BAM-22P

The immunocytochemical distribution of [Leu]enkephalin and an adrenal enkephalin precursor fragment (BAM-22P) immunoreactivity was investigated in the diencephalon and brainstem of rats pretreated with relatively high doses of colchicine (300-400 micrograms/10 microliters intracerebroventricularly). The higher ranges of colchicine pretreatment allowed the visualization of extensive enkephalin-containing systems in these brain regions, some of which are reported for the first time. Immunoreactive perikarya were found in many hypothalamic and thalamic nuclei, interpeduncular nucleus, substantia nigra, the colliculi, periaqueductal gray, parabrachial nuclei, trigeminal motor and spinal nuclei, nucleus raphe magnus and other raphe nuclei, nucleus reticularis paragigantocellularis, vestibular nuclei, several noradrenergic cell groups, nucleus tractus solitarius, as well as in the spinal cord dorsal horn. In addition to the above regions, immunoreactive fibers were also noted in the habenular nuclei, trigeminal sensory nuclei, locus coeruleus, motor facial nucleus, cochlear nuclei, dorsal motor nucleus of the vagus, and hypoglossal nucleus. When adjacent sections of those stained for [Leu]enkephalin were processed for BAM-22P immunoreactivity, it was found that these two immunoreactivities were distributed identically at almost all anatomical locations. BAM-22P immunoreactivity was generally less pronounced and ws preferentially localized to neuronal perikarya. The results of the present as well as the preceding studies (Khachaturian et al., '83) strongly suggest substantial structural similarity between the adrenal proenkephalin precursor and that which occurs in the brain. Also discussed are some differences and parallels between the distribution of [Leu]enkephalin and dynorphin immunoreactivities.

The adrenal gland is known to produce and release endogenous opioids into the circulation. Bovine adrenal medulla docosapeptide (BAM-22P) is a potent opioid agonist, derived from the proenkephalin A gene, which is present in the adrenal medulla. This study was undertaken to determine whether BAM-22P is released into plasma during acute cholestatic liver injury, which increases plasma total opioid activity. Acute cholestasis was induced by bile duct ligation or administration of the hepatotoxin alpha-naphthylisothiocyanate. Plasma levels of BAM-22P were determined by a sensitive radioimmunoassay, and the specificity of the assay was confirmed using high-performance liquid chromatography. Plasma BAM-22P levels was cholestatic rats were significantly higher than those in control rats. This increase in plasma BAM-22P levels was completely prevented by adrenalectomy. Adrenal steady-state levels of proenkephalin mRNA, as determined by Northern blot hybridization analyses, were also increased significantly in cholestatic rats. These increases in proenkephalin mRNA levels were not paralleled by changes in adrenal BAM-22P peptide levels, which were similar in cholestatic rats and their respective controls. Similar levels of proenkephalin mRNA expression were observed in innervated and denervated adrenal glands from cholestatic rats, suggesting that the increase in adrenal proenkephalin mRNA levels in acute cholestasis is not due to splanchnic nerve activation. Thus acute cholestasis in the rat is associated with adrenal secretion and accumulation in plasma of the highly potent opioid peptide BAM-22P and an augmentation of adrenal proenkephalin mRNA expression. The increase in plasma BAM-22P levels may contribute substantially to the increase in total circulating opioid activity documented in cholestatic rats.

The human and canine small intestine exhibit increased contractility when exposed to exogenous or endogenous opioid peptides. The response of the canine small intestine to the proenkephalin A-derived peptide, peptide E and related processing fragments [Met5]enkephalin, BAM-12P, BAM-18P and BAM-22P was investigated by administering each peptide to isolated, small intestine segments which causes a significant increase in intraluminal pressure. Concentration-response curves from intraarterial bolus administration of peptide E, [Met5]enkephalin, BAM-12P, BAM-18P and BAM-22P showed decreasing efficacy with decreasing amino acid chain length while naloxone (305 nM) significantly antagonized the response. Results using the classical guinea pig ileum/myenteric plexus longitudinal muscle and mouse vas deferens bioassays with specific opioid receptor antagonists provide evidence that peptide E and BAM-18P are relatively specific to the mu opioid receptor, [Met5]enkephalin is more delta specific, BAM-22P is both mu and kappa specific and BAM-12P is kappa opioid receptor specific. These studies demonstrate that locally released (and possibly circulating) peptide E and related processing fragments increase contractility in the small intestine and may be active through more than a single receptor mechanism, particularly the mu receptor.