Histotripsy is an extracorporeal ablative technology that utilizes microsecond pulses of intense ultrasound to produce nonthermal, mechanical fractionation of targeted tissue. We have previously demonstrated the feasibility of histotripsy ablation in in vivo renal and prostate models. We sought to assess the chronic tissue response, tolerability, and safety of prostate histotripsy in a chronic in vivo canine model. Five acute and thirteen chronic canine subjects were anesthetized and treated with histotripsy targeting the prostate. Pulses consisting of three cycle bursts of 750‐kHz ultrasound at a repetition rate of 300 Hz were delivered from a highly focused 15‐cm aperture array. Prostates were harvested at 0, 7, 28, or 56 days after treatment. Transrectal ultrasound imaging provided accurate targeting and real‐time monitoring of histotripsy treatment. Consistent mechanical tissue fractionation and debulking of prostate tissue was seen acutely and at delayed time points without collateral injury. Canine subjects tolerated histotripsy with minimal hematuria or discomfort. Only mild transient lab abnormalities were noted. Histotripsy is a promising noninvasive therapy for prostate tissue fractionation and debulking that appears safe and well tolerated without systemic side effects in the canine model.