A prospective clinical study was carried out to evaluate the influence of posture on perineal ultrasound imaging parameters. One hundred and thirty-two consecutive women presenting with symptoms of lower urinary tract dysfunction were examined by multichannel videourodynamics and perineal ultrasound, both supine and standing. Ultrasound included color Doppler imaging when available, i.e. in a subgroup of 99 patients. The position of the bladder neck at rest was higher in the supine position (P<0.001) and it descended further on Valsalva (P < 0.001), to reach an almost identical final position. There was a higher degree of urethral rotation on supine imaging (P = 0.001). Urethral funneling on Valsalva was more likely in the erect position (P < 0.001), as was urine leakage documented by color Doppler imaging (P < 0.001). The effect of a pelvic floor contraction was not significantly influenced by posture. Imaging of the urethrovesical junction should be undertaken in both supine and erect positions to document optimally both hypermobility and funneling or leakage.