The low-fluence 1064-nm Q-switched neodymium:yttrium-aluminium-garnet (QSNY) laser is a widely used treatment for melasma in East Asia, although its mechanism of action is unclear. The aim of this study was to elucidate the mechanism of action of the QSNY laser. We performed a histopathological study on eight Korean women who had considerable improvement in their melasma lesions after a series of low-fluence QSNY laser treatments. Compared with nonlesional skin, samples from melasma lesions showed increased reactivity in melanin (Fontana-Masson staining) and in melanogenesis-associated proteins, including α-melanocyte-stimulating hormone, tyrosinase, tyrosinase-related protein (TRP)-1, TRP-2, nerve growth factor and stem cell factor. After laser treatment, the melasma skin showed a decrease in the number of melanosomes and reduced expression of melanogenesis-associated proteins. Expression levels of the melanogenic proteins were reduced after laser treatment, although the number of melanocytes was unchanged even in hypopigmented areas. Based on these results, we believe that repeated application of low thermal energy via QSNY laser may result in damage to melanocytes and long-lasting hypopigmentation.