In an attempt to assess possible discomfort in rats subjected to orbital puncture while under diethyl-ether anaesthesia, their endocrine stress response was determined. Concentrations of corticosterone, adrenaline and noradrenaline were measured in plasma obtained via a jugular catheter from rats subjected to diethyl-ether anaesthesia with or without orbital puncture. No statistically significant differences were found between the punctured and non-punctured rats as to peak levels of plasma corticosterone and adrenaline as well as for the times required by the increased concentrations to return to baseline values. The rate by which the plasma noradrenaline level returned to baseline values was somewhat decreased by orbital puncture. Diethyl-ether anaesthesia alone produced a marked endocrine response when compared with handling and novelty stress associated with the induction of anaesthesia. It is concluded that diethyl-ether anaesthesia causes pronounced increases in the plasma levels of the selected stress hormones and that orbital puncture does not amplify this response. It is suggested that diethyl-ether anaesthesia masks any effects of orbital puncture.