Copeptin is a stable part of a vasopressin precursor that closely mirrors arginine vasopressin (AVP) secretion. It is known that AVP/copeptin is also released in response to nonosmotic stimuli, such as stress evoked during anterior pituitary dynamic testing. In order to examine the role of AVP in challenging the hypothalamo-pituitary-adrenal axis, we assessed adrenocorticotropic hormone (ACTH), cortisol, copeptin and growth hormone (GH) during a glucagon stimulation test (GST) in 10 patients with satisfactory initial cortisol concentrations (mean ± SD: 20.34 ± 5.10 µg/dL) and failure to show any further cortisol increment on stimulation. For comparison, we measured copeptin in two subjects during an insulin tolerance test (ITT). During GST, there was an increase in copeptin (p = 0.02, average individual increase of 98%, range 10% to 321%). There was a robust increase in GH (p = 0.002, average increase 3300%), a decline in cortisol (p = 0.02, average decline 21.8%) and a fall in ACTH (p = 0.06). The relative increase in copeptin during ITT (176% and 52.2%) overlapped with increments observed during GST; however, here there was an increase in cortisol (20.45→24.26 µg/dL and 4.23→29.29 µg/dL, respectively). There was a moderate correlation between copeptin and GH concentrations (r = 0.4235, p = 0.0007). These results confirm that AVP is not crucial for ACTH-cortisol stimulation, though it might be an important factor in GH secretion.