Delta-opioid Research Articles

Novel histone deacetylase-5 inhibitor T2943 exerts an anti-depressive effect in mice by enhancing GRID1 expression

Histone deacetylase-5 (HDAC5) is implicated in the pathogenesis of depression and the mechanistic pathways underlying the effects of antidepressant medications. We previously identified a novel HDAC5 inhibitor, T2943, with antidepressant properties that promote histone 3 lysine-14 acetylation (H3K14ac) by inhibiting HDAC5 activity. In this study, we identify the core genes promoting transcription and expression following T2943-mediated upregulation of H3K14ac, highlighting Grid1 (GluD1) as a central gene. We used cleavage under targets and tagmentation (CUT&Tag), gene set enrichment analysis, and behavioral tests after GRID1 (glutamate receptor delta-1 subunit) knockdown. Gene ontology and pathway enrichment analysis via CUT&Tag suggested the following mechanism for the antidepressant action of T2943: T2943 inhibits HDAC5 activity to promote H3K14 acetylation. This modification loosens the chromatin structure, allowing transcription factors to bind to the Grid1 promoter region and enhance its transcription and expression. Upregulated GRID1 mediates signal transmission in neural pathways, restores the regenerative ability of hippocampal nerve cells, promotes nerve growth and synaptic formation, increases synapse numbers, and enhances synaptic function. Our findings highlight the therapeutic potential of targeting HDAC5 in depression and clarify the antidepressant mechanism of T2943.

Delta-opioid receptor signaling alleviates neuropathology and cognitive impairment in the mouse model of Alzheimer’s disease by regulating microglia homeostasis and inhibiting HMGB1 pathway

BackgroundRecent studies suggest that opioid receptor signaling may differentially affect Alzheimer’s disease (AD) pathology and the relevant behavioral dysfunctions. However, the precise roles and mechanisms of opioid receptor subtypes in AD pathologies are still unclear with major controversies.MethodsWe compared the delta-opioid receptor (DOR)- and mu-opioid receptor (MOR)-mediated effects on AD-associated cognitive deficits, pathologies, neuroinflammations, cell death using transgenic APP/PS1 mouse model and BV2 cell line at behavioral, molecular, and cellular levels. Unpaired t-test and one/two way analysis for variance (ANOVA) were used to analyze statistical significance of the data.ResultsWe show a distinct role of DOR and its major difference with MOR in AD injury in an APP/PS1 mouse model. DOR activation by UFP-512, but not MOR activation by DAMGO, attenuated cognitive impairment, reduced beta-amyloid (Aβ) production and aggregation, as well as protected the neurons from apoptosis in APP/PS1 mice. DOR and MOR also differentially modulated microglia in APP/PS1 mice and in vitro AD cell model with a DOR-mediated inhibition on the excessive activation of microglia and the release of pro-inflammatory cytokines in AD pathologies. Gene expression profiling further revealed that the alternations in DOR/MOR are closely associated with microglial homeostatic signatures and high mobility group protein B1 (HMGB1) in AD. DOR activation inhibited HMGB1 secretion and its translocation from nuclear to cytoplasm. Our in-vitro studies further confirmed that DOR overexpression mitigated microglial inflammatory response and rescued neurons from AD injury via HMGB1-NF-κB signaling pathway.ConclusionsThese novel findings uncover previously unappreciated roles of DOR in neuroprotection against AD injury via modulating microglia-related inflammatory responses.

Peripherally mediated opioid combination therapy in mouse and pig

The concomitant epidemics of chronic pain and opioid misuse in the United States have led to a call for novel analgesics with limited abuse potential. Previously, we have shown that co-delivery of a novel combination targeting both μ- and δ-opioid receptors in the peripheral and central nervous systems can produce synergistic analgesia. Loperamide, a peripherally restricted μ-opioid agonist, and oxymorphindole, a δ-opioid receptor partial agonist, synergize in multiple mouse models of hyperalgesia. We predicted this effect would generalize across species and therefore assessed this combination for analgesic synergy in a mouse model of post-incisional hypersensitivity. In mice, oxymorphindole and loperamide displayed significant analgesic synergy. Similar synergy was observed with N-benzyl-oxymorphindole and loperamide. In cross-bred pigs, we compared the analgesic effects of either morphine alone or the combinations of oxymorphindole and loperamide or the combination of N-benzyl-oxymorphindole and loperamide. Both combinations showed increased potency as compared to morphine sulfate and effectively reduced hypersensitivity following injury without side effects. From these data we conclude that the combination of oxymorphindole and loperamide or the combination of N-benzyl-oxymorphindole reverse incisional hyperalgesia, likely by acting in the periphery, in a large animal model without adverse effects on respiration or heart rate. PerspectiveThis article presents novel opioid combinations, the μ-opioid agonist loperamide with a δ-opioid agonist, either oxymorphindole (OMI) or N-benzyl-oxymorphindole (BOMI), that relieve pain in mice and pigs without adverse side effects. These therapies could help clinicians manage pain in patients while reducing overall opioid burden and limiting side effects.

Delta opioid receptors engage multiple signaling cascades to differentially modulate prefrontal GABA release with input and target specificity

Delta opioid receptors engage multiple signaling cascades to differentially modulate prefrontal GABA release with input and target specificity

Opioid receptor signaling throughout ontogeny: Shaping neural and behavioral trajectories.

Due to the recent and ongoing opioid crisis in the United States, exposure to opioid drugs in utero is becoming more common, including during medication-assisted therapy used to treat opioid use disorder. As such, careful consideration of opioidergic signaling in utero and beyond, as well as alterations to this signaling via introduction of exogenous opioids, is warranted. This review explores the ontogeny and function of the Mu, Kappa and Delta opioid receptor systems throughout the lifespan, highlighting their importance in guiding neurobehavioral development. We argue for a paradigm shift in conceptualization of opioids as not only contributors within their own system, but also vital regulators of a multitude of downstream neurodevelopmental processes.

D-Serine disrupts Cbln1 and GluD1 interaction and affects Cbln1-dependent synaptic effects and nocifensive responses in the central amygdala

Ionotropic glutamate receptors (iGluRs) mediate fast excitatory neurotransmission in the nervous system. In addition to NMDA receptor co-agonists, D-serine is a ligand for glutamate delta receptors (GluDs) and interacts with the ligand-binding domain with low affinity. However, D-serine binding does not lead to typical ion channel currents in GluD1 or GluD2 but may contribute to synaptic plasticity. In the developing brain, D-serine binding to GluD2 facilitates long-term depression at parallel fiber-Purkinje cell synapses. However, the influence of D-serine on GluD1's interaction with its amino terminal domain synaptogenic ligand Cbln1 and its subsequent impact on synaptic function and behavior remains unexplored. Here, we found that D-serine inhibited the interaction between Cbln1 and GluD1 in an in vitro cell-binding assay. This effect was concentration-dependent, with an IC50 value of ~ 300 µM. Furthermore, in ex vivo central amygdala (CeA) slices application of recombinant Cbln1 (rCbln1), consistent with its synaptogenic property, produced a robust increase in excitatory neurotransmission and GluD1 expression. This effect of rCbln1 was partially blocked by pre-treatment with D-serine. Finally, in behavioral experiments, we observed that the pro-nociceptive effect of intra-CeA injection of rCbln1 was inhibited by pre-treatment with D-serine. In addition, the antinociceptive effect of intra-CeA rCbln1 injection in an inflammatory pain model was blocked by D-serine. Overall, these results demonstrated that D-serine binding to GluD1 reduces its interaction with Cbln1, which may be relevant to synaptic plasticity and behavior.Graphical

Scaled and Weighted Laplacian Matrices as Functional Descriptors for GPCR Ligands.

The G protein-coupled receptor (GPCR) pharmacology accounts for a significant field in research, clinical studies, and therapeutics. Computer-aided drug discovery is an evolving suite of techniques and methodologies that facilitate accelerated progress in drug discovery and repositioning. However, the structure-activity relationships of molecules targeting GPCRs are highly challenging in many cases since slight structural modifications can lead to drastic changes in biological functionality. Numerous molecular descriptors have been described, many of which successfully characterize the structural and physicochemical features of drug sets. Nonetheless, elucidating the structure-functionality relationships over extensive sets of drugs with multiple structural variations and known biological activity remains challenging in various biological systems. This work presents novel topological descriptors using Laplacian matrices, weighted, and scaled by atomic mass and partial charges. We tested these descriptors on three sets of GPCR ligands: muscarinic, β-adrenergic, and δ-opioid receptor ligands, evaluating their potential as functional descriptors of these receptors.

Buprenorphine Pharmacodynamics: A Bridge to Understanding Buprenorphine Clinical Benefits.

Buprenorphine is an agonist at the mu opioid receptor (MOR) and antagonist at the kappa (KOR) and delta (DOR) receptors and a nociceptin receptor (NOR) ligand. Buprenorphine has a relatively low intrinsic efficacy for G-proteins and a long brain and MOR dwell time. Buprenorphine ceiling on respiratory depression has theoretically been related multiple factors such as low intrinsic efficacy at MOR, binding to six-transmembrane MOR and interactions in MOR/NOR heterodimers. Buprenorphine reduces analgesic tolerance by acting as a delta opioid receptor (DOR) antagonist. As a kappa opioid receptor (KOR) antagonist, buprenorphine reduces craving associated with addiction. Buprenorphine is a model opioid for the ordinal bifunctional analogs BU10038, BU08028 which have been shown to be potent analgesics in non-human primates without reinforcing effects and little to no respiratory depression.

Delta opioid receptors affect acoustic features of song during vocal learning in zebra finches

Delta-opioid receptors (δ-ORs) are known to be involved in associative learning and modulating motivational states. We wanted to study if they were also involved in naturally-occurring reinforcement learning behaviors such as vocal learning, using the zebra finch model system. Zebra finches learn to vocalize early in development and song learning in males is affected by factors such as the social environment and internal reward, both of which are modulated by endogenous opioids. Pairs of juvenile male siblings (35-day-old) were systemically administered a δ-OR-selective antagonist naltrindole or vehicle (controls) for a period of 10 days. The acoustic structure of songs differed across treated and control groups at adulthood (120 days). Naltrindole-treated birds had a significantly lower pitch, mean frequency, and frequency modulation than controls, whereas there was no difference in the number of songs in naltrindole-treated and control siblings. Since the opioid and dopaminergic systems interact, we decided to study whether blocking δ-ORs during the sensitive period led to changes in dopaminoceptive neurons in Area X, a song control nucleus in the basal ganglia. Interestingly, compared with controls, naltrindole-treated birds had higher numbers of DARPP-32-positive medium spiny neurons and potentially excitatory synapses in Area X. We show that manipulating δ-OR signaling during the learning phase resulted in alterations in the acoustic features of song and had long term effects on dopaminergic targets within the basal ganglia in adulthood. Our results suggest that endogenous opioids regulate the development of cognitive processes and the underlying neural circuitry during the sensitive period for learning.

P0252 Mild Crohn’s disease is associated with altered sphingolipid metabolism and reduced neutrophilic inflammation

Abstract Background Around 20-30% of patients with Crohn's disease (CD) experience a mild disease course. However, identifying patients likely to maintain a mild course is challenging. This study aimed to identify molecular features of mild CD by exploring biological datasets from two large patient cohorts. Methods Patients with mild CD were analysed from two longitudinal cohorts: the Mount Sinai Crohn's and Colitis Registry (MSSCR) observational cohort and the Ocean State Crohn’s and Colitis Registry (OSCCAR) inception cohort. Baseline assessments in both cohorts included biological/immunomodulator use, history of IBD-related surgeries, and complications (stricturing, penetrating, and perianal CD). Mild CD was defined by the absence of these variables at baseline and after five years of follow-up (Fig. 1A). The remaining patients were mild at baseline but progressed during follow-up. Patients with mild CD and those who progressed after being mild at baseline were evaluated for potential molecular biomarkers including circulating metabolites (metabolomics), immune cell profiles (multiparametric flow cytometry), serum proteomics, and intestinal transcriptomics. Results In total, 100 (MSSCR) and 152 (OSCCAR) patients were characterized, with 55 (55.0%) and 74 (48.7%) classified as mild CD, respectively. Intestinal transcriptomic analysis revealed downregulation of genes associated with neutrophilic inflammation and innate immunity, including SAA, DUOXA2, FCGR3B, LCN2, S100A8, and AQP9 (Fig. 1B). Flow cytometry showed lower frequencies of neutrophils (CD16+), lower neutrophil-to-lymphocyte ratios (NLR), and higher counts of NK and total B cells in mild CD (Fig. 1C). Metabolomic analysis identified ceramide- and sphingomyelin compounds (particularly acyl-CoA chain lengths C14-16 and C22-26) as positively associated with mild CD (odds ratios [OR] >2.0), whereas primary and secondary bile acids and amino acid derivatives from lysine, tyrosine, and proline metabolism were inversely associated (Fig. 1D). Proteomic analysis in OSCCAR demonstrated associations between C-X-C motif chemokine ligand 9 (CXCL9), macrophage colony stimulating factor (CSF-1), and TGF-β1 latency-associated peptide (LAP-TGF-β), and an inverse association for Delta and Notch-like epidermal growth factor-related receptor (DNER) with mild CD (adjusted P<0.05) (Fig. 1E). Conclusion This study provides multi-omics-based biomarker signatures of mild CD, which primarily relate to altered ceramide- and sphingomyelin metabolites and lower systemic neutrophilic inflammation. Future studies focusing on the biological significance and potential clinical impact on disease monitoring and therapeutic strategies are warranted.

Activation of δ-opioid receptors blocks allodynia in a model of headache induced by PACAP.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a human migraine trigger that is being targeted for migraine. The δ-opioid receptor (δ-receptor) is a novel target for the treatment of migraine, but its mechanism remains unclear. The goals of this study were to develop a mouse PACAP-headache model using clinically significant doses of PACAP; determine the effects of δ-receptor activation in this model; and investigate the co-expression of δ-receptors, PACAP and PACAP-PAC1 receptor. Cephalic allodynia to low doses of acute and chronic PACAP were tested. A triptan (sumatriptan) and a CGRP receptor antagonist (olcegepant) were tested in this model. The δ-receptor agonist SNC80 was tested in PACAP and CGRP-induced headache models. Expression of PACAP, PAC1, CRLR and δ-receptors was determined using in situ hybridisation. Low doses of PACAP produced dose-dependent acute and chronic cephalic allodynia, blocked by sumatriptan but not by olcegepant. The PAC1 antagonist (M65) did not inhibit CGRP-induced allodynia. There was moderate co-expression of PAC1 and CRLR transcripts in migraine-related regions. SNC80 blocked PACAP- and CGRP-induced allodynia. There was low co-expression of PACAP and δ-receptors in brain regions measured. However, there was high co-expression of PAC1 and δ-receptors in somatosensory cortex, hippocampus and trigeminal nucleus caudalis. We developed a translationally significant model of PACAP-induced headache, which was mechanistically distinct from CGRP. Activation of δ-receptors blocked PACAP- and CGRP-induced allodynia, and δ-receptors were highly co-expressed with the PACAP-ergic system. Future studies will examine the functional relationship between δ-receptors and PAC1.

Circular mRNA Vaccine against SARS-COV-2 Variants Enabled by Degradable Lipid Nanoparticles.

The emergence of mRNA vaccines offers great promise and a potent platform in combating various diseases, notably COVID-19. Nevertheless, challenges such as inherent instability and potential side effects of current delivery systems underscore the critical need for the advancement of stable, safe, and efficacious mRNA vaccines. In this study, a robust mRNA vaccine (cmRNA-1130) eliciting potent immune activation has been developed from a biodegradable lipid with eight ester bonds in the branched tail (AX4) and synthetic circular mRNA (cmRNA) encoding the trimeric Delta receptor binding domain of the SARS-CoV-2 spike protein. Notably, the cmRNA-1130 vaccine exhibits outstanding stability, remaining effective after six months of storage at 4 °C and multiple freeze-thaw cycles. In comparison with the commercial MC3 lipid, the nanoparticles formed from the degradable AX4 lipid revealed a much faster metabolic rate from the liver and spleen, affording negligible impairment to the hepatorenal function. Following intramuscular administration, cmRNA-1130 generates robust and sustained neutralizing antibodies and induces the activation of Delta RBD-specific CD4+ and CD8+ T effector memory cells (TEM) and Th1-biased T cells in mice. Featured with potent immune activation, high stability, and decent safety, vaccines formed from cmRNA and AX4 hold a huge clinical potential for the prophylaxis and treatment of different diseases.

Novel tertiary diarylethylamines as functionally selective agonists of the kappa opioid receptor.

Novel kappa opioid receptor (KOR) agonists that preferentially activate G-protein signaling versus β-arrestin-2 recruitment are described. Starting from a literature-reported phenol-containing diphenethylamine KOR agonist, structure-activity relationship (SAR) studies revealed replacement of the phenol with various non-hydroxylated bicyclic heteroaromatics led to tertiary diarylethylamines which retained KOR agonist activity and improved metabolic stability in human liver microsomes. Further optimizations produced compound 39, a potent activator of G-protein signaling (GTPγS EC50 = 14 nM, 83 % Emax) that did not elicit a β-arrestin-2 recruitment functional response (Emax < 10 %). Compound 39 demonstrated moderate to high intrinsic clearance in human hepatocytes and low potential for Pgp-mediated efflux when evaluated in the MDR1-MDCK permeability assay. Compound 39 exhibited 60- and 810-fold selectivities versus the related mu (MOR) and delta (DOR) opioid receptors in recombinant radioligand binding (Ki) assays. These findings highlight compound 39 and related structures as potential leads toward safe and tolerable therapeutics that target central nervous system (CNS) disorders for which KOR agonism could provide benefit.

Upregulation of delta opioid receptor by meningeal interleukin-10 prevents relapsing pain

Chronic pain often includes periods of transient amelioration and even remission that alternate with severe relapsing pain. While most research on chronic pain has focused on pain development and maintenance, there is a critical unmet need to better understand the mechanisms that underlie pain remission and relapse. We found that interleukin (IL)-10, a pain resolving cytokine, is produced by resident macrophages in the spinal meninges during remission from pain and signaled to IL-10 receptor-expressing sensory neurons. Using unbiased RNA-sequencing, we identified that IL-10 upregulated expression and antinociceptive activity of δ-opioid receptor (δOR) in the dorsal root ganglion. Genetic or pharmacological inhibition of either IL-10 signaling or δOR triggered relapsing pain. Overall, our findings, from electrophysiology, genetic manipulation, flow cytometry, pharmacology, and behavioral approaches, indicate that remission of pain is not simply a return to the naïve state. Instead, remission is an adapted homeostatic state associated with lasting pain vulnerability resulting from persisting neuroimmune interactions within the nociceptive system. Broadly, this sheds light on the elusive mechanisms underlying recurrence a common aspect across various chronic pain conditions.

Peripheral inflammation enhances opioid-induced gastrointestinal motility inhibition via up-regulating spinal mu opioid receptor.

Opioids are potent analgesics in clinical pain management but exert variable analgesia in different pain types. Opioid-induced constipation is a common side effect of opioid therapy, and whether opioids induce different gastrointestinal motility inhibitions in different pain types is unknown. In this study, we evaluated the antinociceptive effects and inhibition of upper gastrointestinal transit and colonic bead expulsion of morphine, DAMGO, and Deltorphin in mouse CFA chronic inflammatory pain, SNI chronic neuropathic pain, and carrageenan chronic inflammatory pain models. Furthermore, quantitative PCR and immunofluorescence were used to investigate the mechanisms underlying the altered inhibition. Results showed that intrathecal administration of morphine, DAMGO, and Deltorphin produced higher antinociceptive effects in the CFA and carrageenan groups than in the SNI group. Upper gastrointestinal transit inhibition was significantly enhanced in the carrageenan group by morphine and DAMGO; colonic bead expulsion inhibition was also enhanced in the CFA and carrageenan groups by morphine and DAMGO, but not in Deltorphin treatment. Additionally, mu (MOR) opioid receptor mRNA and MOR-expressing cell density in the lumbar spinal cord of CFA and carrageenan mice were increased, whereas delta opioid receptor expression remained unchanged in these groups. Finally, the pharmacological blockade of MOR completely prevented the enhanced upper gastrointestinal transit inhibition in the carrageenan group by morphine and DAMGO. Altogether, our results indicate that gastrointestinal motility inhibition induced by MOR agonists can be enhanced with upregulated spinal MOR expression in chronic inflammatory pain.

Ultrastructural Localization of Glutamate Delta Receptor 1 in the Rodent and Primate Lateral Habenula.

Glutamate delta receptor 1 (GluD1) is a unique synaptogenic molecule expressed at excitatory and inhibitory synapses. The lateral habenula (LHb), a subcortical structure that regulates negative reward prediction error and major monoaminergic systems, is enriched in GluD1. LHb dysfunction has been implicated in psychiatric disorders such as depression and schizophrenia, both of which are associated with GRID1, the gene that encodes GluD1. Thus, disruption in GluD1 synaptic signaling may contribute to LHb dysfunction and the pathophysiology of LHb-associated disorders. Despite its strong cellular expression, little is known about the subsynaptic and subcellular localization of GluD1 in LHb neurons. Given that GluD1 is involved in the development and/or regulation of glutamatergic and GABAergic synapses in various brain regions, a detailed map of GluD1 synaptic localization is essential to elucidate its role in the LHb. To address this issue, we used immunoelectron microscopy methods in rodents and monkeys. In both species, GluD1 immunoreactivity was primarily expressed in dendritic profiles, with lower expression in somata, spines, and glial elements. Pre- and post-embedding immunogold experiments revealed strong GluD1 expression in the core of symmetric GABAergic synapses. Albeit less frequent, GluD1 was also found at the edges (i.e., perisynaptic) of asymmetric, putative glutamatergic synapses. Through the combination of anterograde tracing with immunogold labeling in rats, we showed that axon terminals from the entopeduncular nucleus and the lateral hypothalamus express postsynaptic GluD1 immunolabeling in the LHb. Our findings suggest that GluD1 may play a critical role in modulating GABAergic transmission in the rodent and primate LHb.

Delta opioid receptor agonists activate PI3K-mTORC1 signaling in parvalbumin-positive interneurons in mouse infralimbic prefrontal cortex to exert acute antidepressant-lie effects.

The delta opioid receptor (DOP) is a promising target for novel antidepressants due to its potential for rapid action with minimal adverse effects; however, the functional mechanism underlying acute antidepressant actions remains elusive. We report that subcutaneous injection of the selective DOP agonist KNT-127 reduced immobility in the forced swimming test, and that this antidepressant-like response was reversed by intracerebroventricular injection of the selective mechanistic (or mammalian) target of rapamycin (mTOR) inhibitor rapamycin or the phosphatidylinositol-3 kinase (PI3K) inhibitor LY294002. KNT-127 also alleviated social avoidance and reduced sucrose consumption (anhedonia) among chronic vicarious social defeat stress model mice, which were similarly reversed by PI3K and mTOR inhibitors. In addition, KNT-127 increased phosphorylation levels of the mTOR signaling-related proteins Akt and p70S6 kinase in medial prefrontal cortex as revealed by immunoblotting. In the forced swimming test, a microinfusion of KNT-127 and another DOP agonist SNC80 in the infralimbic prefrontal cortex (IL-PFC) attenuated the immobility, which were blocked by rapamycin and LY294002. Perfusion of KNT-127 onto IL-PFC slices increased miniature excitatory postsynaptic current frequency and reduced miniature inhibitory postsynaptic current frequency in pyramidal neurons as measured by whole-cell patch-clamping, and both responses were reversed by rapamycin. Imaging of brain slices from transgenic mice with DOP-promoter-driven green fluorescent protein revealed that most DOPs were expressed in parvalbumin-positive interneurons in the IL-PFC. These findings suggest that DOP agonists exert antidepressant-like actions by suppressing GABA release from parvalbumin-positive interneurons via the PI3K-Akt-mTORC1-p70S6 kinase pathway, thereby enhancing IL-PFC pyramidal neuron excitation.

Indian Shot (Canna Indica L). Leaves Provide Valuable Insights into the Management of Inflammation and Other Associated Disorders Offering Health Benefits.

Throughout history, plants have played a crucial role in advancing medicinal treatments by providing a diverse range of compounds for the development of innovative therapies. Canna indica L. a tropical herb of the Cannaceae family, also known as Indian shot, has a rich history of traditional use in treating ailments like inflammation, malaria, dysentery, fever, dropsy, and diarrhea. This comprehensive research invesigates the extract preparation of C. indica leaves using multidisciplinary analytical approaches for this extract in order to shed light on its therapeutic potentials. The research, an international collaboration involving researchers from Bangladesh and China, utilized GC-MS/MS analysis to identify bioactive compounds across different C. indica extracts. Biological assays were conducted to assess antimicrobial activity using the disc diffusion method (in vitro), cytotoxicity through the brine shrimp lethality assay (in vitro), analgesic effects via the acetic acid-induced writhing test (in vivo), and antidiarrheal activity with the castor oil-induced diarrhea model (in vivo). Molecular docking studies were performed to determine binding affinities with Epidermal Growth Factor Receptor (EGFR), Dihydrofolate Reductase (DHFR), Delta Opioid Receptor (DOR), Tumor Necrosis Factor-alpha (TNF-α), and Cyclooxygenase-2 (COX-2) receptors. The GC-MS/MS analysis identified 35, 43, 27, and 20 compounds in dichloromethane, aqueous, petroleum ether, and ethyl acetate extracts, respectively. The aqueous (AQSF) and dichloromethane (DCMSF) extracts showed notable antimicrobial activity, particularly against gram-negative bacteria. Cytotoxicity tests indicated that ethyl acetate (EASF) and dichloromethane (DCMSF) fractions were potent. Analgesic activity was highest in DCMSF, and antidiarrheal effects were dose-dependent, with DCMSF showing the greatest efficacy. Molecular docking revealed strong affinities of Ergostane-3,5,6,12,25-pentol, 25-acetate, (3.beta.,5.alpha.,6.beta.,12.beta).- for EGFR and Norgestrel for COX-2. This research provides valuable insights into the bioactivity evaluation of C. indica, bridging the gap between its chemical composition and diverse biological effects. The findings contribute to the growing body of knowledge in natural product-based drug discovery and underscore the significance of C. indica as a potential source of novel therapeutic agents to treat inflammation and other disease states.

Design, Synthesis, and Evaluation of Novel (-)-cis-N-Normetazocine Derivatives: InVitro andMolecular Modeling Insights.

Suitable structural modifications of the functional groups at N-substituent of (-)-cis-N-normetazocine nucleus modulate the affinity and activity profile of related ligands toward opioid receptors. Our research group has developed several compounds and the most interesting ligands, LP1 and LP2, exhibited a dual-target profile for mu-opioid receptor (MOR) and delta-opioid receptor (DOR). Recent structure-affinity relationship studies led to the discovery of novel LP2 analogs (compounds 1 and 2), which demonstrated high MOR affinity in the nanomolar range. Here, we reported the synthesis of the new (-)-cis-N-normetazocine derivatives (3-8) characterized by the absence of the phenyl ring in the N-substituent compared to all previous reported ligands. Compounds 3 and 4, featuring a methyl ester functional group in the N-substituent, retained significant MOR affinity and exhibited similar affinity for the kappa-opioid receptor (KOR). In contrast, compounds 7 and 8, which contain a hydroxamic acid functionality, maintained affinity exclusively toward MOR. Neither of compounds (3-8) showed DOR affinity. Molecular modeling studies confirmed a similar docking pose in the MOR binding pocket for these compounds. Additionally, the in silico ADME profile of the most interesting ligands (3, 4, 7, and 8) was investigated revealing a favorable profile for compound 7 regarding the blood-brain barrier permeability, suggesting its potential as a peripherally restricted opioid ligand.

Kratom Alkaloids for the Treatment of Alcohol Use Disorder.

Alcohol use disorder (AUD) accounts for nearly 4.7% of all deaths and imposes a huge economic burden on society. Despite the magnitude of the problem, only a few Food and Drug Administration (FDA)/European Medicines Agency (EMA)-approved drugs are currently available for AUD treatment. Despite being efficacious, these drugs are not without problems, adverse effects being a major issue. That combined with medication adherence and compliance problems, the discovery of new drugs is imperative. Kratom (Mitragyna speciosa) alkaloids and some of their semisynthetic derivatives reduce alcohol intake and alcohol-induced withdrawal symptoms in animal models. These compounds act as G-protein-biased ligands at the μ-, δ-, and κ-opioid receptors, and their effect in reducing alcohol intake is mediated through the δ-opioid receptor. This article provides a critical overview of recent preclinical studies involving kratom alkaloids for AUD treatment, with a particular focus on the pharmacology and medicinal chemistry of these alkaloids. FDA/EMA approved drugs, repurposed drugs, and plant-based compounds for the treatment of AUD are briefly mentioned. Finally, important caveats and future research directions on this topic are discussed.