Background and Purpose: Yohimbine (YO) is used for lipolysis in mesotherapy since it stimulates the sympathetic nerve system by antagonizing α2-adrenergic receptors, although there is limited evidence on its mechanisms of action and safety. Thus, this study aimed to investigate the acute metabolic effects of a single YO dose on adipose tissue in mice. Experimental Approach: Female Swiss mice received a single subcutaneous injection of 100 µL saline (control) or 100 µL YO (80 µg or 800 µg) in the right inguinal white adipose tissue (iWAT) and were euthanized 2 or 24 h post-injection. We assessed body mass (BM), blood glucose, serum total cholesterol (TC), triglycerides (TG), and glycerol. Left and right iWAT were collected, weighed, and investigated by histopathology (adipocyte phenotype and cell size), and immunohistochemistry (uncoupling protein-1 [UCP-1] and hormone-sensitive lipase [HSL]). Key Results: YO 800 µg reduced BM after 24h and reduced glycaemia 2 and 24 h after injection. TG and TC were also lower, but glycerol was higher than the control group. On the site of YO injection (right iWAT), it reduced white adipocyte diameter time- and dose-dependently. Beige cells positive to UCP-1 were seen in iWAT, but surprisingly the HSL stain was unchanged. Conclusion and Implications: Acutely, YO mesotherapy in mice promoted significant local and systemic effects. It modulated BM and glycaemia, promoted lipid mobilization, and induced iWAT browning. However, further studies are needed to determine its safety dosage in mesotherapy and the metabolic effects when used chronically.