Many host species have evolved sophisticated defences to mitigate the high fitness costs imposed by brood parasitism. Even though the physiological mechanisms behind such defences can offer important insights into the evolutionary relationship between brood parasites and hosts, they have received little attention so far. Hormones play a critical role in the regulation of bird reproduction, which make them a key element when investigating the physiological effects of brood parasitism on hosts. Here, we experimentally parasitized Eurasian blackbird (Turdus merula) nests with non-mimetic eggs to study its impact on the hormonal levels (corticosterone and prolactin) of females during incubation, as well as the magnitude of the response to the standardized stress protocol in parasitized and non-parasitized individuals. Parasitized females had higher baseline corticosterone levels and showed a poorer body condition than non-parasitized birds, while we found no differences for prolactin levels. Both parasitized and non-parasitized females responded to the standardized-stress protocol with a significant increase in corticosterone levels. However, the decrease in prolactin after the standardized stress protocol was significantly more pronounced in parasitized individuals. Our results suggest that the presence of a non-mimetic parasitic egg involves a stressful situation for hosts, negatively affecting the physical state of parasitized females. Unaffected prolactin levels of parasitized individuals could explain the absence of nest desertion found in this species in response to parasitism. Finally, both hormones were not correlated in blackbirds, confirming that their combined study provides valuable pieces of information on the endocrine mechanisms underlying behavioural responses in animals, including hosts of brood parasites. Physiological mechanisms behind avian brood parasitism remain unclear. In this study, we assessed the effect of experimental parasitism on the hormonal profiles of hosts. We found that the presence of a non-mimetic egg in the nest modified baseline corticosterone levels, but not prolactin levels, of parasitized females and negatively impacted their body condition. Moreover, experimental parasitism affected the prolactin response to stress. These results expand previous information on the endocrine consequences of brood parasitism at other stages of the breeding cycle (nestling and fledgling stage) and might shed light on the hormonal mechanisms that underlie the host response against parasitic eggs.