The existence of a local renin-angiotensin system (RAS) in neurons was first postulated forty years ago. Further studies indicated intraneuronal generation of angiotensin II (ANG II). However, the function and signaling mechanisms of intraneuronal ANG II remained elusive. Since angiotensin II type 1 receptor, AT₁, is the major type of receptor mediating the effects of ANG II, we used intracellular microinjection and concurrent calcium and voltage imaging to examine the functionality of intracellular AT₁ receptor in neurons. We show here that intracellular administration of ANG II produces a dose-dependent elevation in cytosolic Ca²⁺ concentration, [Ca²⁺]_i, in hypothalamic neurons, that is sensitive to AT₁ receptor antagonism. Endo-lysosomal, but not Golgi apparatus disruption, prevents the effect of microinjected ANG II on [Ca²⁺]_i. Additionally, the ANG II-induced Ca²⁺ response is dependent on microautophagy and sensitive to inhibition of phospholipase C or antagonism of inositol 1,4,5-trisphosphate receptors. Furthermore, intracellular application of ANG II produces AT₁-mediated depolarization of hypothalamic neurons, which was dependent on [Ca²⁺]_i increase and on cation influx via transient receptor potential canonical channels. In summary, in the present study we provide evidence that intracellular ANG II activates endo-lysosomal AT₁ receptors in hypothalamic neurons. Our results point to the functionality of a novel intraneuronal angiotensinergic pathway extending the current understanding of intracrine angiotensin II signaling.