Consistency and effect of body position change on measurement of upper and lower esophageal sphincter geometry using impedance planimetry in a canine model.

The EndoFLIP (Endolumenal Functional Lumen Imaging Probe, Crospon Inc, Galway, Ireland) device uses the technique of impedance planimetry to evaluate dimensions and distensibility of the upper and lower esophageal sphincter. The null hypotheses for this study were that EndoFLIP variables would be stable between anesthestic episodes and would not be affected by body position when evaluating the upper and lower esophageal sphincters in healthy dogs. During each of three consecutive general anesthesia episodes administered to eight healthy adult research colony dogs with a standardized protocol, the EndoFLIP catheter was positioned to measure cross-sectional area, intrabag pressure, upper and lower esophageal sphincter length at two different balloon fill volumes (30 and 40 mL) and two body positions (lateral and dorsal recumbency). From these measured variables, a distensibility index was also calculated. Mixed effect analysis of variance was used to evaluate the fixed marginal and interaction effects of anesthesia episode, body position, and balloon volume on measured and calculated variables. For the upper esophageal sphincter significant interactions were present between anesthetic episode and body position for all variables except intrabag pressure; adjusting for body position significant differences were present between anesthetic episodes for all variables except distensibility index; adjusting for anesthetic episode cross-sectional area, intrabag pressure, upper esophageal sphincter length and distensibility index were all affected by body position. For the lower esophageal sphincter distensibility index was the only variable where a significant interaction between anesthesia episode and body position occurred; cross-sectional area, intrabag pressure, and lower esophageal length were not significantly affected by anesthesia episode when adjusting for body position; distensibility index was the only variable significantly affected by body position. Measurements of the geometry of the lower esophageal sphincter as measured by the EndoFLIP device were consistent under conditions of general anesthesia. Similar measurements taken at the upper esophageal sphincter displayed greater variability between anesthetic episodes and were affected to a greater extent by body position. Body position should be standardized in studies using the EndoFLIP to assess geometric and functional characteristics of the upper and lower esophageal sphincters.

Engineering plant membranes using droplet interface bilayers.

Droplet interface bilayers (DIBs) have become widely recognised as a robust platform for constructing model membranes and are emerging as a key technology for the bottom-up assembly of synthetic cell-like and tissue-like structures. DIBs are formed when lipid-monolayer coated water droplets are brought together inside a well of oil, which is excluded from the interface as the DIB forms. The unique features of the system, compared to traditional approaches (e.g., supported lipid bilayers, black lipid membranes, and liposomes), is the ability to engineer multi-layered bilayer networks by connecting multiple droplets together in 3D, and the capability to impart bilayer asymmetry freely within these droplet architectures by supplying droplets with different lipids. Yet despite these achievements, one potential limitation of the technology is that DIBs formed from biologically relevant components have not been well studied. This could limit the reach of the platform to biological systems where bilayer composition and asymmetry are understood to play a key role. Herein, we address this issue by reporting the assembly of asymmetric DIBs designed to replicate the plasma membrane compositions of three different plant species; Arabidopsis thaliana, tobacco, and oats, by engineering vesicles with different amounts of plant phospholipids, sterols and cerebrosides for the first time. We show that vesicles made from our plant lipid formulations are stable and can be used to assemble asymmetric plant DIBs. We verify this using a bilayer permeation assay, from which we extract values for absolute effective bilayer permeation and bilayer stability. Our results confirm that stable DIBs can be assembled from our plant membrane mimics and could lead to new approaches for assembling model systems to study membrane translocation and to screen new agrochemicals in plants.

Kinetics and potency of halothane, isoflurane, and desflurane in the Northern Leopard frog Rana pipiens.

Equilibration between delivered and effect site anesthetic partial pressure is slow in frogs. The use of less soluble agents or overpressure delivery may speed equilibration. Ten Northern leopard frogs were exposed to 3-4 constant concentrations of halothane, isoflurane or desflurane and their motor response to noxious electrical stimulation of the forelimb evaluated every 30 minutes until a stable proportion of frogs were immobile. Each frog received each anesthetic and concentration in random order and allowed at least 14 hours to recover between anesthetic exposures. An overpressure technique based upon the kinetics in the first study was then tested with 4 concentrations of desflurane. For halothane, isoflurane and desflurane respectively; the proportion of frogs immobile in response to stimulus became stable after 510, 480 and 180 minutes, and ED50 values were 1.36, 1.67 and 6.78 % atm. Desflurane ED50 delivered by overpressure was not significantly different at 6.85 % atm. Halothane, isoflurane and desflurane are effective general anesthetics in frogs with potencies similar to those reported in mammals. The time required for anesthetic equilibration is fastest with desflurane and can be hastened further by initial delivery of higher partial pressures.

Isoflurane potency in the northern leopard frog Rana pipiens is similar to that in mammalian species and is unaffected by decerebration.

Amphibians are commonly used in biomedical research, including studies of mechanisms of anaesthetic action. There is, however, little published work describing the kinetics of inhaled anaesthetic agents or the potency of isoflurane in amphibians. Ten Northern leopard frogs were exposed to a constant isoflurane concentration of 1.0%, 1.2% or 1.5% atm for 4 h, and their response to a noxious stimulus was tested every 20 min. Each frog was anaesthetized with each concentration in random order and allowed at least 16 h to recover between anaesthetic exposures. Frogs were then pithed and the protocol was repeated. Frogs first displayed immobility during stimulus application at 80 min, and the proportion of animals becoming immobile steadily increased to reach a stable level at 4 h. The 50% effective dose for isoflurane in intact and pithed frogs did not differ, and was 1.15 and 1.25% atm, respectively. The potency of isoflurane in leopard frogs was similar to that reported in mammalian species. Cutaneous uptake of anaesthetic is effective given sufficient time, approximately 4 h in this study. Forebrain structures appear to be unimportant for the immobilizing action of isoflurane in the frog.

Halothane and propofol differentially affect electroencephalographic responses to noxious stimulation.

BACKGROUND: Anaesthetics blunt neuronal responses to noxious stimulation, including effects on electroencephalographic (EEG) responses. It is unclear how anaesthetics differ in their ability to modulate noxious stimulation-evoked EEG activation. We investigated the actions of propofol and halothane on EEG responses to noxious stimuli, including repetitive electrical C-fibre stimulation, which normally evokes neuronal wind-up. METHODS: Rats were anaesthetized with halothane (n=8) or propofol (n=8), at 0.8x or 1.2x the amount required to produce immobility in response to tail clamping [minimum alveolar concentration (MAC) for halothane and median effective dose (ED(50)) for propofol]. We recorded EEG responses to repetitive electrical stimulus trains (delivered to the tail at 0.1, 1 and 3 Hz) as well as supramaximal noxious tail stimulation (clamp; 50 Hz electrical stimulus, each for 30 s). RESULTS: Under halothane anaesthesia, noxious stimuli evoked an EEG activation response manifested by increased spectral edge frequency (SEF) and median edge frequency (MEF). At 0.8 MAC halothane, the tail clamp increased the MEF from approximately 6 to approximately 8.5 Hz, and the SEF from approximately 25.5 to approximately 27 Hz. At both 0.8 and 1.2 MAC halothane, similar patterns of EEG activation were observed with the 1 Hz, 3 Hz and tetanic stimulus trains, but not with 0.1 Hz stimulation, which does not evoke wind-up. Under propofol anaesthesia, noxious stimuli were generally ineffective in causing EEG activation. At 0.8 ED(50) propofol, only the tail clamp and 1 Hz stimuli increased MEF ( approximately 8 to approximately 10-10.5 Hz). At the higher propofol infusion rate (1.2 ED(50)) the repetitive electrical stimuli did not evoke an EEG response, but the tetanic stimulus and the tail clamp paradoxically decreased SEF (from approximately 23 to approximately 21.5 Hz). CONCLUSIONS: Propofol has a more significant blunting effect on EEG responses to noxious stimulation compared with halothane.

Intrathecal picrotoxin minimally alters electro-encephalographic responses to noxious stimulation during halothane and isoflurane anesthesia.

BACKGROUND: Isoflurane and halothane act in the spinal cord to blunt ascending transmission of impulses to the brain resulting from noxious stimulation. Because intrathecal picrotoxin (an antagonist at the gamma-aminobutyric acid-A receptor) partially reverses the immobilizing effect of isoflurane and halothane, we hypothesized that the electroencephalographic response to noxious stimulation would likewise be partially reversed by intrathecal picrotoxin. METHODS: Rats were anesthetized with isoflurane (n = 8) or halothane (n = 8) and a laminectomy performed. Following determination of minimum alveolar concentration (MAC), the electroencephalogram (EEG) was recorded during separate applications of a hindpaw clamp, tail clamp and electrical current to the tail at 0.8 and 1.2 MAC. Picrotoxin was then applied to the exposed spinal cord and the EEG response to noxious stimulation again determined. RESULTS: The EEG was more active during halothane anesthesia than isoflurane (spectral edge frequency for 95% power: 25.6 +/- 2.1 Hz vs. 23.1 +/- 1.6 Hz, P < 0.05). Noxious stimulation usually caused the EEG to shift to higher frequencies (e.g. for 0.8 MAC halothane, median edge frequency for 50% power: from 7.6 +/- 3.1 Hz to 10.7 +/- 2.6 Hz, P < 0.05). Picrotoxin minimally affected this response. CONCLUSIONS: Noxious stimulation evokes an EEG response that is minimally altered by intrathecal picrotoxin. This suggests that isoflurane and halothane do not have GABAergic actions in the spinal cord that indirectly suppress the EEG response.

Animal dependence of inhaled anaesthetic requirements in cats.

BACKGROUND: The minimum alveolar concentration (MAC) of an inhaled anaesthetic describes its potency as a general anaesthetic. Individuals vary in their sensitivity to anaesthetics and we sought to determine whether an individual animal's sensitivity to inhaled anaesthetics would be maintained across different agents. METHODS: Six female mongrel cats, age 2 yr (range 1.8-2.3) and mean weight 3.5 (SD 0.3) kg, were studied on three separate occasions over a 12-month period to determine the MAC of isoflurane, sevoflurane and desflurane. Induction of anaesthesia in a chamber was followed by orotracheal intubation and maintenance of anaesthesia with the inhaled agent in oxygen delivered via a non-rebreathing circuit. MAC was determined in triplicate using standard tail-clamp technique. RESULTS: Mean MAC values for isoflurane, sevoflurane and desflurane were 1.90 (SD 0.18), 3.41 (0.65) and 10.27 (1.06)%, respectively. Body temperature, systolic pressure and Sp(O(2)) recorded at the time of MAC determinations for isoflurane, sevoflurane and desflurane were 38.3 (0.3), 38.6 (0.1) and 38.3 (0.3) degrees C; 71.2 (8.3), 74.6 (15.9) and 88.0 (12.0) mmHg; 99.2 (1.1), 99.1 (1.3) and 99.4 (0.8)%, respectively. Both the anaesthetic agent and the individual cat had significant effects on MAC. Correlation coefficients for comparisons between desflurane and isoflurane, desflurane and sevoflurane, and sevoflurane and isoflurane were 0.90, 0.89 and 0.97, respectively. CONCLUSIONS: These findings show that an individual has a consistent degree of sensitivity to a variety of inhaled anaesthetics, suggesting a genetic basis for sensitivity to inhaled anaesthetic effects.

Relationship between excitation energy transfer, trapping, and antenna size in photosystem II.

We present a systematic study of the effect of antenna size on energy transfer and trapping in photosystem II. Time-resolved fluorescence experiments have been used to probe a range of particles isolated from both higher plants and the cyanobacterium Synechocystis 6803. The isolated reaction center dynamics are represented by a quasi-phenomenological model that fits the extensive time-resolved data from photosystem II reaction centers and reaction center mutants. This representation of the photosystem II "trapping engine" is found to correctly predict the extent of, and time scale for, charge separation in a range of photosystem II particles of varying antenna size (8-250 chlorins). This work shows that the presence of the shallow trap and slow charge separation kinetics, observed in isolated D1/D2/cyt b559 reaction centers, are indeed retained in larger particles and that these properties are reflected in the trapping dynamics of all larger photosystem II preparations. A shallow equilibrium between the antennae and reaction center in photosystem II will certainly facilitate regulation via nonphotochemical quenching, and one possible interpretation of these findings is therefore that photosystem II is optimized for regulation rather than for efficiency.

A pilot study of protein sparing in healthy dogs using peripheral parenteral nutrition.

Total parenteral nutrition is the standard nutritional support of dogs when the enteral route is contraindicated, but it can be difficult because of cost, technical difficulties, and potential complications. Peripheral parenteral nutrition (PPN) may be a feasible option for short-term support in some cases. The objectives of this study were to determine the effect of PPN on nitrogen balance (as an indicator of the effect on protein sparing), serum folate concentrations and serum insulin-like growth factor-I (IGF - I) concentrations in fasting dogs. The effect of PPN on these parameters has not previously been reported in dogs. Using a cross-over design, three healthy adult fasting dogs were randomly assigned to three treatments: 5 per cent amino acid solution, 5 per cent glucose solution, and a control electrolyte solution. The solutions were administered into a peripheral vein at 60 ml kg(-1)per day for 4 days. The amino acid infusion resulted in a positive nitrogen balance and the glucose infusion produced less nitrogen loss than the control. Amino acid, but not glucose or electrolyte infusions, decreased serum folate concentrations. Amino acid and glucose infusions resulted in higher serum IGF -I concentrations than electrolyte infusions, although the differences were small and IGF -I decreased in all cases. In conclusion, these findings suggest that PPN increases nitrogen balance in healthy dogs undergoing short-term fasting.

Modulation of quantum yield of primary radical pair formation in photosystem II by site-directed mutagenesis affecting radical cations and anions.

Pigment-protein interactions play a significant role in determining the properties of photosynthetic complexes. Site-directed mutants of Synechocystis PCC 6803 have been prepared which modify the redox potential of the primary radical pair anion and cation. In one set of mutants, the environment of P680, the primary electron donor of Photosystem II, has been modified by altering the residue at D1-His198. It has been proposed that this residue is an axial ligand to the magnesium cation. In the other set, the D1-Gln130 residue, which is thought to interact with the C9-keto group of the pheophytin electron acceptor, has been changed. The effect of these mutations is to alter the free energy of the primary radical pair state, which causes a change in the equilibrium between excited singlet states and radical pair states. We show that the free energy of the primary radical pair can be increased or decreased by modifications at either the D1-His198 or the D1-Gln130 sites. This is demonstrated by using three independent measures of quantum yield and equilibrium constant, which exhibit a quantitative correlation. These data also indicate the presence of a fast nonradiative decay pathway that competes with primary charge separation. These results emphasize the sensitivity of the primary processes of PS II to small changes in the free energy of the primary radical pair.

Comparison of methods to assess dog owners' therapeutic compliance.

OBJECTIVE: To compare different methods for assessing the compliance of veterinary clients administering medication to their dogs. PROCEDURE: Thirty-one owners whose dogs were prescribed amoxycillin-clavulanate, twice and thrice daily, for a duration of five to seven days were recruited from three Sydney veterinary hospitals. Compliance was assessed by electronic monitoring devices, return medication counts, client self-reports and veterinarians' estimation of likely compliance. RESULTS: Electronic monitoring showed owners administered on average 84% (range 7 to 104%) of prescribed medication to their dogs. Both return medication counts and client self-reports tended to overestimate therapeutic compliance when compared with electronic monitoring. When questioned, the majority of owners (71%) claimed perfect compliance with the prescribed regimen. No correlation was found between veterinarians' estimates of owner compliance and that assessed electronically. CONCLUSION: Electronic monitoring provided valuable information on dose timing and variation, but proved costly and difficult to employ in veterinary practice. Simplicity and low cost of return medication counts makes this method attractive for use in veterinary compliance studies. Client self-reports reliably detected some noncompliers and permitted identification of individual problems or errors. For practical purposes a combination of return medication counts and client self-reports may have merit in future veterinary compliance studies.

Owner compliance with short term antimicrobial medication in dogs.

The degree to which dog owners complied with instructions to administer a 5 to 10 day course of antimicrobial medication to their pets was assessed using microprocessor based monitoring devices. Twenty two clients gave an average of 84% of prescribed doses of amoxycillin-clavulanate. No difference was found between twice and thrice daily dosing regimens in the overall percentage of prescribed doses given. However, timing of doses was far from ideal in many cases and only 34% of doses were given within the designated optimum time period. Adherence to desired dosing intervals tended to be better with twice daily than with thrice daily dosing, although the difference was statistically insignificant.