The effects of synthetic GHRH-(1-44) (1 microgram/kg, iv), clonidine (0.15 mg/m2, orally), L-dopa (0.5 g, orally), and insulin (0.1 IU/kg, iv) on plasma immunoreactive (ir) GHRH and GH levels were determined in normal men, aged 31-46 yr (n = 4-8). In addition, plasma ir-GHRH and GH concentrations were determined before and after the administration of clonidine in six younger men, aged 19-25 yr. GHRH was extracted from plasma using Sep-Pak C18 cartridges and measured with a mid-portion-specific GHRH antiserum. The mean plasma ir-GHRH and GH levels ranged from 9-11 ng/L and 0.5-1.5 microgram/L, respectively, in the older men during a 2-h control study. After GHRH administration, the mean plasma ir-GHRH concentration increased to a peak of 512.5 ng/L at 3 min and GH to a peak of 9.2 micrograms/L at 10 min. Clonidine resulted in a significant increase in mean plasma GH levels (P less than 0.05) in the younger men, but not in the older men. Plasma ir-GHRH concentrations did not change after clonidine. L-Dopa increased plasma ir-GHRH at 60 min (P less than 0.05) and GH at 60-120 min (P less than 0.05). Insulin-induced hypoglycemia increased plasma GH levels (to a mean of 23.8 micrograms/L at 60 min; P less than 0.001), whereas plasma ir-GHRH levels did not change. We conclude that the mechanisms of the various GH stimulation tests differ. Some GH responses, including those induced by insulin, do not appear to be mediated by GHRH.