The phasic contraction to phenylephrine of the rat isolated portal vein was investigated using functional studies. Phasic contractions to phenylephrine and caffeine could be produced after several minutes in Ca²⁺-free Krebs solution, which were inhibited by cyclopiazonic acid or ryanodine. The phenylephrine and caffeine contractions were abolished, however, within 10 min in Ca²⁺-free Krebs solution and by nifedipine. This indicated the Ca²⁺ stores were depleted in the absence of Ca²⁺ influx through voltage-gated channels. The phasic contraction to phenylephrine was also abolished by niflumic acid even in Ca²⁺-free Krebs solution. This showed that the response depended on intracellular Ca²⁺ release stimulated directly by depolarization, resulting from opening of Ca²⁺-activated Cl channels, but did not require Ca²⁺ influx. In support of this, K⁺-induced phasic contractions were also produced in Ca²⁺-free Krebs solution. The phenylephrine but not K⁺-induced phasic contractions in Ca²⁺-free Krebs solution were inhibited by ryanodine or cyclopiazonic acid. This would be consistent with Ca²⁺ release from more superficial intracellular stores (affected most by these agents), probably by inositol 1,4,5-trisphospate, being required to stimulate the phenylephrine depolarization.