Our objective was to investigate the effects of topically applied neuropeptide Y (NPY) on ischemic wounds. Initially, the animal model for ischemic wound healing was validated using 16 male Sprague Dawley albino rats. In the intervention study, an additional 28 rats were divided into three groups: NPY (0.025%), the positive control insulin-like growth factor-I (IGF-I, 0.0025%), and the hydrogel carrier alone (control). The hydrogel was selected due to its capacity to prolong NPY release (p < 0.001), as demonstrated in a Franz diffusion cell. In the animals, an 8 mm full-thickness wound was made in a pedunculated dorsal ischemic skin flap. Wounds were then treated and assessed for 14 days and collected at the end of the experiment for in situ hybridization analysis (RNAscope^®) targeting NPY receptor Y2R and for meticulous histologic examination. Wound healing rates, specifically the percentage changes in wound area, did not show an increase with NPY (p = 0.907), but there was an increase with rhIGF-I (p = 0.039) compared to the control. Y2R mRNA was not detected in the wounds or adjacent skin but was identified in the rat brain (used as a positive control). Light microscopic examination revealed trends of increased angiogenesis and enhanced inflammatory cell infiltration with NPY compared to control. An interesting secondary discovery was the presence of melanophages in the wounds. Our findings suggest the potential of NPY to enhance neovascularization under ischemic wound healing conditions, but further optimization of the carrier and dosage is necessary. The mechanism remains elusive but likely involves NPY receptor subtypes other than Y2R.