Aquatic and terrestrial cyanobacteria produce methane.

Evidence is accumulating to challenge the paradigm that biogenic methanogenesis, considered a strictly anaerobic process, is exclusive to archaea. We demonstrate that cyanobacteria living in marine, freshwater, and terrestrial environments produce methane at substantial rates under light, dark, oxic, and anoxic conditions, linking methane production with light-driven primary productivity in a globally relevant and ancient group of photoautotrophs. Methane production, attributed to cyanobacteria using stable isotope labeling techniques, was enhanced during oxygenic photosynthesis. We suggest that the formation of methane by cyanobacteria contributes to methane accumulation in oxygen-saturated marine and limnic surface waters. In these environments, frequent cyanobacterial blooms are predicted to further increase because of global warming potentially having a direct positive feedback on climate change. We conclude that this newly identified source contributes to the current natural methane budget and most likely has been producing methane since cyanobacteria first evolved on Earth.

Transcripts from amplified sequences of an inherited homogeneously staining region in chromosome 1 of the house mouse (Mus musculus).

Several populations of the house mouse, Mus musculus, are polymorphic for the presence or absence of an inherited homogeneously staining region (HSR) in chromosome 1. The HSR consists of highly amplified DNA sequences, present in low copy numbers in the HSR- genome. A cloned HSR-derived genomic sequence detected transcripts of about 1.3 and 4.5 kb on blots of poly(A)+ RNA from liver of HSR+ mice but not from that of HSR- mice. A cDNA library was established from RNA of HSR+ mice and screened with the HSR-derived genomic clone. Positive clones were isolated and shown to be complementary to the 1.3-kb RNA species and to amplified DNA sequences in the HSR+ genome. The combined sequence of four overlapping cloned cDNAs is 959 nucleotides long and includes an open reading frame encoding a putative protein of 208 amino acids. The pertinent gene is unidentified. No homologous sequence is stored in the EMBL data base. A stretch of 109 nucleotides at the 3' end of the 1.3-kb RNA homology region in the same genomic fragment, as indicated by hybridization data and sequence motifs resembling promoter elements. Thus, our data suggest that at least two genes or gene families are encoded in the HSR.

Insensitivity to pain induced by a potent selective closed-state Nav1.7 inhibitor.

Pain places a devastating burden on patients and society and current pain therapeutics exhibit limitations in efficacy, unwanted side effects and the potential for drug abuse and diversion. Although genetic evidence has clearly demonstrated that the voltage-gated sodium channel, Nav1.7, is critical to pain sensation in mammals, pharmacological inhibitors of Nav1.7 have not yet fully recapitulated the dramatic analgesia observed in Nav1.7-null subjects. Using the tarantula venom-peptide ProTX-II as a scaffold, we engineered a library of over 1500 venom-derived peptides and identified JNJ63955918 as a potent, highly selective, closed-state Nav1.7 blocking peptide. Here we show that JNJ63955918 induces a pharmacological insensitivity to pain that closely recapitulates key features of the Nav1.7-null phenotype seen in mice and humans. Our findings demonstrate that a high degree of selectivity, coupled with a closed-state dependent mechanism of action is required for strong efficacy and indicate that peptides such as JNJ63955918 and other suitably optimized Nav1.7 inhibitors may represent viable non-opioid alternatives for the pharmacological treatment of severe pain.

Aspen shaving versus chip bedding: effects on breeding and behavior.

The choice of laboratory cage bedding material is often based on both practical and husbandry issues, whereas behavioral outcomes rarely appear to be considered. It has been noted that a breeding success difference appears to be associated with the differential use of aspen chip and aspen shaving bedding in our facility; therefore, we sought to analyze breeding records maintained over a 20-month period. In fact, in all four mouse strains analyzed, shaving bedding was associated with a significant increase in average weanlings per litter relative to chip bedding. To determine whether these bedding types also resulted in differences in behaviors associated with wellbeing, we examined nest building, anxiety-like, depressive-like (or helpless-like), and social behavior in mice housed on chip versus shaving bedding. We found differences in the nests built, but no overall effect of bedding type on the other behaviors examined. Therefore, we argue that breeding success, perhaps especially in more challenging strains, is improved on shaving bedding and this is likely due to improved nest-building potential. For standard laboratory practices, however, these bedding types appear equivalent.

Regulation of rRNA metabolism in Tetrahymena pyriformis. I. Nutritional shift-down.

Changes of rates of rRNA transcription and accumulation as well as of stability of rRNA precursor fractions and mature cytoplasmic rRNAs were determined in Tetrahymena after shift-down to a non-nutrient buffer. During the initial period (4-6 hours) an enhanced degradation of pre-existing cytoplasmic rRNA and a successive reduction of rRNA transcription, processing and nucleo-cytoplasmic transport was detected. Thereafter, a "residual" rRNA metabolism at low turnover is maintained in the cells which has the following characteristics: (1) The rate of pre-rRNA transcription, as measured in vitro in isolated macronuclei, is about 3 to 5% of the rate in optimally growing cells, indicating regulatory processes at the level of initiation of the nucleolar RNA polymerases. The strong reduction of in vitro pre-rRNA synthesis can partially be reversed by pre-treatment of the starved cells with low concentrations of cycloheximide, but not with puromycin.--(2) The processing time of nuclear pre-rRNA is considerably prolonged and the introduction of the central hidden break into newly synthesized cytoplasmic 26S rRNA is strongly delayed, as shown by methionine pulse-chase experiments.--(3) The accumulation rate of cytoplasmic rRNA is 1 to 2% of the rate in optimally growing cells, as determined from the specific radioactivity of ATP at saturation with labelled exogeneous adenosine and the changes of the specific radioactivity of the AMP residues in rRNA as well as from the rRNA turnover rate.

Characterization of pre-rRNA components in ribosomal precursor particles from macronuclei of Tetrahymena thermophila.

Ribosomal precursor particles were extracted from purified macronuclei of Tetrahymena thermophila and separated in sucrose gradients. The RNA components of major particle fractions were isolated and analyzed by Northern blot hybridization using cloned rDNA fragments. A tentative scheme of preribosome maturation was established based on the RNA constituents and corresponding processing steps occurring in the different particle classes. The primary transcript of ribosomal genes, the unspliced precursor rRNA, was found in some experiments in the upper region (less than 40S) of sucrose gradients run for short times. This is in accordance with earlier results by others indicating that slowly sedimenting ribonucleoprotein (RNP) structures may exist as a transitory stage of preribosome formation. Usually, however, unspliced pre-rRNA was only found in 80S preribosomes, where splicing occurred as indicated by the presence of splice intermediates and products only in this fraction. In addition, the further processing of spliced pre-rRNA at three major sites in variable temporal order took place in the 80S preribosomes, i.e., (i) the cleavage at or near the 5'end of the 17S rRNA sequence, (ii) the central cleavage in the internal transcribed spacer (ITS2) between the 5.8S and 26S rRNA sequence, and (iii) the cleavage in the ITS1 at or near the 3' end of the 17S rRNA sequence. Only the latter event was found to result more or less immediately in the division of the 80S preribosomes into separate precursors (p40S and 60S) of the small and large ribosomal subunits. If the alternative pre-rRNA cleavage site in the ITS2 was used first the 80S preribosomes retained their integrity. The conversion of the p40S precursors into nuclear 40S subribosomal particles was correlated with the processing of pre-17S rRNA into 17S rRNA. In the 60S ribosomal precursor particles the processing of pre-26S rRNA, including the formation of precursors (ITS and 7S RNA) to 5.8S rRNA, occurred. A substantial proportion of 26S rRNA molecules isolated from these particles already contained the central hidden break as indicated by the presence of 26S rRNA alpha- and beta-subfragments. The major pre-rRNA processing by-products, IVS and ETS RNA, were partly associated with preribosomes and partly present as free RNAs in the supernatant of sucrose gradients. This indicates that they are liberated and degraded mainly outside the particles in which they are formed. In contrast, the initiation fragment (IF), a small promoter-proximal transcript, was exclusively associated with large particles.(ABSTRACT TRUNCATED AT 400 WORDS)

In vitro synthesis and processing of pre-rRNA in isolated macronuclei from Tetrahymena.

Macronuclei were isolated from logarithmically growing Tetrahymena cells in isoosmotic medium containing the weak detergent n-octanol and were purified in sucrose gradients. Electron microscopy revealed good structural preservation including intact nuclear envelopes. Initial rates of [3H]UTP incorporation into these nuclei were relatively high (2-4 pmol UMP/micrograms DNA per min), and 70 to 80% of transcription was resistant to alpha-amanitin, which is similar to the percentage of nuclear label associated with the nucleoli in electron microscopic autoradiograms. The use of transcription initiation inhibitors indicated that elongation of in vivo initiated pre-rRNA chains had essentially occurred in vitro. The radioactivity profiles of in vitro synthesized RNA in gels exhibit a heterogeneous pattern with the exception of a small peak corresponding to the length of pre-rRNA molecules. Detailed analysis of the extent and specificity of pre-rRNA processing was performed by RNA transfer hybridizations using cloned rDNA fragments as probes. The results show that the early processing events, i.e., splicing, 5'terminal and central cleavage of pre-rRNA, proceed faithfully, but at reduced rates and efficiencies. Furthermore, processing of pre-17S rRNA at the 3'end, and pre-26S rRNA at the 5'end, including the formation of immediate 5.8S rRNA precursors (ITS and 7S RNA), occurred. In contrast to previous in vivo results, a central hidden break was also introduced into part of nuclear 26S rRNA molecules. In addition to the known intermediates and by-products of processing, a large number of distinct fragments due to non-random cleavages of rRNA precursors appeared during in vitro incubation of macronuclei. Most prominent were two novel small RNA fragments from the 5'terminal end of pre-rRNA which may be products of alternative processing sites in the external transcribed spacer. Another small promoter-proximal RNA which is present in substantial amounts in vivo, was not formed under in vitro processing conditions, but degraded rapidly. This is further support to the notion that this RNA species may represent a product of premature transcription termination.

Pericardial angiosarcoma simulating pericardial effusion by echocardiography.

A fast-growing angiosarcoma caused incapacitation of a boy and death in a period of three months. The growth of the tumor was well documented by a series of echocardiograms. The heart was well encased by a thick layer of purplish vascular neoplasm enveloped mostly by thin pericardium, with some additional few foci of extracardiac metastasis. The heart weighed 2,000 gm. It is worthy to note that another cause of an echo-free space may be the presence of a pericardial tumor, rather than pericardial effusion.

Corticotropin-releasing hormone neurons in the paraventricular nucleus project to the external zone of the median eminence: a study combining retrograde labeling with immunocytochemistry.

Corticotropin-releasing hormone (CRH) is the major regulator of the pituitary-adrenal axis. CRH-immunoreactive perikarya are widely distributed in the central nervous system; however, only those which participate directly in the regulation of adrenocorticotropin are connected to the portal circulation in the external zone of the median eminence. The present study describes the identification of these hypophysiotropic neurons using retrograde labeling and CRH immunocytochemistry. Fluoro-Gold was injected peripherally then, 5 days later, the animals were treated with colchicine. Twenty-four hours later the animals were sacrificed, and their brains were immunostained for CRH with the indirect immunofluorescence technique. The results indicate that the vast majority of the Fluoro-Gold-accumulating and CRH-immunopositive perikarya (hypophysiotropic neurons) are located in the medial parvicellular subdivision of the paraventricular nucleus (PVN). However, not each CRH-immunoreactive neuron contains Fluoro-Gold, i.e. a small portion of these neurons project to areas of the brain other than the median eminence. The anterior, lateral and periventricular subdivisions of the PVN also contain hypophysiotropic CRH-immunoreactive perikarya, however, their number is much less than in the medial parvicellular subdivision. Scattered double-labeled cells are also present in the medial preoptic area and the dorsal hypothalamus, just behind the PVN. These results support previous observations that the PVN, particularly the medial parvicellular subdivision, is the predominant source of the hypophysiotropic CRH neurons.

Tyrosine phosphorylation in rat spinal cord after sciatic nerve transection.

Immunocytochemistry using antibodies against phosphotyrosine was employed to identify changes in tyrosine phosphorylation in the rat spinal cord consequent to sciatic nerve injury. Increased immunostaining in the spinal gray matter, dorsal columns and gracile nucleus on the side of the lesion became evident after 3 days and was more pronounced with longer survival times up to 3 weeks (the longest survival tested). This increase was most prominent in the fourth lumbar segment (the focus of termination of sciatic nerve afferents). Immunostaining was ain astroglial cells and their processes in the dorsal horn; stained microglia were also seen. Immunopositivity also increased in glial cells surrounding motoneurons at the same levels. These changes suggest that a diffusible growth factor released centrally by injured nerve fibers activates tyrosine phosphorylation in glial cells via receptor tyrosine kinases.

Controlled-release levodopa/benserazide (Madopar HBS): clinical observations and levodopa and dopamine plasma concentrations in fluctuating parkinsonian patients.

In five levodopa (L-dopa)-treated patients with Parkinson's disease with severe fluctuations of motor performance, plasma L-dopa as well as dopamine levels were measured during 2 days, first under optimal standard L-dopa with peripheral decarboxylase inhibitor (PDI) and then after a dose adjustment period using slow-release L-dopa/benserazide (Madopar HBS) in an open inpatient trial. Three patients benefited from the slow-release preparation; two patients deteriorated with a tendency to have an unpredictable response, a delay to turn "on" with the first dose in the morning, as well as an increase in dyskinesia corresponding to L-dopa cumulation during the day. These problems were subsequently also seen during the follow-up period of 1 year in those patients who benefited from Madopar HBS as inpatients. This might indicate that patient compliance is more difficult with the new formulation. After 1 year all patients had returned to their previous standard L-dopa/PDI treatment. L-Dopa levels continued to fluctuate, but to a lesser degree with Madopar HBS. The equivalent L-dopa dosage had to be increased by 56% (29-100%) with Madopar HBS while mean dopamine levels increased in four patients (by 47-257%) without the occurrence of peripheral side-effects. This implies that with the new formulation more L-dopa is metabolized to dopamine and explains the necessity to increase the equivalent L-dopa dosage.

Interference of biocytin with opioid-evoked hyperpolarization and membrane properties of rat spinal substantia gelatinosa neurons.

In our laboratory, preliminary whole-cell, tight seal recordings of rat spinal substantia gelatinosa neurons including biocytin in the patch pipette yielded a significantly smaller proportion of neurons hyperpolarized by selective opioid agonists compared with recordings without biocytin. Therefore, we investigated the effects of biocytin inclusion on opioid responses and other membrane properties during whole-cell, tight seal recordings of these neurons. The percentage of neurons hyperpolarized by mu-, delta(1)-, and kappa-selective opioids was significantly reduced when 1% but not < or =0.2% biocytin was included in the recording pipette, compared with neurons recorded without biocytin. However, a significantly higher proportion of neurons fired spontaneous action potentials with either 0.05-0.2 or 1% biocytin compared to no biocytin. Resting membrane potential, input impedance and the proportion of neurons displaying transient outward rectification were each significantly altered for neurons recorded with 1% but not 0.05-0.2% biocytin. These effects may be due to a relatively specific blockade of diverse potassium channel types. Because efficient labeling can be achieved with 0.1% biocytin with whole-cell recording, higher concentrations are contraindicated.

Acute rupture of the peroneal retinaculum.

The anatomy of the superior peroneal retinaculum at the ankle includes a sheath somewhat distinct from a fibrous lip extending from the periosteum of the fibula at the lateral aspect of the lip of the groove. In seventy-three cases of injury to the retinaculum, three grades of injury were recognized. Surgical repair was quite successful except in three cases.

[99m-Tc-MIBI for recurrent and metastatic differentiated thyroid carcinoma]. / 99mTc-MIBI zur Rezidiv- und Metastasensuche bei differenzierten Schilddrüsenkarzinomen.

AIM: The aim of the study has been the examination of the diagnostic value of 99mTc-MIBI scintigraphy for the detection of local tumor or metastases following total thyroidectomy and 131I ablation therapy in differentiated thyroid carcinoma. METHODS: MIBI-scintigraphy has been indicated in 85 patients because of ascending thyroglobulin values or suspected local recurrencies by ultrasonography. The results have been compared to cytology or histology or ultrasonography, computed tomography, X-ray and radioiodine scanning. RESULTS: MIBI scintigraphy was found positive in 32 of 40 metastases. Only 18 metastases have been seen by radioiodine. MIBI scintigraphy was most effective in detecting local tumor recurrencies and lymph node metastases (94%). The specificity of MIBI and radioiodine was similar (100%). In inflammatory enlarged lymph nodes no MIBI uptake was found, so it is possible to differentiate reactive lymph node enlargement from metastatic disease. CONCLUSION: In conclusion scintigraphy with 99mTc-MIBI is advisable in suspected local recurrencies and negative radioiodine scan. It is favourable that withdrawing TSH-suppressive hormone medication is not necessary.

The pathology of self-mutilation and destructive acts: a forensic study and review.

Self-destructive behavior of man and its consequences may be presented in various forms, including self-mutilation, injury, and malingering, and with various manifestations, including the Munchausen syndrome, polysurgery, purposeful accidents, impotence, and frigidity. The general subject of focal suicide has been a relatively unknown and unrecognized entity for different reasons, including the fact that most physicians are trained to approach a case in a manner which treats the patient-physician relationship as one of honor and faith regarding the authenticity of the complaints of the patient. The maturity of years and experience may be required before the physician becomes aware of the motivation of patients' behavior. It is hoped that this article will provide new insight into areas which may be unfamiliar to the forensic practitioner and clinician but which might enable them to learn about the background of an unexpected death or the possibilities of motivation in civil suits alleging professional negligence or malpractice.