The relationship between Motorcycle Rider Behaviour Questionnaire scores and crashes for riders in Australia.

Motorcycle riders are over-represented in road fatalities in Australia. While riders represent 18% of the road users killed each year, motorcycle registrations constitute only 4.5% of the registered vehicle fleet. The Motorcycle Rider Behaviour Questionnaire (MRBQ) was developed with a view toward understanding behaviours likely to be associated with crash risk. These include behaviours that are either intentional (such as violations of road and speed regulations and stunts) or unintentional (such as errors relating to traffic or control of the motorcycle), as well as protective behaviours related to use of safety equipment. The dual aims of the current study were, first, to determine the appropriate structure of a modified version of the MRBQ for use in a representative sample of riders in Australia and, second, to understand which MRBQ factors are associated with crash involvement. A stratified sampling procedure was undertaken to ensure the socio-economic status of local government area, age and gender of the sample was representative of the broader population of riders in New South Wales, Australia. The sample consisted of 470 riders (males=89%). Exploratory factor analysis revealed a 29-item, five factor structure was suitable on the Australian data encompassing traffic errors, speed violations, protective gear, control errors and stunts. Overall, riders reported relatively safe behaviours, with frequent use of protective gear and infrequent aberrant behaviours. However, even though infrequent, violations of speed and errors related to control of the motorcycle increased the odds of near-crash involvement, whilst stunt behaviours were associated with increased odds of crash involvement. Interventions and countermeasures need to target these specific behaviours.

The place of occupational therapy in rehabilitation strategies of complex regional pain syndrome: Comparative study of 60 cases.

The purpose of the study was to assess the value of combining occupational therapy (OT) with physical therapy (PT) for the rehabilitation of complex regional pain syndrome (CRPS) and to measure its effectiveness on activities of daily life. Sixty patients with CRPS type 1 were recruited and interviewed between September 1, 2014 and February 1, 2015. Thirty patients had undergone PT and thirty had undergone PT+OT. They were administered the short-form of the "Assessment of Life Habits" questionnaire (v.3.0 LIFE-H) created in Canada. This questionnaire consists of 16 items exploring activities of daily living, which were used to compare the effectiveness of the two rehabilitation protocols. The results of each test were submitted to the Wilcoxon test. After confirming the complexity of CRPS in terms of its etiology, clinical signs and progression, rehabilitation was effective, especially for pain. The patients who received PT+OT had on average 10% better dressing and undressing function, 25% better for meal preparation, and 20% better on personal care than those who underwent PT only. In CRPS, OT combined with PT brings a real benefit in restoring the essential activities of daily life. This strategy could be implemented as soon the diagnosis confirmed and continued for a very long time. It helps to avoid the risk of dependence on third parties.

Effectiveness of motorcycle protective clothing: riders' health outcomes in the six months following a crash.

BACKGROUND: Little is known about the contribution of protective clothing worn in motorcycle crashes to subsequent health-related outcomes, impairment and quality of life. METHODS: A prospective cohort of 212 adult motorcyclists were recruited following presentations to hospitals or crash repair services in a defined geographic area in Australia between June 2008 and July 2009. Data was obtained from participant interviews and medical records at baseline, then by mailed survey two and six months post-crash (n=146, 69%). The exposure factor was usage of protective clothing classified as full protection (motorcycle jacket and pants), partial protection (motorcycle jacket) and unprotected (neither). Outcomes of interest included general health status (Short Form SF-36), disability (Health Assessment Questionnaire) treatment and recovery progress, quality of life and return to work in the six months post-crash. Odds ratios (OR) were estimated for categorical outcomes using multiple logistic regression to assess differences in outcomes associated with levels of protection adjusted for potential confounders including age, sex, occupation, speed and type of impact. Non-parametric procedures were used for data that was not normally distributed. RESULTS: Compared to unprotected riders, both fully and partially protected riders had fewer days in hospital and reported less pain immediately post-crash; at two months both protection groups were less likely to have disabilities or reductions in physical function. By six months there were no significant differences in disability or physical function between groups, but both protection groups were more likely to be fully recovered and returned to pre-crash work than unprotected riders. Fully protected riders achieved better outcomes than either partially or unprotected riders on most measures. There were few significant differences between the full and partial protection groups although the latter showed greater impairment in physical health two months post-crash. CONCLUSIONS: We found strong associations between use of protective clothing and mitigation of the consequences of injury in terms of post-crash health and well-being. Given this evidence it seems likely that the use of protective clothing will confer significant benefits to riders in the event of a crash.

Nonresolving pneumonia and mimics of pneumonia.

Physicians caring for patients with community-acquired pneumonia are often faced with the dilemma of how to approach a patient with slowly resolving or even nonresolving pneumonia. When the radiograph has failed to resolve by 50% in 2 weeks or completely in 4 weeks, the pneumonia should be considered to be nonresolving or slowly resolving. The causes of a nonresolving pneumonia and an approach to the work-up are presented.

Mutually exclusive muscle designs: the power output of the locomotory and sonic muscles of the oyster toadfish (Opsanus tau).

Animals perform a vast array of motor activities. Although it has generally been accepted that muscles are well suited to the function that they must perform, specialization for performing one function may compromise their ability for carrying out another. We examined this principle in the toadfish muscular system: slow-twitch red and fast-twitch white myotomal muscles are used for powering swimming at relatively low frequencies, while the superfast swimbladder muscle powers mating calls by contracting at 100 Hz. We measured muscle power output over a wide range of frequencies. The red and white locomotory muscles could not generate power over ca. 2.2 and 12 Hz, respectively and, hence, could not power sound production. In contrast, the swimbladder muscle has many specializations that permit it to generate power at frequencies in excess of 100 Hz. However, these specializations drastically reduce its power output at low frequencies: the swimbladder muscle generated only one-twentieth of the power of the red muscle and one-seventh of the power of the white muscle at the frequencies used during swimming. To generate the same total power needed for swimming would require unfeasibly large amounts of swimbladder muscle that could not fit into the fish. Hence, the designs of the swimbladder and locomotory muscles are mutually exclusive.

cAMP-dependent fluid secretion in rat inner medullary collecting ducts.

We used an unambiguous in vitro method to determine if inner medullary collecting ducts (IMCD) have intrinsic capacities to absorb and secrete solutes and fluid in an isotonic medium. IMCD(1), IMCD(2), and IMCD(3) were dissected from kidneys of young Sprague-Dawley rats. 8-Bromo-3',5'-cyclic monophosphate (8-BrcAMP) stimulated lumen formation and progressive dilation in all IMCD subsegments; lumen formation was greatest in IMCD(1.) Benzamil potentiated the rate of lumen expansion in response to 8-BrcAMP. Fluid entered tubule lumens by transcellular secretion rather than simple translocation of intracellular fluid. Secreted lumen solutes were osmometrically active. Inhibition of protein kinase A with H-89 and Rp diastereomer of adenosine 3',5'-cyclic monophosphorothioate blocked fluid secretion. The rate of lumen expansion was reduced by the selective addition of ouabain, barium, diphenyl-2-carboxylate, bumetanide, glybenclamide, or DIDS, or reduction of extracellular Cl(-). We conclude that IMCD absorb and secrete electrolytes and fluid in vitro and that secretion is accelerated by cAMP. We suggest that salt and fluid secretion by the terminal portions of the renal collecting system may have a role in modulating the composition and volume of the final urine.

Assembly of vault-like particles in insect cells expressing only the major vault protein.

Vaults are the largest (13 megadalton) cytoplasmic ribonucleoprotein particles known to exist in eukaryotic cells. They have a unique barrel-shaped structure with 8-fold symmetry. Although the precise function of vaults is unknown, their wide distribution and highly conserved morphology in eukaryotes suggests that their function is essential and that their structure must be important for their function. The 100-kDa major vault protein (MVP) constitutes approximately 75% of the particle mass and is predicted to form the central barrel portion of the vault. To gain insight into the mechanisms for vault assembly, we have expressed rat MVP in the Sf9 insect cell line using a baculovirus vector. Our results show that the expression of the rat MVP alone can direct the formation of particles that have biochemical characteristics similar to endogenous rat vaults and display the distinct vault-like morphology when negatively stained and examined by electron microscopy. These particles are the first example of a single protein polymerizing into a non-spherically, non-cylindrically symmetrical structure. Understanding vault assembly will enable us to design agents that disrupt vault formation and hence aid in elucidating vault function in vivo.

Up-regulation of vaults may be necessary but not sufficient for multidrug resistance.

Vaults are ribonucleoprotein complexes comprised of the 100 kDa major vault protein (MVP), the 2 high m.w. vault proteins p193 (VPARP) and p240 (TEP1) and an untranslated small RNA (vRNA). Increased levels of MVP, vault-associated vRNA and vaults have been linked directly to non-P-glycoprotein-mediated multidrug resistance (MDR). To further characterize the putative role of vaults in MDR, expression levels of all of the vault proteins were examined in various MDR cell lines. Subcellular fractionation of vault particles revealed that all 3 vault proteins are increased in MDR cells compared to the parental, drug-sensitive cells. Furthermore, protein analysis of subcellular fractions of the drug-sensitive, MVP-transfected AC16 cancer cell line indicated that vault levels are increased, in this stable line. Since TEP1 is shared by both vaults and the telomerase complex, TEP1 protein (and vault) levels were compared with telomerase activity in a variety of cell lines, including various MDR lines. Our studies demonstrate that while vault levels may be a good predictor of drug resistance, their up-regulation alone is not sufficient to confer the drug-resistant phenotype. This implies a requirement of an additional factor(s) for vault-mediated MDR.

Quantitative electrophoretic analysis of myosin heavy chains in single muscle fibers.

To better understand the molecular basis of the large variation in mechanical properties of different fiber types, there has been an intense effort to relate the mechanical and energetic properties measured in skinned single fibers to those of their constituent cross bridges. There is a significant technical obstacle, however, in estimating the number of cross bridges in a single fiber. In this study, we have developed a procedure for extraction and quantification of myosin heavy chains (MHCs) that permits the routine and direct measurement of the myosin content in single muscle fibers. To validate this method, we also compared MHC concentration measured in single fibers with the MHC concentration in whole fast-twitch (psoas and gracilis) and slow-twitch (soleus) muscles of rabbit. We found that the MHC concentration in intact psoas (184 microM) was larger than that in soleus (144 microM), as would be expected from their differing mitochondrial content and volume of myofibrils. We obtained excellent agreement between MHC concentration measured at the single fiber level with that measured at the whole muscle level. This not only verifies the efficacy of our procedure but also shows that the difference in concentration at the whole muscle level simply reflects the concentration differences in the constituent fiber types. This new procedure should be of considerable help in future attempts to determine kinetic differences in cross bridges from different fiber types.

The influence of thermal acclimation on power production during swimming. I. In vivo stimulation and length change pattern of scup red muscle.

Ectothermal animals are able to locomote in a kinematically similar manner over a wide range of temperatures. It has long been recognized that there can be a significant reduction in the power output of muscle during swimming at low temperatures because of the reduced steady-state (i.e. constant activation and shortening velocity) power-generating capabilities of muscle. However, an additional reduction in power involves the interplay between the non-steady-state contractile properties of the muscles (i.e. the rates of activation and relaxation) and the in vivo stimulation and length change pattern the muscle undergoes during locomotion. In particular, it has been found that isolated scup (Stenotomus chrysops) red muscle working under in vivo stimulus and length change conditions (measured in warm-acclimated scup swimming at low temperatures) generates very little power for swimming. Even though the relaxation of the muscle has slowed greatly, warm-acclimated fish swim with the same tail-beat frequencies and the same stimulus duty cycles at cold temperatures, thereby not affording the slow-relaxing muscle any extra time to relax. We hypothesize that considerable improvement in the power output of the red muscle at low temperatures could be achieved if cold acclimation resulted in either a faster muscle relaxation rate or in the muscle being given more time to relax (e.g. by shortening the stimulus duration or reducing the tail-beat frequency). We test these hypotheses in this paper and the accompanying paper. Scup were acclimated to 10 degrees C (cold-acclimated) and 20 degrees C (warm-acclimated) for at least 6 weeks. Electromyograms (EMGs) and high-speed cine films were taken of fish swimming steadily at 10 degrees C and 20 degrees C. At 10 degrees C, we found that, although there were no differences in tail-beat frequency, muscle strain or stimulation phase between acclimation groups, cold-acclimated scup had EMG duty cycles approximately 20 % shorter than warm-acclimated scup. In contrast at 20 degrees C, there was no difference between acclimation groups in EMG duty cycle, nor in any other muscle length change or stimulation parameter. Thus, in response to cold acclimation, there appears to be a reduction in EMG duty cycle at low swimming temperatures that is probably due to an alteration in the operation of the pattern generator. This novel acclimation probably improves muscle power output at low temperatures compared with that of warm-acclimated fish, an expectation we test in the accompanying paper using the work-loop technique.

The influence of thermal acclimation on power production during swimming. II. Mechanics of scup red muscle under in vivo conditions.

We have previously shown that the power output of red muscle from warm-acclimated scup is greatly reduced when the fish swim at low temperatures. This reduction occurs primarily because, despite the slowing of muscle relaxation rate at cold temperatures, warm-acclimated scup swim with the same tail-beat frequency and the same stimulation durations, thereby not affording the slower-relaxing muscle any extra time to relax. We hypothesize that power output during swimming could be increased if the stimulus duration were reduced or if the relaxation rate of the red muscle were increased during cold acclimation. Scup were acclimated to 10 degrees C (cold-acclimated) and 20 degrees C (warm-acclimated) for at least 6 weeks. Cold acclimation dramatically increased the ability of scup red muscle to produce power at 10 degrees C. Power output measured from cold-acclimated muscle bundles driven through in vivo conditions measured from cold-acclimated scup swimming at 10 degrees C (i.e. work loops) was generally much greater than that from warm-acclimated muscle driven through its respective in vivo conditions at 10 degrees C. The magnitude of the increase depended both on the anatomical location of the muscle and on swimming speed. Integrated over the length of the fish, the red musculature from cold-acclimated fish generated 2.7, 8.9 and 5.8 times more power than the red musculature from warm-acclimated fish while swimming at 30 cm s(-)(1), 40 cm s(-)(1) and 50 cm s(-)(1), respectively. Our analysis suggests that the cold-acclimated fish should be able to swim in excess of 40 cm s(-)(1) with just their red muscle whereas the warm-acclimated fish must recruit their pink muscle well below this speed. Because the red muscle is more aerobic than the pink muscle, cold acclimation may increase the sustained swimming speed at which scup perform their long seasonal migrations at cool temperatures. We then explored the underlying mechanisms for the increase in muscle power output in cold-acclimated fish. Contrary to our expectations, cold-acclimated muscle did not have a faster relaxation rate; instead, it had an approximately 50 % faster activation rate. Our work-loop studies showed that this faster activation rate, alone, can increase the mechanical power production during cyclical contractions to a surprising extent. By driving cold-acclimated muscle through warm- and cold-acclimated in vivo conditions, we were able to partition the improvement in power production associated with increased activation rate and the approximately 20 % reduction in the duration of electromyographic activity found in the accompanying study. Depending on the position and swimming speed, approximately 60 % of the increase in power output was due to the change in the red muscle's contractile properties (i.e. faster activation); the remainder was due to the shorter stimulus duty cycle of cold-acclimated scup. Thus, by both shortening the in vivo stimulation duration and speeding up the rate of muscle activation as part of cold-acclimation, scup achieve a very large increase in the power output of their red muscle during swimming at low temperature. This increase in power output probably results in an increase in muscle efficiency and, hence, a reduction in the energetic cost of swimming. This increase in power output also reduces reliance on the less aerobic and less fatigue-resistant pink muscle. Both these abilities may increase the swimming speed at which prolonged aerobic muscle activity can occur and thus reduce the travel time for the long seasonal migrations in which scup engage.

The Telomerase/vault-associated protein TEP1 is required for vault RNA stability and its association with the vault particle.

Vaults and telomerase are ribonucleoprotein (RNP) particles that share a common protein subunit, TEP1. Although its role in either complex has not yet been defined, TEP1 has been shown to interact with the mouse telomerase RNA and with several of the human vault RNAs in a yeast three-hybrid assay. An mTep1(-/-) mouse was previously generated which resulted in no apparent change in telomere length or telomerase activity in six generations of mTep1-deficient mice. Here we show that the levels of the telomerase RNA and its association with the telomerase RNP are also unaffected in mTep1(-/-) mice. Although vaults purified from the livers of mTep1(-/-) mice appear structurally intact by both negative stain and cryoelectron microscopy, three-dimensional reconstruction of the mTep1(-/-) vault revealed less density in the cap than previously observed for the intact rat vault. Furthermore, the absence of TEP1 completely disrupted the stable association of the vault RNA with the purified vault particle and also resulted in a decrease in the levels and stability of the vault RNA. Therefore, we have uncovered a novel role for TEP1 in vivo as an integral vault protein important for the stabilization and recruitment of the vault RNA to the vault particle.

Superfast contractions without superfast energetics: ATP usage by SR-Ca2+ pumps and crossbridges in toadfish swimbladder muscle.

1. The rate at which an isometrically contracting muscle uses energy is thought to be proportional to its twitch speed. In both slow and fast muscles, however, a constant proportion (25-40 %) of the total energy has been found to be used by SR-Ca2+ pumps and the remainder by crossbridges. We examined whether SR-Ca2+ pumps account for a larger proportion of the energy in the fastest vertebrate muscle known (the toadfish swimbladder), and whether the swimbladder muscle utilizes energy at the superfast rate one would predict from its mechanics. 2. The ATP utilization rates of the SR-Ca2+ pumps and crossbridges were measured using a coupled assay system on fibres skinned with saponin. Surprisingly, despite its superfast twitch speed, the ATP utilization rate of swimbladder was no higher than that of much slower fast-twitch amphibian muscles. 3. The swimbladder achieves tremendous twitch speeds with a modest steady-state ATP utilization rate by employing two mechanisms: having a small number of attached crossbridges and probably utilizing intracellular Ca2+ buffers (parvalbumin) to spread out the time over which Ca2+ pumping can occur. 4. Finally, although the total ATP utilization rate was not as rapid as expected, the relative proportions used by SR-Ca2+ pumps and the crossbridges were similar to other muscles.

RNA location and modeling of a WD40 repeat domain within the vault.

The vault complex is a ubiquitous 13-MDa ribonucleoprotein assembly, composed of three proteins (TEP1, 240 kDa; VPARP, 193 kDa; and MVP, 100 kDa) that are highly conserved in eukaryotes and an untranslated RNA (vRNA). The vault has been shown to affect multidrug resistance in cancer cells, and one particular component, MVP, is thought to play a role in the transport of drug from the nucleus. To locate the position of the vRNA, vaults were treated with RNases, and cryo-electron microscopy (cryo-EM) was performed on the resulting complexes. Using single-particle reconstruction techniques, 3,476 particle images were combined to generate a 22-A-resolution structure. Difference mapping between the RNase-treated vault and the previously calculated intact vault reconstructions reveals the vRNA to be at the ends of the vault caps. In this position, the vRNA may interact with both the interior and exterior environments of the vault. The finding of a 16-fold density ring at the top of the cap has allowed modeling of the WD40 repeat domain of the vault TEP1 protein within the cryo-EM vault density. Both stoichiometric considerations and the finding of higher resolution for the computationally selected and refined "barrel only" images indicate a possible symmetry mismatch between the barrel and the caps. The molecular architecture of the complex is emerging, with 96 copies of MVP composing the eightfold symmetric barrel, and the vRNA together with one copy of TEP1 and four predicted copies of VPARP comprising each cap.

Prevalence of cocaine use and its impact on asthma exacerbation in an urban population.

OBJECTIVES: To assess the prevalence of cocaine use, and its impact on severity of presentation, among adults presenting to the emergency department (ED) with asthma. A secondary aim was to assess the use of various asthma treatment modalities, with reference to the 1997 National Asthma Education and Prevention Program (NAEPP) guidelines. METHODS: All adults aged 18 to 55 years who presented to the ED of this institution with an asthma attack, were approached about participating in the study, which required giving informed consent, answering a facilitated questionnaire, and giving a urine sample for drug screening. RESULTS: Patients were enrolled during a 7-month period. A total of 163 patients were approached to enter the study; 116 patients consented to participate in the study, with 103 submitting complete urine samples. Thirty-seven patients refused to participate, and 10 were excluded. Sixty-eight percent of the patients were women, with a mean age of 33 years. African-Americans made up 89% of the total group. Thirty-five percent were cigarette smokers. Urine cocaine tests were positive in 13 of 103 (13%); 6 of 103 (5.8%) were positive for opiates. In the cocaine-positive group, 5 of 13 patients (38%) were admitted to the hospital, including two patients requiring intubation and mechanical ventilation. Of the total group, 23 of 103 patients (22%) were admitted, and 5 of those 23 admitted patients (22%) were cocaine-positive. Length of stay was significantly longer (5 vs 2.5 days, p < 0.05) in the cocaine-positive admitted patients. Forty-six percent of all patients reported using inhaled corticosteroids (ICS), with 39% of admitted patients using them. Thirty-two percent of all patients had obtained three or more refills of their beta(2)-agonist inhaler in the previous month. CONCLUSIONS: The prevalence of cocaine use may be much higher than the 13% shown in this study, because of patients' refusal to participate in the study. Second, the severity of exacerbation appears to be worse in the cocaine-positive group. Finally, the majority of patients presenting did not use ICS in accordance with the NAEPP guidelines.

cAMP stimulates the in vitro proliferation of renal cyst epithelial cells by activating the extracellular signal-regulated kinase pathway.

BACKGROUND: : Cellular proliferation is a key factor in the enlargement of renal cysts in autosomal dominant polycystic kidney disease (ADPKD). We determined the extent to which adenosine 3':5'-cyclic monophosphate (cAMP) may regulate the in vitro proliferation of cyst epithelial cells derived from human ADPKD cysts. METHODS: : Epithelial cells from cysts of individuals with ADPKD and from normal human kidney cortex (HKC) of individuals without ADPKD were cultured. The effects of agonists and inhibitors on the rate of cellular proliferation and the activation of extracellular signal-regulated kinase (ERK1/2) were determined. RESULTS: : 8-Br-cAMP (100 micromol/L) stimulated the proliferation of cells from eight different ADPKD subjects to 99.0% above baseline; proliferation was inhibited by protein kinase A (PKA) antagonists H-89 (97%) and Rp-cAMP (90%). Forskolin (10 micromol/L), which activates adenylyl cyclase, increased proliferation 124%, and receptor-mediated agonists arginine vasopressin, desmopressin, secretin, vasoactive intestinal polypeptide, and prostaglandin E2 stimulated proliferation 54.2, 56.3, 46.7, 37.1, and 48.3%, respectively. The mitogen extracellular kinase (MEK) inhibitor PD98059 completely inhibited ADPKD cell proliferation in response to cAMP agonists, but genistein, a receptor tyrosine kinase inhibitor, did not block cAMP-dependent proliferation. cAMP agonists increased the activity of ERK above control levels within five minutes. In contrast to ADPKD, proliferation and ERK activity of cells derived from normal HKC were not stimulated by cAMP agonists, although electrogenic Cl- secretion was increased by these agonists in both ADPKD and HKC cell monolayers. CONCLUSIONS: : We conclude that cAMP agonists stimulate the proliferation of ADPKD but not HKC epithelial cells through PKA activation of the ERK pathway at a locus distal to receptor tyrosine kinase. We suggest that the adenylyl cyclase signaling pathway may have a unique role in determining the rate of cyst enlargement in ADPKD through its actions to stimulate cellular proliferation and transepithelial solute and fluid secretion.

The Mr 193,000 vault protein is up-regulated in multidrug-resistant cancer cell lines.

Vaults are 13 megadalton ribonucleoprotein particles composed largely of the major vault protein (MVP) and two high molecular weight proteins, p240 and p193, and a small vault RNA (vRNA). Increased levels of MVP expression, vault-associated vRNA, and vaults have been linked directly to multidrug resistance (MDR). To further define the putative role of vaults in MDR, we produced monoclonal antibodies against the Mr 193,000 vault protein and studied its expression levels in various multidrug-resistant cell lines. We find that, like MVP, p193 mRNA and protein levels are increased in various multidrug-resistant cell lines. Subcellular fractionation of vault particles revealed that vault-associated p193 levels are increased in multidrug-resistant cells as compared with the parental, drug-sensitive cells. Furthermore, protein analysis of postnuclear supernatants and co-immunoprecipitation studies show that drug-sensitive MVP-transfected tumor cells lack this up-regulation in vault-associated p193. Our observations indicate that vault formation is limited not only by the expression of the MVP but also by the expression or assembly of at least one of the other vault proteins.

Analysis of sulfatide from rat cerebellum and multiple sclerosis white matter by negative ion electrospray mass spectrometry.

The accumulation of sulfatide (sulfatogalactosyl cerebroside) and changes in the sulfatide species present have been examined in the cerebellum of day 6-32 aged rats and in multiple sclerosis (MS) tissue samples. Negative ion electrospray mass spectrometry with daughter and parent ion analyses were used to distinguish the fatty acyl character in the amide linkage of sulfatide; measurement was done by selected ion and multiple reaction monitoring of individually identified sulfatide molecules. Sulfatide accumulation in rat cerebellum shows that 18:0- and hydroxylated 18:0-sulfatide are the first sulfatide molecules detectable. Very long fatty acyl chain sulfatide molecules (>20:0) are present at day 7 and the ratio of non-hydroxylated compared to hydroxylated sulfatide rises as the amount of non-hydroxylated sulfatide increases. 24:1-sulfatide accumulates at a ratio of about 3:1 over 24:0-sulfatide during active myelination. Analyses of the sulfatide in human tissue have shown differences between MS plaque tissues, normal appearing adjacent white matter and control tissues. The findings show that total sulfatide is reduced by 60% in the plaque matter and decreased 25% in adjacent normal appearing white matter. There are significant increases (P=0.05) in the amount of hydroxylation of sulfatide, demonstrated by an increase in the percentage of hydroxylated h24:0-sulfatide (hydroxy-lignoceroyl sulfatide).

The influence of temperature on power production during swimming. I. In vivo length change and stimulation pattern.

Ectothermal animals are able to locomote effectively over a wide range of temperatures despite low temperature reducing the power output of their muscles. It has been suggested that animals recruit more muscle fibres and faster fibre types to compensate for the reduced power output at low temperature, but it is not known how much low temperature actually reduces power output in vivo. 'Optimized' work-loop measurements, which are thought to approximate muscle function in vivo, give a Q(10) of approximately 2.3 for power output of scup (Stenotomus chrysops) red muscle between 10 degrees C and 20 degrees C. However, because of the slower muscle relaxation rate at low temperatures, 'optimizing' work loops requires stimulation duration to be reduced and oscillation frequency to be decreased to obtain maximal power output. Previous fish swimming experiments suggest that similar optimization may not occur in vivo, and this may have substantial consequences in terms of muscle power generation and swimming at low temperatures. To assess more precisely the effects of temperature on muscle performance and swimming, in the present study, we measured the length change, stimulation duration and stimulus phase of red muscle at various positions along scup swimming at several speeds at 10 degrees C and 20 degrees C. In a companion study, we determined the effects of temperature on in vivo power generation by driving muscle fibre bundles through these in vivo length changes and stimulation conditions, and measuring the resulting power output. The most significant finding from the present study is that, despite large differences in the in vivo parameters along the length of the fish (a decrease in stimulus duration, an increase in strain and a negative shift in phase) moving posteriorly, these parameters do not change with temperature. Thus, although the nervous system of fish could, in theory, compensate for slow muscle relaxation by greatly reducing muscle stimulation duration at low temperatures, it does not. This lack of compensation to low temperatures might reflect a potential limitation in neural control.

The influence of temperature on power production during swimming. II. Mechanics of red muscle fibres in vivo.

We found previously that scup (Stenotomus chrysops) reduce neither their stimulation duration nor their tail-beat frequency to compensate for the slow relaxation rates of their muscles at low swimming temperatures. To assess the impact of this 'lack of compensation' on power generation during swimming, we drove red muscle bundles under their in vivo conditions and measured the resulting power output. Although these in vivo conditions were near the optimal conditions for much of the muscle at 20 degrees C, they were far from optimal at 10 degrees C. Accordingly, in vivo power output was extremely low at 10 degrees C. Although at 30 cm s(-)(1), muscles from all regions of the fish generated positive work, at 40 and 50 cm s(-)(1), only the POST region (70 % total length) generated positive work, and that level was low. This led to a Q(10) of 4-14 in the POST region (depending on swimming speed), and extremely high or indeterminate Q(10) values (if power at 10 degrees C is zero or negative, Q(10) is indeterminate) for the other regions while swimming at 40 or 50 cm s(-)(1). To assess whether errors in measurement of the in vivo conditions could cause artificially reduced power measurements at 10 degrees C, we drove muscle bundles through a series of conditions in which the stimulation duration was shortened and other parameters were made closer to optimal. This sensitivity analysis revealed that the low power output could not be explained by realistic levels of systematic or random error. By integrating the muscle power output over the fish's mass and comparing it with power requirements for swimming, we conclude that, although the fish could swim at 30 cm s(-)(1) with the red muscle alone, it is very unlikely that it could do so at 40 and 50 cm s(-)(1), thus raising the question of how the fish powers swimming at these speeds. By integrating in vivo pink muscle power output along the length of the fish, we obtained the surprising finding that, at 50 cm s(-)(1), the pink muscle (despite having one-third the mass) contributes six times more power to swimming than does the red muscle. Thus, in scup, pink muscle is crucial for powering swimming at low temperatures. This overall analysis shows that Q(10) values determined in experiments on isolated tissue under arbitrarily selected conditions can be very different from Q(10) values in vivo, and therefore that predicting whole-animal performance from these isolated tissue experiments may lead to qualitatively incorrect conclusions. To make a meaningful assessment of the effects of temperature on muscle and locomotory performance, muscle performance must be studied under the conditions at which the muscle operates in vivo.