The muscular anatomy of the tongue consists of a complex three-dimensional array of fibers, which together produce the variations of shape and position necessary for deglutition. To define the myoarchitecture of the intact mammalian tongue, we have utilized NMR techniques to assess the location and orientation of muscle fiber bundles through measurement of the direction-specific diffusional properties of water molecules. Whole sheep tongues were excised and imaged with a slice-selective stimulated-echo diffusion sequence in the midline sagittal plane, and three-dimensional diffusion tensors were determined for each voxel. The derived diffusion tensors were depicted graphically as octahedra whose long axes indicate local muscle fiber orientation. Two distinct groups of midline fibers were identified: 1) in-plane sagittal fibers originating in the posteroinferior region of the tongue, radiating with a fanlike projection anteriorly and superiorly and merging with vertically oriented fibers, and 2) cross-plane (transverse) fibers, oriented at right angles to the vertically aligned fibers, predominantly in the anterior and superior regions of the tongue. Regional comparison of diffusion anisotropy revealed uniform and parallel alignment (high anisotropy) in the posteroinferior region of the tongue, corresponding to the base of the genioglossus, and less uniform, orthogonally aligned fibers (low anisotropy) in the anterosuperior region of the tongue, corresponding to the core intrinsic muscles. These data indicate that lingual myoarchitecture, determined through direction-dependent mobility of water molecules, can be depicted as discrete regions of muscle fibers, whose orientation and extent of diffusion anisotropy predict local contractility.