Traits influencing plant water use eventually define the fitness of genotypes for specific rainfall environments. We assessed the response of several water use traits to vapour pressure deficit (VPD) in pearl millet (Pennisetum glaucum (L.) R.Br.) genotypes known to differ in drought adaptation mechanisms: PRLT 2/89-33 (terminal drought-adapted parent), H 77/833-2 (terminal drought-sensitive parent) and four near-isogenic lines introgressed with a terminal drought tolerance quantitative trait locus (QTL) from PRLT 2/89-33 (ICMR01029, ICMR01031, ICMR02042, and ICMR02044). Plant water use traits at various levels of plant organisation were evaluated in seven experiments in plants exposed either transiently or over the long term to different VPD regimes: biomass components, transpiration (water usage per time unit) and transpiration rate (TR) upon transient VPD increase (gH2Ocm-2h-1)), transpiration efficiency (g dry biomass per kg H2O transpired), leaf expansion rate (cm per thermal time unit) and root anatomy (endodermis dimensions)). High VPD decreased biomass accumulation by reducing tillering, the leaf expansion rate and the duration of leaf expansion; decreased root endodermis cell size; and increased TR and the rate of TR increase upon gradual short-term VPD increases. Such changes may allow plants to increase their water transport capacity in a high VPD environment and are genotype-specific. Some variation in water use components was associated with terminal drought adaptation QTL. Knowledge of water use traits' plasticity in growth environments that varied in evaporative demand, and on their genetic determinacy, is necessary to develop trait-based breeding approaches to complex constraints.