We used the fluorescent Ca2+ indicator Fura-2 in cultured porcine aortic smooth muscle cells (PASMC) to study effects of the sympathetic neurotransmitters norepinephrine (NE) and neuropeptide Y (NPY) on free intracellular Ca2+ (Cai). Both transmitters transiently increased intracellular Ca2+ in a concentration-dependent manner. Selective agonists and antagonists demonstrated that the NE-stimulated Cai increase is predominantly (if not exclusively) mediated by alpha 2-adrenoceptors, whereas the NPY response appears to be mediated by the peptide YY-insensitive Y3-like receptor subtype. Pretreatment of cells with pertussis toxin abolished NPY and alpha-adrenoceptor agonist-stimulated intracellular Ca2+ elevations (but not those stimulated by angiotensin II) suggesting involvement of a Gi-like G-protein. alpha 2-Adrenoceptor-stimulated Ca2+ increases resulted from mobilization from intracellular stores, whereas Y3-like NPY receptors mobilized Ca2+ from intracellular stores and also promoted Ca2+ influx.