It is currently unknown in which ways activity of the ab- and adductor shoulder muscles affects shoulder biomechanics (scapular kinematics and glenohumeral translation), and whether these changes are relevant for alterations of the subacromial space width. The objective of this experimental in vivo study was thus to test the hypotheses that potential changes of the subacromial space width (during antagonistic muscle activity) are caused by alterations of scapular kinematics and/or glenohumeral translation. The shoulders of 12 healthy subjects were investigated with an open MRI-system at 30 degrees, 60 degrees, 90 degrees, 120 degrees and 150 degrees of arm elevation. A force of 15N was applied to the distal humerus, once causing isometric contraction of the abductors and once contraction of the adductors. The scapulo-humeral rhythm, scapular tilting and glenohumeral translation were calculated from the MR image data for both abducting and adducting muscle activity. Adducting muscle activity led to significant increase of the subacromial space width in all arm positions. The scapulo-humeral rhythm (2.2-2.5) and scapular tilting (2-4 degrees) remained relatively constant during elevation, no significant difference was found between abducting and adducting muscle activity. The position of the humerus relative to the glenoid was, however, significantly (p < 0.05) different (inferior and anterior) for adducting versus abducting muscle activity in midrange elevation (60-120 degrees). These data show that the subacromial space can be effectively widened by adducting muscle activity, by affecting the position of the humerus relative to the glenoid. This effect may be employed for conservative treatment of the impingement syndrome.