Cerebrovascular dysfunction contributes to the pathology and progression of Alzheimer's disease (AD), but the mechanisms are not completely understood. Using transgenic mouse models of AD (TgCRND8, PDAPP, and Tg2576), we evaluated blood–brain barrier damage and the role of fibrin and fibrinolysis in the progression of amyloid-β pathology. These mouse models showed age-dependent fibrin deposition coincident with areas of blood–brain barrier permeability as demonstrated by Evans blue extravasation. Three lines of evidence suggest that fibrin contributes to the pathology. First, AD mice with only one functional plasminogen gene, and therefore with reduced fibrinolysis, have increased neurovascular damage relative to AD mice. Conversely, AD mice with only one functional fibrinogen gene have decreased blood–brain barrier damage. Second, treatment of AD mice with the plasmin inhibitor tranexamic acid aggravated pathology, whereas removal of fibrinogen from the circulation of AD mice with ancrod treatment attenuated measures of neuroinflammation and vascular pathology. Third, pretreatment with ancrod reduced the increased pathology from plasmin inhibition. These results suggest that fibrin is a mediator of inflammation and may impede the reparative process for neurovascular damage in AD. Fibrin and the mechanisms involved in its accumulation and clearance may present novel therapeutic targets in slowing the progression of AD.

Cerebral amyloid ␤-protein angiopathy (CAA) is a key pathological feature of patients with Alzheimer's disease and certain related disorders. In these conditions the CAA is characterized by the deposition of A␤ within the cerebral vessel wall and, in severe cases, hemorrhagic stroke. Several mutations have been identified within the A␤ region of the A␤ protein precursor (A␤PP) gene that appear to enhance the severity of CAA. We recently described a new mutation within the A␤ region (D23N) of A␤PP that is associated with severe CAA in an Iowa kindred (Grabowski, T. J., Cho, H. S., Vonsattel, J. P. G., Rebeck, G. W., and Greenberg, S. M. (2001) Ann. Neurol. 49, 697-705). In the present study, we investigated the effect of this new D23N mutation on the processing of A␤PP and the pathogenic properties of A␤. Neither the D23N Iowa mutation nor the E22Q Dutch mutation affected the amyloidogenic processing of A␤PP expressed in H4 cells. The A21G Flemish mutation, in contrast, resulted in a 2.3-fold increase in secreted A␤ peptide. We also tested synthetic wild-type and mutant A␤40 peptides for fibrillogenesis and toxicity toward cultured human cerebrovascular smooth muscle (HC-SM) cells. The E22Q Dutch, D23N Iowa, and E22Q,D23N Dutch/Iowa double mutant A␤40 peptides rapidly assembled in solution to form fibrils, whereas wild-type and A21G Flemish A␤40 peptides exhibited little fibril formation. Similarly, the E22Q Dutch and D23N Iowa A␤40 peptides were found to induce robust pathologic responses in cultured HCSM cells, including elevated levels of cell-associated A␤PP, proteolytic breakdown of smooth muscle cell ␣-actin, and cell death. Double mutant E22Q,D23N Dutch/Iowa A␤40 was more potent than either single mutant form of A␤ in causing pathologic responses in HCSM cells. These data suggest that the different CAA mutations in A␤PP may exert their pathogenic effects through different mechanisms. Whereas the A21G Flemish mutation appears to enhance A␤ production, the E22Q Dutch and D23N Iowa mutations enhance fibrillogenesis and the pathogenicity of A␤ toward HCSM cells. Cerebral amyloid angiopathy (CAA)1 is a common pathology found at increased frequency in patients with Alzheimer's disease (AD) and related disorders such as Down's syndrome and hereditary cerebral hemorrhage with amyloidosis of the Dutch type (1-7). A␤ is a 39 -43-amino acid peptide that is proteolytically derived from its larger parent transmembrane molecule amyloid-␤ precursor protein (A␤PP) (8). Although A␤PP exhibits a variety of biological activities, its role as the parent molecule of the A␤ peptide has drawn the most interest (9). In this regard, full-length A␤PP can undergo proteolytic cleavage by ␤-and ␥-secretases to liberate the A␤ peptide. Alternatively, fulllength A␤PP can be proteolytically processed by an enzyme termed ␣-secretase at position 16 of the A␤ domain, resulting in a non-amyloidogenic membrane-spanning carboxyl-terminal fragment and truncated secretory forms of A␤PP that are released into the extracellular environment. Mutati...

Although neuronal stress circuits have been identified, little is known about the mechanisms that underlie the stress-induced neuronal plasticity leading to fear and anxiety. Here we found that the serine protease tissue-plasminogen activator (tPA) was upregulated in the central and medial amygdala by acute restraint stress, where it promoted stress-related neuronal remodeling and was subsequently inhibited by plasminogen activator inhibitor-1 (PAI-1). These events preceded stress-induced increases in anxiety-like behavior of mice. Mice in which the tPA gene has been disrupted did not show anxiety after up to three weeks of daily restraint and showed attenuated neuronal remodeling as well as a maladaptive hormonal response. These studies support the idea that tPA is critical for the development of anxiety-like behavior after stress.

Accumulation of the amyloid-beta (Abeta) peptide depends on both its generation and clearance. To better define clearance pathways, we have evaluated the role of the tissue plasminogen activator (tPA)-plasmin system in Abeta degradation in vivo. In two different mouse models of Alzheimer's disease, chronically elevated Abeta peptide in the brain correlates with the upregulation of plasminogen activator inhibitor-1 (PAI-1) and inhibition of the tPA-plasmin system. In addition, Abeta injected into the hippocampus of mice lacking either tPA or plasminogen persists, inducing PAI-1 expression and causing activation of microglial cells and neuronal damage. Conversely, Abeta injected into wild-type mice is rapidly cleared and does not cause neuronal degeneration. Thus, the tPA-plasmin proteolytic cascade aids in the clearance of Abeta, and reduced activity of this system may contribute to the progression of Alzheimer's disease.

Repeated stress can impair function in the hippocampus, a brain structure essential for learning and memory. Although behavioral evidence suggests that severe stress triggers cognitive impairment, as seen in major depression or posttraumatic stress disorder, little is known about the molecular mediators of these functional deficits in the hippocampus. We report here both pre-and postsynaptic effects of chronic stress, manifested as a reduction in the number of NMDA receptors, dendritic spines, and expression of growth-associated protein-43 in the cornu ammonis 1 region. Strikingly, the stress-induced decrease in NMDA receptors coincides spatially with sites of plasminogen activation, thereby predicting a role for tissue plasminogen activator (tPA) in this form of stress-induced plasticity. Consistent with this possibility, tPA؊͞؊ and plasminogen؊͞؊ mice are protected from stress-induced decrease in NMDA receptors and reduction in dendritic spines. At the behavioral level, these synaptic and molecular signatures of stressinduced plasticity are accompanied by impaired acquisition, but not retrieval, of hippocampal-dependent spatial learning, a deficit that is not exhibited by the tPA؊͞؊ and plasminogen؊͞؊ mice. These findings establish the tPA͞plasmin system as an important mediator of the debilitating effects of prolonged stress on hippocampal function at multiple levels of neural organization.dendritic spines ͉ learning ͉ NMDA receptor P sychological stress induces neuronal responses that can be either adaptive and directed toward maintaining homeostasis or maladaptive, leading to severe behavioral abnormalities (1). Posttraumatic stress disorder (PTSD) is a devastating disease triggered by a severe traumatic event(s) and characterized by cognitive impairment, depression, fear, and anxiety (2). Although little is known about the cellular mechanisms of PTSD, its different aspects are mediated by different brain structures (3). Animal and human studies suggest that stress-induced fear and anxiety are mediated by the amygdala (4, 5), and cognitive decline is a result of hippocampal dysfunction (6, 7). It has been hypothesized that the decrease in complexity of the hippocampal dendritic tree contributes to learning deficits (8), but molecular mechanisms underlying this dendritic plasticity are poorly understood.One molecule strategically positioned to control neuronal activity, dendritic remodeling, and learning is the NMDA receptor. It is located on dendritic spines and is critically involved in spine motility (9) and experience-induced neuronal plasticity (10). Although the decrease in the number of NMDA receptors leads to memory deficits (10), overexpression of some of its subunits results in more efficient learning (11).There is evidence that NMDA receptor function is linked to stress-induced neuronal and cognitive changes, because stressinduced remodeling is blocked by NMDA-receptor antagonists (12). The NMDA receptor has numerous ligands and modulators, and it is likely that the above processes may involve a n...