Pharmacological profiles of opioid ligands at Kappa opioid receptors

BACKGROUND: The aim of the present study was to describe the activity of a set of opioid drugs, including partial agonists, in a human embryonic kidney cell system stably expressing only the mouse κ-opioid receptors. Receptor activation was assessed by measuring the inhibition of cyclic adenosine mono phosphate (cAMP) production stimulated by 5 μM forskolin. Intrinsic activities and potencies of these ligands were determined relative to the endogenous ligand dynorphin and the κ agonist with the highest intrinsic activity that was identified in this study, fentanyl. RESULTS: Among the ligands studied naltrexone, WIN 44,441 and dezocine, were classified as antagonists, while the remaining ligands were agonists. Intrinsic activity of agonists was assessed by determining the extent of inhibition of forskolin-stimulated cAMP production. The absolute levels of inhibition of cAMP production by each ligand was used to describe the rank order of intrinsic activity of the agonists; fentanyl = lofentanil ≥ hydromorphone = morphine = nalorphine ≥ etorphine ≥ xorphanol ≥ metazocine ≥ SKF 10047 = cyclazocine ≥ butorphanol > nalbuphine. The rank order of affinity of these ligands was; cyclazocine > naltrexone ≥ SKF 10047 ≥ xorphanol ≥ WIN 44,441 > nalorphine > butorphanol > nalbuphine ≥ lofentanil > dezocine ≥ metazocine ≥ morphine > hydromorphone > fentanyl. CONCLUSION: These results elucidate the relative activities of a set of opioid ligands at κ-opioid receptor and can serve as the initial step in a systematic study leading to understanding of the mode of action of these opioid ligands at this receptor

Hm1 muscarinic cholinergic receptor internalization requires a domain in the third cytoplasmic loop

Selected regions of the Hm1 muscarinic cholinergic receptor were mutated to analyze the molecular mechanisms of agonist-induced receptor internalization (or sequestration). The wild-type and mutant Hm1 genes were expressed, using pSG5, in U293 human kidney cells. Whereas surface receptor density measured with the polar tracer N-[3H]methylscopolamine was rapidly reduced by carbachol exposure, total receptor content measured with [3H]quinuclidinyl benzilate did not decline for at least 24 h, indicating the absence of extensive receptor down-regulation in U293 cells. Carbachol stimulation of phosphatidylinositol turnover paralleled receptor internalization, both with EC50 values of 10-20 µM. Furthermore, a D71N point mutation that prevented receptor activation also abolished carbachol-induced receptor internalization, indicating that receptor activation (but not necessarily second messenger stimulation) was required for internalization. Truncation of the COOH-terminal tail (K447 trunc) and point mutations of several potential Ser and Thr phosphorylation sites to Ala failed to affect receptor activation and internalization. In contrast, partial deletions of the third intracellular loop (i3) (Tyr208-Thr366) resulted in receptor mutants deficient in agonist-induced receptor internalization/sequestration. Various deletions caused either complete loss of internalization (d 232-358) or impaired internalization, ranging from 10 to 30% over 2 h, whereas wild-type Hm1 internalized to approximately ~50%. Whereas the reason for the observed differences among the deficient deletion mutants remains unclear, the initial rate of N-[3H]methylscopolamine binding loss from the cell surface was much slower than that of wild-type Hm1 in each case. The deletion of only one single domain, 284-292 (SMESLTSSE), in the middle of i3 was consistently associated with impaired internalization. Domain 284-292 is partially conserved among closely related muscarinic receptors, whereas most of the remainder of i3 is not (except for the i3 membrane junctions), and similar Ser- and Thr-rich regions are present in many other G protein-coupled receptors. We propose that a small receptor domain in the middle of the i3 loop of Hm1 is involved in agonist-induced receptor internalization

Minimum marginal abatement cost curves (Mini-MAC) for CO2 emissions reduction planning

The economic impact of CO2 emissions reduction requirements demands strategic planning to identify low-cost CO2 mitigation pathways from combinations of the many available CO2 emissions reduction options. Different tools have been developed to plan minimal cost CO2 reduction pathways taking into consideration various options such as CO2 capture, utilization, and sequestration (CCUS), shifting from fossil to renewable energy sources, as well as adopting sector-specific low emissions technologies. Current methods used to support strategic planning include high-level tools that cannot account for many possible options or fail to incorporate cost objective, and complex optimization approaches that are capable of identifying detailed low-cost solutions yet are demanding to use and often yield complex solutions in terms of processing schemes that are not easily understood by strategic planners. To address these limitations, a simple and clear methodology is proposed that allows to determine minimum cost CO2 reduction pathways from the rich set of available options. The novel methodology employs an algebraic targeting technique that yields minimum marginal abatement cost (Mini-MAC) curves to clearly represent the low-cost CO2 emissions reduction pathway available. The application of the methodology is illustrated with an example to develop minimum cost emissions reduction pathways considering CCUS, power shifting options, and negative emissions technologies. The benefits of the proposed Mini-MAC curves over alternative methods stem from their richness in terms of assessing CCUS, energy management options, and various integration options. Further, the clarity of the proposed Mini-MAC curves enables planners to easily understand available minimum cost pathways when developing strategies aimed at achieving low-cost CO2 emissions reduction. Graphical abstractOther Information Published in: Clean Technologies and Environmental Policy License: https://creativecommons.org/licenses/by/4.0See article on publisher's website: http://dx.doi.org/10.1007/s10098-021-02095-y</p

Characterization of the RDC1 Gene Which Encodes the Canine Homolog of a Proposed Human VIP Receptor Expression Does Not Correlate with an Increase in VIP Binding Sites

We have isolated a portion of the canine gene encoding the orphan receptor RDC1 [1]. The complete coding sequence is contained in a single exon, and an intron divides the 5′ untranslated region of RDC1 mRNA. The RDC1 protein is 94% homologous to the gene product of GPRN1, which has been proposed to serve as a VIP receptor when expressed in CHO-K1 and COS-7 cells (Sreedharan, S.P. et al. (1991) Proc. Natl. Acad. Sci. USA 88, 4986–4990). Northern analysis indicates that CHO-K1 cells endogenously express a 2.1 kb RDC1 mRNA. However, while CHO-K1 cells possess detectable low affinity [125I]VIP binding sites, VIP binding is not altered in membranes of CHO-K1 cells expressing varying amounts of the RDC1 gene construct. Further, endogenous VIP binding is not increased by transient expression of RDC1 in COS-7 cells. Taken together, the data suggest that RDC1 is not a canine homolog of the proposed VIP receptor

Structure and function of G protein coupled receptors

The G protein coupled receptors (GPC-Rs) comprise a large superfamily of genes encoding numerous receptors which all show common structural features, e.g., seven putative membrane spanning domains. Their biological functions are extremely diverse, ranging from vision and olfaction to neuronal and endocrine signaling. The GPC-Rs couple via multiple G proteins to a growing number of recognized second messenger pathway, e.g., cAMP and phosphatidyl inositol turnover. This review summarizes our current knowledge of the molecular mechanisms by which the GPC-Rs activate second messenger systems, and it addresses their regulation and structure

Mutations of MYO6 Are Associated with Recessive Deafness, DFNB37

Cosegregation of profound, congenital deafness with markers on chromosome 6q13 in three Pakistani families defines a new recessive deafness locus, DFNB37. Haplotype analyses reveal a 6-cM linkage region, flanked by markers D6S1282 and D6S1031, that includes the gene encoding unconventional myosin VI. In families with recessively inherited deafness, DFNB37, our sequence analyses of MYO6 reveal a frameshift mutation (36-37insT), a nonsense mutation (R1166X), and a missense mutation (E216V). These mutations, along with a previously published missense allele linked to autosomal dominant progressive hearing loss (DFNA22), provide an allelic spectrum that probes the relationship between myosin VI dysfunction and the resulting phenotype