Effect of three peptidase inhibitors on antinociceptive potential and toxicity with intracerebroventricular administration of dynorphin A (1–17) or (1–13) in the rat

Peptide fragments from Dyn A (1–17) following incubation with membrane preparation under pretreatment with a mixture of the three PIs was identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometer (MALDI–TOF–MS). Infusion of drugs and peptides into the third ventricle in rats was performed via indwelling cannulae. Induction of antinociception and toxicity by Dyn A (1–17), Dyn A (1–13), Dyn A (1–6), or Dyn A (7–17) were determined by the tail-flick test and induction of barrel rotation, respectively. The effects of the PIs on antinociception and toxicity were evaluated by a dose−response study and a comparison of differences among various combinations of Dyn A (1–17) or Dyn A (1–13) and the three PIs and p-hydroxymercuribenzoate.

MALDI–TOF–MS analysis identified Dyn A (1–6) and Dyn A (1–10) fragments as products following incubation of Dyn A (1–17) with membrane preparation of rat midbrain under pretreatment with a mixture of the three PIs. Pretreatment with a mixture of the three PIs produced an approximately 30-fold augmentation in antinociception induced by low-dose intracerebroventricular administration of Dyn A (1–17) or (1–13) in a μ-, δ- and κ-opioid receptor antagonist-reversible manner, but without signs of toxicity such as barrel rotation in the rat. Dyn A (1–17)-induced antinociception and toxicity was greater than that of Dyn A (1–6), Dyn A (1–13), or Dyn A (7–17) at the same dose. Dyn A (1–17)-induced antinociception and toxicity under pretreatment with various combinations of the three PIs and p-hydroxymercuribenzoate was greater than that with a mixture of the three PIs alone.