Age-related loss of muscle mass may result from reduced protein synthesis stimulation in response to anabolic stimuli, such as amino acid (AA) supplementation. The exact etiology of anabolic resistance to AA remains unclear. Therefore, the aim of this study was to investigate the anabolic response [cell size, protein synthesis and mechanistic target of rapamycin (mTOR) pathway] to the AA glutamine (a strong anabolic AA highly present in skeletal muscle) in myotubes obtained from 8 young (YW; 21-35 yrs) and 8 older (OW; 65-70 yrs) healthy women. This in vitro model of human primary myogenic cells explores the intrinsic behavior of muscle cells, while excluding potential influences of external factors. We showed that despite lower muscle mass, strength and cardiorespiratory fitness in OW compared to YW, myotube size (myotube diameter and area) and protein synthesis were not altered in OW, and glutamine-induced myotube hypertrophy and protein synthesis were preserved in OW. Apart from a lower glutamine-induced increase in P70S6 kinase phosphorylation in OW, no significant differences in other components of the mTOR pathway were observed between groups. Altogether, our data support the idea that the intrinsic capacity of muscle cells to respond to glutamine stimulation is preserved in healthy older women.