Laying hens are thought to express an expectation of a rewarding event through a specific vocalisation, the gakel-call. It has been suggested that the gakel-call is related to frustration, i.e. the thwarting of behaviour. We investigated if frustrative nonreward (nonreinforcement in a situation that previously was consistently reinforcing) in laying hens is expressed through this gakel-call. Twenty hens of two commercial strains, ten ISA White Leghorn and ten ISA Brown Warren were subjected to a classical conditioning procedure. After 23 h of food-deprivation they were trained, in automated Skinnerboxes, to use red lights as a signal (unconditioned stimulus) for a food reward. After this the hens of each strain were equally divided into two groups of five. They were subjected to a control session (light as conditioned stimulus followed by food reward) and to a frustration session (conditioned stimulus followed by nonreward). Both during the frustration and control session behaviour and vocalisations were recorded. In the White Leghorn strain high levels of alarm-calls are found as indicators of anxiety. In the Brown Warren strain more gakel-calls and an increased locomotor activity are found after frustrative nonreward. In conclusion, both findings suggest that vocalisations could serve as indicators of a laying hen's welfare.