Ultrastructural studies of endocrine-like cells in the fundic gastric mucosa of the bullfrog,Rana catesbeiana

Summary This study is concerned with electron-microscopic observations on endocrine or paracrine cells in the fundic gastric mucosa of the bullfrog. Also, an attempt was made to identify the histamine-releasing cells involved in the secretagogue response. At least three distinct endocrine-like cell types were found. The classification is based on the appearance of secretory granules and other organelles, and the relationship of endocrine-like cells with other cells in the tissue. The amphibian endocrine-like cells resemble the ECL, D and EC cells of mammals. Type-I (ECL) cells showed degranulation after repeated stimulation with tetragastrin (TG), acetylcholine (ACh) and K⁺ depolarizing solution, all of which release histamine.     

AFDX-116 discriminates between muscarinic M2 receptors of the heart and the iris smooth muscle

Summary Studies with the atypical muscarinic antagonist pirenzepine provide convincing evidence for the classification of muscarinic acetylcholine receptors (mAChRs) into two subtypes, M₁ and M₂. The present study examines the heterogeneity of the M₂ subtype employing the newly developed competitive muscarinic antagonist, AFDX-116. Comparison of the binding affinities of pirenzepine, atropine, and AFDX-116 to mAChRs in microsomes from the rabbit cerebral cortex, heart, and iris smooth muscle shows that iris mAChRs, which are pharmacologically of the M₂ subtype, can be distinguished from M₂ cardiac receptors based on their affinity for AFDX-116. These results are consistent with the hypothesis that the M₂ receptor subtype consists of a heterogeneous population of receptors.Abbreviations mAChRs Muscarinic Acetylcholine Receptors - CCh Carbachol - NMS N-Methylscopolamine - AFDX-116 11-[[2-[(diethylamino)methyl]-1-piperidinyl]acetyl]-5,11-dihydro-6Hpyrido[2,3-b][1,4]benzodiazepine-6-one     

Human acetylcholinesterase and butyrylcholinesterase are encoded by two distinct genes

1. Various hybridization approaches were employed to investigate structural and chromosomal interrelationships between the human cholinesterase genes CHE and ACHE encoding the polymorphic, closely related, and coordinately regulated enzymes having butyrylcholinesterase (BuChE) and acetylcholinesterase (AChE) activities. 2. Homologous cosmid recombination with a 190-base pair 5' fragment from BuChEcDNA resulted in the isolation of four overlapping cosmid clones, apparently derived from a single gene with several introns. The Cosmid CHEDNA included a 700-base pair fragment known to be expressed at the 3' end of BuChEcDNA from nervous system tumors and which has been mapped by in situ hybridization to the unique 3q26-ter position. In contrast, cosmid CHEDNA did not hybridize with full-length AChEcDNA, proving that the complete CHE gene does not include AChE-encoding sequences either in exons or in its introns. 3. The chromosomal origin of BuChE-encoding sequences was further examined by two unrelated gene mapping approaches. Filter hybridization with DNA from human/hamster hybrid cell lines revealed BuChEcDNA-hybridizing sequences only in cell lines including human chromosome 3. However, three BuChEcDNA-homologous sequences were observed at chromosomal positions 3q21, 3q26-ter, and 16q21 by a highly stringent in situ hybridization protocol, including washes at high temperature and low salt. 4. These findings stress the selectivity of cosmid recombination and chromosome blots, raise the possibility of individual differences in BuChEcDNA-hybridizing sequences, and present an example for a family highly similar proteins encoded by distinct, nonhomologous genes.     

Coexistence of cytoplasmic and nuclear genes for male sterility in Petunia

Summary Cytoplasmic male sterility (cms) and nuclear male sterility (nms) in Petunia were described respectively as possible autonomous and integrated states of the same genetic element by Frankel (1971). In the present study we describe genetic analysis of the interaction between the cms, the nuclear gene for male sterility (e) and the fertility restorer allele (Rf). The main findings in this study are: (1) The nuclear sterility allele can coexist in one or two dosages with the cytoplasmic male sterility elements (ste) in somatic cells or female gametes; (2) the presence of the fertility restorer allele Rf is not required for the coexistence of ste and e and (3) Rf does not interact epistatically with e, e.g., the expression of e is independent of Rf—the genotypes (S) RfRfee and (S) Rfrfee are male sterile.Contribution from the Agricultural Research Organization. The Volcani Center, Bet Dagan, Israel. 1983 series No. 846 E     

Molecular phylogenetic study of flavonoids in medicinal plants: a case study family Apiaceae

Journal of Plant Research - The current study examined the phylogenetic pattern of medicinal species of the family Apiaceae based on flavonoid groups production, as well as the overall mechanism of...     

Properties of monovalent nickel complexes with tetraaza-macrocyclic ligands in aqueous solutions

The effects of N-alkylation on the redox potential of the couples NiLⁱ²⁺/NiLⁱ⁺, L = tetraaza-14-membered-macrocyclic ligands, and on the properties of the monovalent nickel complexes in aqueous solutions are reported for 14 complexes. The spectra and lifetimes of the NiLⁱ⁺ complexes are reported. The self-exchange rates for the couples NiLⁱ²⁺/NiLⁱ⁺ were determined. Two of the ligands were synthesized for the first time for this study. Cyclic voltammetry and pulse radiolysis were used. The results point out that: (i) N-alkylation always shifts the redox potential to a less cathodic one; this effect stems to a large degree from the decrease in the solvation energy of the complex caused by the N-alkylation of the ligand. (ii). The lifetime of the monovalent complexes is not linearly related to the redox potential of the NiLⁱ²⁺/NiLⁱ⁺ couples. (iii) The NiLⁱ⁺ complexes exist in several isomeric forms; the rate of the isomerization depends on the structure of the ligand. (iv) Different isomers of NiLⁱ⁺ may be formed when the complex NiLⁱ²⁺ is reduced by different reagents; therefore, the pulse-radiolytically formed NiLⁱ⁺ complexes might have different properties than those formed electrochemically.     

Requirement for calcium ions in acetylcholine-stimulated phosphodiesteratic cleavage of phosphatidyl-myo-inositol 4,5-bisphosphate in rabbit iris smooth muscle

1. The mechanism of acetylcholine-stimulated breakdown of phosphatidyl-myo-inositol 4,5-bisphosphate and its dependence on extracellular Ca(2+) was investigated in the rabbit iris smooth muscle. 2. Acetylcholine (50mum) increased the breakdown of phosphatidylinositol bisphosphate in [(3)H]inositol-labelled muscle by 28% and the labelling of phosphatidylinositol by 24% of that of the control. Under the same experimental conditions there was a 33 and 48% increase in the production of (3)H-labelled inositol trisphosphate and inositol monophosphate respectively. Similarly carbamoylcholine and ionophore A23187 increased the production of these water-soluble inositol phosphates. Little change was observed in the (3)H radioactivity of inositol bisphosphate. 3. Both inositol trisphosphatase and inositol monophosphatase were demonstrated in subcellular fractions of this tissue and the specific activity of the former was severalfold higher than that of the latter. 4. The acetylcholine-stimulated production of inositol trisphosphate and inositol monophosphate was inhibited by atropine (20mum), but not tubocurarine (100mum); and it was abolished by depletion of extracellular Ca(2+) with EGTA, but restored on addition of low concentrations of Ca(2+) (20mum). 5. Calcium-antagonistic agents, such as verapamil (20mum), dibenamine (20mum) or La(3+) (2mm), also abolished the production of the water-soluble inositol phosphates in response to acetylcholine. 6. Release of inositol trisphosphate from exogenous phosphatidylinositol bisphosphate by iris muscle microsomal fraction (;microsomes') was stimulated by 43% in the presence of 50mum-Ca(2+). 7. The results indicate that increased Ca(2+) influx into the iris smooth muscle by acetylcholine and ionophore A23187 markedly activates phosphatidylinositol bisphosphate phosphodiesterase and subsequently increases the production of inositol trisphosphate and its hydrolytic product inositol monophosphate. The marked increase observed in the production of inositol monophosphate could also result from Ca(2+) activation of phosphatidylinositol phosphodiesterase. However, there was no concomitant decrease in the (3)H radioactivity of this phospholipid.     

Functional membrane vesicles from the nervous system of insects: I. Sodium- and chloride-dependent γ-aminobutyric acid transport

(1) A synaptosomal fraction obtained from locust nervous tissue has been shown to possess an active γ-aminobutyric acid transport mechanism. This activity is preserved and even enriched by the membrane vesicles derived from osmotically shocked synaptosomes. (2) Electron-microscopy examination indicates that the above membrane vesicles are derived predominantly from the neuronal plasma membrane and are devoid of any internal cellular organelles and components. Active transport of γ-aminobutyric acid into these vesicles has been demonstrated with artificially imposed ion gradients as the sole energy source. (3) γ-Aminobutyric acid transport can be driven by an Na⁺ gradient (out>in) and /or by a gradient of Cl^? (out>in). This process is absolutely dependent on the simultaneous presence of both types of ion in the external medium. The stimulation of the process by valinomycin indicates that γ-aminobutyric acid transport is an electrogenic process which is stimulated by a membrane potential (interior negative).     

The ultrastructure of normal and glycerol treated muscle in the ghost crab,Ocypode cursor

The ultrastructure of normal and glycerol treated fibers of the closer muscle of the ghost crab, Ocypode cursor, was studiedmthe muscle is composed of presumably phasic (short sarcomeres) and tonic (long sarcomeres) fibers, the latter greatly predominating. Horseradish peroxidase (HRP) was used as an extracellular tracer to delineate the tubular system (TS), and to determine to what extent this system becomes detached from the extracellular space as a result of glycerol treatment. Sarcolemmal clefts invade deeply into the muscle at Z-lines and I-bands; tubules invaginate into the muscle from the clefts and from the surface sarcolemma at the Z-lines, A-I overlaps and A-bands. A tubules are in frequent diadic or tetradic contact with the sarcoplasmic reticulum (SR), whereas Z tubules appear to be randomly associated with SR, terminal cisterns (TC) and Z-line fibrils. When HRP was administered to normal muscle, black reaction product was found adjacent to the outer surface of the sarcolemma, within the clefts and within profiles of the TS throughout the tissue. In glycerol treated muscle peripheral vacuolation frequently occurred; black reaction product penetrated only as far as the vacuoles and into dilated Z-line tubules, but was virtually absent from the rest of the TS. This lack of continuity between the extracellular space and the A tubules indicated disruption or constriction of the A tubules as a result of glycerol treatment, although Z tubule contact with the extracellular space appeared unimpaired. These findings provide ultrastructural correlates of the electrophysiological changes produced by glycerol treatment of the closer muscle of the ghost crab (Papir, 1973), namely, interference with excitation-contraction (e-c) coupling. The random association of the Z tubules with SR and TC, and their resistance to disruption by glycerol treatment, tend to endorse the claims that the Z tubules in crustacean muscle are not directly involved in e-c coupling (Brandt et al., 1965; Peachey, 1967; Selverston, 1967).     

Time-resolved fluorescence of 2-aminopurine as a probe of base flipping in M.HhaI-DNA complexes

DNA base flipping is an important mechanism in molecular enzymology, but its study is limited by the lack of an accessible and reliable diagnostic technique. A series of crystalline complexes of a DNA methyltransferase, M.HhaI, and its cognate DNA, in which a fluorescent nucleobase analogue, 2-aminopurine (AP), occupies defined positions with respect the target flipped base, have been prepared and their structures determined at higher than 2 Å resolution. From time-resolved fluorescence measurements of these single crystals, we have established that the fluorescence decay function of AP shows a pronounced, characteristic response to base flipping: the loss of the very short (~100 ps) decay component and the large increase in the amplitude of the long (~10 ns) component. When AP is positioned at sites other than the target site, this response is not seen. Most significantly, we have shown that the same clear response is apparent when M.HhaI complexes with DNA in solution, giving an unambiguous signal of base flipping. Analysis of the AP fluorescence decay function reveals conformational heterogeneity in the DNA–enzyme complexes that cannot be discerned from the present X-ray structures.