and a reduced participation of ATP-sensitive K(+) channels (KATP) and voltage-sensitive K(+) channels (KV).


Subject(s)
Carotid Arteries/drug effects , Diabetes Mellitus/metabolism , Epoprostenol/metabolism , Natriuretic Peptide, Brain/pharmacology , Potassium Channels/metabolism , Animals , Blood Glucose/metabolism , Body Weight/drug effects , Carotid Arteries/metabolism , Diabetes Mellitus/blood , Diabetes Mellitus/physiopathology , Dose-Response Relationship, Drug , Male , Natriuretic Peptide, Brain/blood , Nitric Oxide/metabolism , Peptide Fragments/blood , Potassium/metabolism , Rabbits , Receptors, Atrial Natriuretic Factor/metabolism
15.
Pharmacol Res ; 66(5): 392-400, 2012 Nov.
Article in English | MEDLINE | ID: mdl-22898325

ABSTRACT

Diabetes is associated with increased prevalence of hypertension, cardiovascular and renal disease. Atrial natriuretic peptide (ANP) plays an important role in cardiovascular pathophysiology and is claimed to have cardioprotective and renoprotective effect in diabetic patients. The working hypothesis was that alloxan-induced diabetes might modify the vascular effects of ANP in isolated rabbit renal arteries and the mechanisms involved in such actions. Plasma ANP levels were higher in diabetic rabbits than in control rabbits. ANP (10(-12)-10(-7)M) induced a relaxation of precontracted renal arteries, which was lower in diabetic than in control rabbits. In arteries from both groups of animals, endothelium removal decreased the ANP-induced relaxation but inhibition of NO-synthesis did not modify ANP-induced relaxations. In KCl-depolarised arteries, relaxation to ANP was almost abolished both in control and diabetic rabbits. Tetraethylammonium (TEA) partly inhibited the relaxation to ANP in control rabbits but did not modify it in diabetic rabbits. Glibenclamide and 4-aminopyridine inhibited the relaxation to ANP, and these inhibitions were lower in diabetic than in control rabbits. Indomethacin potentiated the relaxation to ANP, more in control than in diabetic rabbits. In the presence of ANP the renal artery released thromboxane A(2) and prostacyclin, and the release of prostacyclin resulted decreased in diabetic rabbits. The present results suggest that diabetes produces hyporeactivity of the rabbit renal artery to ANP by mechanisms that at least include the reduced modulation by prostacyclin and a lower participation of ATP-sensitive K(+) channel (K(ATP)), voltage-sensitive K(+) channels (K(V)) and TEA-sensitive K(+) channels (K(Ca)).


Subject(s)
Atrial Natriuretic Factor/pharmacology , Diabetes Mellitus, Experimental/physiopathology , Epoprostenol/physiology , Renal Artery/drug effects , Vasodilation/drug effects , Animals , Atrial Natriuretic Factor/blood , Endothelium, Vascular/physiology , In Vitro Techniques , Male , Potassium Channels/physiology , Rabbits , Renal Artery/physiology , Tetraethylammonium/pharmacology , Thromboxane A2/physiology , Vasodilation/physiology
16.
Front Neuroendocrinol ; 32(3): 336-52, 2011 Aug.
Article in English | MEDLINE | ID: mdl-21163293

ABSTRACT

This review highlights our investigations into the neuroprotective efficacy of estradiol and other estrogenic agents in a clinically relevant animal model of transient global ischemia, which causes selective, delayed death of hippocampal CA1 neurons and associated cognitive deficits. We find that estradiol rescues a significant number of CA1 pyramidal neurons that would otherwise die in response to global ischemia, and this is true when hormone is provided as a long-term pretreatment at physiological doses or as an acute treatment at the time of reperfusion. In addition to enhancing neuronal survival, both forms of estradiol treatment induce measurable cognitive benefit in young animals. Moreover, estradiol and estrogen analogs that do not bind classical nuclear estrogen receptors retain their neuroprotective efficacy in middle-aged females deprived of ovarian hormones for a prolonged duration (8weeks). Thus, non-feminizing estrogens may represent a new therapeutic approach for treating the neuronal damage associated with global ischemia.


Subject(s)
Estradiol Congeners/therapeutic use , Estradiol/therapeutic use , Ischemia/drug therapy , Neuroprotective Agents/therapeutic use , Animals , Humans , Ischemia/pathology , Ischemia/physiopathology
17.
Pharmacol Res ; 63(3): 190-8, 2011 Mar.
Article in English | MEDLINE | ID: mdl-21073954

ABSTRACT

Atrial natriuretic peptide (ANP) plays an important role in the pathophysiology of the vascular complications in diabetes. The working hypothesis was that diabetes might modify the vascular actions of ANP in isolated rabbit carotid arteries and the mechanisms involved in these actions. ANP (10(-12)-10(-7)M) induced a relaxation of precontracted carotid arteries, which was lower in diabetic than in control rabbits. In arteries from both groups of animals, endothelium removal increased the ANP-induced relaxation. Isatin inhibited the relaxation to ANP both in arteries with and without endothelium. Carotid arteries from diabetic rabbits showed a decreased natriuretic peptide receptor (NPR)-A expression and an enhanced NPR-C expression. Inhibition of NO-synthesis did not modify ANP-induced relaxation in control rabbits but inhibited it in diabetic rabbits. In arteries with endothelium indomethacin enhanced the relaxation to ANP in control rabbits but did not modify it in diabetic rabbits. In endothelium-denuded arteries indomethacin inhibited the relaxation to ANP in both groups of animals. In KCl-depolarised arteries, relaxation to ANP was almost abolished both in control and diabetic rabbits. Tetraethylammonium inhibited the relaxation to ANP, and this inhibition was higher in diabetic than in control rabbits. These results suggest that diabetes produces hyporeactivity of the rabbit carotid artery to ANP by a mechanism that at least includes a reduced expression of NPR-A, an enhanced expression of NPR-C and a reduced participation of K(+)-channels. Furthermore, diabetes enhances endothelial NO release and diminishes the ratio thromboxane A(2)/prostacyclin. This increase of vasodilators could result from compensatory mechanisms counteracting the arterial hyporeactivity to ANP.


Subject(s)
Atrial Natriuretic Factor/physiology , Carotid Arteries/metabolism , Diabetes Mellitus, Experimental/metabolism , Down-Regulation/physiology , Vasodilation/physiology , Animals , Diabetes Mellitus, Experimental/physiopathology , Male , Rabbits , Random Allocation
18.
Brain Res ; 1321: 1-12, 2010 Mar 19.
Article in English | MEDLINE | ID: mdl-20114038

ABSTRACT

Global ischemia arising during cardiac arrest or cardiac surgery causes highly selective, delayed death of hippocampal CA1 neurons. Exogenous estradiol ameliorates global ischemia-induced neuronal death and cognitive impairment in male and female rodents. However, the molecular mechanisms by which a single acute injection of estradiol administered after the ischemic event intervenes in global ischemia-induced apoptotic cell death are unclear. Here we show that acute estradiol acts via the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling cascade to protect CA1 neurons in ovariectomized female rats. We demonstrate that global ischemia promotes early activation of glycogen synthase kinase-3beta (GSK3beta) and forkhead transcription factor of the O class (FOXO)3A, known Akt targets that are related to cell survival, and activation of caspase-3. Estradiol prevents ischemia-induced dephosphorylation and activation of GSK3beta and FOXO3A, and the caspase death cascade. These findings support a model whereby estradiol acts by activation of PI3K/Akt signaling to promote neuronal survival in the face of global ischemia.


Subject(s)
Apoptosis/drug effects , Brain Ischemia/drug therapy , Estradiol/pharmacology , Estrogens/pharmacology , Neurons/drug effects , Signal Transduction/drug effects , Animals , Blotting, Western , Brain Ischemia/pathology , Female , Hippocampus/drug effects , Hippocampus/metabolism , Hippocampus/pathology , Nerve Degeneration/prevention & control , Neurons/metabolism , Neurons/pathology , Ovariectomy , Phosphatidylinositol 3-Kinases/drug effects , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/drug effects , Proto-Oncogene Proteins c-akt/metabolism , Rats
19.
Pharmacol Res ; 61(2): 149-56, 2010 Feb.
Article in English | MEDLINE | ID: mdl-19755160

ABSTRACT

Kidney disease is a frequent complication in diabetes, and significant differences have been reported between male and female patients. Our working hypothesis was that diabetes might modify the vascular actions of testosterone in isolated rabbit renal arteries and the mechanisms involved in these actions. Testosterone (10(-8) to 10(-4)M) induced relaxation of precontracted arteries, without significant differences between control and diabetic rabbits. Both in control and diabetic rabbits endothelium removal inhibited testosterone relaxant action. In arteries with endothelium, incubation with indomethacin (10(-5)M), N(G)-nitro-l-arginine (10(-5)M) or tetraethylammonium (10(-5)M) did not modify relaxations to testosterone neither in control nor in diabetic rabbits. In endothelium-denuded arteries indomethacin enhanced the relaxant action of testosterone, both in control and diabetic rabbits. In arteries from diabetic rabbits, eNOS, iNOS and COX-1 expression and testosterone-induced release of thromboxane A(2) and prostacyclin were not significantly different from those observed in control rabbits. However, COX-2 expression was significantly lower in diabetic rabbits that in control rabbits. In nominally Ca(2+)-free medium, cumulative addition of CaCl2 (10(-5) to 3x10(-2)M) contracted previously depolarized arteries. Testosterone (10(-4)M) inhibited CaCl2 contractions of the renal artery both in control and diabetic rabbits. These results show that testosterone relaxes the renal artery both in control and diabetic rabbits. This relaxation is modulated by muscular thromboxane A(2), it is partially mediated by endothelial prostacyclin, and it involves the blocking of extracellular Ca2+ entry. Diabetes does not modify the mechanisms involved in the relaxant action of testosterone in the rabbit renal artery.


Subject(s)
Diabetes Mellitus, Experimental/metabolism , Renal Artery/metabolism , Testosterone/metabolism , Vasodilation , Animals , Blood Glucose/metabolism , Blotting, Western , Calcium/metabolism , Cyclooxygenase 1/metabolism , Cyclooxygenase 2/metabolism , Cyclooxygenase Inhibitors/pharmacology , Diabetes Mellitus, Experimental/physiopathology , Dose-Response Relationship, Drug , Endothelium, Vascular/metabolism , Endothelium, Vascular/physiopathology , Immunoenzyme Techniques , Indomethacin/pharmacology , Male , Muscle, Smooth, Vascular/metabolism , Muscle, Smooth, Vascular/physiopathology , Nitric Oxide Synthase Type II/antagonists & inhibitors , Nitric Oxide Synthase Type II/metabolism , Nitric Oxide Synthase Type III/antagonists & inhibitors , Nitric Oxide Synthase Type III/metabolism , Nitroarginine/pharmacology , Potassium/metabolism , Potassium Channel Blockers/pharmacology , Prostaglandins I/metabolism , Rabbits , Renal Artery/physiopathology , Signal Transduction , Testosterone/blood , Tetraethylammonium/pharmacology , Thromboxane A2/metabolism
20.
Pharmacol Res ; 61(1): 62-70, 2010 Jan.
Article in English | MEDLINE | ID: mdl-19573602

ABSTRACT

Cardiovascular disease is the major cause of morbidity and mortality in diabetic patients, which in turn is also associated with low levels of serum testosterone. The working hypothesis was that diabetes might modify the mechanisms involved in the vascular actions of testosterone in isolated rabbit carotid arteries. Testosterone (10(-8)-3x10(-4)M) induced a concentration-dependent relaxation of precontracted carotid arteries, which was higher in diabetic than in control rabbits. In control rabbits neither endothelium removal nor the nitric oxide synthase (NOS) inhibitor N(G)-nitro-l-arginine (l-NOArg, 10(-5)M) modified the relaxant action of testosterone, and the cyclooxygenase (COX) inhibitor indomethacin (10(-5)M) enhanced this relaxation. In contrast, in diabetic rabbits endothelium removal, l-NOArg (10(-5)M) or indomethacin (10(-5)M) inhibited the testosterone induced relaxation. In arteries from diabetic rabbits, eNOS, iNOS and COX-2 expression and testosterone induced release of prostacyclin resulted enhanced in comparison with arteries from control rabbits. Testosterone (10(-4)M) strongly inhibited CaCl(2) (10(-5)-3x10(-2)M) concentration-related contractions of the carotid artery both in control and diabetic rabbits. These results suggest that testosterone relaxes the rabbit carotid artery by blocking the extracellular calcium entry. Diabetes enhances the vasodilator response of the rabbit carotid artery to testosterone by a mechanism that at least includes an increased modulatory activity of the endothelial nitric oxide and an augmented release of COX-2 vasodilator, prostacyclin rather than the absence of COX-1 vasoconstrictor, thromboxane A(2). The hypotestosteronemia observed in diabetic rabbits could be a consequence of the increased expression of iNOS and could contribute to the hyperreactivity of the rabbit carotid artery to testosterone.


Subject(s)
Carotid Arteries/enzymology , Carotid Artery Diseases/etiology , Cyclooxygenase 2/metabolism , Diabetes Mellitus, Experimental/enzymology , Diabetic Angiopathies/etiology , Nitric Oxide Synthase Type III/metabolism , Nitric Oxide Synthase Type II/metabolism , Testosterone/metabolism , Vasodilation , Animals , Apamin/pharmacology , Blood Glucose/metabolism , Blotting, Western , Calcium/metabolism , Carotid Arteries/drug effects , Carotid Arteries/physiopathology , Carotid Artery Diseases/enzymology , Carotid Artery Diseases/physiopathology , Charybdotoxin/pharmacology , Cyclooxygenase Inhibitors/pharmacology , Diabetes Mellitus, Experimental/complications , Diabetes Mellitus, Experimental/physiopathology , Diabetic Angiopathies/enzymology , Diabetic Angiopathies/physiopathology , Dose-Response Relationship, Drug , Endothelium, Vascular/enzymology , Endothelium, Vascular/physiopathology , Epoprostenol/metabolism , Immunoenzyme Techniques , Indomethacin/pharmacology , Male , Nitric Oxide/metabolism , Nitric Oxide Synthase Type II/antagonists & inhibitors , Nitric Oxide Synthase Type III/antagonists & inhibitors , Nitroarginine/pharmacology , Potassium/metabolism , Potassium Channel Blockers/pharmacology , Rabbits , Thromboxane A2/metabolism , Vasodilation/drug effects
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