Abstract: The formation of macrospores and microspores, along with the development of female and male gametophytes in Paeonia lactiflora Pall., was systematically investigated to elucidate the anatomical basis of its low seed-setting rate. A combination of paraffin sectioning, scanning electron microscopy, I₂-KI staining, and in vitro pollen culture was employed to assess pollen morphology and germination characteristics. Results showed that the anther wall followed a dicotyledonous developmental pattern, with tetrads exhibiting both tetrahedral and decussate configurations. Mature pollen grains were bicellular and tricolporate. However, premature tapetum degradation during the uninucleate stage led to widespread microspore abortion, leaving only a minority to develop normally. Viable pollen grains appeared spherical and turgid, while non-viable pollen grains often appeared collapsed and empty. Ovules were anatropous with a biregonic, crassinucellate structure and contained multiple megaspore mother cells and a distinct nucellar cap. Tetrads were linearly arranged, and embryo sac development followed a polygonum-type pathway, with most mature ovules bearing one or two embryo sacs. The findings indicate that defective microspore development is a critical factor leading to pollen abortion, which significantly reduces the likelihood of successful pollination and ultimately affects the seed-setting rate.