Objective— Earlier in vitro studies suggested a putative role for the plasma phospholipid transfer protein (PLTP) in the modulation of blood coagulation. The effect of PLTP expression on blood coagulation under both basal and oxidative stress conditions was compared here in wild-type and PLTP-deficient (PLTP−/−) mice. Methods and Results— Under basal conditions, PLTP deficiency was associated with an extended tail bleeding time despite a significant depletion of vascular α-tocopherol content and an impairment of endothelial function. When acute oxidative stress was generated in vivo in the brain vasculature, the steady state levels of oxidized lipid derivatives, the extent of blood vessel occlusion, and the volume of ischemic lesions were more severe in wild-type than in PLTP−/− mice. Conclusion— In addition to its recognized hyperlipidemic, proinflammatory, and proatherogenic properties, PLTP increases blood coagulation and worsens the extent of ischemic lesions in response to acute oxidative stress. Thus, PLTP arises here as a cardiovascular risk factor for the late thrombotic events occurring in the acute phase of atherosclerosis.

The pathogenesis of ischemic stroke is a complex sequence of events including inflammatory reaction, for which the microglia appears to be a major cellular contributor. However, whether post-ischemic activation of microglial cells has beneficial or detrimental effects remains to be elucidated, in particular on long term brain plasticity events. The objective of our study was to determine, through modulation of post-stroke inflammatory response, to what extent microglial cells are involved in some specific events of neuronal plasticity, neurite outgrowth and synaptogenesis. Since microglia is a source of neurotrophic factors, the identification of the brain-derived neurophic factor (BDNF) as possible molecular actor involved in these events was also attempted. As a means of down-regulating the microglial response induced by ischemia, 3-aminobenzamide (3-AB, 90 mg/kg, i.p.) was used to inhibit the poly(ADP-ribose) polymerase-1 (PARP-1). Indeed, PARP-1 contributes to the activation of the transcription factor NF-kB, which is essential to the upregulation of proinflammatory genes, in particular responsible for microglial activation/proliferation. Experiments were conducted in rats subjected to photothrombotic ischemia which leads to a strong and early microglial cells activation/proliferation followed by an infiltration of macrophages within the cortical lesion, events evaluated at serial time points up to 1 month post-ictus by immunostaining for OX-42 and ED-1. Our most striking finding was that the decrease in acute microglial activation induced by 3-AB was associated with a long term down-regulation of two neuronal plasticity proteins expression, synaptophysin (marker of synaptogenesis) and GAP-43 (marker of neuritogenesis) as well as to a significant decrease in tissue BDNF production. Thus, our data argue in favour of a supportive role for microglia in brain neuroplasticity stimulation possibly through BDNF production, suggesting that a targeted protection of microglial cells could represent an innovative approach to potentiate post-stroke neuroregeneration.

Whereas iron chelators were shown to induce neuroprotection against brain injury, the effect of iron chelators on ischemia-induced damage of cerebral endothelium is largely unknown. Our objective was to explore the endothelioprotective effect of the lipophilic iron chelator dipyridyl (DP) (i) in vitro on the death of cerebral endothelial cells (CECs) subjected to intracellular iron loading and (ii) in vivo on the ischemia-induced blood–brain barrier (BBB) disruption. When given shortly after iron exposure or brain ischemia, DP prevented the death of CECs and diminished BBB disruption, respectively, whereas a delayed administration of DP was associated with a lower CECs protection. Interestingly, when given preventively, DP also abrogated the death of CECs and reduced BBB disruption. However, a long delay between DP treatment and iron exposure led to a higher protection, suggesting a preconditioning effect of DP. Accordingly, prevention of hydroxyl radical formation through iron chelation cannot explain alone the beneficial effect of preventive DP treatment. Our findings showing that DP failed to induce the potentially cytoprotective proteins, heme oxygenase-1 and manganese superoxide dismutase, suggest that other proteins participate to the preconditioning effect of DP. To conclude, the curative and preventive effects of DP evidenced in this study suggest that iron chelation therapy represents a favorable and effective approach to increase BBB resistance towards ischemic injury.

Hypoxia-inducible factor 1 α (HIF-1α) is a transcription factor that was suggested in vitro to promote cell death by modulation of proapoptotic genes. In this report, we tested the hypothesis of an in vivo proapoptotic role of HIF-1α after an ischemic insult. For this purpose, HIF-1α and procaspase-3 mRNA and protein expressions were examined in rat brain subjected to 12- and 24-h permanent focal ischemia and the presence of an HIF-1 binding activity to the caspase-3 gene promoter was explored. The results showed that HIF-1α and procaspase-3 expressions increased with a similar pattern in response to ischemia. In addition, caspase-3 activation was observed in cells that express HIF-1α. Moreover, electrophoretic mobility assay revealed a specific HIF-1 binding activity to the caspase-3 gene promoter. Altogether the present data provide strong arguments for a causative relationship between HIF-1α and caspase-3 inductions through a functional binding activity to the caspase-3 gene promoter.

To evaluate the effect of body iron stores on the vulnerability of the brain to ischemia, a focal permanent brain ischemia was induced by photothrombotic occlusion of cortical vessels in rats with or without chronic treatment with iron dextran (25 mg iron/kg, every other day for 20 days, intraperitoneally). Iron dextran induced systemic iron overload as evidenced by high ferritin (Ft) (x 5) and total iron levels (x 3) in serum as well as increased Ft expression in the liver and heart. Conversely, neither serum free iron levels nor Ft expression in the brain were changed by iron dextran. Finally, infarct volume was not modified by iron dextran. In addition, induction of ischemia in rats treated with FeCl3 (560 μg iron/kg, intravenously) as a means of increasing serum free iron levels during the ischemic period did not enlarge infarct volume. We then explored the effect of brain ischemia itself on serum Ft by measuring serum Ft before and after induction of brain ischemic insults with different neurologic outcomes in rats (brain embolization with microspheres, photothrombotic occlusion of cortical vessels, four-vessel occlusion). Serum Ft levels were found higher at day 1 after ischemia than before ischemia only in rats subjected to the most severe insult (brain embolization). In conclusion, our study showed that increased body iron stores do not increase the vulnerability of the brain to ischemia and that brain ischemia, if severe, results in the elevation of serum Ft levels.

X-linked adrenoleukodystrophy (X-ALD) is a demyelinating disease due to mutations in the ABCD1 (ALD) gene, encoding a peroxisomal ATP-binding cassette transporter (ALDP). Overexpression of adrenoleukodystrophy-related protein, an ALDP homologue encoded by the AB CD2 (adrenoleukodystrophy-related) gene, can compensate for ALDP deficiency. 4-Phenyl-butyrate (PBA) has been shown to induce both ABCD2 expression and peroxisome proliferation in human fibroblasts. We show that peroxisome proliferation with unusual shapes and clusters occurred in liver of PBA-treated rodents in a PPAR alpha-independent way. PBA activated Abcd2 in cultured glial cells, making PBA a candidate drug for therapy of X-ALD. The Abcd2 induction observed was partially PPAR alpha independent in hepatocytes and totally independent in fibroblasts. We demonstrate that a GC box and a CCAAT box of the Abcd2 promoter are the key elements of the PBA-dependent Abcd2 induction, histone deacetylase (HDAC) 1 being recruited by the GC box. Thus, PBA is a nonclassical peroxisome proliferator inducing pleiotropic effects, including effects at the peroxisomal level mainly through HDAC inhibition.

The present work aimed 1) to evaluate whether an increase in galanin or galanin receptors could be induced in the nucleus basalis magnocellularis (nbm) by degeneration of the basalocortical neurons from the cortex and 2) to analyze the consequences of such an increase on cortical activity. First, a mild ischemic insult to the frontoparietal cortex was performed to induce the degeneration of the basalocortical system; galanin immunoreactivity, galanin binding sites, and cholinergic muscarinic receptors were quantified through immunocytochemistry and autoradiography. Second, galanin infusions in the nbm were undertaken to mimic a local increase of the galaninergic innervation; cortical acetylcholine release, cerebral glucose use, and cerebral blood flow were then measured as indices of cortical activity. As a result of the cortical ischemic lesion, the postsynaptic M1 and presynaptic M2 muscarinic receptors were found to be reduced in the altered cortex. In contrast, galaninergic binding capacity and fiber density were found to be increased in the ipsilateral nbm in parallel with a local decrease in the cholinergic markers such as the muscarinic M1 receptor density. Galanin infusion into the nbm inhibited the cortical acetylcholine release and cerebral blood flow increases elicited by the activation of the cholinergic basalocortical system but failed to affect acetylcholine release, cerebral blood flow, and cerebral glucose use when injected alone in the nbm. These results demonstrate that degeneration of the basalocortical system from the cortex induces an increase in galaninergic markers in the nbm, a result that might suggest that the galaninergic overexpression described in the basal forebrain of patients with Alzheimer's disease can result from a degeneration of the cholinergic basalocortical system from the cortex. Because galanin was found to reduce the activity of the basalocortical cholinergic system only when this one is activated, galanin might exert its role rather during activation deficits than under resting conditions such as the resting cortical hypometabolism, which is characteristic of Alzheimer's disease.

Cerebral tissue with T2 magnetic resonance imaging (MRI) abnormalities following stroke is generally considered infarcted, while surrounding regions with normal MRI appearance are believed to be healthy. To assess whether these surrounding regions consist of normal tissue, we explored the distribution of N-acetylaspartate (NAA) and lactate within and around the hyperintense area on T2-weighted MRI using proton MR spectroscopy. The study was carried out in 25 patients with middle cerebral artery occlusion imaged between 1 and 42 days after stroke onset. NAA/choline (Cho) ratios were significantly reduced in both areas of T2 hyperintensity and in surrounding tissue. The reduction was greater in the region of T2 hyperintensity than in the surrounding region (-50% vs. -28%, respectively) and was unrelated to the delay after the ictus. Lactate/Cho ratios increased massively within the abnormal T2 area, but did not differ from control values beyond the margin of hyperintensity. Overall data indicate that T2 visible lesions on MRI do not infer the entire injured tissue.

Neurochemical activation of the substantia innominata (SI) in the rat, through the direct injection of the cholinergic agonist carbachol, has been reported to induce large increases in cerebral blood flow (CBF) throughout cortical and subcortical projection regions. The present study aimed to determine whether the vasomotor responses to cholinergic stimulation of the SI were, or were not, the consequence of an increase in metabolic activity. To this end, coupled measurements of CBF and cerebral glucose use (CGU) were undertaken during carbachol-elicited stimulation of the SI. Infusion of carbachol into the basal forebrain induced significant CBF increases in several ipsilateral cortical and subcortical areas including the amygdala. In contrast, CGU increased only in the ipsilateral amygdala and SI. Thus, we tested the hypothesis of a direct neurogenic, rather than metabolic, contribution of the basalocortical system. In this respect, carbachol-elicited stimulation resulted in significant increases in extracellular acetylcholine concentrations in the ipsilateral parietal cortex; systemic pretreatment with the muscarinic receptor antagonist scopolamine completely abolished the increase in cortical CBF elicited by cholinergic stimulation of the SI in the ipsilateral frontoparietal motor cortex while it failed to affect the increase observed in the ipsilateral temporal cortex. Several conclusions can be drawn from the present study. The stimulation of the SI by carbachol induces an increase in CBF that can be dissociated from changes in underlying glucose metabolism. Secondly, these induced changes in cortical CBF are paralleled by an increase in acetylcholine release. Lastly, the failure of scopolamine to block the flow response in all cortical regions would suggest that SI stimulation will evoke the release of vasodilatatory neurotransmitter(s) as well as acetylcholine itself.

Unilateral cerebral microembolism was performed in the rat by injecting calibrated, 50 μm in diameter, carbonized microspheres into the internal carotid artery. The events that follow brain ischemia due to cerebral embolization were studied by the analysis of the blood-brain barrier (BBB) function, the degree of regional cerebral blood flow (CBF) and the development of brain edema. Two hours after embolization there was no change in the brain water content. The local CBF (14C-ethanol technique) was only reduced in the ipsilateral hemisphere. Twenty-four hours after embolization the brain water content was increased significantly in the ipsilateral, but not in the contralateral hemisphere. Local CBF further decreased in the ipsilateral hemisphere and a reduction in flow was also observed in the contralateral hemisphere. Embolization led to an increase in the BBB permeability, analysed as regional penetrability of 3H-dextran and of Evans blue-albumin complexes, which was restricted to the side of the injection of the microspheres.

The synthesis of noradrenaline (NA) was studied in vivo in the hypothalamus, the brain stem and the “rest of brain” of rats which were exposed to 4° C for 0.5, 2.5 or 24 h. The rate of NA synthesis was estimated 30 min after an i.v. injection of3H tyrosine (TY) by evaluation of the ratio:3H-NA specific activity/3H-TY specific activity.

The synthesis of catecholamines (CA) has been studied in the heart, spleen, submaxillary glands and adrenals of rats exposed to 4° C for 2.5, 24 or 48 h. The synthesis rate has been estimated 30 min after an i.v. injection of3H tyrosine (TY) by the evaluation of the ratio:3H-CA specific activity/3H-TY specific activity.

Le taux de renouvellement de la noradrénaline qui reflète l'intensité du tonus sympathique a été evalue à partir de la vitesse de disparition de la NA3H dans huit organes périphériques chez des rats témoins et traités de façon chronique par la thyroxine ou le propylthiouracile. Dans le cceur, la rate et le muscle fémoral, on observe une relation inverse entre le degré d'activité thyroïdienne et l'intensité du tonus sympathique, les taux de renouvellement de la NA étant réduits après surcharge en thyroxine et accrus après administration de PTU: l'effet le plus important est observé au niveau du cœur, le taux de renouvellement du médiateur sympathique étant doublé chez les rats traités par le PTU comparativement aux animaux surchargés en thyroxine. Dans les autres organes étudiés (rein, poumon, intestin, estomac, glande sous-maxillaire) on ne retrouve pas cette relation inverse entre l'activité thyroïdienne et celle du système sympathique et les modifications de taux de renouvellement consécutives à l'administration de thyroxine et de PTU ne sont pas statistiquement significatives. L'ensemble des résultats obtenus montre que le contrôle du tonus sympathique par la thyroïde est limité à certains territoires périphériques.Les mécanismes par lesquels les hormones thyroïdiennes peuvent influencer l'activité du système sympathique sont discutés. Il est suggéré que l'action de ces hormones s'exerce d'une façon indirecte par l'intermédiaire de leurs effets sympathomimétiques cardiovasculaires.

Summary The turnover times and the synthesis rates of tissue norepinephrine were determined in the vas deferens and in the seminal vesicle of the rat from the rate of disappearance of H3-norepinephrine after an i.v. injection of

The turnover times of tissue norepinephrine (NE) were determined in various organs of the rat from the rate of disappearance of H3-NE after an intravenous injection of a same dose ofl ordl H3-NE. In all cases, the labelled amine disappeared by a single exponential decline thus behaving kinetically as though it was stored in a single pool.