ABSTRACT The objective of this application is to investigate glymphatic impairment and cognitive deficits during progression of aging with and without diabetes. Emerging data1-5 indicate that the glymphatic system in the brain mediates the cerebrospinal fluid (CSF)-interstitial (ISF) exchange and solute clearance from the brain parenchyma. However, despite the well-described dysfunction of the glymphatic system in the development of neurodegenerative conditions, there is still no reported study that focuses on the role of the glymphatic system in the development of cognitive impairment during aging and aging with type-2 diabetes (DM). Using non- invasive MRI methodologies to investigate cerebral solute waste clearance in middle-age control and type-2 diabetic (DM) rats, we have found increased impairment of the glymphatic system, as indicated by reduced clearance of interstitial Gd-DTPA in brain parenchyma, primarily in the hippocampus and hypothalamus in DM rats (Fig.2&3)6. In parallel, 3D confocal microscopic analysis of the brain-wide distribution of fluorescent tracers revealed increased delayed clearance of ISF in the hippocampus and hypothalamus from DM rats (Fig.2&3)6. Impairment of the glymphatic system in DM rats was shown to be highly correlated with cognitive deficits as measured by an array of cognitive tests including the Morris Water Maze (MWM) for hippocampal related learning and memory. Importantly, histopathological analysis shows that delayed clearance of interstitial solutes is associated with sporadic cerebral microvascular thrombosis in the hippocampus 2 months after hyperglycemia (15 months from birth), while extensive microvascular thrombosis and para-vascular accumulation of beta- amyloid (A?) are detected at 4 months after induction of hyperglycemia (17 months from birth), suggesting that the impairment of the glymphatic system leads to A? accumulation. Collectively, our preliminary data, for the first time, demonstrate that non-invasive MRI methodologies can detect DM-induced early impairment of the glymphatic system which is highly correlated with hippocampal related dysfunction of learning and memory. Based on our novel preliminary data, we will employ MRI and 3D confocal microscopy to evaluate and quantitatively measure kinetic clearance parameters of the glymphatic system during progression of aging with and without DM (Aim 1). We will then investigate: whether impairment of the glymphatic system predicts cognitive dysfunction, the sensitivity and association between impairment of the glymphatic system, the onset of brain vascular dysfunction, and cognitive deficits during aging with and without DM (Aim 2). Data generated from this application will provide new insights into aging and age-matched DM associated impairment of the glymphatic system and the relationship of the glymphatic system with vascular and cognitive dysfunction.