The goal of the proposed studies is to understand the mechanisms linking whole animal Ca2+ homeostasis and vascular function. Four specific aims will test the hypothesis that elevation of extracellular Ca2+, as occurs in the interstitial compartment of tissues involved in transcellular Ca2+ movement, activates a perivascular nerve Ca2+ sensing receptor (CaSR) that is coupled with the production and release of a endocannabinoid vasodilator that induces local vasorelaxation. Preliminary data that support this hypothesis include our demonstration that the concentration of interstitial Ca2+ in the duodenal submucosa and renal cortex undergoes dynamic changes over a physiologic range; our finding that dorsal root ganglia (DRG) and perivascular sensory nerves express a functional CaSR; our demonstration that Ca2+ induces the release of an endocannabinoid dilator from the vessel wall with activity at the putative anandamide receptor. Additional work has revealed that nitric oxide (NO) negatively modulates Ca2+-induced relaxation and that N18TG2 neuroblastoma cells express a CaSR. Specific aim 1 will use the parathyroid deficient Gcm2 -/- mouse to rescue the CaSR knockout phenotype which will be used to test the hypothesis that a neuronal CaSR mediates Ca2+-induced relaxation. Specific aim 2 will use isolated arteries and cultured DRG neurons coupled with GC-mass spec analysis of endocannabinoids to understand the role of these compounds in mediating Ca2+-induced relaxation. Specific aim 3 will use pharmacologic and molecular genetic approaches to determine the mechanism by which vessel wall NO modulates Ca2+-induced relaxation. Specific aim 4 will use molecular, physiological and pharmacological approaches to pursue our finding that N18TG2 cells expresses a CaSR and to test the hypothesis that this cell line can serve as a model to study neuronal CaSR signaling and transmitter production. We anticipate that these studies will lead to a more complete understanding of the molecular mechanisms that link Ca2+ homeostasis with vascular function; provide detailed mechanistic information about sensory nerve mediated relaxation and the vascular endocannabinoid system, increase our understanding of CaSR sensing function in neuronal tissue, and may reveal new targets for the development of novel vasodilator compounds.