Previous work from our laboratory showed that injection of 1,25-(OH)2 vitamin D3 (1,25-vitD) into the rat over a 3-day period increases force generating capacity of subsequently isolated resistance arteries. To better understand the mechanism of this vascular inotropic action, we examined the time course of its development and Ca2+-associated mechanism of action. Mesenteric resistance arteries were isolated from 12-week-old male spontaneously hypertensive rats (SHR) 6 and 24 h after a single injection and 24 h after three and seven consecutive days of injection of 1,25-vitD (20 ng/100 g) or vehicle. Serum 1,25-vitD was increased nearly threefold at 6 h, but not at later times. Serum total and ionized Ca2+ were not affected at any time point. No changes in contractile force generation were detected at 6 and 24 h, but a significant increase in the active stress response to norepinephrine and arginine vasopressin was observed after both 3 and 7 days of 1,25-vitD treatment. No effect on sensitivity to either agonist was observed at any time point. To determine whether the vascular inotropic effects of 1,25-vitD might translate into changes in blood pressure, 11-week-old male Wistar rats were given daily injections of 1,25-vitD (20 ng/100 g) over a 4-week period, and blood pressure and body weight were measured. While body weight did not differ at any time point in the two groups, systolic blood pressure was elevated in the 1,25-vitD group ν control by 7 days, and continued to be elevated over the period of observation. We conclude that the vascular inotropic action of 1,25-vitD presents with a time course that is consistent with a genomic mechanism rather than a rapid, nonnuclear action, and that the hormone increases blood pressure in the normotensive rat. Am J Hypertens 1993;6:388–396