Outlining the Hidden Curriculum: Perspectives on Successfully Navigating Scientific Conferences.

Scientific conferences and meetings are valuable opportunities for researchers to network, communicate, and develop knowledge. For early career scientists, conferences can also be intimidating, confusing, and overwhelming, especially without having adequate preparation or experience. In this Perspective, we provide advice based on previous experiences navigating scientific meetings and conferences. These guidelines outline parts of the hidden curriculum around preparing for and attending meetings, navigating conference sessions, networking with other scientists, and participating in social activities while upholding a recommended code of conduct.

Barbamide Displays Affinity for Membrane-Bound Receptors and Impacts Store-Operated Calcium Entry in Mouse Sensory Neurons.

Marine cyanobacteria are a rich source of bio-active metabolites that have been utilized as leads for drug discovery and pharmacological tools for basic science research. Here, we describe the re-isolation of a well-known metabolite, barbamide, from Curaçao on three different occasions and the characterization of barbamide's biological interactions with targets of the mammalian nervous system. Barbamide was originally discovered as a molluscicidal agent from a filamentous marine cyanobacterium. In our hands, we found little evidence of toxicity against mammalian cell cultures. However, barbamide showed several affinities when screened for binding affinity for a panel of 45 receptors and transporters known to be involved in nociception and sensory neuron activity. We found high levels of binding affinity for the dopamine transporter, the kappa opioid receptor, and the sigma receptors (sigma-1 and sigma-2 also known as transmembrane protein 97; TMEM97). We tested barbamide in vitro in isolated sensory neurons from female mice to explore its functional impact on calcium flux in these cells. Barbamide by itself had no observable impact on calcium flux. However, barbamide enhanced the effect of the TRPV1 agonist capsaicin and enhanced store-operated calcium entry (SOCE) responses after depletion of intracellular calcium. Overall, these results demonstrate the biological potential of barbamide at sensory neurons with implications for future drug development projects surrounding this molecule.

Identification of PXR Activators from Uncaria Rhynchophylla (Gou Teng) and Uncaria Tomentosa (Cat's Claw).

Uncaria rhynchophylla (Gou Teng) and Uncaria tomentosa (cat's claw) are frequently used herbal supplements in Asia and America, respectively. Despite their common usage, information is limited regarding potential herb-drug interactions associated with Gou Teng and cat's claw. The pregnane X receptor (PXR) is a ligand-dependent transcription factor that regulates cytochrome P450 3A4 (CYP3A4) expression and contributes to some known herb-drug interactions. A recent study found that Gou Teng induces CYP3A4 expression, but its mechanism is unknown. Cat's claw has been determined as a PXR-activating herb, but the PXR activators in cat's claw have not been identified. Using a genetically engineered PXR cell line, we found that the extracts of Gou Teng and cat's claw can dose-dependently activate PXR and induce CYP3A4 expression. We next used a metabolomic approach to profile the chemical components in the extracts of Gou Teng and cat's claw followed by screening for PXR activators. Four compounds, isocorynoxeine, rhynchophylline, isorhynchophylline, and corynoxeine, were identified as PXR activators from both Gou Teng and cat's claw extracts. In addition, three more PXR activators were identified from the extracts of cat's claw, including isopteropodine, pteropodine, and mitraphylline. All seven of these compounds showed the half-maximal effective concentration <10 µM for PXR activation. In summary, our work determined Gou Teng as a PXR-activating herb and discovered novel PXR activators from Gou Teng as well as cat's claw. SIGNIFICANCE STATEMENT: This study's data can be used to guide the safe use of Gou Teng and cat's claw by avoiding PXR-mediated herb-drug interactions.

Novel TRPV1 Modulators with Reduced Pungency Induce Analgesic Effects in Mice.

Capsaicin, the compound in hot chili peppers responsible for their pungency and an agonist of the transient receptor potential cation channel, subfamily V, member 1 (TRPV1), has long been known to promote the desensitization of nociceptors at high concentrations. This has led to the utilization and implementation of topical capsaicin cream as an analgesic to treat acute and chronic pain. Critically, the application of capsaicin cream is limited due to capsaicin's high pungency, which is experienced prior to analgesia. To combat this issue, novel capsaicin analogues were developed to provide analgesia with reduced pungency. Analogues reported in this paper add to and show some differences from previous structure-activity relationship (SAR) studies of capsaicin-like molecules against TRPV1, including the necessity of phenol in the aromatic "A-region", the secondary amide in the "B-region", and modifications in the hydrophobic "C-region". This provided a new framework for de novo small-molecule design using capsaicin as the starting point. In this study, we describe the synthesis of capsaicin analogues, their in vitro activity in Ca2+ assays, and initial in vivo pungency and feasibility studies of capsaicin analogues YB-11 and YB-16 as analgesics. Our results demonstrate that male and female mice treated with YB capsaicin analogues showed diminished pain-associated behavior in the spontaneous formalin assay as well as reduced thermal sensitivity in the hotplate assay.

Gabapentinoid Pharmacology in the Context of Emerging Misuse Liability.

This article will review the epidemiology and pharmacology of gabapentinoids (gabapentin and pregabalin) relevant to their emerging misuse potential and provide guidance for clinical and regulatory management. Gabapentinoids are γ-aminobutyric acid analogues that produce their therapeutic effects by inhibiting voltage-gated calcium channels and decreasing neurotransmitter release. Recently gabapentinoid prescribing and use have increased tremendously. Although traditionally thought to possess a favorable safety profile, gabapentinoid misuse has also risen significantly. Gabapentinoid misuse generally occurs in combination with other substances, most notably opioids, and may be for purposes of eliciting euphoric effects, enhancing the effects of other substances, or self-treating conditions such as withdrawal, pain, anxiety, or insomnia. Given its faster onset, increased bioavailability and potency, and nonsaturable absorption, pregabalin's pharmacokinetics theoretically enhance its misuse liability versus gabapentin. However, gabapentin can produce similar euphoric effects, and epidemiologic studies have identified higher rates of gabapentin misuse in the United States, likely because of greater availability and less regulated prescribing. Although adverse events of gabapentinoid-only ingestion are relatively benign, a growing body of evidence indicates that gabapentinoids significantly increase opioid-related morbidity and mortality when used concomitantly. In addition, significant withdrawal effects may occur on abrupt discontinuation. As a result of these trends, several US states have begun to further regulate gabapentinoid prescribing, reclassifying it as a controlled substance or mandating reporting to local prescription drug-monitoring programs. Although increased regulation of gabapentin prescribing may be warranted, harm reduction efforts and increased patient and provider education are necessary to mitigate this concerning gabapentinoid misuse trend.

Analysis of the antiparasitic and anticancer activity of the coconut palm (Cocos nucifera L. ARECACEAE) from the natural reserve of Punta Patiño, Darién.

Cocos nucifera (C. nucifera) (the coconut palm tree) has been traditionally used to fight a number of human diseases, but only a few studies have tested its components against parasites such as those that cause malaria. In this study, C. nucifera samples were collected from a private natural reserve in Punta Patiño, Darien, Panama. The husk, leaves, pulp, and milk of C. nucifera were extracted and evaluated against the parasites that cause Chagas' disease or American trypanosomiasis (Trypanosoma cruzi), leishmaniasis (Leishmania donovani) and malaria (Plasmodium falciparum), as well as against a line of breast cancer cells. While there was no activity in the rest of the tests, five and fifteen-minute aqueous decoctions of leaves showed antiplasmodial activity at 10% v/v concentration. Removal of some HPLC fractions resulted in loss of activity, pointing to the presence of synergy between the components of the decoction. Chemical molecules were separated and identified using an ultra-performance liquid chromatography (UPLC) approach coupled to tandem mass spectrometry (LC-MS/MS) using atmospheric pressure chemical ionization quadrupole-time of flight mass spectrometry (APCI-Q-TOF-MS) and molecular networking analysis, revealing the presence of compounds including polyphenol, flavone, sterol, fatty acid and chlorophyll families, among others.

Aqueous and methanol extracts of Paullinia pinnata L. (Sapindaceae) improve inflammation, pain and histological features in CFA-induced mono-arthritis: Evidence from in vivo and in vitro studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Paullinia pinnata L. (Sapindaceae) is an African woody vine, traditionally used for the treatment of itch and pain-related conditions such as rheumatoid arthritis. AIM: This work evaluates, in vitro and in vivo, the anti-inflammatory and analgesic effects of aqueous (AEPP) and methanol (MEPP) extracts from Paullinia pinnata leaves. METHODS: AEPP and MEPP (100, 200 and 300 mg/kg/day) were administered orally in monoarthritic rats induced by a unilateral injection of 50 µl of Complete Freund's Adjuvant (CFA) in the ankle joint. During the 14 days of treatment, pain and inflammation were evaluated alternatively in both ankle and paw of the CFA-injected leg. Malondialdehyde (MDA) and glutathione (GSH) levels were assessed in serum and spinal cord. Histology of soft tissue of the ankle was also analyzed. For in vitro studies, AEPP and MEPP (10, 30 and 100 µg/ml) were evaluated against nitric oxide (NO) production by macrophages that were either non-stimulated or stimulated with LPS, 8-Br-AMPc and the mixture of both substances after 8 h exposure. These extracts were also evaluated on TNF-α and IL-1ß production in cells stimulated with LPS for 8 h. Finally, the ability of the extracts to bind to neuroactive receptors was evaluated in vitro using competitive binding assays with >45 molecular targets. RESULTS: AEPP and MEPP significantly reduced by 20-98% (p < 0.001) the inflammation and pain sensation in both the ankle and paw. AEPP significantly increased glutathione levels (p < 0.05) in serum. Both extracts reduced MDA production in serum and spinal cord (p < 0.001), and significantly improved tissue reorganization in treated arthritic rats. P. pinnata extracts did not affect NO production in non-stimulated macrophages but significantly reduced it by 47-88% in stimulated macrophages. AEPP and MEPP also significantly inhibited TNF-α (35-68%) and IL-1ß (31-36%) production in LPS stimulated macrophages. No cytotoxic effect of plant extracts was observed. MEPP showed concentration-dependent affinity for Sigma 2 receptors with an IC50 of 50 µg/ml. CONCLUSION: These results demonstrate the analgesic and anti-inflammatory effects of P. pinnata extracts on monoarthritis and further support its traditional use for pain and inflammation. These activities are at least partly due to the ability of these extracts to inhibit the production of NO, TNF-α, IL-1ß and to likely modulate Sigma 2 receptors.

Evaluating Marine Cyanobacteria as a Source for CNS Receptor Ligands.

Natural products have a long history as a source of psychoactive agents and pharmacological tools for understanding the brain and its circuitry. In the last two decades, marine cyanobacteria have become a standard source of natural product ligands with cytotoxic properties. The study of cyanobacterial metabolites as CNS modulatory agents has remained largely untapped, despite the need for new molecules to treat and understand CNS disorders. We have generated a library of 301 fractions from 37 field collected cyanobacterial samples and screened these fractions against a panel of CNS receptors using radiolabeled ligand competitive-binding assays. Herein we present an analysis of the screening data collected to date, which show that cyanobacteria are prolific producers of compounds which bind to important CNS receptors, including those for 5-HT, DA, monoamine transporters, adrenergic, sigma, and cannabinoid receptors. In addition to the analysis of our screening efforts, we will also present the isolation of five compounds from the same cyanobacterial collection to illustrate how pre-fractionation followed by radioligand screening can lead to rapid identification of selective CNS agents. The systematic screening of natural products sources, specifically filamentous marine cyanobacteria, will yield a number of lead compounds for further development as pharmacological tools and therapeutics.

Cyanobacterial extract with serotonin receptor subtype 7 (5-HT7 R) affinity modulates depression and anxiety-like behavior in mice.

Marine cyanobacteria represent a unique source in the field of drug discovery due to the secondary metabolites they produce and the structural similarity these compounds have to endogenous mammalian receptor ligands. A series of cyanobacteria were subjected to extraction, fractionation by column chromatography and screened for affinity against CNS targets with a focus on serotonin receptors (5-HTRs). Out of 276 fractions screened, 21% had activity at 5-HTRs and/or the 5-HT transporter (SERT). One sample, a cyanobacterium identified by 16S rRNA sequencing as Leptolyngbya from Las Perlas archipelago in Panama, contained a fraction with noted affinity for the 5-HT7 receptor (5-HT7 R). This fraction (DUQ0002I) was screened via intracerebroventricular (ICV) injections in mice using depression and anxiety assays including the forced swim, tail suspension, elevated zero maze, and light-dark preference tests. DUQ0002I decreased depression and anxiety-like behaviors in males and did not have effects in 5-HT7 R knockout or female mice. Administration of DUQ0002I to the CA1 of the hippocampus induced antidepression-like, but not anxiolytic-like behaviors. Testing of further purified materials showed no behavioral effects, leading us to hypothesize that the behavioral effects are likely caused by a synergistic effect between multiple compounds in the fraction. Finally, DUQ0002I was used in a model of neuropathic pain with comorbid depression (spared nerve injury-SNI). DUQ0002I had a similar antidepressant effect in animals with SNI, suggesting a role for the 5-HT7 R in the development of comorbid pain and depression. These results demonstrate the potential that cyanobacterial metabolites have in the field of neuropharmacognosy.

Pharmacophore Comparison and Development of Recently Discovered Long Chain Arylpiperazine and Sulfonamide Based 5-HT7 Ligands.

The serotonin system exerts its effects on the CNS and many peripheral systems. Of the 14 serotonin receptors, the 5-HT7 receptor is the most recently discovered. The 5-HT7 receptor has been shown to be involved in stress reduction, depression, and nociceptive control. Despite the 20 years since the discovery of 5-HT7R, there are still few truly selective ligands. Two of the common scaffolds for 5-HT7R ligands are long chain arylpiperazines (LCAPs) and sulfonamide containing compounds. This review focuses on recently developed (2014-2016) 5-HT7R ligands, their selectivity for the receptor, and suggests the possible new pharmacophore models for these ligands.

Development and Implementation of a Global Health Elective with a Drug Discovery Game for Pharmacy Students.

Interest in global health education within the pharmacy curriculum has increased significantly in recent years. However, discussion of different models and methods to evaluate course structures are limited. The overall objective was to (1) describe the structure of our global health elective for pharmacy students, and (2) assess educational outcomes related to perceived/formal knowledge and attitudes associated with global health. Our elective was designed using a competency-centered approach to global health education, incorporating reflection, projects, service and game-learning. In addition to course assessments, a pre-post survey questionnaire assessing attitudes, knowledge perception, formalized knowledge and opinions was utilized. Overall, students demonstrated appropriate performance on course assessments, temporally improving throughout longitudinal projects. The survey demonstrated significant increases in knowledge perception as a result of the course; however, no change in formalized knowledge was evident through the survey assessment. Additionally, the incorporation of game-learning into the course was well received by students. Future iterations of the course will focus on utilization of different assessment methods to meet learning outcomes.

Summer undergraduate research: A new pipeline for pain clinical practice and research.

BACKGROUND: Most medical schools fail to provide adequate training of clinicians in the treatment of pain. Similarly, despite the fact that over 1/3 of Americans suffer from chronic pain, National Institutes of Health (NIH) funding for pain represents only ~1% of the NIH budget. These issues may dissuade students from pursing pain in their clinical and research careers. To address these gaps in training and funding, we argue that exposing students to pain science early in their careers, at the undergraduate level, may be an effective method to develop a pipeline for future pain clinicians and scientists. To highlight our argument, we will describe our recent successful implementation of a cross-disciplinary and community-engaged biomedical summer research program. The Pain Undergraduate Research Experience (PURE) summer program involved both off-site and on-site experiences to expose undergraduate students to the range of careers in the pain field from basic science to clinical practice. The objective of the 10-week long PURE program was to evaluate whether a combination of basic science research, clinical practice visits, and patient interactions would increase student understanding of and exposure to the underlying science of pain. METHODS: A pre-post cohort study was used without a comparison group. Entry and exit surveys were used to evaluate students' perceptions about pain clinical practice and research, student interest in pain, and student confidence about communicating about pain and doing basic science pain research. RESULTS: Students reported significant increases to a number of questions in the survey. Questions were scored on 5 point Likert scales and there was significant increases in student understanding of what life is like with chronic pain (2.6 vs 4.3 post survey), their confidence in explaining pain to a patient (2.8 vs 4.1) or researcher (2.8 vs 4), and their comfort with pain terminology(2.8 vs 3.9). CONCLUSIONS: With the PURE program, we wanted to entice top undergraduates to consider pain as a future area of study, practice, and/or research. We present a model that can be easily implemented at research universities throughout the United States.

Marine cyanobacteria-derived serotonin receptor 2C active fraction induces psychoactive behavioral effects in mice.

CONTEXT: Marine cyanobacteria offer a robust resource for natural products drug discovery due to the secondary metabolites they produce. OBJECTIVE: To identify novel cyanobacterial compounds that exhibit CNS psychoactive effects. MATERIALS AND METHODS: Cyanobacteria were collected from Las Perlas Archipelago, Panama and subjected to dichloromethane/methanol extraction and fractionation by column chromatography before being screened for affinity against a panel of CNS targets. A 50:50 ethyl acetate:methanol fraction of one cyanobacterial extract (2064H) was subjected to HPLC and the major peak was isolated (2064H3). At a dose of 20 µg per animal, 2064H and 2064H3 were tested in mice using behavioral assays that included the forced swim, open field and formalin tests. RESULTS: 2064H was shown to bind to the serotonin 2C (5-HT2C) receptor, a known target for depression and pain treatment. 2064H showed 59.6% inhibition of binding of [3H]-mesulergine with an IC50 value of 179 ng/mL and did not show inhibition of binding greater than 45% with any other receptors tested. Both 2064H and 2064H3 decreased immobility time in the first minute of the tail suspension test. 2064H increased time, distance and number of entries in the center region in the first half of the open field test. 2064H increased overall nocifensive behaviors in the formalin test. DISCUSSION AND CONCLUSION: Overall, manipulating the 5-HT2C receptor with these receptor-specific ligands derived from cyanobacteria altered pain, depression and anxiety-like behaviors, illustrating the importance of this receptor in affective behaviors. These results demonstrate the potential of cyanobacteria as a source for CNS active compounds.

Coibacins A-D, antileishmanial marine cyanobacterial polyketides with intriguing biosynthetic origins.

Four unsaturated polyketide lactone derivatives, coibacins A-D, were isolated from a Panamanian marine cyanobacterium, cf. Oscillatoria sp. The two different types of termini observed in these co-occurring metabolites, either a methyl cyclopropyl ring as seen in curacin A or a methyl vinyl chloride similar to that observed in the jamaicamides, suggest an intriguing flexibility in the "beta branch" forming biosynthetic process. The coibacins possess selective antileishmanial activity as well as potent anti-inflammatory activity.

Antinociceptive effects of herkinorin, a MOP receptor agonist derived from salvinorin A in the formalin test in rats: new concepts in mu opioid receptor pharmacology: from a symposium on new concepts in mu-opioid pharmacology.

Herkinorin is the first µ opioid (MOP) selective agonist derived from salvinorin A, a hallucinogenic natural product. Previous work has shown that, unlike other opioids, herkinorin does not promote the recruitment of ß-arrestin-2 to the MOP receptor and does not lead to receptor internalization. This paper presents the first in vivo evaluation of herkinorin's antinociceptive effects in rats, using the formalin test as a model of tonic inflammatory pain. Herkinorin was found to produce a dose-dependent decrease in the number of flinches evoked by formalin. These antinociceptive effects were substantially blocked by pretreatment with the nonselective antagonist naloxone, indicating that the antinociception is mediated by opioid receptors. Contralateral administration of herkinorin did not attenuate the number of flinches evoked by formalin, indicating that its effects are peripherally restricted to the site of injection. Following chronic administration (5-day), herkinorin maintained antinociceptive efficacy in both phases of the formalin test. Furthermore, unlike morphine, herkinorin was still able to inhibit flinching in both phases of the formalin test in animals made tolerant to chronic systemic morphine treatment. Collectively, these results suggest that herkinorin may produce peripheral antinociception with decreased tolerance liability and thereby represents a promising template for the development of agents for the treatment of a variety of pain states.

Evolved diversification of a modular natural product pathway: apratoxins F and G, two cytotoxic cyclic depsipeptides from a Palmyra collection of Lyngbya bouillonii.

A collection of Lyngbya bouillonii from Palmyra Atoll in the Central Pacific, a site several thousand kilometers distant from all previous collections of this chemically prolific species of cyanobacterium, was found to contain two new cancer cell cytotoxins of the apratoxin family. The structures of the new compounds, apratoxins F and G, were determined by 1D and 2D NMR techniques in combination with mass spectrometric methods. Stereochemistry was explored by using chromatographic analyses of the hydrolytically released fragments in combination with NMR and optical rotation comparisons with known members of the apratoxin family. Apratoxins F and G add fresh insights into the SAR of this family because they incorporate an N-methyl alanine residue at a position where all prior apratoxins have possessed a proline unit, yet they retain high potency as cytotoxins to H-460 cancer cells with IC(50) values of 2 and 14 nM, respectively. Additional assays using zone inhibition of cancer cells and clonogenic cells give a comparison of the activities of apratoxin F to apratoxin A. Additionally, the clonogenic studies in combination with maximum tolerated dose (MTD) studies provided insights as to dosing schedules that should be used for in vivo studies, and preliminary in vivo evaluation validated the predicted in vivo efficacy for apratoxin A. These new apratoxins are illustrative of a mechanism (the modification of an NRPS adenylation domain specificity pocket) for evolving a biosynthetic pathway so as to diversify the suite of expressed secondary metabolites.

Malyngolide dimer, a bioactive symmetric cyclodepside from the panamanian marine cyanobacterium Lyngbya majuscula.

Fractionation of the extract of the marine cyanobacterium Lyngbya majuscula collected from Panama led to the isolation of malyngolide dimer (1). The planar structure of 1 was determined using 1D and 2D NMR spectroscopy and HRESI-TOFMS. The absolute configuration was established by chemical degradation followed by chiral GC-MS analyses and comparisons with an authentic sample of malyngolide seco-acid (4). Compound 1 showed moderate in vitro antimalarial activity against chloroquine-resistant Plasmodium falciparum (W2) (IC(50) = 19 microM) but roughly equivalent toxicity against H-460 human lung cell lines. Furthermore, because the closely related cyanobacterial natural product tanikolide dimer (5) was a potent SIRT2 inhibitor, compound 1 was evaluated in this assay but found to be essentially inactive.

Dragonamide E, a modified linear lipopeptide from Lyngbya majuscula with antileishmanial activity.

Tropical parasitic and infectious diseases, such as leishmaniasis, pose enormous global health threats, but are largely neglected in commercial drug discovery programs. However, the Panama International Cooperative Biodiversity Group (ICBG) has been working to identify novel treatments for malaria, Chagas' disease, and leishmaniasis through an investigation of plants and microorganisms from Panama. We have pursued activity-guided isolation from an extract of Lyngbya majuscula that was found to be active against leishmaniasis. A new modified linear peptide from the dragonamide series was isolated, dragonamide E (1), along with two known modified linear peptides, dragonamide A (2) and herbamide B (3). Dragonamides A and E and herbamide B exhibited antileishmanial activity with IC50 values of 6.5, 5.1, and 5.9 microM, respectively. Spectroscopic and stereochemical data for dragonamide E (1) and herbamide B (3; the spectroscopic and stereochemical data for this substance is incomplete in the literature) are presented as well as comparisons of biological activity within the dragonamide compound family. Biosynthetic differences among marine compounds with a terminal free amide are also discussed.

Herkinorin analogues with differential beta-arrestin-2 interactions.

Salvinorin A is a psychoactive natural product that has been found to be a potent and selective kappa opioid receptor agonist in vitro and in vivo. The activity of salvinorin A is unusual compared to other opioids such as morphine in that it mediates potent kappa opioid receptor signaling yet leads to less receptor downregulation than observed with other kappa agonists. Our initial chemical modifications of salvinorin A have yielded one analogue, herkinorin ( 1c), with high affinity at the microOR. We recently reported that 1c does not promote the recruitment of beta-arrestin-2 to the microOR or receptor internalization. Here we describe three new derivatives of 1c ( 3c, 3f, and 3i) with similar properties and one, benzamide 7b, that promotes recruitment of beta-arrestin-2 to the microOR and receptor internalization. When the important role micro opioid receptor regulation plays in determining physiological responsiveness to opioid narcotics is considered, micro opioids derived from salvinorin A may offer a unique template for the development of functionally selective mu opioid receptor-ligands with the ability to produce analgesia while limiting adverse side effects.

Synthetic studies of neoclerodane diterpenes from Salvia divinorum: exploration of the 1-position.

Modification of the C-1 ketone of salvinorin A (2a) produces analogues with opioid antagonist properties. Of particular significance is the finding that 1-deoxo-1,10-dehydrosalvinorin A (11a) is a moderately potent antagonist at all three opioid receptor subtypes, and that herkinorin (2b), a mu agonist, is converted to a weak antagonist by removal of the C-1 ketone (3b and 11b). These observations suggest that the ketone of 2b is a key structural feature responsible for mu agonist activity.