Nitric oxide-cGMP pathway is involved in endotoxin-induced contractile dysfunction in rat hearts.

The mechanisms by which endotoxemia causes cardiac depression have not been fully elucidated. The present study examined the involvement of nitric oxide (NO) in this pathology. Rats were infused with lipopolysaccharide (LPS) or saline, and the plasma and myocardial NO(2)(-) and NO(3)(-) (NOx) concentrations were measured before or 3, 6, and 24 h after treatment. The hearts were then immediately isolated and mounted in a Langendorff apparatus, and left ventricular developed pressure (LVDP) was determined before biochemical analysis of the myocardium. LPS injection effected the expression of inducible NO synthase (iNOS) in the myocardium, a marked increase in plasma and myocardial NOx levels, and a significant decline in LVDP compared with saline controls. The LPS-induced NO production and concomitant cardiac depression were most pronounced 6 h after LPS injection and were accompanied by a significant increase in myocardial cGMP content. Myocardial ATP levels were not significantly altered after LPS injection. Significant negative correlation was observed between LVDP and myocardial cGMP content, as well as between LVDP and plasma NOx levels. Aminoguanidine, an inhibitor of iNOS, significantly attenuated the LPS-induced NOx production and contractile dysfunction. Furthermore, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, an inhibitor of soluble guanylate cyclase, significantly decreased myocardial cGMP content and attenuated the contractile depression, although aminoguanidine or 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one was not able to completely reverse myocardial dysfunction. Our data suggest that endotoxin-induced contractile dysfunction in rat hearts is associated with NO production by myocardial iNOS and a concomitant increase in myocardial cGMP.     

Immunohistochemical demonstration of c-fos protein in neurons of the medulla oblongata of the musk shrew (Suncus murinus) after veratrine administration.

We subcutaneously injected 0.5 mg/kg veratrine into the musk shrew (Suncus murinus), observed the presence or absence, latency, and the incidence of vomiting in each animal for 90 min, and selected animals that frequently vomited (FV group) and those that did not vomit (NV group). Subsequently, animal brains were removed, and the induction of c-fos protein (Fos) was immunohistochemically examined to evaluate neuronal activity in the medulla oblongata. The distribution of Fos-positive neurons in the medulla oblongata was similar between FV and NV groups, with numerous neurons along the entire length of the nucleus of the solitary tract and in the ventrolateral reticular formation. Both veratrine-injected groups showed higher numbers of positive neurons than the saline administered group. However, while the FV group showed a high concentration of positive neurons in the dorsal-dorsomedial reticular formation of the nucleus ambiguus in the rostral medulla, the NV group showed few positive neurons in this area. Fos activity in neurons in this area appeared to be higher in animals with a higher incidence of vomiting.     

An attempt to restore suitable conditions for demersal fishes and crustaceans in the Port of Sakai-Semboku, north Osaka Bay, Japan

Hypoxia and/or anoxia developed in the summer of 1995 and 1996 in the lower water layer in the Port of Sakai-Semboku, north Osaka Bay, causing an increase in sulfide content and a decrease in the redox potential of the bottom sediment. As a result, the number of benthic animal species was reduced and in the innermost part of the port no benthic animals were found. A jet stream pump system was installed at the Dejima Fishing Port, the innermost part of the Port of Sakai-Semboku to improve the oxygen conditions in the bottom waters. This led to an increase in percentage saturation of oxygen (ca. 10% increase) of the lower seawater layer along with an increase in richness and abundance of benthic fishes and crustaceans in the summer. These findings indicate that dissolved oxygen is a crucial environmental factor in the functioning of the port as a habitat for various benthic animals in north Osaka Bay. Based on the results of field surveys, the possibility of restoration of the benthic fauna by using a jet stream pump system is also discussed.     

Prediction of the coding sequences of mouse homologues of KIAA gene: II. The complete nucleotide sequences of 400 mouse KIAA-homologous cDNAs identified by screening of terminal sequences of cDNA clones randomly sampled from size-fractionated libraries.

We have accumulated information of the coding sequences of uncharacterized human genes, which are known as KIAA genes, and the number of these genes exceeds 2000 at present. As an extension of this sequencing project, we recently have begun to accumulate mouse KIAA-homologous cDNAs, because it would be useful to prepare a set of human and mouse homologous cDNA pairs for further functional analysis of the KIAA genes. We herein present the entire sequences of 400 mouse KIAA cDNA clones and 4 novel cDNA clones which were incidentally identified during this project. Most of clones entirely sequenced in this study were selected by computer-assisted analysis of terminal sequences of the cDNAs. The average size of the 404 cDNA sequences reached 5.3 kb and that of the deduced amino acid sequences from these cDNAs was 868 amino acid residues. The results of sequence analyses of these clones showed that single mouse KIAA cDNAs bridged two different human KIAA cDNAs in some cases, which indicated that these two human KIAA cDNAs were derived from single genes although they had been supposed to originate from different genes. Furthermore, we successfully mapped all the mouse KIAA cDNAs along the genome using a recently published mouse genome draft sequence.     

Prevalence of Pandemic Thermostable Direct Hemolysin-Producing Vibrio parahaemolyticus O3:K6 in Seafood and the Coastal Environment in Japan

Although thermostable direct hemolysin (TDH)-producing Vibrio parahaemolyticus has caused many infections in Asian countries, the United States, and other countries, it has been difficult to detect the same pathogen in seafoods and other environmental samples. In this study, we detected and enumerated tdh gene-positive V. parahaemolyticus in Japanese seafoods with a tdh-specific PCR method, a chromogenic agar medium, and a most-probable-number method. The tdh gene was detected in 33 of 329 seafood samples (10.0%). The number of tdh-positive V. parahaemolyticus ranged from <3 to 93/10 g. The incidence of tdh-positive V. parahaemolyticus tended to be high in samples contaminated with relatively high levels of total V. parahaemolyticus. TDH-producing strains of V. parahaemolyticus were isolated from 11 of 33 tdh-positive samples (short-necked clam, hen clam, and rock oyster). TDH-producing strains of V. parahaemolyticus were also isolated from the sediments of rivers near the coast in Japan. Representative strains of the seafood and sediment isolates were examined for the O:K serovar and by the PCR method specific to the pandemic clone and arbitrarily primed PCR and pulsed-field gel electrophoresis techniques. The results indicated that most O3:K6 tdh-positive strains belonged to the pandemic O3:K6 clone and suggested that serovariation took place in the Japanese environment.     

Cultivation and biogeochemical analyses reveal insights into methanogenesis in deep subseafloor sediment at a biogenic gas hydrate site

Gas hydrates deposited in subseafloor sediments are considered to primarily consist of biogenic methane. However, little evidence for the occurrence of living methanogens in subseafloor sediments has been provided. This study investigated viable methanogen diversity, population, physiology and potential activity in hydrate-bearing sediments (1–307 m below the seafloor) from the eastern Nankai Trough. Radiotracer experiments, the quantification of coenzyme F430 and molecular sequencing analysis indicated the occurrence of potential methanogenic activity and living methanogens in the sediments and the predominance of hydrogenotrophic methanogens followed by methylotrophic methanogens. Ten isolates and nine representative culture clones of hydrogenotrophic, methylotrophic and acetoclastic methanogens were obtained from the batch incubation of sediments and accounted for 0.5–76% of the total methanogenic sequences directly recovered from each sediment. The hydrogenotrophic methanogen isolates of Methanocalculus and Methanoculleus that dominated the sediment methanogen communities produced methane at temperatures from 4 to 55 °C, with an abrupt decline in the methane production rate at temperatures above 40 °C, which is consistent with the depth profiles of potential methanogenic activity in the Nankai Trough sediments in this and previous studies. Our results reveal the previously overlooked phylogenetic and metabolic diversity of living methanogens, including methylotrophic methanogenesis.Subject terms: Biogeochemistry, Biodiversity, Environmental microbiology     

Roles of Abscisic Acid and Indoleacetic Acid in the Stunted Growth of Water-Stressed, Etiolated Squash Hypocotyls

The endogenous contents of indoleacetic acid (IAA) and abscisicacid (ABA) were assayed simultaneously in etiolated squash hypocotylsthat had been subjected to water stress applied to squash rootsby immersing them in 60 mM polyethylene glycol solution. Thegrowth rate and suction force of the hypocotyl under this water-stresscondition were determined for comparison with the two hormonecontents. Water stress retarded hypocotyl growth and increased the suctionforce of hypocotyl cells, as the ABA content increased, by 5-foldof the initial content one day after treatment with polyethyleneglycol. The ABA contents thereafter remained high up to day4. The increase in ABA content was detected in stressed hypocotylas early as 6 h after treatment, when water stress had not affectedhypocotyl growth. When the water stress was terminated one dayafter treatment began, hypocotyl growth immediately recovered,and the ABA content decreased to the value for unstressed hypocotylsin one day. The IAA content gradually decreased in unstressedhypocotyls, but was maintained at a high value in stressed hypocotyls. Logarithmic concentrations of endogenous ABA (ng/g fr wt) werecorrelated significantly with the suction force of the hypocotyls(r=0.92) and with the growth rates (r=–0.78). IAA contentwas correlated neither with the growth rate nor the suctionforce. These results suggest that the endogenous ABA contentwas associated with the stunted growth of the etiolated squashhypocotyls produced by water stress but that the endogenousIAA content did not have an important role in growth regulationunder water stress conditions. (Received April 11, 1984; Accepted October 2, 1984)     

SIK2 is a key regulator for neuronal survival after ischemia via TORC1-CREB

The cAMP responsive element-binding protein (CREB) functions in a broad array of biological and pathophysiological processes. We found that salt-inducible kinase 2 (SIK2) was abundantly expressed in neurons and suppressed CREB-mediated gene expression after oxygen-glucose deprivation (OGD). OGD induced the degradation of SIK2 protein concomitantly with the dephosphorylation of the CREB-specific coactivator transducer of regulated CREB activity 1 (TORC1), resulting in the activation of CREB and its downstream gene targets. Ca(2+)/calmodulin-dependent protein kinase I/IV are capable of phosphorylating SIK2 at Thr484, resulting in SIK2 degradation in cortical neurons. Neuronal survival after OGD was significantly increased in neurons isolated from sik2(-/-) mice, and ischemic neuronal injury was significantly reduced in the brains of sik2(-)(/-) mice subjected to transient focal ischemia. These findings suggest that SIK2 plays critical roles in neuronal survival, is modulated by CaMK I/IV, and regulates CREB via TORC1.