Photostimulation initiates and maintains the rise in circulating prolactin (PRL) observed during the reproductive cycle of the female turkey. Vasoactive intestinal peptide (VIP) is the principal PRL-releasing factor. This study tested the hypothesis that gonadal stimulatory photoperiods stimulate PRL secretion by releasing hypothalamic VIP. Therefore, an experiment was designed to determine if VIP immunoneutralization altered photo-induced PRL secretion. Reproductively quiescent female turkeys were divided into two groups comprising turkeys actively immunized with synthetic VIP conjugated to keyhole limpet hemocyanin (VIP-KLH; immunized; n = 48) or KLH alone (control; n = 48). The first immunization was administered 6 weeks before photostimulation. Blood samples were collected at frequent intervals prior to and following photostimulation, and plasma PRL concentrations were determined. Vasoactive intestinal peptide antibody titer was estimated from the percentage of 125I labeled VIP bound to plasma diluted 1:1000. At the onset of photostimulation (Day 0), plasma PRL levels were similar for immunized and control turkeys (9.1 ± 0.3 versus 8.9 ± 0.3 ng/ml, respectively). Plasma PRL of control birds increased (P < 0.05) by Day 16 of photostimulation, reaching a peak value of 724.9 ± 90.1 ng/ml on Day 84. In contrast, plasma PRL remained essentially unchanged in immunized birds. Titer of anti-VIP antibodies expressed as 125I-VIP bound by plasma in immunized birds was 10.9 ± 1.5% on the day of photostimulation. Incubation behavior was blocked in immunized birds, whereas 75% of controls exhibited incubation behavior. The control group laid 1.83 eggs/week/hen compared to 3.40 eggs/week/hen in immunized hens. These findings suggest that photoperiodic modulation of PRL secretion in the turkey is influenced by hypothalamic VIP neuronal system.