Systemic osmoregulation is a homeostatic process whereby endocrine (e.g. vasopressin release) and behavioral responses (e.g. thirst) maintain a constant proportion of solutes to water (osmolality) in the extracellular fluid. This process is controlled by central osmosensory neurons (ONs), notably those in the organum vasculosum laminae terminalis (OVLT) and the supraoptic nucleus (SON). ONs display a cell-autonomous response to hypertonicity whereby shrinking induces an increase in non-selective cation conductance which causes membrane depolarization and contributes to the excitation of ONs that mediates thirst and vasopressin secretion. The molecular identity of the osmoreceptor channel is not known, but involves a capsaicin-insensitive variant of the transient receptor potential vanilloid type 1 (Trpv1) channel that lacks amino acids located in the protein’s amino terminus (DN-Trpv1; Ciura and Bourque, 2006; Sharif-Naeini et al, 2006). We isolated high quality mRNA from OVLTs and SONs of Long Evans rats and reversed transcribed this material into cDNA using oligo-DT primers. Probing this material by Polymerase chain reaction (PCR) with specific primers we identified a 2 kb amplicon whose size was consistent with that expected of a defined N-terminal variant. Sequencing analysis identified the complete nucleotide sequence of the variant, providing important information concerning its secondary structure. Moreover, specific primers verified the expression of this specific variant by RT-PCR in single OVLT and SON neurons. Heterologous expression of DN-Trpv1 by transfection into human embryonic kidney (HEK293) cells, conferred osmosensory characteristics similar to those of SON and OVLT neurons. These results suggest that DN-Trpv1 operates as an osmoreceptor in SON and OVLT neurons.