Sleep is a critical factor that affects overall health, and humans typically spend one-third of their lives sleeping. Sleep consists of two states: rapid eye movement (REM) sleep and non-REM (NREM) sleep. The brainstem pontine tegmental area contains neurons that either positively or negatively regulate REM sleep (Vanni-Mercier et al., 1989; Lu et al., 2006; Hayashi et al., 2015; Kashiwagi et al., 2020). Additionally, some neurons in this area regulate urination (Hou et al., 2016; Keller et al., 2018) and fear (Yang et al., 2016). Thus, the pontine tegmental area consists of multiple subpopulations of neurons with diverse roles, and the current characterization of individual subpopulations is likely insufficient.By conducting a screen for neuronal subpopulations in the mouse pontine tegmental area based on expression of marker genes, we identified and focused on a novel subpopulation of neurons. Within the pontine tegmental area, these neurons are distributed in the medial area. Single nucleus RNA-sequencing suggested that the novel subpopulation mainly consists of GABAergic neurons. We hypothesized that these neurons might play a role in sleep regulation. To examine this, we performed chemogenetic activation of these neurons. Results revealed that chemogenetic activation led to an increase in the amount of NREM sleep and a decrease in the amount of wake, without affecting the amount of REM sleep. This study suggests the presence of novel NREM sleep-promoting neurons. This finding will lead to a deeper understanding of the role and heterogeneity of neuronal populations in the pontine tegmental area in sleep regulation.