1. Acetylation of morphine at the 6-position changes its pharmacology. To see if similar changes are seen with codeine, we examined the analgesic actions of codeine and 6-acetylcodeine.2. Like codeine, 6-acetylcodeine is an effective analgesic systemically, supraspinally and spinally, with a potency approximately a third that of codeine.3. The sensitivity of 6-acetylcodeine analgesia to the mu-selective antagonists beta-FNA and naloxonazine confirmed its classification as a mu opioid. However, it differed from the other mu analgesics in other paradigms.4. Antisense mapping revealed the sensitivity of 6-acetylcodeine to probes targeting exons 1 and 2 of the mu opioid receptor gene (Oprm), a profile distinct from either codeine or morphine. Although heroin analgesia also is sensitive to antisense targeting exons 1 and 2, heroin analgesia also is sensitive to the antagonist 3-O-methylnaltrexone, while 6-acetylcodeine analgesia is not.5. Thus, 6-acetylcodeine is an effective mu opioid analgesic with a distinct pharmacological profile.

14-Methoxymetopon is a potent opioid analgesic. When given systemically, it is approximately 500-fold more active than morphine. However, this enhanced potency is markedly increased with either spinal or supraspinal administration, where its analgesic activity is more than a million-fold greater than morphine. It was μ-opioid receptor selective in binding assays and its analgesia was blocked only by μ-opioid receptor-selective antagonists. Yet, it had a different selectivity profile than either morphine or morphine-6β-glucuronide. Unlike morphine, 14-methoxymetopon was antagonized by 3-O-methylnaltrexone, it was sensitive to antisense probes targeting exons 1, 2 and 8 of the opioid receptor gene and was inactive both spinally and supraspinally in CXBK mice. Although it retarded gastrointestinal transit, it displayed a ceiling effect with no dose lowering transit by more than 65%, in contrast to the complete inhibition of transit by morphine. These finding demonstrate that 14-methoxymetopon is a highly potent μ-opioid with a pharmacological profile distinct from that of the traditional μ-opioid morphine.

Many peptides and transmitters found within the brain also have peripheral sites of action. We now demonstrate that the brain releases functionally active neurotransmitters/neuromodulators directly from the brain into the blood through a saturable P-glycoprotein (Pgp) transport system. Downregulating Pgp1 expression with antisense reduced the brain-to-blood transport of morphine, β-endorphin and other opioids. Lowering Pgp expression significantly enhanced systemic morphine analgesia and prevented tolerance, but diminished the analgesic activity of centrally administered morphine, implying that supraspinal analgesia resulted from a combination of central and peripheral mechanisms activated by morphine transported from the brain to the blood. Similarly, mice with a disruption of the Mdr1a gene were more sensitive to systemic morphine and less sensitive to morphine given centrally. This ability of the Pgp transport system to pump functionally active compounds from the brain to periphery defines a potentially important mechanism for the central nervous system to modulate peripheral systems.

In mice, 3-O-methylnaltrexone blocks the analgesic actions of morphine-6beta-glucuronide and heroin at doses which are inactive against morphine. We found a similar selectivity in rats. 3-O-Methylnaltrexone antagonized the analgesic actions of 6-acetylmorphine in Sprague-Dawley rats and heroin in Wistar rats at doses that were inactive against morphine. Inclusion of a fixed dose of 3-O-methylnaltrexone significantly shifted the analgesic dose-response curves for 6-acetylmorphine and heroin without altering the morphine dose-response curves. In a self-administration model, 3-O-methylnaltrexone treatment significantly increased both heroin and morphine intake during the first hour, suggestive of an antagonist effect. This effect at doses of 3-O-methylnaltrexone which were inactive against morphine analgesia implied a role for the morphine-6beta-glucuronide opioid receptor in the reinforcing properties of heroin and morphine.

Morphine produces analgesia by activating mu opioid receptors encoded by the MOR–1 gene. Although morphine–6β–glucuronide (M6G), heroin and 6–acetylmorphine also are considered mu opioids, recent evidence suggests that they act through a distinct receptor mechanism. We examined this question in knockout mice containing disruptions of either the first or second coding exon of MOR–1. Mice homozygous for either MOR–1 mutation were insensitive to morphine. Heroin, 6–acetylmorphine and M6G still elicited analgesia in the exon–1 MOR–1 mutant, which also showed specific M6G binding, whereas M6G and 6–acetylmorphine were inactive in the exon–2 MOR–1 mutant. These results provide genetic evidence for a unique receptor site for M6G and heroin analgesia.

Orphanin FQ/nociceptin (OFQ/N) is a recently identified neuropeptide with high affinity for the orphan opioid receptor. OFQ/N blocked morphine analgesia in mice in a dose-dependent manner, as well as the analgesic actions of [d-Pen2, d-Pen5]enkephalin (DPDPE), morphine-6β-glucuronide, trans-3,4-dichloro-N-[2-(1-pyrrolindinyl)-cyclohexyl]-benzeneacetamide, methane sulfonate hydrate (U50,488H) and naloxone benzoylhydrazone. These actions are anti-analgesic, because OFQ/N also blocked clonidine analgesia and OFQ/N was inactive against the inhibition of gastrointestinal transit by morphine. Although OFQ/N was quite potent in these paradigms, two truncated forms, OFQ/N(1-11) and OFQ/N(1-7), were inactive. An antisense oligodeoxynucleotide targeting the first coding exon of KOR-3, the mouse homolog of the orphan opioid receptor, effectively prevented the anti-opioid actions of OFQ/N, confirming the importance of the orphan opioid receptor in this action.

In the current study, we used an antisense oligodeoxynucleotide targeting the recently cloned σ1 receptor to assess its functions within the nervous system. σ1 antagonists potentiate the analgesic actions of opioids. Similarly, the antisense probe targeting the σ1 receptor enhanced the analgesic activity of the κ1-opioid receptor agonist U50,488H (trans-3,4-dichloro-N-[2-(1-pyrrolidinyl)cyclohexyl]benzeacetamidel) and the κ3-opioid receptor agonist naloxone benzoylhydrazone. A mismatch control was inactive. These results confirm the role of σ1 receptors in an anti-opioid analgesic system and illustrate the utility of antisense approaches towards the elucidation of σ receptor functions.

In the current study, we used an antisense oligodeoxynucleotide targeting the recently cloned sigma1 receptor to assess its functions within the nervous system. Sigma1 antagonists potentiate the analgesic actions of opioids. Similarly, the antisense probe targeting the sigma1 receptor enhanced the analgesic activity of the kappa1-opioid receptor agonist U50,488H (trans-3,4-dichloro-N-[2-(1-pyrrolidinyl)cyclohexyl]benzeacetamidel++ +) and the kappa3-opioid receptor agonist naloxone benzoylhydrazone. A mismatch control was inactive. These results confirm the role of sigma1 receptors in an anti-opioid analgesic system and illustrate the utility of antisense approaches towards the elucidation of sigma receptor functions.