Aim: Calcium-activated chloride channels (CACCs) play an important role in smooth muscle contractility. Despite the wealth of data, the molecular architecture of CACCs in smooth muscle has yet to be resolved. Recently a new gene family (TMEM16) has been identified, which when expressed produces currents displaying biophysical properties similar to native calcium-activated chloride currents (I_ClCa). There is no information on whether members of the TMEM16 family are expressed in smooth muscle. The aim of this study was to determine if TMEM16A & B was expressed in vascular smooth muscle so it could be considered a molecular correlate for CACCs. Methods: Whole-cell patch clamp electrophysiological recordings, semi-quantitative and quantitative reverse transcriptase polymerase chain reaction experiments and immunological experiments were performed on murine (BALB/c; 6 – 8 weeks) smooth muscle tissues. Results: Cl- currents with distinctive voltage-dependent characteristics were recorded from myocytes isolated from murine portal vein (mPV), thoracic aorta (mTA) and carotid artery (mCA) (n>=5). TMEM16A was expressed in all three blood vessels with expression higher in the portal vein (mPV>mTA>mCA; n=3). All vessels expressed more TMEM16A mRNA than murine brain. In contrast, expression of TMEM16B was much less than TMEM16A in mPV, mTA and mCA; n=3. TMEM16A protein expression was detected by various TMEM16A antibodies and was shown to localise to the smooth muscle layer in mPV, mCA and mTA by immunohistochemistry (n>=3) and immunocytochemistry (n>=3). In mPV discontinuous sucrose fractionation studies, TMEM16A expression was shown to be confined to lipid rafts (n=3). Conclusion: This study is the first to demonstrate the presence of TMEM16A mRNA and protein in a variety of vascular smooth muscle tissues, and show that TMEM16A exists predominantly in discrete micro-environments. Consequently, TMEM16A can now be considered a strong candidate for CACCs in smooth muscle vasculature.