Abstract: Neuroendocrine response to stress stimuli is influenced by previous stimuli of different nature. The aim of the study was to test whether antecedent orthostatic stress may affect the neuroendocrine response to subsequent hypoglycemia. A group of 12 (6 men, 6 women) nonobese, healthy volunteers aged 19 to 27 y (mean 24 ± 0.8) participated in the study in two sessions: controlled insulin-induced hypoglycemia to 2.7 mmol/L for 15 min either with or without antecedent orthostatic stress (30 min of 60° head-up tilt before insulin administration). Orthostatic stress caused a significant decrease in plasma volume (−9.6%; P < 0.001) and a significant increase in plasma renin activity, aldosterone, norepinephrine (P < 0.01), and adrenocorticotropic hormone (ACTH) concentrations (P < 0.05) in all subjects. Growth hormone response to hypoglycemia was diminished in women (P < 0.01). The epinephrine response to hypoglycemia was diminished in women in comparison to men (P < 0.001), but was unaffected by antecedent orthostatic stress. Hypoglycemia failed to induce the ACTH release after its elevation during orthostatic stress. ACTH response to moderate hypoglycemia without previous orthostatic stress was evident only in men in comparison to women (P < 0.05). We conclude that the epinephrine, growth hormone, and ACTH responses to hypoglycemia were diminished in women. Except ACTH, the neuroendocrine response to mild hypoglycemia was not affected by previous orthostatic stress in healthy subjects. In the case of ACTH, the first stress stimulus is consequential for the subsequent response of this hormone, probably due to short-loop negative feedback effects.