In Vivo Regulation of the μ Opioid Receptor: Role of the Endogenous Opioid Agents

It is well known that genotypic differences can account for the subject-specific responses to opiate administration. In this regard, the basal activity of the endogenous system (either at the receptor or ligand level) can modulate the effects of exogenous agonists as morphine and vice versa. The μ opioid receptor from zebrafish, dre-oprm1, binds endogenous peptides and morphine with similar affinities. Morphine administration during development altered the expression of the endogenous opioid propeptides proenkephalins and proopiomelanocortin. Treatment with opioid peptides (Met-enkephalin [Met-ENK], Met-enkephalin-Gly-Tyr [MEGY] and β-endorphin [β-END]) modulated dre-oprm1 expression during development. Knocking down the dre-oprm1 gene significantly modified the mRNA expression of the penk and pomc genes, thus indicating that oprm1 is involved in shaping penk and pomc expression. In addition, the absence of a functional oprm1 clearly disrupted the embryonic development, since proliferation was disorganized in the central nervous system of oprm1-morphant embryos: mitotic cells were found widespread through the optic tectum and were not restricted to the proliferative areas of the mid- and hindbrain. Transferase-mediated dUTP nick-end labeling (TUNEL) staining revealed that the number of apoptotic cells in the central nervous system (CNS) of morphants was clearly increased at 24-h postfertilization. These findings clarify the role of the endogenous opioid system in CNS development. Our results will also help unravel the complex feedback loops that modulate opioid activity and that may be involved in establishing a coordinated expression of both receptors and endogenous ligands. Further knowledge of the complex interactions between the opioid system and analgesic drugs will provide insights that may be relevant for analgesic therapy.     

Muscarinic receptor-linked elevation of cAMP in SH-SY5Y neuroblastoma cells is mediated by Ca2+ and protein kinase C.

The mechanisms of muscarinic receptor-linked increase in cAMP accumulation in SH-SY5Y human neuroblastoma cells has been investigated. The dose-response relations of carbachol-induced cAMP synthesis and carbachol-induced rise in intracellular free Ca2+ were similar. The stimulated cAMP synthesis was inhibited by about 50% when cells were entrapped with the Ca2+ chelator BAPTA or in the presence of the protein kinase C (PKC) inhibitor staurosporine. Production of cAMP could be induced also by the Ca2+ ionophore, ionomycin and by TPA, an activator of PKC. When added together TPA and ionomycin had a synergistic effect. When cAMP synthesis was activated with cholera toxin, PGE1 or PGE1 + pertussis toxin carbachol stimulated cAMP production to the same extent as in control cells. Ca2+ and protein kinase C thus seem to be the mediators of muscarinic-receptor linked cAMP synthesis by a direct action on adenylate cyclase.     

Reproductive pattern among women in 19th century Sweden

Among the married female population in rural areas of 19th century Sweden infertility and subfertility were found in 7.5% and 6.1% respectively with increasing incidence during the latter half of the century. In relation to the age group 20-24, fecundity declined by 42% and 92% in the age groups 35-39 and 40-45 respectively. One-third of married women died before age 50. Re-marriages and step-parents were common. Less than half of the married women brought up the majority of the children in the community.     

Interphase epichromatin: last refuge for the 30-nm chromatin fiber?

“Interphase epichromatin” describes the surface of chromatin located adjacent to the interphase nuclear envelope. It was discovered in 2011 using a bivalent anti-nucleosome antibody (mAb PL2-6), now known to be directed against the nucleosome acidic patch. The molecular structure of interphase epichromatin is unknown, but is thought to be heterochromatic with a high density of “exposed” acidic patches. In the 1960s, transmission electron microscopy of fixed, dehydrated, sectioned, and stained inactive chromatin revealed “unit threads,” frequently organized into parallel arrays at the nuclear envelope, which were interpreted as regular helices with ~ 30-nm center-to-center distance. Also observed in certain cell types, the nuclear envelope forms a “sandwich” around a layer of closely packed unit threads (ELCS, envelope-limited chromatin sheets). Discovery of the nucleosome in 1974 led to revised helical models of chromatin. But these models became very controversial and the existence of in situ 30-nm chromatin fibers has been challenged. Development of cryo-electron microscopy (Cryo-EM) gave hope that in situ chromatin fibers, devoid of artifacts, could be structurally defined. Combining a contrast-enhancing phase plate and cryo-electron tomography (Cryo-ET), it is now possible to visualize chromatin in a “close-to-native” situation. ELCS are particularly interesting to study by Cryo-ET. The chromatin sheet appears to have two layers of ~ 30-nm chromatin fibers arranged in a criss-crossed pattern. The chromatin in ELCS is continuous with adjacent interphase epichromatin. It appears that hydrated ~ 30-nm chromatin fibers are quite rare in most cells, possibly confined to interphase epichromatin at the nuclear envelope.

     

Antigens in immune complexes from patients with breast cancer

Summary Sera and effusion fluids of patients with breast cancer (BC) contain immune complexes (IC). Antigens present in these complexes were isolated as follows: a pool of effusions from patients with BC was fractionated with ammonium sulfate. The proteins precipitating at 40% saturation were further fractionated by filtration through a Sephadex G-200 column. The material recovered in the first peak (molecules larger than monomeric IgG) was brought to pH 3.0 to dissociate the IC, and the mixture was filtered through a column of Sephacryl S-300 at pH 3.0. Proteins smaller than monomeric IgG were collected, radioiodinated, and used as antigens (¹²⁵Ag) to search for corresponding antibodies in sera of patients with BC (BCS) and of healthy individuals (NHS). ¹²⁵Ag was reacted with the sera and the immune complexes obtained were precipitated with an antiserum to human Ig and analyzed by SDS-polyacrylamide gel electrophoresis followed by autoradiography. Both NHS and BCS contained antibodies against two antigens; one of these appeared as a strong band of 17KD, the other as a doublet of approximately 25KD. It is concluded that some of the proteins in the IC from patients with BC are auto-antigens. No BC-specific antigens were identified.