Acetylcholine produces venoconstriction of isolated vein strip preparations. However, the effect of acetylcholine on overall vascular capacity is not known. To investigate this effect and to elucidate the mechanisms involved, 38 anesthetized dogs were placed on total cardiopulmonary bypass, splenectomized, and given intraarterial infusions of acetylcholine. Almost all of the effect on vascular volume was found to be in the splanchnic circulation, because in four eviscerated animals there was no significant change in capacity. In the animals in which the mesenteric arteries were cannulated to provide constant inflow, and the hepatic vein was cannulated to measure splanchnic venus outflow, acetylcholine infusion for 5 and 21 min increased splanchnic vascular capacity in all animals by 107 +/- 28 (SEM)ml (p less than 0.01) and 291 +/- 132 ml (p less than 0.05), respectively. This increase in splanchnic vascular volume was associated with a rapid and sustained increase in transhepatic resistance to portal blood flow for the duration of the infusions (p less than 0.01). In the animals in which the portal vein was vented proximal to the liver, no significant volume change occurred in the splanchnic vasculature with acetylcholine infusion. Increasing hepatic venous pressure to elevate portal venous pressure to the same level as that achieved with acetylcholine resulted in a similar increase in splanchnic vascular volume. Atropine, but not adrenergic blockade, blocked the acetylcholine-induced volume retention, indicating that the effect of acetylcholine was direct. Substantial volume retention was also achieved by stimulation of the distal ends of the sectioned cervical vagi. Thus, acetylcholine administration directly increases transhepatic resistance and is associated with a pooling of volume in the splanchnic vasculature that would, in the intact animal, result in a decrease in venous return to the heart.