The gut-brain axis is known to modulate behavioral and immune responses in animals; evidence supporting this modulation in chickens, however, is elusive. Here, we analyzed the effects of heat stress and/or Clostridium perfringens (CP) infection on behavior, intestinal morphology, brain activity, and corticosterone serum levels in chickens. Broilers were randomly divided into 5 equal groups: a naïve group (N), a thioglycolate group (T), a thioglycolate heat-stressed group (T/HS35), an infected group (I), and an infected/stressed (I/HS35) group. Broilers in the I and I/HS35 groups were experimentally infected with Clostridium perfringens from the 15th to the 19th day of life. Heat stress (35 ± 1°C) was constantly applied to the broilers in the stressed groups from the 14th to the 19th day of life. Our data showed that heat stress and C. perfringens infection produced significant differential responses in the chickens' behavior and in c-fos expression in the paraventricular nucleus of the hypothalamus (PVN), nucleus taenia of the amygdala (Tn), medial preoptic area (POM), and globus pallidus (GP) of the chickens. Heat stress ameliorated some of the intestinal lesions and the neuroendocrine changes induced by C. perfringens in the birds. Our results suggest the existence of clear relationships between the degree of intestinal lesions, the chickens' behavioral outcomes, brain activity, and serum levels of corticosterone. Together, they reinforce the importance of neuroimmunomodulation and especially of brain-gut axis interactions.