Phylogeography of the leaf beetle Chrysolina virgata in wetlands of Japan inferred from the distribution of mitochondrial haplotypes

The genetic differentiation among populations of the leaf beetle Chrysolina virgata living in wetlands of Japan was studied based on the sequence data of the mitochondrial cytochrome oxidase subunit I gene region (750 bp). Two distinct lineages of mitochondrial haplotypes were found: one (clade A) consisted of 26 haplotypes distributed over the distribution range of C. virgata between north‐east Honshu and Kyushu, whereas the other (clade B) was monotypic and confined to a small region in north‐east Honshu where it coexisted with clade A. Nested clade analysis for these haplotypes suggested that range expansion and following differentiation due to isolation by distance might have resulted in the present distribution pattern of the haplotypes in clade A. We discuss the evolutionary process leading to the occurrence of two distinct haplotype clades in Japan in terms of repeated colonization from the continent and range expansion and contraction during climatic changes.     

Biochemistry and neurotoxicology of guanidino compounds. History and recent advances

Guanidino compounds are known to have important biological roles, such as the participation of arginine in ureagenesis, and of creatine in muscular contraction. On the other hand, the high toxicity of guanidino compounds, such as methylguanidine and guanidine, has been under study for quite a long time in the biochemical as well as clinical fields. In this review, the author summarizes the experimental results of neurophysiological and neurochemical studies on guanidino compound-induced seizures, conducted by his colleagues since 1966, and introduces several topics arising from their recent investigations on guanidino compounds and seizure mechanism, i.e., (1) alpha-guanidino-glutaric acid in the cobalt epileptic focus and its convulsive activity; (2) guanidino-ethanesulfonic acid and epilepsy; (3) delta-guanidinovaleric acid, and endogenous and specific GABA receptor antagonist; and (4) guanidino compounds as radical generators.     

Inhibition of ovulation in PMSG/hCG-treated immature rats by rotenone, a specific inhibitor of mitochondrial oxidation

Immature Wistar rats were induced to ovulate by treatment with PMSG and hCG. Control animals ovulated 43.5 +/- 0.36 ova/rat. Intraperitoneal injection of rotenone doses of 0.125, 0.25 and 0.50 mg/kg reduced the ovulation rate to 24.0 +/- 3.08, 8.0 +/- 0.88 and 1.5 +/- 0.44 ova/rat, respectively. The rotenone significantly reduced ovarian cytochrome oxidase activity and progesterone production, but not production of oestradiol or testosterone. Thyroxine treatment at a dose of 5 mg/kg s.c. reversed the rotenone inhibition of ovulation. The results suggest that an increase in mitochondrial respiration is an essential feature of the ovulation process in mammals.     

NAD(P)H oscillation in relation to the rhythmic contraction in thePhysarum plasmodium

Summary ThePhysarum plasmodium shows rhythmic contractile activities with a period of a few min. Phases of the oscillation in the plasmodium migrating unindirectionally agreed sideways throughout at the frontal part. So, time course of an intracellular chemical component was determined by analyzing small pieces cut off successively from the frontal part of the large plasmodium. Intracellular NAD(P)H concentration oscillated with the same period as the rhythmic contraction but with a different phase advancing about 1/3 of the period. UV irradiation suppressed the rhythmic contraction without affecting the rhythmic variation of NAD(P)H. Thus, the NAD(P)H oscillator works independently of the rhythmic contractile system, but seems entraining with each other.Abbreviations UV ultraviolet - NADH nicotinamide adenine dinucleotide, reduced form - NADPH nicotinamide adenine dinucleotide phosphate, reduced form - ATP adenosine 5-triphosphate - cAMP cyclic adenosine 3, 5-monophosphate - FMNH₂ flavin mononucleotide, reduced form - TCA tricarboxylic acid - BSA bovine serum albumin - DTT dithiothreitol     

Preferential synthesis of diacyl and alkenylacyl ethanolamine and choline glycerophospholipids in rabbit platelet membranes

In rabbit platelet membranes, the contents of alkenylacyl phospholipids (plasmalogen) were 56% of phosphatidylethanolamine and 3% of phosphatidylcholine. This uneven distribution of plasmalogens in each phospholipid class could be attributed to the different substrate specificity of ethanolaminephosphotransferase (EC 2.7.8.1) and cholinephosphotransferase (EC 2.7.8.2). The properties of the enzymes were studied, using endogenous diglycerides and CDP-[3H]ethanolamine or CDP-[14C]choline as substrates. The newly formed phospholipids were mainly diacyl and alkenylacyl and only rarely alkylacyl type. The ratios of the labeled alkenylacyl to diacyl type of phospholipids clearly varied with the concentrations of CDP-ethanolamine or CDP-choline. When 1, 10, and 30 microM CDP-[3H]ethanolamine were used, the labeled phospholipids contained 53, 37, and 27% of the alkenylacyl type, respectively. The apparent Km for CDP-ethanolamine to synthesize alkenylacyl and diacyl types were 2.2 and 8.1 microM. On the other hand, when 1, 10, and 30 microM CDP-[14C]choline were used, the labeled lipids contained 10, 17, and 24% alkenylacyl type, respectively. The apparent Km for CDP-choline to synthesize alkenylacyl and diacyl types were 24 and 4.3 microM. Further, the syntheses of diacyl type of phosphatidylethanolamine and the alkenylacyl type of phosphatidylcholine were markedly inhibited by unlabeled CDP-choline and CDP-ethanolamine, respectively. The two enzymes had opposite substrate specificities, and ethanolaminephosphotransferase showed a high preference to plasmalogen synthesis, especially in the presence of CDP-choline.     

Characterization of murine monoclonal antibodies to human interferon-gamma (IFN-gamma) and their application for sandwich enzyme-linked immunosorbent assay (ELISA)

The three murine monoclonal antibodies (MAb), D1G2, D9D10, and D13C8, are specific for human interferon-gamma (IFN-gamma), but not human IFN-alpha and IFN-beta. They react weakly with heat-treated IFN-gamma. The three antibodies recognize different epitopes of the IFN-gamma molecule, as evaluated by antibody-binding inhibition experiments. We have used these three monoclonal antibodies to construct a sandwich enzyme-linked immunosorbent assay (ELISA). The best result was obtained when we used D1G2 or D9D10 MAb as a solid-phase immunosorbent and D1G2 or D9D10 MAb as a tracer. When we measured IFN-gamma in sera by a combination of D1G2 (a solid-phase) and D1G2 (a tracer), a result similar to the one by a combination of D9D10 (a solid-phase) and D1G2 (a tracer), was obtained. This may suggest that human IFN-gamma exists in oligomeric form. Recombinant human IFN-gamma expressed in E. coli is detectable at a concentration of 1 ng/ml in this sandwich ELISA. This assay can be employed for the analysis of the structural characteristics of the human IFN-gamma molecule as well as measurement of IFN-gamma in human sera and tissue culture fluids.     

Immunoregulatory abnormalities in autoimmune thyroid diseases

We have investigated the ability of lymphocytes from normal subjects and patients with autoimmune thyroid diseases to respond to a thyroidal antigen (human thyroglobulin, hTG) and a non-thyroidal antigen (Keyhole limpet hemocyanin, KLH) in vitro, using a hapten (trinitrophenol, TNP)-carrier system. This system is based on the concept that the T helper cells which respond to hTG or KLH should stimulate anti-TNP antibody producing B cells in the presence of TNP conjugated hTG (TNP-hTG) or KLH (TNP-KLH). After 5 or 6 days of culture of peripheral blood mononuclear cells with pokeweed mitogen (PWM), PWM and TNP-hTG, or PWM and TNP-KLH, IgM anti-TNP and IgM anti-sheep red blood cell (SRBC) plaque forming cells (PFC) were enumerated. The results showed that (1) in normal controls, hTG caused only suppression in both TNP and SRBC response, and KLH caused dose-related enhancement and suppression in TNP response without a change in SRBC response, and (2) in patients, both hTG and KLH resulted in dose-related enhancement in TNP response without a change in SRBC response. These data suggest that patients with autoimmune thyroid diseases have regulatory cell abnormalities confined to a thyroid antigen.