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Thoracotomy, chest surgery, causes severe pain and impaired lung function after surgery, especially in the elderly, the obese, smokers, and those with pre-existing heart or lung problems. After surgery, proper inspiration and adequate deep breathing and coughing is necessary to prevent lung complications, but these necessary breathing functions may be limited by the patient's severe pain. Effective relief of pain can minimize impaired lung function and speed recovery, as well as prevent lung complications. Because most lung procedures are performed on only one side, pain control may be possible by nerve block of the intercostals, nerves located between the ribs. Continuous intercostal nerve block can be achieved by administrating an anesthetic through an indwelling catheter placed at the time of surgery. To assess the efficacy of this technique, a prospective random study was carried out with 56 patients undergoing thoracotomy; 29 patients received the anesthetic bupivacaine and 27 received a placebo (saline solution). Infusion was continued after surgery for five days, and levels of pain and lung function were evaluated during this period. The group receiving bupivacaine reported less pain and required fewer doses of other pain medication than did the control group. The postoperative reduction in breathing capacity of the anesthetic group was significantly less than that of the control group. There were no complications related to the infusion of either anesthetic or placebo. These findings indicate that continuous intercostal nerve block is a safe and effective method of pain relief following thoracotomy. Its use significantly reduces the loss of lung function in the immediate period following operation. (Consumer Summary produced by Reliance Medical Information, Inc.)

Thermally activated delayed fluorescence (TADF) has shown great potential as a mechanism for harvesting low-lying triplet excited states in organic molecules and is therefore of great interest in the context of organic electronics, especially organic light emitting diodes (OLEDs). Herein we study the mechanism for triplet harvesting in triquinolonobenzene (TQB), which instead of relying upon the well-established donor–acceptor (D-A) scheme uses excited-state intramolecular proton transfer (ESIPT). We demonstrate that upon photoexcitation into the lowest singlet excited state the proton is transferred within 20 fs, suggesting it plays little role in triplet harvesting, which occurs on the nano- to microsecond time scale. However, TQB exhibits multiple low-lying triplet states that are strongly coupled along this proton transfer coordinate. The majority of these states favor the structure prior to proton transfer (TQB-TA) and this means that the proton transfer dynamics (3TQB-TA → 1TQB-TB) plays a crucial role in triplet harvesting. This mechanism yields an energy gap in good agreement with that reported experimentally and is consistent with previous photophysical characterization. Finally, a discussion upon extending this understanding into a device context is also presented.

Background: Brodifacoum poisoning is a health concern. Scenarios in which it occurs include: accidental ingestion in children, suicide attempts, and Munchausen syndrome. This case reports a patient who attempted suicide with a brodifacoum product who was treated with Bebulin[R] VH Immuno (Factor IX Complex). Case Report: A 52-year-old man presented to the emergency department by ambulance in acute respiratory distress. He complained of throat swelling. The patient had an expanding hematoma to the anterior neck region superior to the supraclavicular notch. He also had blood present around his lips. Further exam revealed blood on the oral mucosa. The patient's respiratory status declined rapidly. Oral intubation was unsuccessful due to the large amount of blood in the oral pharynx and the loss of anatomical landmarks. Cricothyrotomy was not attempted in the emergency department due to the expansion of the neck hematoma. The patient was emergently taken to the operating room for placement of a tracheostomy. Initial laboratory values included a PT [is greater than] 100 seconds, INR [is greater than] 98.5, aPTT 111.3 seconds, Hb 10.7, Hct 31.6, Factor VII [is less than] 4, and Factor VI [is less than] 4. The patient was treated with Bebulin[R] 2000 units IV and vitamin K. He was subsequently discharged for extensive psychiatric evaluation and treatment with a PT [is less than] 12 seconds. He was maintained on oral vitamin K. Conclusion: There have been many case reports of brodifacoum poisonings in the literature. These cases have been treated routinely with vitamin K therapy. We report the first case of brodifacoum poisoning treated with Bebulin[R] VH Immuno (Factor IX Complex) with an excellent patient outcome., Eng J, Ramstack T. Michigan State University Emergency Medicine Residency, Ingham Regional Medical Center, Lansing, [...]

Background: Suicidal ingestions of yew (Taxus) have been reported to cause fatal arrhythmia. The literature describes both tachyarrhythmias and bradyarrhythmias, but most reports confirm the presence of a prolonged ventricular complex. Animal studies of the toxic component, taxine, show it acts as a Class I antiarrhythmic agent, by blocking sodium channels. This case illustrates the use of sodium bicarbonate in treatment of a nearfatal ingestion of Taxus. Case Report: A 32-year-old male stated he combined unknown quantities of dehydrated yew, paroxetine, brodificoum, and shellac, which he then baked. After eating one dozen 'cookies' he experienced repeated vomiting. He was taken the hospital approximately three hours postingestion. Upon arrival he became unresponsive and suffered sudden cardiac arrest. ACLS measures were initiated. ABG at the time of arrest showed a pH of 7.34. Bicarbonate was given as the original ECG demonstrated a prolonged QRS complex. Circulation was reestablished and wide complex rhythm remained. Bicarbonate therapy was therefore continued. Original epinephrine and dopamine drips were converted to norepinephrine for persistent hypotension. Serial ABGs reveal that as the serum pH became greater than 7.5, the QRS complex narrowed. Urine analyzed by GCMS revealed only the presence of phenylpropanolamine. Detection of taxine would not be expected due to the technique used. The presence of taxine in the 'dough' however, was confirmed by GCMS. The patient eventually recovered and was discharged to a psychiatric facility. Conclusions: Sodium bicarbonate is an accepted standard of care in treatment of wide complex arrhythmias associated with sodium channel blocking agents. This case describes the first use of sodium bicarbonate in the treatment of yew toxicity., Miller MB, Eng J, Curry SC. Ingham Regional Medical Center, Dewitt, [...]

Objective: In 1997, the AACT/EAPCCT issued a position statement regarding the use of ipecac stating it 'should not be administered routinely in the management of poisoned patients.' Our study was designed to see if Poison Centers (PCs) are still recommending the use of ipecac, and under what circumstances. Methods: All PCs in the US were contacted by phone and questioned about their policies for ipecac use in both the home and the Emergency Department (ED). Five factitious cases were presented to evaluate what type of ingestion by a toddler might prompt ipecac use: 1) four unidentified red berries, 2) a potentially toxic acetaminophen ingestion, 3) a suspected imipramine ingestion, 4) a potentially toxic iron ingestion, and 5) a toothpaste ingestion. Results: 58 PCs responded to the survey; 15 could not be reached. Annual calls for human exposures ranged from 11,000-135,000, with pediatric calls (age [is less than] 6 years) ranging from 33-100%. Of these 58 centers, 41 (70.7%) are AAPCC certified, 28 (48.3%) are associated with a pediatric hospital, and 56 (96.6%) have some type of physician support. Ipecac was recommended in the home always (3.4%), sometimes (89.7%), and never (6.9%); while in the ED, it was used always (0%), sometimes (29.3%) and never (69.0%). While responses to specific cases varied, iron was the most common ingestion in which ipecac was advised. 93.1% of PCs were aware of the 1997 guidelines, but only 37.9% had changed their protocols. No demographic variable was statistically able to predict the likelihood of a PC recommending ipecac for use either at home or in the ED (CI of 95%). Conclusions: Despite the AACT/EAPCCT position statement, a substantial amount of PCs still routinely recommended ipecac use. The frequency for which this occurs varied with the type of ingestion., Miller MB, Eng J, Schneiderman E. Michigan State University Emergency Medicine Residency Program, Lansing, [...]

Background: The financial performance for the practice of clinical toxicology is modest. A clinical toxicologist may be affiliated with a Poison Control Center (PCC) or a fellowship training program and receive financial support. Many toxicologists also volunteer time to the clinical practice. We would like to present data regarding patient demographics and financial performance from our toxicology service established approximately two and a half years ago. Case Report: The toxicology service was established in August 1996 with practicing emergency physicians forming a private consultation and admitting toxicology service not affiliated with a PCC. Consulting privileges were obtained at two area hospitals with admitting privileges at one of these hospitals. Patients for occupational exposures are occasionally seen in an outpatient clinic. Results: For the period August 1996 through March 1999, 275 patients were seen. Of these, 209 were adults ([is greater than] 18 yo) and 66 were children. Two hundred sixty-two patients were seen in consultation and 13 patients were admitted to the clinical toxicology service. As of March 1999, the total amount billed was $117,009.00 with collections equaling $31,295.87. To date, the aging amount is $35,578.14 with an adjusted amount of $50,134.99. Our total expenses equal $6,629.58 with a remaining net income of $24,666.29. Conclusion: Our clinical toxicology service is small but already it shows evidence of growth. Although the net income our service generated thus far cannot support all of our personal finances, we hope to see continued growth and return. This report serves as an example that establishing a clinical toxicology practice independent of a PCC is possible; however, its development may take time., Eng J, Miller MB. Ingham Regional Medical Center and Sparrow Hospital, Lansing, [...]

Study design:Longitudinal observational study.Objective:To quantify the amount of upper- and lower-extremity movement repetitions (that is, voluntary movements as part of a functional task or specific motion) occurring during inpatient spinal cord injury (SCI), physical (PT) and occupational therapy (OT), and examine changes over the inpatient rehabilitation stay.Setting:Two stand-alone inpatient SCI rehabilitation centers.Methods:Participants: A total of 103 patients were recruited through consecutive admissions to SCI rehabilitation. Interventions: Trained assistants observed therapy sessions and obtained clinical outcome measures in the second week following admission and in the second to last week before discharge. Main outcome measures: PT and OT time, upper- and lower-extremity repetitions and changes in these outcomes over the course of rehabilitation stay.Results:We observed 561 PT and 347 OT sessions. Therapeutic time comprised two-thirds of total therapy time. Summed over PT and OT, the median upper-extremity repetitions in patients with paraplegia were 7 repetitions and in patients with tetraplegia, 42 repetitions. Lower-extremity repetitions and steps primarily occurred in ambulatory patients and amounted to 218 and 115, respectively (summed over PT and OT sessions at discharge). Wilcoxon-signed rank tests revealed that most repetition variables did not change significantly over the inpatient rehabilitation stay. In contrast, clinical outcomes for the arm and leg improved over this time period.Conclusions:Repetitions of upper- and lower-extremity movements are markedly low during PT and OT sessions. Despite improvements in clinical outcomes, there was no significant increase in movement repetitions over the course of inpatient rehabilitation stay.

Bone marrow lesions (BMLs) are the MRI hallmark of bone involvement in knee OA and correlate with knee pain. While they may have great prognostic values, their assessment in large-scale datasets can be time-consuming and subject to inter-reader variability.

Accumulating evidence suggests that white matter lesions are associated with vascular cognitive impairment. The authors investigated the relationships between white matter lesions, cognitive impairment, and risk of recurrent hemorrhage in a prospectively identified cohort of patients with lobar intracerebral hemorrhage (ICH).

We report the results of our work to facilitate protein identification using tandem mass spectra and protein sequence databases. We describe a parallel version of SEQUEST (SEQUEST-PVM) that is tolerant toward arithmetic exceptions. The changes we report effectively separate search processes on slave nodes from each other. Therefore, if one of the slave nodes drops out of the cluster due to an error, the rest of the cluster will carry the search process to the end. SEQUEST has been widely used for protein identifications. The modifications made to the code improve its stability and effectiveness in a high-throughput production environment. We evaluate the overhead associated with the parallelization of SEQUEST. A prior version of software to preprocess LC/MS/MS data attempted to differentiate the charge states of ions. Singly charged ions can be accurately identified, but the software was unable to reliably differentiate tandem mass spectra of +2 and +3 charge states. We have designed and implemented a computational approach to narrow charge states of precursor ions from nominal resolution ion-trap tandem mass spectra. The preprocessing code, 2to3, determines the charge state of the precursor ion using its mass-to-charge ratio (m/z) and fragment ions contained in the tandem mass spectrum. For each possible charge state the program calculates the expected fragment ions that account for precursor ion m/z vlues. If any one of the numbers is less than an empirically determined threshold value then the spectrum corresponding to that charge state is removed. If both numbers are higher than the threshold value then +2 and +3 copies of the spectrum are kept. We present the comparison of results from protein identification experiments with and without using 2to3. It is shown that by determining the charge state and eliminating poor quality spectra 2to3 decreases the number of spectral files to be searched without affecting the search results. The decrease reduces computer requirements and researcher efforts for analysis of the results. Keywords: mass spectrometry • protein identification • database search • charge determination

The effectiveness of proteome-wide protein identification and quantitative expression profiling is dependent on the ability of the analytical methodologies employed to routinely obtain information on low-abundance proteins, as these are frequently of great biological importance. Two-dimensional gel electrophoresis, the traditional method for proteome analysis, has proven to be biased toward highly expressed proteins. Recently, two-dimensional chromatography of the complex peptide mixtures generated by the digestion of unseparated protein samples has been introduced for the identification of their components, and isotope-coded affinity tags (ICAT) have been introduced to allow for accurate quantification of the components of protein mixtures by mass spectrometry. Here, we demonstrate that the combination of isotope coded affinity protein tags and multidimensional chromatography/mass spectrometry of tryptic peptide mixtures is capable of detecting and quantifying proteins of low abundance in complex samples. Keywords: gene expression • functional genomics • proteomics • protein profiling • mass spectrometry • isotope-coded affinity tags

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The present study explores the potential utility of peripheral versus central administration of glucagon-like peptide-1 (GLP-1) receptor agonists in the regulation of feeding behavior in Wistar and Zucker obese rats. Acute central (intracerebroventricular [ICV]) and peripheral (subcutaneous [SC]) administration of both GLP-1 (7-36) amide and exendin-4 resulted in a reduction in food intake for at least 4 hours, exendin-4 being much more potent than GLP-1 (7-36) amide, especially after peripheral administration. Both Zucker obese rats (fa/fa) and their lean littermates (Fa/-) responded to acute central and peripheral administration of exendin-4. Moreover, in situ hybridization revealed specific labeling for the mRNA for GLP-1 receptors in several brain areas of both the obese and lean rats. The presence of this receptor was also detected by affinity cross-linking assays. Long-term SC administration of exendin-4 (1 single injection per day, 1 hour prior to the onset of the dark phase of the cycle) decreased daily food intake and practically blocked weight gain in obese rats. In contrast to previous studies, these findings show that peripheral (SC) administration of both GLP-1 receptor agonists also induces satiety and weight loss in rats, and suggest the potential usefulness of exendin-4 as a therapeutic tool for the treatment of diabetes and/or obesity.

Vanadium carbides were prepared via the temperature-programmed reaction of V2O5with a CH4/H2mixture and evaluated for the dehydrogenation of n-butane. Thermogravimetric analysis coupled with X-ray diffraction indicated that the solid-state reaction proceeded by the following sequential reaction: V2O5→V2O3→V8C7. The space velocity and heating rates had insignificant effects on the surface areas; however, the use of a high-temperature H2post-treatment caused a reduction in the surface area and carbon content. Temperature-programmed reduction results indicated that oxygen was more strongly bound to the substoichiometric vanadium carbide than to the stoichiometric material. The results were also consistent with the presence of an oxycarbide near surfaces of the passivated vanadium carbides. The passivated vanadium carbides were sufficiently activated by reduction in H2at 500°C for 3 h. Oxygen chemisorptive uptakes on the reduced vanadium carbides corresponded to an O/V ratio of 0.28. This oxygen-to-metal ratio is half that measured for the vanadium nitrides, suggesting that some excess carbon may have been present on surfaces of the carbides. The butane dehydrogenation turnover frequency for the vanadium carbide catalyst was 10−3s−1at 450°C. The corresponding turnover frequency for a Pt-Sn/Al2O3catalyst was 6.3×10−2s−1. Near-edge X-ray absorption fine structure spectroscopy indicated that the vanadium carbide and nitride catalysts were partially ionic, with charge transfer being from vanadium to carbon or nitrogen. This degree of ionic bonding distinguishes the vanadium compounds from other carbides and nitrides and could partly explain their high dehydrogenation selectivities. Similarities between catalytic properties of the vanadium carbides and nitrides were likely a consequence of their similar electronic structures. The p-projected density of unoccupied states near the carbon and nitrogen K-edges nearly identical.

Twenty‐one mycobacterial type strains and 334 clinical isolates of mycobacteria were identified by standardized sequence analysis using part of the gene encoding 16S rRNA. Apart from two clinical isolates, the resulting sequences corresponded to previously published sequences. The results of the molecular determinations of the type strains completely overlapped the identities obtained using conventional techniques (cultural characteristics, biochemical tests, commercial DNA probes, and gas chromatographic lipid profiles). Of 323 isolates conventionally identified as slow‐growing mycobacteria, 318 (98.5%) were identified to the same species or group level by 16S rDNA sequence analysis, while 6 of the 11 strains of rapid growers obtained a corresponding identity with the two approaches. The sequencing protocol combined with a few cultural characteristics (i.e. growth rate, pigmentation and susceptibility testing) offers a rapid, reliable and usually definite identification of mycobacterial isolates.

Specifications of about eight-hundred colored glasses made by thirteen manufacturers are examined and intercompared. The internal spectral transmittances of typical glasses are plotted in a series of forty-four diagrams. A log (internal density) vs wavelength representation is used. Curves with similar characteristics are grouped together in the same diagram. The internal spectral transmittances for a number of different thicknesses of each glass type can be read directly from the diagram. The curves are identified in a table, which lists the equivalent thicknesses of glasses yielding approximately the same transmittance. Such information facilitates the choice of filter components of reasonable thickness.

We have investigated the surface reactivities of Ni/Pt(111) bimetallic model catalysts using ethylene and cyclohexene as probing molecules. The bimetallic surfaces were generated by evaporating Ni onto a Pt(111) single- crystal surface held at 600 K. The surface chemistry was investigated using high-resolution electron energy loss spectroscopy (HREELS), Auger electron spectroscopy (AES), temperature-programmed desorption (TPD) and low-energy electron diffraction (LEED). The reactivities of the bimetallic surfaces were compared with those of the clean Pt(111) surface and a thick Ni(111) film on the Pt(111) substrate. Formation of the bimetallic surface led to a significantly reduced reactivity towards the decomposition of ethylene when compared to either Pt(111) or Ni(111)/Pt(111) surfaces. Furthermore, although the surface reactivity towards cyclohexene was retained for the bimetallic surface, the decomposition mechanism was distinctly altered from that of either Pt(111) or Ni(111)/Pt(111) surfaces.

As a part of a continuing investigation of acetylene reactions on tungsten surfaces covered by ultrathin metal films, the chemistry of acetylene on clean and carbide-modified W(211) surfaces has been investigated using high-resolution electron energy-loss spectroscopy (HREELS) and temperature-programmed desorption (TPD) measurements. Acetylene decomposes between 100 and 450 K on the clean W(211) surface, ultimately forming carbidic carbon and gaseous hydrogen. During this decomposition process, a mixture of different hydrocarbon fragments are formed, most likely vinylidene (CCH2) or vinyl (HCCH2) and acetylide (CCH). On the carbide-modified W(211) surface, acetylene also decomposes to produce carbon and gaseous hydrogen. However, compared to clean W(211), the CH bond activation is suppressed on the carbide-modified W surface, as evidenced by the broadening and significant shifting of the H2 TPD features toward higher temperatures as well as by the HREEL spectra monitoring the thermal decomposition of acetylene on the carbide-modified W(211) surface.

In an ongoing investigation to study structure−reactivity relationships on bimetallic surfaces, acetylene cyclotrimerization to form benzene is of particular interest:  in this structure-sensitive catalytic reaction, C−C and C−H bonds can be formed readily under ultrahigh vacuum (UHV) conditions without C−C bond breaking. In this paper, we present results for acetylene cyclization and hydrogenation on Pd/W(211). Pd on W is chosen because it is a morphologically unstable system, and W(211) facets develop after annealing Pd/W(111) to ≥700 K. Temperature-programmed desorption (TPD) results exhibit negligible amounts of benzene detected from acetylene adsorption on clean W(211). A single monolayer (ML) of Pd on W(211) decreases the high reactivity toward acetylene decomposition and several different reaction pathways are accessed, including hydrogenation of C2H2 to C2H4 and cyclotrimerization of C2H2 to form C6H6. The cyclotrimerization reaction produces three benzene desorption states at ~340, ~390, and ~430 K. In addition, the detection of C4H6 during TPD provides evidence that an elusive C4H4 intermediate is present on the surface. Furthermore, ethylene is observed in substantial yields, lending insight into the activity of the bimetallic system. The use of high-resolution electron energy-loss spectroscopy (HREELS) provides complementary information regarding the reaction mechanisms of acetylene on the Pd/W(211) surfaces.

Molecular cloning has established the structure of preprocholecystokinin in porcine cerebral cortex and duodenal mucosa. This precursor is 114 amino acids long, is identical in brain and gut, contains all the cholecystokinin (CCK) peptides previously isolated, and has the characteristics of a prohormone. It contains a putative amino-terminal signal peptide, basic processing sites, and a carboxyl-terminal amidation signal. The CCK mRNAs from brain and gut are approximately 850 nucleotides long and differ by only a few single base changes. This analysis establishes by a strict criterion that CCK is synthesized in both brain and gut and that the different distributions of molecular forms of CCK in the two tissues are most probably a consequence of tissue-specific posttranslational events.

Surface photoabsorption (SPA) has been applied to monitor, in real time, the surface of GaAs(100) during chemical dry etching by a molecular beam of HCl. Changes in the HCl flux to the surface at a constant temperature (840 K) have been used to induce changes in the Ga:As ratio on the surface. These changes in surface stoichiometry have been detected in situ by SPA measurements of the transient fractional change in the reflectance of p-polarized, 488-nm light that is incident onto the surface near the pseudo-Brewster angle. On the basis of results from prior applications of SPA to the study of the atomic layer deposition of GaAs, the changes in the SPA signal as a function of the etching parameters can be correlated with changes in the relative surface densities of Ga and As. The findings are confirmed by independent determinations of the changes in surface stoichiometry made by measuring the time-integrated difference in the fluxes of Ga- and As-containing etching products evolved from the surface as a function of the HCl flux.

The dehydrogenation and thermal decomposition mechanisms of cyclohexene and 1,3-cyclohexadiene on clean Mo(110) and carbide-modified (4×4)-C/Mo(110) surfaces have been studied using temperature-programmed desorption (TPD) and high-resolution electron energy loss spectroscopy (HREELS). On the clean Mo(110) surface, partial dehydrogenation of a fraction of the cyclohexene molecules occurs at temperatures as low as 80 K. When the surface is heated to 150 K, the HREEL spectra obtained are characteristic of a C6H9 intermediate, as seen by a comparison with HREEL spectra reported for C6H9 on Pt(111).^1,2 At higher temperatures, competing C−C and C−H bond cleavage reactions lead to the formation of surface carbon and the evolution of hydrogen. In contrast, on the carbide-modified surface, the primary reaction pathway for cyclohexene is selective dehydrogenation to form benzene and hydrogen. In the case of 1,3-cyclohexadiene, the HREEL results suggest that dehydrogenation to form benzene occurs at 80 K on the clean Mo(110) surface, based on a comparison with the HREEL spectrum for benzene directly dosed onto Mo(110) at 80 K. However, upon heating, most of the benzene decomposes to form surface carbon and hydrogen, as shown by TPD studies. On the carbide-modified surface, the primary reaction pathway for 1,3-cyclohexadiene is selective dehydrogenation to form benzene, which desorbs at 313 K. Furthermore, the HREEL results also indicate that a competing reaction pathway occurs to form a surface intermediate which most likely has an tilted aromatic c-C6 ring, such as a surface phenyl species.

A sequential method employing methanol extraction of the COOH-terminal fragment of cholecystokinin (CCK) from pig brain followed by HCl extraction of the more basic CCK peptides was used as the first step in purification of these peptides. Recovery was monitored with two different assays, one directed to the COOH terminus of CCK and the other to the NH2 terminus. The amino acid content and sequence were determined for each of five peptides after purification. The only peptide containing COOH-terminal immunoreactivity was CCK-octapeptide (CCK8). The other four peptides did not contain CCK8 and had lost one or two additional amino acids, perhaps as a consequence of the action of carboxypeptidases. These peptides were shown to be CCK33-desnonapeptide, CCK39-desnonapeptide and -desdecapeptide, and a large molecular weight precursor, CCK58-desnonapeptide, containing 19 amino acids (Ala-Val-Gln-Lys-Val-Asp-Gly-Glu-Ser-Arg-Ala-His-Leu-Gly-Ala-Leu-Leu-Ala-Arg) NH2-terminal to CCK39. The three NH2-terminal fragments of CCK58, CCK39, and CCK33 were about equally prominent. The brain, unlike the gut, appears to cleave CCK8 rapidly from a precursor peptide but to process the NH2-terminal portions of the molecule more slowly and incompletely.

To find mammalian analogues of exendin-4, a peptide from Helodermatidae venoms that interacts with newly discovered exendin receptors on dispersed acini from guinea pig pancreas, we examined the actions of recent additions to the vasoactive intestinal peptide/secretin/glucagon family of regulatory peptides. In every respect tested, the truncated form of glucagon-like peptide-1, GLP-1(7-36)NH2, mimicked the actions of exendin-4. Like exendin-4, GLP-1(7-36)NH2 caused an increase in acinar cAMP without stimulating amylase release. GLP-1(7-36)NH2-induced increases in cAMP were inhibited progressively by increasing concentrations of the specific exendin-receptor antagonist, exendin(9-39)NH2. In dispersed acini from guinea pig and rat pancreas, concentrations of GLP-1(7-36)NH2 that stimulated increases in cAMP caused potentiation of cholecystokinin-induced amylase release. Binding of 125I-[Y39]exendin-4 or 125I-GLP-1(7-36)NH2 to dispersed acini from guinea pig pancreas was inhibited by adding increasing concentrations of unlabeled exendin-4 or GLP-1(7-36)NH2. We conclude that the mammalian peptide GLP-1(7-36)NH2 interacts with exendin receptors on dispersed acini from guinea pig pancreas. Exendin(9-39)NH2, a competitive antagonist of the actions of GLP-1(7-36)NH2 in pancreatic acini, may be a useful tool for examining the physiological actions of this peptide.

Abstract: Agonists that elevate calcium in T84 cells stimulate chloride secretion by activating KBIC, an inwardly rectifying K channel in the basolateral membrane. We have studied the regulation of this channel by calcium, nucleotides and phosphorylation using patch clamp and short-circuit current (I SC) techniques. Open probability (P 0) was independent of voltage but declined spontaneously with time after excision. Rundown was slower if patches were excised into a bath solution containing ATP (10 mgrm–5 mm), ATP (0.1 mm) + protein kinase A (PKA; 180 nm), or isobutylmethylxanthine (IBMX; 1 mm). Analysis of event durations suggested that the channel has at least two open and two closed states, and that rundown under control conditions is mainly due to prolongation of the long closed time. Channel activity was restimulated after rundown by exposure to ATP, the poorly hydrolyzable ATP analogue AMP-PNP, or ADP. Activity was further enhanced when PKA was added in the presence of MgATP, but only if free calcium concentration was elevated (400 nm). Nucleotide stimulation and inward rectification were both observed in nominally Mg-free solutions. cAMP modulation of basolateral potassium conductance in situ was confirmed by measuring currents generated by a transepithelial K gradient after permeabilization of the apical membrane using agr-toxin. Finally, protein kinase C (PKC) inhibited single KBIC channels when it was added directly to excised patches. These results suggest that nonhydrolytic binding of nucleotides and phosphorylation by PKA and PKC modulate the responsiveness of the inwardly rectifying K channel to Ca-mediated secretagogues.

Abstract: Cholinergic stimulation of chloride secretion involves the activation of a basolateral membrane potassium conductance, which maintains the electrical gradient favoring apical Cl efflux and allows K to recycle at the basolateral membrane. We have used transepithelial short-circuit current (I SC), fluorescence imaging, and patch clamp studies to identify and characterize the K channel that mediates this response in T84 cells. Carbachol had little effect on I SC when added alone but produced large, transient currents if added to monolayers prestimulated with cAMP. cAMP also enhanced the subsequent I SC response to calcium ionophores. Carbachol (100 mgrm) transiently elevated intracellular free calcium ([Ca2+] i ) by sim3-fold in confluent cells cultured on glass coverslips with a time course resembling the I sc response of confluent monolayers that had been grown on porous supports. In parallel patch clamp experiments, carbachol activated an inwardly rectifying potassium channel on the basolateral aspect of polarized monolayers which had been dissected from porous culture supports. The same channel was transiently activated on the surface of subconfluent monolayers during stimulation by carbachol. Activation was more prolonged when cells were exposed to calcium ionophores. The conductance of the inward rectifier in cell-attached patches was 55 pS near the resting membrane potential (–54 mV) with pipette solution containing 150 mm KCl (37°C). This rectification persisted when patches were bathed in symmetrical 150 mm KCl solutions. The selectivity sequence was 1 K > 0.88 Rb > 0.18 Na Gt Cs based on permeability ratios under bi-ionic conditions. The channel exhibited fast block by external sodium ions, was weakly inhibited by external TEA, was relatively insensitive to charybdotoxin, kaliotoxin, 4-aminopyridine and quinidine, and was unaffected by external 10 mm barium. It is referred to as the KBIC channel based on its most distinctive properties (Ba-insensitive, inwardly rectifying, Ca-activated). Like single KBIC channels, the carbachol-stimulated I SC was relatively insensitive to several blockers on the basolateral side and was unaffected by barium. These comparisons between the properties of the macroscopic current and single channels suggest that the KBIC channel mediates basolateral membrane K conductance in T84 cell monolayers during stimulation by cholinergic secretagogues.

GLP-1-(7-36)-amide and exendin-4-(1-39) are glucagon-like peptide-1 (GLP-1) receptor agonists, whereas exendin-(9-39) is the only known antagonist. To analyze the transition from agonist to antagonist and to identify the amino acid residues involved in ligand activation of the GLP-1 receptor, we used exendin analogs with successive N-terminal truncations. Chinese hamster ovary cells stably transfected with the rat GLP-1 receptor were assayed for changes in intracellular cAMP caused by the test peptides in the absence or presence of half-maximal stimulatory doses of GLP-1. N-terminal truncation of a single amino acid reduced the agonist activity of the exendin peptide, whereas N-terminal truncation of 3-7 amino acids produced antagonists that were 4-10-fold more potent than exendin-(9-39). N-terminal truncation of GLP-1 by 2 amino acids resulted in weak agonist activity, but an 8-amino acid N-terminal truncation inactivated the peptide. Binding studies performed using 125I-labeled GLP-1 confirmed that all bioactive peptides specifically displaced tracer with high potency. In a set of exendin/GLP-1 chimeric peptides, substitution of GLP-1 sequences into exendin-(3-39) produced loss of antagonist activity with conversion to a weak agonist. The results show that receptor binding and activation occur in separate domains of exendin, but they are more closely coupled in GLP-1.

A radioimmunoassay was developed to detect the cholecystokinin (CCK)-associated nonapeptide (CAP-9) that forms the COOH terminus of pig preproCCK. This peptide (Ser-Ala-Glu-Glu-Tyr-Glu-Tyr-Thr-Ser) is presumably produced at the time that the tyrosine-sulfated octapeptide CCK8(s) is cleaved from preproCCK. Radioimmunoassay of a dried methanol extract of pig brain revealed no detectable CAP-9 immunoreactivity, whereas acid desulfation of the dried methanol extract prior to radioimmunoassay resulted in easily measurable concentrations of CAP-9 immunoreactivity. Two peptides, CAP-9 and des-Ser9-CAP-9, were purified from a methanol extract of 8 kg of commercially obtained whole pig brains. Amino acid analysis showed that each peptide has both tyrosines sulfated. Thus, the likely sequence of CCK post-translational processing events is sulfation of the three tyrosines in the COOH terminus of preproCCK followed by peptide cleavage and appearance of CCK8(s) and CAP-9(s,s).

Fifteen-hundred hospital blood cultures were made in duplicate, with and without 0.05% sodium polyanethol sulfonate in the broth medium. A significantly higher rate and speed of recovery of both gram-positive cocci and gram-negative bacilli was accomplished in sodium polyanethol sulfonate broth. The effect was independent of the content of 0.1% agar in the growth medium. In the cases of Neisseria meningitidis septicemia examined, however, a detrimental result on recoveries was observed. The addition of sodium polyanethol sulfonate also resulted in an increased frequency of recoveries of contaminating organisms.

For more than 15 years, Norway has had the highest incidence of meningococcal disease in northern Europe, with 80% of cases being due to serogroup B meningococci. The case-fatality has remained high, at about 10%. In this study, an outer membrane vaccine, which had previously been shown to induce an increase in bactericidal antibodies to the parent strain, was assessed in a large-scale, randomised, double-blind trial. From October, 1988, 171 800 students in secondary schools volunteered to take part in a double-blind, placebo-controlled, efficacy trial with school as the randomisation unit. Hospitals and clinics that routinely receive patients with infectious disease were asked to report urgently all cases of suspected meningitis and/or septicaemia in 13-21-year-old students in Norway. These cases were registered and further investigated according to a detailed protocol. 89 out of the 221 cases investigated by June 3, 1991, were shown to be severe systemic disease due to group B meningococci. 36 cases in 35 schools took part in the trial (11 schools with vaccinated students and 24 with students given placebo). The calculated rate of protection was thus 57·2% (p=0·012, one-sided test). The findings suggest that, although the vaccine conferred protection against group B meningococcal disease, the effect was insufficient to justify a public vaccination programme.

Using previously cloned cDNAs to pig brain prepro-cholecystokinin mRNA and slot blot and S1 nuclease protection assays, the relative cholecystokinin mRNA levels in different regions of the pig brain were measured. The relative amounts of cholecystokinin mRNA generally correlated well with the levels of cholecystokinin-immunoreactive peptides in the various regions tested. One clear exception was noted in the cerebellum; in this region, levels of cholecystokinin mRNA were about 20% of the levels in brain cortex (or second highest level in all areas tested) whereas the mature forms of cholecystokinin peptides (cholecystokinin 58, cholecystokinin 8) were undetectable (less than 3 pmol/g). In vitro translation of cerebellar and cortical cholecystokinin mRNA indicated that there was no difference in the efficiency with which these two RNAs were translated into immunoreactive prepro-cholecystokinin. DNA sequence analysis confirmed that a cloned full-length cerebellar cholecystokinin cDNA was indistinguishable from its cortical counterpart and, therefore, must encode an identical prepro-cholecystokinin. We conclude that there are pronounced regional differences in cholecystokinin expression in pig brain. The apparent discrepancy between levels of immunoreactive cholecystokinin peptides and cholecystokinin mRNA in the cerebellum could be explained by a high turnover rate for the peptides, differential processing of the peptides, or tissue-specific inhibition of cholecystokinin mRNA translation.

An amino-terminal histidyl structure (His1) is characteristic of most peptides in the glucagon superfamily. An assay for His1 peptides performed by amino-terminal amino acid sequencing was used to screen venom from the Gila monster lizard, Heloderma horridum. Two His1 peptides were identified: helospectin and a new His1 peptide that has been named exendin-3 to indicate that it is the third peptide to be found in an exocrine secretion of Heloderma lizards which has endocrine activity, the first two being helospectin (exendin-1) and helodermin (exendin-2). In the lot of H. horridum venom tested, exendin-3 was 5-10-fold more abundant in molar concentration than helospectin. The structure of exendin-3 was analyzed by amino acid sequencing and mass spectrometry. Exendin-3 is a 39-amino acid peptide with a mass of 4200. It contains a carboxyl-terminal amide and has a strong homology with secretin at its amino-terminal 12 amino acids. The complete structure of exendin-3 is His-Ser-Asp-Gly-Thr-Phe-Thr-Ser-Asp-Leu-Ser-Lys-Gln-Met-Glu-Glu-Glu-Ala- Val-Arg - Leu-Phe-Ile-Glu-Trp-Leu-Lys-Asn-Gly-Gly-Pro-Ser-Ser-Gly-Ala-Pro-Pro-Pro- Ser- amide. It is 32 and 26% homologous with helospectin and helodermin, respectively. It has greatest homology with glucagon (48%) and human glucagon-like peptide-1 (50%). Exendin-3 (3 microM) stimulated increases in cellular cAMP and amylase release from dispersed guinea pig pancreatic acini.

Two major classes of immunoreactive cholecystokinin peptides (iCCK) have been identified in rat and pig brains: (i) large basic peptides (big iCCK) resembling the 33-amino acid porcine cholecystokinin (pCCK33) in size and charge; (ii) small acidic peptides (small iCCK) resembling the COOH-terminal fragments of CCK. Boiling 0.1 M HCl maximally extracts big iCCK; boiling 0.1 M NaOH maximally extracts small iCCK. The differences in hormonal forms removed by these extractants are not likely to be due to enzymatic conversion during the extraction procedures. Fractionation on Sephadex G-50 and starch gel electrophoresis combined with radioimmunoassay using three antisera of different specificities--(i) directed towards the NH2 terminus of pCCK33, (ii) produced by immunization with COOH-terminal fragment CCK8, (iii) produced by immunization with COOH-terminal fragment CCK4--are consistent with the hypothesis that a major fraction of big iCCK may represent intact cholecystokinin with a COOH-terminal extension, as has recently been suggested for gastrin, a molecule having a COOH-terminal pentapeptide identical with that of cholecystokinin.

Twenty-nine species (76 strains) of members of the genus Legionella were analyzed for their cellular hydroxylated fatty acids (OH-FAs). The individual patterns were unusually complex and included both monohydroxylated and dihydroxylated chains of unbranched or branched (iso and anteiso) types. Comparison of the strain profiles by SIMCA (Soft Independent Modelling of Class Analogy) principal component analysis revealed four main groups. Group 1 included Legionella pneumophila plus L. israelensis strains, and group 2 included L. micdadei and L. maceacherneii strains. These two closely related groups were characterized by the occurrence of di-OH-FAs and differed mainly in the amounts of 3-OH-a21:0, 3-OH-n21:0, 3-OH-n22:0, and 3-OH-a23:0. Group 3 (13 species) was distinguished by i14:0 at less than 3%, 3-OH-3-OH-n14:0 at greater than 5%, 3-OH-n15:0 at greater than 2%, and minute amounts of OH-FAs with chains longer than 21:0. Group 4 (12 species) was heterogeneous. Its main characteristics were the presence of 3-OH-n12:0 and 3-OH-n13:0, 3-OH-i14:0 at greater than 5%, as well as significant amounts of 3-OH-a21:0 and 3-OH-n21:0. The groupings obtained by OH-FA profiles were found to reflect DNA-DNA homology groupings reasonably well, and the profiles appear to be useful for differentiation of Legionella species.

The aim of this study was to elucidate the mechanisms by which sucrose improves growth in a hypertonic medium for isolating aerobes from blood. Clinical blood cultures were made routinely in duplicate in plain broth consisting of brain heart infusion broth with sodium polyanetholesulfonate, gelatin, and penicillinase and the same broth with 20% sucrose added. The growth patterns of Staphylococcus aureus and Enterobacteriaceae from plain and from hypertonic broth were correlated with the presence or absence of antimicrobial therapy in patients when the blood cultures were collected. In S. aureus bacteremias, 58.7% of the positive cultures collected during treatment of patients with beta-lactam antibiotics showed earlier growth or growth only in hypertonic broth, compared with 16.7% of the cultures taken during treatment with other antimicrobial agents (P less than 0.05) and 17.6% of the cultures made in antibiotic-free intervals (P less than 0.01). In the group of cultures yielding growth of Enterobacteriaceae, growth occurred earlier or solely in hypertonic broth in 28.9% of the cultures taken during treatment with beta-lactam antibiotics, compared with 15.7% of the cultures taken during treatment with other antimicrobial agents and 21.6% of the cultures collected in antibiotic-free intervals (differences not statistically significant). It is concluded that treatment with beta-lactam antibiotics is an important reason for the improved growth of S. aureus from hypertonic broth, but other factors are also involved.