Vascular smooth muscle cells (VSMC) are able to switch from a contractile to a proliferative phenotype, in response to environmental factors. This phenotypic switch (PM) involves changes in the expression of contractile proteins and also membrane receptors and ion channels. Our previous work provides evidence that PM of VSMCs from different vascular beds and different species associated with stereotyped changes in voltage-dependent potassium channels (Kv channels). Specifically, we observed an increased functional expression of Kv1.3 channels and a complete disappearance of Kv1.5 channels, so that the changes in the Kv1.3 to Kv1.5 ratio can define the phenotype. These results were also validated in heterologous expression system. Kv1.5 overexpression decreased proliferation of transfected HEK293 cells, while Kv1.3 expression leads to an increase in cell proliferation that was sensitive to selective Kv1.3 blockers and independent of potassium flux 1. Our data suggest that the movement of the voltage-sensor of Kv1.3 channels could be coupled to signalling pathways leading to proliferation. In the present work, we propose 1) to confirm the anti-proliferative effect of Kv1.5 channels in human VSMCs cultures and 2) to identify the Kv1.3 domains involved in the signalling mechanism(s) associated with proliferation. For the first objective, lentiviral vectors carrying a Kv1.5-GFP fusion protein have been expressed in both HEK cells and human VSMCs in primary culture, and we tested the rate of proliferation of Kv1.5 expressing cells as compared with GFP- transduced and mock transfected cells. We obtained around 90-100% infectivity both in HEK and in VSMCs, and our preliminary results confirm the anti-proliferative effect of Kv1.5 overexpression. For the second objective, taking advantage of the opposite roles of Kv1.3 and Kv1.5 on cell proliferation, we designed chimeric Kv1.3-Kv1.5 channels by substituting either NH2- or COOH- terminal domain on Kv1.5 backbone with the corresponding domains of Kv1.3, creating the K5-N3 and the K5-C3 constructs. We expressed these chimeras as GFP- or cherry-fusion proteins in HEK cells. Immunocytochemical analysis was used to confirm their expression and subcellular distribution and electrophysiological studies were used to determine their ability to form functional channels and their pharmacological profile. Finally, we explored the effects on the rate of proliferation of HEK cells of the expression of these chimeric channels compared to wild type Kv1.3 and Kv1.5 channels and GFP-transfected cells. Altogether, our data validate Kv1.3/Kv1.5 ratio as a good indicator of the PM and can help to define the molecular interactions of the channels with intracellular proteins related to proliferation