Purpose: The neck extensor muscles contribute to spinal support and posture while performing head and neck motion. Muscle stiffness relates to passive elasticity (support) and active tensioning (posture and movement) of muscle. It was hypothesized that support and motion requirements are reflected in the distribution of stiffness between superficial and deep neck extensor muscles. Methods: In ten healthy participants, shear modulus (stiffness) of five neck extensor muscles was determined in prone at rest and during isometric head lift at three intensities using shear wave elastography. Results: Shear modulus differed between muscles (P < 0.001), and was larger for the deeper muscles: (median (interquartile range)) trapezius 7.7 kPa (4.4), splenius capitis 6.5 kPa (2.5), semispinalis capitis 8.9 kPa (2.8), semispinalis cervicis 9.5 kPa (2.5), multifidus 14.9 kPa (1.4). Shear modulus differed between the resting condition and head lift (P < 0.001) but not between levels of head lift intensity. Conclusion: Shear wave elastography revealed highest passive and active stiffness of the deep neck extensor muscles most close to the spine. The highest active increase of stiffness during the head lift was found in the semispinalis cervicis muscle. The non-invasive, clinically applicable estimates of muscle stiffness have potential for the assessment of muscular changes associated with neck pain/injury.