Background: The quadratus lumborum block (QLB) has recently been used frequently for postoperative analgesia after abdominal surgery. The aim of this study was to investigate the effect of pressure changes between the middle thoracolumbar fascia layers to which the anterior QLB (aQLB) is applied at the level of the sensory block. Methods: A total of 67 patients planned to undergo laparoscopic cholecystectomy were evaluated in the context of this prospective study. Bilateral aQLBs were administered to all patients, and a peripheral nerve block catheter was placed between the psoas and quadratus lumborum muscles. The correlation of interfascial pressures (IFPs) during block application, 30 min intraoperatively, and 30 min postoperatively with the sensory block level was determined as the primary outcome measure. Secondary outcome measures were the sensory block levels 30 min after block application and 30 min postoperatively; visual analog scale scores 30 min and 6, 12, and 24 h postoperatively; and 24 h tramadol consumption. Results: The preoperatively and intraoperatively measured IFPs differed significantly between the right and left sides (P < 0.05). The preoperative and postoperative block levels also differed significantly between the right and left sides (P < 0.05). The preoperative and postoperative block levels and preoperatively measured IFPs showed a weakly negative correlation (right: r = 0.374; left: r = 0.470). Conclusions The results of this study show that pressure changes between the fasciae may be effective in aQLB application.