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Background: Cortical spreading depolarization (CSD) is a massive neuro-glial depolarization wave, which propagates across the cerebral cortex. In stroke, CSD is a necessary and ubiquitous mechanism for the development of neuronal lesions that initiates in the ischemic core and propagates through the penumbra extending the tissue injury. Although CSD propagation induces dramatic changes in cerebral blood flow, the vascular responses in different ischemic regions and their consequences on reperfusion and recovery remain to be defined., Methods: Ischemia was performed using the thrombin model of stroke and reperfusion was induced by r-tPA (recombinant tissue-type plasminogen activator) administration in mice. We used in vivo electrophysiology and laser speckle contrast imaging simultaneously to assess both electrophysiological and hemodynamic characteristics of CSD after ischemia onset. Neurological deficits were assessed on day 1, 3, and 7. Furthermore, infarct sizes were quantified using 2,3,5-triphenyltetrazolium chloride on day 7., Results: After ischemia, CSDs were evidenced by the characteristic propagating DC shift extending far beyond the ischemic area. On the vascular level, we observed 2 types of responses: some mice showed spreading hyperemia confined to the penumbra area (penumbral spreading hyperemia) while other showed spreading hyperemia propagating in the full hemisphere (full hemisphere spreading hyperemia). Penumbral spreading hyperemia was associated with severe stroke-induced damage, while full hemisphere spreading hyperemia indicated beneficial infarct outcome and potential viability of the infarct core. In all animals, thrombolysis with r-tPA modified the shape of the vascular response to CSD and reduced lesion volume., Conclusions: Our results show that different types of spreading hyperemia occur spontaneously after the onset of ischemia. Depending on their shape and distribution, they predict severity of injury and outcome. Furthermore, our data show that modulating the hemodynamic response to CSD may be a promising therapeutic strategy to attenuate stroke outcome.

Following stroke, the survival of neurons and their ability to reestablish connections is critical to functional recovery. This is strongly influenced by the balance between neuronal excitation and inhibition. In the acute phase of experimental stroke, lethal hyperexcitability can be attenuated by positive allosteric modulation of GABA A receptors (GABA A Rs). Conversely, in the late phase, negative allosteric modulation of GABA A R can correct the suboptimal excitability and improves both sensory and motor recovery. Here, we hypothesized that octadecaneuropeptide (ODN), an endogenous allosteric modulator of the GABA A R synthesized by astrocytes, influences the outcome of ischemic brain tissue and subsequent functional recovery. We show that ODN boosts the excitability of cortical neurons, which makes it deleterious in the acute phase of stroke. However, if delivered after day 3, ODN is safe and improves motor recovery over the following month in two different paradigms of experimental stroke in mice. Furthermore, we bring evidence that, during the subacute period after stroke, the repairing cortex can be treated with ODN by means of a single hydrogel deposit into the stroke cavity. SIGNIFICANCE STATEMENT Stroke remains a devastating clinical challenge because there is no efficient therapy to either minimize neuronal death with neuroprotective drugs or to enhance spontaneous recovery with neurorepair drugs. Around the brain damage, the peri-infarct cortex can be viewed as a reservoir of plasticity. However, the potential of wiring new circuits in these areas is restrained by a chronic excess of GABAergic inhibition. Here we show that an astrocyte-derived peptide, can be used as a delayed treatment, to safely correct cortical excitability and facilitate sensorimotor recovery after stroke., (Copyright © 2021 the authors.)

26RFa is a neuropeptide that activates the rhodopsin-like G protein-coupled receptor QRFPR/GPR103. This peptidergic system is involved in the regulation of a wide array of physiological processes including feeding behavior and glucose homeostasis. Herein, the pharmacological profile of a homogenous library of QRFPR-targeting peptide derivatives was investigated in vitro on human QRFPR-transfected cells with the aim to provide possible insights into the structural determinants of the Phe residues to govern receptor activation. Our work advocates to include in next generations of 26RFa (20-26) -based QRFPR agonists effective substitutions for each Phe unit, i.e., replacement of the Phe 22 residue by a constrained 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid moiety, and substitution of both Phe 24 and Phe 26 by their para -chloro counterpart. Taken as a whole, this study emphasizes that optimized modifications in the C-terminal part of 26RFa are mandatory to design selective and potent peptide agonists for human QRFPR.

Days and weeks after an ischemic stroke, the peri-infarct area adjacent to the necrotic tissue exhibits very intense synaptic reorganization aimed at regaining lost functions. In order to enhance functional recovery, it is important to understand the mechanisms supporting neural repair and neuroplasticity in the cortex surrounding the lesion. Brain oscillations of the local field potential (LFP) are rhythmic fluctuations of neuronal excitability that synchronize neuronal activity to organize information processing and plasticity. Although the oscillatory activity of the brain has been probed after stroke in both animals and humans using electroencephalography (EEG), the latter is ineffective to precisely map the oscillatory changes in the peri-infarct zone where synaptic plasticity potential is high. Here, we worked on the hypothesis that the brain oscillatory system is altered in the surviving peri-infarct cortex, which may slow down the functional repair and reduce the recovery. In order to document the relevance of this hypothesis, oscillatory power was measured at various distances from the necrotic core at 7 and 21 days after a permanent cortical ischemia induced in mice. Delta and theta oscillations remained at a normal power in the peri-infarct cortex, in contrast to low gamma oscillations that displayed a gradual decrease, when approaching the border of the lesion. A broadband increase of power was also observed in the homotopic contralateral sites. Thus, the proximal peri-infarct cortex could become a target of therapeutic interventions applied to correct the oscillatory regimen in order to boost post-stroke functional recovery., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Octadecaneuropeptide (ODN) and its precursor diazepam-binding inhibitor (DBI) are peptides belonging to the family of endozepines. Endozepines are exclusively produced by astroglial cells in the central nervous system of mammals, and their release is regulated by stress signals and neuroactive compounds. There is now compelling evidence that the gliopeptide ODN protects cultured neurons and astrocytes from apoptotic cell death induced by various neurotoxic agents. In vivo , ODN causes a very strong neuroprotective action against neuronal degeneration in a mouse model of Parkinson's disease. The neuroprotective activity of ODN is based on its capacity to reduce inflammation, apoptosis, and oxidative stress. The protective effects of ODN are mediated through its metabotropic receptor. This receptor activates a transduction cascade of second messengers to stimulate protein kinase A (PKA), protein kinase C (PKC), and mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase (ERK) signaling pathways, which in turn inhibits the expression of proapoptotic factor Bax and the mitochondrial apoptotic pathway. In N2a cells, ODN also promotes survival and stimulates neurite outgrowth. During the ODN-induced neuronal differentiation process, numerous mitochondria and peroxisomes are identified in the neurites and an increase in the amount of cholesterol and fatty acids is observed. The antiapoptotic and neurotrophic properties of ODN, including its antioxidant, antiapoptotic, and pro-differentiating effects, suggest that this gliopeptide and some of its selective and stable derivatives may have therapeutic value for the treatment of some neurodegenerative diseases., (Copyright © 2020 Masmoudi-Kouki, Namsi, Hamdi, Bahdoudi, Ghouili, Chuquet, Leprince, Lefranc, Ghrairi, Tonon, Lizard and Vaudry.)

The existence of specific binding sites for benzodiazepines (BZs) in the brain has prompted the search for endogenous BZ receptor ligands designated by the generic term « endozepines ». This has led to the identification of an 86-amino acid polypeptide capable of displacing [ 3 H]diazepam binding to brain membranes, thus called diazepam-binding inhibitor (DBI). It was subsequently found that the sequence of DBI is identical to that of a lipid carrier protein termed acyl-CoA-binding protein (ACBP). The primary structure of DBI/ACBP has been well preserved, suggesting that endozepines exert vital functions. The DBI/ACBP gene is expressed by astroglial cells in the central nervous system, and by various cell types in peripheral organs. Endoproteolytic cleavage of DBI/ACBP generates several bioactive peptides including a triakontatetraneuropeptide that acts as a selective ligand of peripheral BZ receptors/translocator protein, and an octadecaneuropeptide that activates a G protein-coupled receptor and behaves as an allosteric modulator of the GABA A R. Although DBI/ACBP is devoid of a signal peptide, endozepines are released by astrocytes in a regulated manner. Consistent with the diversity and wide distribution of BZ-binding sites, endozepines appear to exert a large array of biological functions and pharmacological effects. Thus, intracerebroventricular administration of DBI or derived peptides induces proconflict and anxiety-like behaviors, and reduces food intake. Reciprocally, the expression of DBI/ACBP mRNA is regulated by stress and metabolic signals. In vitro, endozepines stimulate astrocyte proliferation and protect neurons and astrocytes from apoptotic cell death. Endozepines also regulate neurosteroid biosynthesis and neuropeptide expression, and promote neurogenesis. In peripheral organs, endozepines activate steroid hormone production, stimulate acyl chain ceramide synthesis and trigger pro-inflammatory cytokine secretion. The expression of the DBI/ACBP gene is enhanced in addiction/withdrawal animal models, in patients with neurodegenerative disorders and in various types of tumors. We review herein the current knowledge concerning the various actions of endozepines and discuss the physiopathological implications of these regulatory gliopeptides., (Copyright © 2019. Published by Elsevier Inc.)

26RFa, the endogenous QRFPR ligand, is implicated in several physiological and pathological conditions such as the regulation of glucose homeostasis and bone mineralization; hence, QRFPR ligands display therapeutic potential. At the molecular level, functional interaction occurs between residues Arg 25 of 26RFa and Gln 125 of QRFPR. We have designed 26RFa (20-26) analogues incorporating arginine derivatives modified by alkylated substituents. We found that the Arg 25 side chain length was necessary to retain the activity of 26RFa (20-26) and that N-monoalkylation of arginine was accommodated by the QRFPR active site. In particular, [(Me) ω Arg 25 ]26RFa (20-26) (5b, LV-2186) appeared to be 25-fold more potent than 26RFa (20-26) and displayed a position in a QRFPR homology model slightly different to that of the unmodified heptapeptide. Other peptides were less potent than 26RFa (20-26) , exhibited partial agonistic activity, or were totally inactive in accordance to different ligand-bound structures. In vivo, [(Me) ω Arg 25 ]26RFa (20-26) exerted a delayed 26RFa-like hypoglycemic effect. Finally, N-methyl substituted arginine-containing peptides represent lead compounds for further development of QRFPR agonists.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by a progressive loss of dopamine (DA) neurons through apoptotic, inflammatory and oxidative stress mechanisms. The octadecaneuropeptide (ODN) is a diazepam-binding inhibitor (DBI)-derived peptide, expressed by astrocytes, which protects neurons against oxidative cell damages and apoptosis in an in vitro model of PD. The present study reveals that a single intracerebroventricular injection of 10 ng ODN 1 h after the last administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) prevented the degeneration of DA neurons induced by the toxin in the substantia nigra pars compacta of mice, 7 days after treatment. ODN-mediated neuroprotection was associated with a reduction of the number of glial fibrillary acidic protein-positive reactive astrocytes and a strong inhibition of the expression of pro-inflammatory genes such as interleukins 1β and 6, and tumor necrosis factor-α. Moreover, ODN blocked the inhibition of the anti-apoptotic gene Bcl-2, and the stimulation of the pro-apoptotic genes Bax and caspase-3, induced by MPTP in the substantia nigra pars compacta. ODN also decreased or even in some cases abolished MPTP-induced oxidative damages, overproduction of reactive oxygen species and accumulation of lipid oxidation products in DA neurons. Furthermore, DBI knockout mice appeared to be more vulnerable than wild-type animals to MPTP neurotoxicity. Taken together, these results show that the gliopeptide ODN exerts a potent neuroprotective effect against MPTP-induced degeneration of nigrostriatal DA neurons in mice, through mechanisms involving downregulation of neuroinflammatory, oxidative and apoptotic processes. ODN may, thus, reduce neuronal damages in PD and other cerebral injuries involving oxidative neurodegeneration.

Aim: Develop modified dextran nanoparticles showing potential to assist with drug permeation across the blood-brain barrier for the delivery of neuropeptides., Methods: Nanoparticles loaded by emulsification with model macromolecular actives were characterized in terms of stability, cytotoxicity and drug-release behavior. Peptide-loaded nanoformulations were tested in an in vivo trout model and in food-deprived mice., Results: Nanoformulations loaded with model peptides showed good stability and appeared nontoxic in low concentration against human brain endothelial cells. They were found to preserve the bioactivity of loaded peptides (angiotensin II) as demonstrated in vivo using a trout model, and to induce a transient reduction of food consumption in mice when loaded with an anorexigenic octaneuropeptide., Conclusion: Octylglyceryl dextran-graft-poly(lactic acid) nanoparticles formulated by emulsification demonstrate potential for peptide delivery.

Astrocytes synthesize and release endozepines, a family of regulatory neuropeptides, including diazepam-binding inhibitor (DBI) and its processing fragments such as the octadecaneuropeptide (ODN). At the molecular level, ODN interacts with two types of receptors, i.e. it acts as an inverse agonist of the central-type benzodiazepine receptor (CBR), and as an agonist of a G protein-coupled receptor (GPCR). ODN exerts a wide range of biological effects mediated through these two receptors and, in particular, it regulates astrocyte activity through an autocrine/paracrine mechanism involving the metabotropic receptor. More recently, it has been shown that Müller glial cells secrete phosphorylated DBI and that bisphosphorylated ODN ([bisphospho-Thr(3,9)]ODN, bpODN) has a stronger affinity for CBR than ODN. The aim of the present study was thus to investigate whether bpODN is released by mouse cortical astrocytes and to compare its potency to ODN. Using a radioimmunoassay and mass spectrometry analysis we have shown that bpODN as well as ODN were released in cultured astrocyte supernatants. Both bpODN and ODN increased astrocyte calcium event frequency but in a very different range of concentration. Indeed, ODN stimulatory effect decreased at concentrations over 10(-10)M whereas bpODN increased the calcium event frequency at similar doses. In vivo effects of bpODN and ODN were analyzed in two behavioral paradigms involving either the metabotropic receptor (anorexia) or the CBR (anxiety). As previously described, ODN (100ng, icv) induced a significant reduction of food intake. Similar effect was achieved with bpODN but at a 10 times higher dose (1000 ng, icv). Similarly, and contrasting with our hypothesis, bpODN was also 10 times less potent than ODN to induce anxiety-related behavior in the elevated zero maze test. Thus, the present data do not support that phosphorylation of ODN is involved in receptor selectivity but indicate that it rather weakens ODN activity., (Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.)

Background and Purpose: The neuropeptide 26RFa and its cognate receptor GPR103 are involved in the control of food intake and bone mineralization. Here, we have tested, experimentally, the predicted ligand-receptor interactions by site-directed mutagenesis of GPR103 and designed point-substituted 26RFa analogues., Experimental Approach: Using the X-ray structure of the β2 -adrenoceptor, a 3-D molecular model of GPR103 has been built. The bioactive C-terminal octapeptide 26RFa(19-26) , KGGFSFRF-NH2 , was docked in this GPR103 model and the ligand-receptor complex was submitted to energy minimization., Key Results: In the most stable complex, the Phe-Arg-Phe-NH2 part was oriented inside the receptor cavity, whereas the N-terminal Lys residue remained outside. A strong intermolecular interaction was predicted between the Arg(25) residue of 26RFa and the Gln(125) residue located in the third transmembrane helix of GPR103. To confirm this interaction experimentally, we tested the ability of 26RFa and Arg-modified 26RFa analogues to activate the wild-type and the Q125A mutant receptors transiently expressed in CHO cells. 26RFa (10(-6)  M) enhanced [Ca(2+) ]i in wild-type GPR103-transfected cells, but failed to increase [Ca(2+) ]i in Q125A mutant receptor-expressing cells. Moreover, asymmetric dimethylation of the side chain of arginine led to a 26RFa analogue, [ADMA(25) ]26RFa(20-26) , that was unable to activate the wild-type GPR103, but antagonized 26RFa-evoked [Ca(2+) ]i increase., Conclusion and Implications: Altogether, these data provide strong evidence for a functional interaction between the Arg(25) residue of 26RFa and the Gln(125) residue of GPR103 upon ligand-receptor activation, which can be exploited for the rational design of potent GPR103 agonists and antagonists., (© 2014 The British Pharmacological Society.)

26RFa, a novel RFamide neuropeptide, is the endogenous ligand of the former orphan receptor GPR103. Intracerebroventricular injection of 26RFa and its C-terminal heptapeptide, 26RFa((20-26)), stimulates food intake in rodents. To develop potent, stable ligands of GPR103 with low molecular weight, we have designed a series of aza-β(3)-containing 26RFa((20-26)) analogues for their propensity to establish intramolecular hydrogen bonds, and we have evaluated their ability to increase [Ca(2+)](i) in GPR103-transfected cells. We have identified a compound, [Cmpi(21),aza-β(3)-Hht(23)]26RFa((21-26)), which was 8-fold more potent than 26RFa((20-26)) in mobilizing [Ca(2+)](i). This pseudopeptide was more stable in serum than 26RFa((20-26)) and exerted a longer lasting orexigenic effect in mice. This study constitutes an important step toward the development of 26RFa analogues that could prove useful for the treatment of feeding disorders.

Astrocytes communicate with synapses by means of intracellular calcium ([Ca(2+)](i)) elevations, but local calcium dynamics in astrocytic processes have never been thoroughly investigated. By taking advantage of high-resolution two-photon microscopy, we identify the characteristics of local astrocyte calcium activity in the adult mouse hippocampus. Astrocytic processes showed intense activity, triggered by physiological transmission at neighboring synapses. They encoded synchronous synaptic events generated by sparse action potentials into robust regional (∼12 μm) [Ca(2+)](i) elevations. Unexpectedly, they also sensed spontaneous synaptic events, producing highly confined (∼4 μm), fast (millisecond-scale) miniature Ca(2+) responses. This Ca(2+) activity in astrocytic processes is generated through GTP- and inositol-1,4,5-trisphosphate-dependent signaling and is relevant for basal synaptic function. Thus, buffering astrocyte [Ca(2+)](i) or blocking a receptor mediating local astrocyte Ca(2+) signals decreased synaptic transmission reliability in minimal stimulation experiments. These data provide direct evidence that astrocytes are integrated in local synaptic functioning in adult brain., (© 2011 Nature America, Inc. All rights reserved.)

Glucose is the primary energetic substrate of the brain, and measurements of its metabolism are the basis of major functional cerebral imaging methods. Contrary to the general view that neurons are fueled solely by glucose in proportion to their energetic needs, recent in vitro and ex vivo analyses suggest that glucose preferentially feeds astrocytes. However, the cellular fate of glucose in the intact brain has not yet been directly observed. We have used a real-time method for measuring glucose uptake in astrocytes and neurons in vivo in male rats by imaging the trafficking of the nonmetabolizable glucose analog 6-deoxy-N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-aminoglucose (6-NBDG) using two-photon microscopy. During resting conditions we found that astrocytes and neurons both take up 6-NBDG at the same rate in the barrel cortex of the rat. However, during intense neuronal activity triggered by whisker stimulation, astrocytes rapidly accelerated their uptake, whereas neuronal uptake remained almost unchanged. After the stimulation period, astrocytes returned to their preactivation rates of uptake paralleling the neuronal rate of uptake. These observations suggest that glucose is taken up primarily by astrocytes, supporting the view that functional imaging experiments based on glucose analogs extraction may predominantly reflect the metabolic activity of the astrocytic network.

Urotensin-II (U-II) is a cyclic peptide identified recently in many mammalian species including man. U-II and its receptor are expressed in the central nervous system, in the cardiovascular system and in other peripheral tissues. Although this peptide has been reported initially to be a potent vasoconstrictor, increasing evidence shows that its vascular actions strongly depend on species and vascular beds. Here we analyzed the effects of U-II administration on cerebral blood flow (CBF) under physiological conditions and following cerebral ischemia in rats. Although intravenous injection of U-II had minimal effects on CBF as measured by the technique of laser Doppler flowmetry, its administration (10 nmol) into the lateral cerebral ventricle induced gradual and long lasting increase in CBF (+61% at 1 h post-injection, p<0.05). These U-II-mediated CBF increases were not related to the transient systemic pressor actions of the peptide and were reduced by nitric oxide synthase inhibition (61 vs 17%, p<0.05). Intracerebroventricular administration of U-II following the induction of cerebral ischemia, failed to alter residual CBF in the affected cerebral hemisphere. Nonetheless, following reperfusion (90 min after ischemia), U-II-treated animals displayed a remarkable hyperperfusion compared to vehicle-treated rats (+168%, p<0.05). The volume of infarction was significantly increased in U-II-treated rats (+40%, p<0.05). These results provide the first evidence that U-II increases cerebral blood flow when administered into the cerebral ventricle and exacerbates brain damage following an ischemic insult.

Background and Purpose: Whereas the effects of chronic arterial hypertension on the cerebral vasculature have been widely studied, its effects on brain tissue have been studied less so. Here we examined if spontaneously hypertensive rats (SHRs) or the normotensive control Wistar Kyoto rats (WKYs) made hypertensive by renal artery stenosis (R-WKYs) are vulnerable to an excitotoxic brain lesion provoked by an overactivation of glutamate receptors., Methods: Lesion volumes were quantified by histology in WKYs and SHRs subjected to striatal administration of N-methyl-d-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA). The expression of AMPA receptors subunits and calcium/calmodulin kinase-II alpha was analyzed by real-time polymerase chain reaction and Western blot., Results: NMDA (50 and 75 nmol) induced similar lesions in both SHRs (10+/-2 mm(3) and 16+/-4 mm(3), respectively) and WKYs (11+/-2 mm(3) and 19+/-7 mm(3), respectively). However, AMPA-induced (2.5 and 5 nmol) lesions were significantly greater in 14-week-old SHRs (14+/-3 mm(3) and 20+/-5 mm(3), respectively) than WKYs (4+/-2 mm(3), P<0.05 and 7+/-4 mm(3), P<0.001, respectively). Furthermore, normotensive 7-week-old SHRs also displayed an aggravated AMPA-induced lesion compared with age-matched WKYs (10+/-3 mm(3) vs 6+/-3 mm(3); P<0.05). Neither NMDA nor AMPA produced increased lesion volumes in R-WKYs (12+/-3 mm(3) and 5+/-4 mm(3), respectively) compared with WKYs. Striatal levels of AMPA receptors subunits, GluR1 and GluR2, were not different between SHRs and WKYs. However, SHRs displayed an increase in phosphorylated form of GluR1 at Ser-831 (P<0.05), as well as in calcium/calmodulin kinase-II alpha (P<0.002). Selective inhibition of this kinase by KN-93 reduced AMPA-induced damage in SHRs (P<0.01 vs vehicle)., Conclusions: These findings show that an increase in phosphorylated GluR1, which increases AMPA receptor conductance, may be involved in the vulnerability of SHRs to AMPA.

Spreading depression (SD) is a propagating wave of neuronal depolarization and ionic shifts, seen in stroke and migraine. In vitro, SD is associated with astrocytic [Ca2+] waves, but it is unclear what role they play and whether they influence cerebral blood flow, which is altered in SD. Here we show that SD in vivo is associated with [Ca2+] waves in astrocytes and neurons and with constriction of intracortical arterioles severe enough to result in arrest of capillary perfusion. The vasoconstriction is correlated with fast astrocytic [Ca2+] waves and is inhibited when they are reduced. [Ca2+] waves appear in neurons before astrocytes, and inhibition of astrocytic [Ca2+] waves does not depress SD propagation. This suggests that astrocytes do not drive SD propagation but are responsible for the hemodynamic failure seen deep in the cortex. Similar waves occur in anoxic depolarizations (AD), supporting the notion that SD and AD are related processes.

Background: Accumulating evidence demonstrates a critical involvement of tissue-type plasminogen activator (tPA) in pathological and physiological brain conditions. Determining whether and how vascular tPA can cross the blood-brain barrier (BBB) to enter the brain is thus important, not only during stroke but also in physiological conditions., Methods and Results: In the present work, we provide evidence in vivo that intravenous injection of tPA increases NMDA-induced striatal lesion in the absence of BBB leakage. Accordingly, we show that tPA crosses the BBB both after excitotoxic lesion and in control conditions. Indeed, vascular injected tPA can be detected within the brain parenchyma and in the cerebrospinal fluid. By using an in vitro model of BBB, we have confirmed that tPA can cross the intact BBB. Its passage was blocked at 4 degrees C, was saturable, and was independent of its proteolytic activity. We have shown that tPA crosses the BBB by transcytosis, mediated by a member of the LDL receptor-related protein family., Conclusions: We demonstrate that blood-derived tPA can reach the brain parenchyma without alteration of the BBB. The molecular mechanism of the passage of tPA from blood to brain described here could represent an interesting target to improve thrombolysis in stroke.

Background and Purpose: Endothelins act through 2 receptors, namely, ET(A) and ET(B). In the cerebral circulation, ET(A) mediates marked and prolonged vasoconstriction, and its blockade increases cerebral blood flow (CBF) and reduces ischemic brain damage. However, the role of ET(B) receptors remains unclear. In this study we examined, in rats, the kinetics of expression of ET(B) and the effects of ET(B) blockade on changes in CBF and brain damage after focal cerebral ischemia and N-methyl-D-aspartate (NMDA)-induced excitotoxic injury., Methods: Rats were subjected to transient (60 minutes) focal cerebral ischemia or cortical injection of NMDA. The selective ET(B) antagonist BQ-788 was injected intracerebroventricularly 30 minutes before and 30 minutes after the onset of ischemia. Cortical perfusion was monitored by laser-Doppler flowmetry. The volume of infarction or NMDA-induced cortical lesion was assessed at 24 hours after the insult. The reverse transcription-polymerase chain reaction technique was used to assess ET(B) expression., Results: Cerebral ischemia failed to alter the expression of ET(B) mRNA in both acute and chronic stages. Administration of BQ-788 resulted in an increase in infarction volume (178%; P<0.05) accompanied by a decrease in residual CBF (-26.7% versus control; P<0.01). In these animals we found a positive correlation between the volume of infarction and the severity of the decrease in CBF. NMDA-induced cortical lesions were not affected by the administration of BQ-788., Conclusions: Our results suggest that the ET(B) antagonist BQ-788 induces deleterious effects that are mediated by the reduction of residual blood flow after ischemia and argue that the optimal therapeutic strategy in stroke would be to target the use of selective ET(A) antagonists and not mixed ET(A)/ET(B) antagonists.

To correlate brain metabolic status with the molecular events during cerebral ischemia, a cDNA array was performed after positron emission tomography scanning in a model of focal cerebral ischemia in baboons. Cluster analysis for the expression of 74 genes allowed the identification of 4 groups of genes. In each of the distinct groups, the authors observed a marked inflection in the pattern of gene expression when the CMRo was reduced by 48% to 66%. These patterns of coordinated modifications in gene expression could define molecular checkpoints for the development of an ischemic infarct and a molecular definition of the penumbra.

Interleukin-1 (IL-1) has pleiotropic actions in the central nervous system. During the last decade, a growing corpus of evidence has indicated an important role of this cytokine in the development of brain damage following cerebral ischaemia. The expression of IL-1 in the brain is dramatically increased during the early and chronic stage of infarction. The most direct evidence that IL-1 contributes significantly to ischaemic injury is that (1) central administration of IL-1beta exacerbates brain damage, and (2) injection or over-expression of interleukin-1 receptor antagonist, and blockade of interleukin-1beta converting enzyme activity reduce, dramatically, infarction and improve behavioural deficit. The mechanisms underlying IL-1 actions in stroke are not definitively elucidated, and it seems likely that its effects are mediated through stimulation and inhibition of wide range of pathophysiological processes.