The long-term goal of this study is to identify and characterize functional, biochemical and molecular abnormalities of the cerebral microcirculation in aging and dementia. Examination of blood-brain barrier function is critical to understanding the pathogenesis of neuronal cell death in aging and Alzheimer's Disease because the cerebral endothelium actively regulates the neuronal ionic and nutrient microenvironment. Although the extent and importance of blood-brain barrier abnormalities in aging and Alzheimer's disease are controversial, considerable evidence exists for a dysfunctional blood-brain barrier. It is our hypothesis that abnormalities in microvascular receptor-mediated signaling pathways may contribute to an altered blood-brain barrier responsiveness in Alzheimer's disease. Abnormalities of the BBB may be age- and/or Alzheimer's disease disease- related. Consequently, microvascular responsiveness in Alzheimer's disease in age-matched, non-demented elderly patientsand i adult control patients will be assessed. In addition, experiments in young and aged rodents are necessary an an immediate and ready source of material on which methods, storage, and age-related changes can be evaluated. Specifically, microvessel adrenergic and cholinergic receptors, adenylate cyclase, and protein kinase C, under both basal and agonist stimulated conditions, will be assessed from the above groups in order to identify- age and disease-related alterations. Experiments are planned to analyze, (1) receptor binding parameters using radioligand binding techniques, receptor-linked Gs function via reconstitution in S49 cyc (G protein deficient) cells, and levels of Gs and Gi using toxin ribosylation; (2) adenylate cyclase levels by measuring cAMP accumulation and [3H]forskolin binding, specificity and activity of this enzyme by phosphoprotein analysis, and mRNA expression by RNA blot analysis; and (3) protein kinase C activity, basal distribution and agonist-stimulated translocation, isoform levels by Western blots, and message levels by Northern analysis. The results will define the function and regulation of intracellular signalling pathways at the blood-brain barrier and may provide a basis for the development of rational therapeutic strategies for improved cerebrovascular and neuronal function in Alzheimer's disease.
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