The aim of this study was to establish the involvement of acetylcholine (Ach) and calcium ions in modulating contractions induced by Bay K8644 (an agonist of calcium channels located in the cell membrane) and KCl (at depolarizing concentrations), and also to examine the importance of the vascular endothelium in the activity of Bay K8644. The study was performed on perfused Wistar rat tail arteries. Contraction induced by Bay K8644 with the participation of intracellular (in calcium‑free physiological salt solution, FPSS) and extracellular (in physiological salt solution, PSS, following the emptying of the cellular Ca2+ stores) pools of Ca2+ and the addition of nitro-L-arginine (L‑NNA; nitric oxide synthase inhibitor) or 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one (ODQ; an inhibitor of soluble guanylyl cyclase) was studied. In addition, the effect of Ach on the contraction response was analyzed and the results were compared with the depolarizing action of KCl. The effects of 8Br‑cGMP on the artery contraction induced by Bay K8644 prior to and following removal of the endothelium were compared. Bay K8644 and KCl in PSS induced vascular contraction, which was reduced with the addition of Ach. The spasmolytic Ach action did not occur in the presence of L‑NNA and ODQ. 8Br‑cGMP reduced the contraction of arterial walls (with and without endothelium) induced by Bay K8644. The increase in vascular tone induced by Bay K8644 and KCl was independent of the intracellular calcium ion pool. The relaxant effect of Ach on the responses stimulated by Bay K8644 and KCl indicated the participation of nitric oxide in modulating the reactivity of the arteries to the factors examined, resulting in an influx of Ca2+ into the cell.