Effects of chronic exposure to 12‰ saltwater on the endocrine physiology of juvenile American alligator (Alligator mississippiensis).

American alligator (Alligator mississippiensis) habitats are prone to saltwater intrusion following major storms, hurricanes or droughts. Anthropogenic impacts affecting hydrology of freshwater systems may exacerbate saltwater intrusion into freshwater habitats. The endocrine system of alligators is susceptible to changes in the environment but it is currently not known how the crocodilian physiological system responds to environmental stressors such as salinity. Juvenile alligators were exposed to 12‰ saltwater for 5 weeks to determine the effects of chronic exposure to saline environments. Following 5 weeks, plasma levels of hormones [e.g. progesterone, testosterone, estradiol, corticosterone, aldosterone (ALDO), angiotensin II (ANG II)] were quantified using liquid chromatography and tandem mass spectrometry. Compared with freshwater-kept subjects, saltwater-exposed alligators had significantly elevated plasma levels of corticosterone, 11-deoxycortisol, 17α-hydroxyprogesterone, testosterone, 17ß-estradiol, estrone and estriol whereas pregnenolone and ANG II were significantly depressed and ALDO levels were unchanged (slightly depressed). On the one hand, saltwater exposure did not affect gene expression of renal mineralocorticoid and glucorticoid and angiotensin type 1 (AT-1) receptors or morphology of lingual glands. On the other hand, saltwater exposure significantly reduced plasma glucose concentrations whereas parameters diagnostic of perturbed liver function (aspartate aminotransferase and alanine aminotransferase) and kidney function (creatinine and creatine kinase) were significantly elevated. Except for plasma potassium levels (K+), plasma ions Na+ and Cl- were significantly elevated in saltwater alligators. Overall, this study demonstrated significant endocrine and physiological effects in juvenile alligators chronically exposed to a saline environment. Results provide novel insights into the effects of a natural environmental stressor (salinity) on the renin-angiotensin-aldosterone system and steroidogenesis of alligators.

Measuring endogenous changes in serotonergic neurotransmission in humans: a [11C]CUMI-101 PET challenge study.

Serotonin (5-HT) neurotransmission is implicated in cognitive and emotional processes and a number of neuropsychiatric disorders. The use of positron emission tomography (PET) to measure ligand displacement has allowed estimation of endogenous dopamine release in the human brain; however, applying this methodology to assess central 5-HT release has proved more challenging. The aim of this study was to assess the sensitivity of a highly selective 5-HT(1A) partial agonist radioligand [(11)C]CUMI-101 to changes in endogenous 5-HT levels induced by an intravenous challenge with the selective 5-HT re-uptake inhibitor (SSRI), citalopram, in healthy human participants. We studied 15 healthy participants who underwent PET scanning in conjunction with [(11)C]CUMI-101 after receiving an intravenous infusion of citalopram 10 mg or placebo in a double-blind, crossover, randomized design. Regional estimates of binding potential (BP(ND)) were obtained by calculating total volumes of distribution (V(T)) for presynaptic dorsal raphe nucleus (DRN) and postsynaptic cortical regions. Relative to placebo, citalopram infusion significantly increased [(11)C]CUMI-101 BP(ND) at postsynaptic 5-HT(1A) receptors in several cortical regions, but there was no change in binding at 5-HT(1A) autoreceptors in the DRN. Across the postsynaptic brain regions, citalopram treatment induced a mean 7% in [(11)C]CUMI-101 BP(ND) (placebo 1.3 (0.2); citalopram 1.4 (0.2); paired t-test P=0.003). The observed increase in postsynaptic [(11)C]CUMI-101 availability identified following acute citalopram administration could be attributable to a decrease in endogenous 5-HT availability in cortical terminal regions, consistent with preclinical animal studies, in which acute administration of SSRIs decreases DRN cell firing through activation of 5-HT(1A) autoreceptors to reduce 5-HT levels in postsynaptic regions. We conclude that [(11)C]CUMI-101 may be sensitive to changes in endogenous 5-HT release in humans.

Identification and characterization of the v-cath gene of the baculovirus, CfMNPV.

The v-cath gene of the Autographa californica multi-nucleocapsid nuclear polyhedrosis virus (AcMNPV) encodes a cathepsin L-like proteinase which plays a role in the liquefaction of host tissues during a viral infection [1]. We have identified a homologous gene in the spruce budworm virus, Choristoneura fumiferana MNPV (CfMNPV). The CfMNPV v-cath gene is 74% identical to AcMNPV v-cath at the nucleotide sequence level and 80% identical at the level of predicted amino acid sequence. Transcription analysis of the CfMNPV v-cath gene revealed that it is expressed late in infection and that transcription initiates within the consensus baculovirus late-promoter motif.

Nucleotide sequence of the p74 gene of a baculovirus pathogenic to the spruce budworm, Choristoneura fumiferana multicapsid nuclear polyhedrosis virus.

Polypeptide p74 has been found to be essential for production of virulent occlusion bodies of the baculovirus Autographa californica multicapsid nuclear polyhedrosis virus (AcMNPV). Hybridization with AcMNPV-derived probes has led to the location of the p74 gene in the spruce budworm virus, Choristoneura fumiferana MNPV. Sequence data indicate that CfMNPV p74 is 73% identical to AcMNPV at the nucleotide level and 77% identical at the amino acid level. Elements of predicted secondary structure are also conserved.

Recombinant beta-galactosidase production in serum-free medium by insect cells in a 14-L airlift bioreactor.

Spodoptera frugiperda (Sf9) insect cells were successfully cultured in serum-free medium in a 14-L airlift bioreactor. Cell densities as high as 1 x 10(7) cells/mL were achieved with specific growth rates of approximately 0.0286 h-1 (doubling time of 24 h). This system was also used to demonstrate the expression of a reported gene, beta-galactosidase (beta-gal), when cells were infected with a recombinant baculovirus. Approximately 0.33 mg of beta-gal/mL (i.e., 104,000 units/mL) of medium were obtained at the 14-L scale, while about 0.95 mg of beta-gal/mL (i.e., 285,000 units/mL) of medium were obtained in small-scale shaker flasks. The difference was attributed to a suboptimal infection in the large scale. Specific oxygen consumption rates decreased from 5.58 x 10(-17) mol O2/cell.s in early exponential growth to 3.13 x 10(-17) mol O2/cell.s at 3 days post-infection.

Protective effects of polymer additives on animal cells exposed to rapidly falling liquid films.

The protective effects of polymer additives on insect cells against fluid mechanical damage was investigated in a falling film-flow device. The falling liquid film creates rapidly moving air-liquid interfaces and high fluid shear stress, mimicking the characteristics of a bursting bubble in aerated cell culture. The additives tested included a group of surface-active polymers, (i.e., Pluronic F68, poly(ethylene glycol)s, and Tween 80) and a group of viscosity-enhancing polymers (i.e., dextrans, methyl-cellulose, and (carboxymethyl)cellulose). We found that methylcellulose, which was previously considered a viscosity-enhancing polymer, actually had significant surface-active properties. All of the surface-active polymers exhibited significant protective effects, with Pluronic F68 and the higher molecular weight poly(ethylene glycol), PEG 20M, providing the best protection. In contrast, the viscosity-enhancing polymers, with the exception of methylcellulose, showed little or no protection for insect cells in the film flow. All of the protective polymers had surface-active properties, even though some of them did not change the surface tension in the actual insect cell medium. There was no correlation between the protective effect and the changes in liquid viscosity and surface tension due to the polymer additives. The level of protection was shown to be dependent upon the type of polymer, its concentration in the culture medium, and the polymer molecular weight. We concluded that the mechanism of protection of these surface-active polymers was through interaction of the polymer molecules with the cell plasma membranes: a fast-acting biological mechanism.

Determination of SB 216469-S during tablet production using near-infrared reflectance spectroscopy.

A near-infrared method was developed for analyzing SB 216469-S tablets at various stages of tablet processing, particularly after (i) high shear granulation, (ii) lubrication, (iii) core tablet compression, and (iv) aqueous film coating. Tablets with three different drug concentrations ranging from 1.5% (w/w) to 6.0% (w/w) were examined along with a placebo. Similarly, moisture levels during the granulation drying process were measured, along with the thickness of the tablet coating. Tablet identification inside blister packaging for clinical supplies was also demonstrated.

Predicting the unpredictable: issues for output-based funding in psychiatric services.

In this study a number of important issues relating to diagnosis related groups and psychiatry are highlighted. Retrospective analysis of admissions to a rural psychiatric admission unit was undertaken utilising two differing approaches to statistical analysis. Results revealed that diagnosis alone is not highly predictive of cost. However, when linked to other variables, greater explanation in variation of length of stay was achieved. Data from social variables yielded sufficiently interesting results to warrant further investigation. The authors conclude by reporting the resultant groups from the classification analysis and offer some suggestions for further research.

The development of a mental health service patient information management system.

In this paper we describe the development of the Mental Health-Patient Information Management System (MH-PIMS), which is an information management system designed for use in a modern, primarily community-based, mental health service. MH-PIMS is a computerised database which was designed by clinicians and is supported by a case management system and complementary patient record set-together called the Assessment and Care Evaluation (ACE) system. The paper also describes the ACE system. MH-PIMS can generate patient reports of use to case managers and teams in managing their caseloads and is of use to senior clinicians and service managers for audit and strategic planning purposes.

A national dosimetry audit of intraoperative radiotherapy.

OBJECTIVE: National dosimetry audits are a fundamental part of quality assurance in radiotherapy, especially for new techniques. Intraoperative radiotherapy with a compact mobile kilovoltage X-ray source is a novel approach for the treatment of breast and other cancers. All seven current clinical sites in the UK were audited by a single visiting group and set of measurement equipment. METHODS: Measurements of output, isotropy and depth doses were performed using an ion chamber in solid water, thermoluminescent dosemeters and radiochromic film, respectively. RESULTS: The mean difference between measured and planned dose across all centres was -3.2±2.7%. Measured isotropy was within ±3% around the lateral plane of the X-ray source and +11±4% in the forward direction compared with the lateral plane. Measured depth doses were agreed within 5±2% of manufacturer-provided calibration values or a mean gamma index of 97% at a tolerance of 7%/0.5 mm. CONCLUSION: Agreement within measurement uncertainties was found for all three parameters except forward anisotropy, which is unlikely to be clinically significant. Steep dose gradients increase the sensitivity to small variations in positioning, but these tests are practical for use in interdepartmental audits and local baseline comparison. ADVANCES IN KNOWLEDGE: The first UK interdepartmental audit of intraoperative radiotherapy builds confidence in the delivery of this treatment.

Regions of repeated DNA in the genome of Choristoneura fumifefana nuclear polyhedrosis virus.

The genome of the nuclear polyhedrosis virus of the spruce budworm, Choristoneura fumiferana (CfMNPV), was shown, by Southern blot hybridization, to contain regions of repeated DNA sequences. These homologous regions have been located at four sites on the physical map of CfMNPV. In addition, the DNA at one of these sites is expanded, possibly via tandem duplication, in a fraction of the total viral DNA population.

A structural polypeptide of the baculovirus Autographa californica nuclear polyhedrosis virus contains O-linked N-acetylglucosamine.

A structural glycopeptide, gp41, derived from the occluded virus of the baculovirus Autographa californica nuclear polyhedrosis virus was characterized. The peptide specifically bound wheat germ agglutinin but was not recognized by a panel of seven other lectins. Reactivity with wheat germ agglutinin was eliminated by treatment of gp41 with beta-N-acetylglucosaminidase, indicating that N-acetylglucosamine (GlcNAc) was present as terminal residues. gp41 was efficiently galactosylated by galactosyltransferase only in the presence of Nonidet P-40, suggesting that GlcNAc residues are not exposed on the surface of the virion. Metabolic labelling of gp41 with [3H]GlcNAc occurred in the presence of tunicamycin. The carbohydrate was released by alkaline borohydride treatment and comigrated with N-acetylglucosaminitol in descending paper chromatography. The data indicate that gp41 contains single residues of GlcNAc O glycosidically linked to the polypeptide chain. Evidence suggesting that gp41 is located in the region between the envelope membrane and the capsid (defined here as the tegument) of the occluded virus is also presented.

Plaque assay of nuclear polyhedrosis viruses in cell culture.

The nuclear polyhedrosis virus of Autographa californica has been titrated in Spodoptera frugiperda cells by the plaque method, using a solid overlay which does not require either the use of modified culture medium or expensive purified agarose or the addition of culture medium as a liquid layer above the solid agarose. This assay is more sensitive than that using a viscous methyl cellulose overlay but less sensitive than the end-point dilution technique. Neither Trichoplusia ni nor Bombyx mori cells were satisfactory as indicators for the assay as described, since they failed to form a stable monolayer. Manduca sexta cells could be utilized for assay of A. californica nuclear polyhedrosis virus, but the sensitivity was lower than with S. frugiperda cells.

The prime time diet: a content analysis of eating behavior and food messages in television program content and commercials.

The purpose of this study was to identify and analyze messages related to food and eating behavior as presented on prime time television (8:00-11:00 pm) both in programming and commercials. Food references occurred an average of 4.8 times per 30 minutes of programming time. Over half (60 percent) of all food references in programs were for low nutrient beverages and sweets. The prime time diet is inconsistent with dietary guidelines for healthy Americans.

Nucleotide sequence and transcriptional analysis of a gene encoding gp41, a structural glycoprotein of the baculovirus Autographa californica nuclear polyhedrosis virus.

We have identified and sequenced a region of the Autographa californica nuclear polyhedrosis virus (AcMNPV) genome encoding the major polyhedron-derived virus structural glycoprotein gp41. The open reading frame is located entirely within the AcMNPV SstII-M fragment. The protein sequence does not have hydrophobic regions characteristic of integral membrane proteins and contains a putative O-linked GlcNAc glycosylation site. gp41 is expressed as a late gene, with transcripts starting within two consensus late transcription start sites (TAAG) located immediately upstream of the first methionine codon. A major transcription termination signal is bypassed, possibly generating a bicistronic message. Finally, the nucleotide and protein sequences of AcMNPV and Bombyx mori nuclear polyhedrosis virus are highly conserved.

Properties of 42S and 26S Sindbis viral ribonucleic acid species.

Two species of ribonuclease-sensitive Sindbis viral ribonucleic acids which sedimented at 42S and 26S were studied. 42S RNA, derived either from virions or from viral nucleoids extracted from infected cultures, was converted by heating to an RNA which sedimented at 26S. The sedimentation patterns of 42S RNA and "derived" 26S RNA were similarly affected in low ionic strength buffers. 42S RNA ran as a homogeneous fraction on polyacrylamide gels; the "derived" 26S RNA as well as "natural" 26S RNA from infected cultures showed similar electrophoretic patterns of heterogeneity. A doubling of 3' polynucleotide termini was observed when 42S RNA was heated. Two possibilities concerning the structure of 42S RNA are considered. (i) It may consist of an aggregate of subunits, joined by means of hydrogen bonds to form a complex molecule. (ii) A heat-labile covalent bond of unknown type may link viral RNA subunits. Although 26S RNA from infected cultures and "derived" 26S RNA from 42S RNA behaved in a similar qualitative manner on gels, their sedimentation characteristics were affected differently in low ionic strength buffers. "Natural" and "derived" 26S RNA appear to consist of a population of fragments. and their behavior in gradients and in gels is probably dictated by the experimental conditions of the analytical methods used.

Molecular weight of Sindbis virus ribonucleic acid as measured by polyacrylamide gel electrophoresis.

The molecular weight (MW) of Sindbis virion ribonucleic acid (RNA) determined by gel electrophoresis was in a range of 3.89 x 10(6) to 4.45 x 10(6) daltons. Upon denaturation with urea, it separated into fragments having an MW of 1.76 x 10(6) daltons. Viral-specific Sindbis 26S RNA contained a major species having an MW of 1.76 x 10(6) daltons.