Background: Stress urinary incontinence (SUI) is a common disorder with high prevalence in women across their life span, but there are no non-surgical curative options for the condition. Stem cell-based therapy, especially endogenous stem cell therapy may be a potential treatment method for SUI. The aims of this study are to identify, isolate, and assay the function of urethral striated muscle derived stem/progenitor cells (uMDSCs) and to assess uMDSC response to microenergy acoustic pulses (MAP). Methods: Urethral striated muscle was identified utilizing 3D imaging of solvent organs (3DISCO) and immunofluorescence (IF). uMDSCs were isolated and purified from Zucker Lean (ZL) (ZUC-LEAN) (ZUC-Leprfa 186) rats, with magnetic-activated cell sorting (MACS) and pre-plating methods. The stemness and differentiation potential of the uMDSCs were measured by cell proliferation, EdU, flow cytometry, IF, and Western blot. Results: Comparison of the cell proliferation assays between MACS and pre-plating reveals the advantage of MACS over pre-plating. In addition, the study reveals that uMDSCs form myotubes when treated with MAP. Conclusions: The uMDSCs within female rat urethral striated muscle could be a therapeutic target of MAP in managing SUI.