Project Summary/Abstract In cerebrovascular disorders, such as Alzheimer’s Disease and stroke, the ability to maintain normal cerebral blood flow is compromised. Neurovascular coupling (NVC), the temporal relationship between neural activity and cerebral blood flow, is thought to be disrupted in these conditions resulting in cerebral hypoperfusion and cognitive dysfunction. As nitric oxide (NO) mediates vasodilation, the neurons that release (NO) are good candidates as a major regulator of cerebral blood flow (CBF). We recently developed a new genetic tool— NK1R-creER knockin mouse— that allows us to target and manipulate a specific subset of NO-generating neurons (nNOS Type 1 neurons), and thus test the involvement of these cells in neurovascular coupling for the first time. Here, we propose to test the specific hypothesis that nNOS Type 1 neurons receive excitatory pyramidal input and mediate vasodilation. Aim 1 will investigate whether NK1R-creER cortical interneurons receive excitatory pyramidal input using immunohistochemical approaches. Aim 2 will investigate whether pyramidal neurons form functional synapses onto NK1R-creER cortical interneurons using optogenetic approaches and slice electrophysiology. Lastly, Aim 3 will test whether NK1R-creER interneuron activity is necessary and sufficient to increase cerebral blood flow in vivo using laser Doppler flowmetry. Together, these experiments will investigate the circuits coupling neural activity and hemodynamics. This insight into NVC is fundamental to our understanding of the pathogenesis of common cerebrovascular diseases and the advancement of pharmacotherapeutics targeting cerebral perfusion.