The aim of this study is to evaluate changes in vaginal pH and epithelium maturation after erbium-doped yttrium aluminum garnet (Er-YAG) laser treatment, and to assess its safety and efficacy on the symptoms of genitourinary syndrome of menopause (GSM). This was a retrospective study conducted between November 2019 and April 2022 and included 32 women diagnosed with GSM who had not benefitted from lubrication treatment and could not or would not use estrogen. Patients received three sessions of Er- YAG laser. All patient data before and after treatment were obtained from computer records. Vaginal maturation index (VMI), maturation value (MV) and vaginal pH values of the patients before and after laser treatment were compared. We also evaluated post-procedural complications and symptoms. Mean age was 59.72 ± 5.66 years. After laser therapy, there was a significant decrease in vaginal pH ( p  < 0.001) and the proportion of parabasal cells in VMI ( p  < 0.001), while there was a significant increase in MV ( p  < 0.001) and the proportion of superficial cells in VMI ( p  < 0.001). In 84.4% of the patients, GSM-related symptoms regressed completely or decreased to a tolerable level. Patients in which symptoms disappeared completely had significantly lower mean age (p = 0.002) and duration of menopause (p = 0.009). The laser procedure resulted in complications including mucosal injury in 5 (15.6%) patients (all recovered spontaneously) and vaginal burning sensation in 2 (6.3%) patients. Vaginal Er-YAG laser treatment may be a safe and effective alternative treatment method in a population of women with GSM who do not want to or cannot use estrogen therapy.