Threatened species frequently have a J-shaped population structure, which indicates a reduction in seed set and poor or nonexistent recruitment. Altered population structures may be due to disrupted demographic processes that result in low reproductive success or small population size. Peyote—Lophophora diffusa Croizat (Bravo)—is a rare, threatened cactus species that is subject to overexploitation because of psychedelic tourism and medicinal and religious uses that decrease its effective population size. We analyzed peyote population structure and identified attributes of its reproductive biology that may limit population persistence. The population's size structure (based on plant size in square centimeters) was determined by census in 2014 and 2015 (n = 420 individuals). Determination of the breeding system of peyote was based on floral morphology and evaluation of herkogamy, dichogamy, and pollen/ovule ratio, and controlled pollination experiments were used to determine the outcrossing rate and whether the species was pollen limited. Additionally, behavior and frequency of floral visitors were recorded to establish the pollinator guild. Peyote's population structure showed the presence of seedlings, juveniles, and adults in both years, indicating some recruitment and low adult mortality. Flowers were herkogamous, homogamous, and diurnal, with a 2-day longevity and high pollen/ovule ratios. These floral traits suggest that peyote is xenogamous, and pollination experiments indicated that it is a facultative outcrossing species, which needs pollinators to set fruit. The time of pistil receptivity coincided with the maximum activity of floral visitors (small solitary bees and small beetles) searching for floral rewards. Evidence indicates that peyote has an outcrossing system with partial self-incompatibility and is pollen limited. Positive factors, such as moderate seedling recruitment, a similar population structure during the two study periods, and a large number of reproductive individuals (close to 50%), indicate natural regeneration and increase the likelihood of population persistence. However, low fruit set and strong dependence upon efficient pollinators reduce reproductive success and increase species vulnerability.