In vitro maturation is associated with increased early embryo arrest without impairing morphokinetic development of useable embryos progressing to blastocysts.

STUDY QUESTIONS: Does polycystic ovarian syndrome (PCOS) or in vitro maturation (IVM) treatment affect embryo development events and morphokinetic parameters after time-lapse incubation? SUMMARY ANSWER: There was an increase in some abnormal phenotypic events in PCOS-IVM embryos as well as an increase in early arrest of PCOS-IVM and PCOS-ICSI embryos; however, IVM treatment or PCOS status did not alter morphokinetic development of embryos suitable for transfer of vitrification. WHAT IS KNOWN ALREADY: IVM has been less successful than standard IVF in terms of clinical pregnancy, implantation and live birth rates. There is currently no information available about the development of IVM embryos according to time-lapse analysis. STUDY DESIGN, SIZE AND DURATION: This article represents a prospective case-control study. The study involved 93 participants who underwent 93 treatment cycles. Cycles were completed between January 2013 and July 2014. PARTICIPANTS/MATERIALS, SETTING AND METHODS: Participants were recruited for the study at Fertility Specialists of WA and Fertility Specialists South, Perth, Western Australia. Of the PCOS diagnosed patients, 32 underwent IVM treatment (PCOS-IVM) and 23 had standard ICSI treatment (PCOS-ICSI). There were 38 patients without PCOS who underwent standard ICSI treatment comprising the control group (control-ICSI). MAIN RESULTS AND THE ROLE OF CHANCE: The PCOS-IVM group showed significantly more embryos with multinucleated two cells (P = 0.041), multinucleated four cells (P = 0.001) and uneven two cells (P = 0.033) compared with the control-ICSI group, but not the PCOS-ICSI group. There were no significant differences in the rates of any abnormal events between the PCOS-ICSI and control-ICSI groups. Embryo arrest between Days 2 and 3 was higher in the PCOS-IVM and PCOS-ICSI groups compared with the control-ICSI group (P < 0.001 and P = 0.001). Embryo arrest from Days 3 to 4 was higher in the PCOS-IVM group compared with both the PCOS-ICSI and control-ICSI groups (P < 0.001). There were no differences in embryo arrest rates across all three groups at the compaction or blastulation stages. Cumulative rates of embryo arrest, from the time to second polar body extrusion (tPB2) to the time to formation of a blastocyst (tB), result in a decreased proportion of useable PCOS-IVM blastocysts compared with the other two treatment groups; however, of the embryos remaining, there was no significant difference in morphokinetic development between the three groups. LIMITATIONS AND REASONS FOR CAUTION: This was a small study using time-lapse analysis of embryo development as the primary end-point. Larger, randomized, clinical trials are required to clarify the implications of time-lapse incubation of IVM embryos and the effects on implantation and ongoing pregnancy. WIDER IMPLICATIONS OF THE FINDINGS: This is the first study to compare the time-lapse analysis of IVM with standard ICSI for patients with and without PCOS. This allows for a more detailed and specific timeline of events from embryos generated using this approach for patients diagnosed with PCOS and shows that embryos generated from IVM have an increased rate of early embryo arrest, however; morphokinetic development is not impaired in embryos that progress to the useable blastocyst stage. STUDY FUNDING/COMPETING INTERESTS: The study was supported by the Women's and Infant's Research Foundation of Western Australia. R.H. is the Medical Director of Fertility Specialists of Western Australia and a shareholder in Western IVF. He has received educational sponsorship from MSD, Merck-Serono and Ferring Pharmaceuticals. The other authors have no competing interests.

In vitro maturation as an alternative to standard in vitro fertilization for patients diagnosed with polycystic ovaries: a comparative analysis of fresh, frozen and cumulative cycle outcomes.

STUDY QUESTION: Is in vitro maturation (IVM) as successful as standard in vitro fertilization (IVF) for the treatment of patients with polycystic ovaries (PCO) in terms of fresh, frozen and cumulative pregnancy outcomes? SUMMARY ANSWER: There was no difference in clinical pregnancy rates in fresh or frozen embryo transfer (FET) cycles between the two treatment groups however, the IVM group showed a lower clinical pregnancy rate cumulatively. There was significantly fewer live births resulting from IVM treatment for both fresh and cumulative cycle outcomes however, there was no difference in live birth rates resulting from FETs between IVM and IVF treatment. WHAT IS KNOWN ALREADY: IVM is well recognized as the only treatment option to eliminate completely the incidence of ovarian hyperstimulation syndrome. However, historically IVM has been less successful than standard IVF in terms of clinical pregnancy, implantation and live birth rates. STUDY DESIGN, SIZE, AND DURATION: This paper represents a retrospective case-control study. The study involved 121 participants who underwent 178 treatment cycles. Cycles were completed between March 2007 and December 2012. All fresh cycles and subsequent FET cycles were included in the analysis to calculate cumulative outcomes. PARTICIPANTS/MATERIALS, SETTING, AND METHODS: All participants were prospectively diagnosed with PCO morphology or polycystic ovarian syndrome (PCOS) and underwent either IVM or standard IVF treatment. Their treatment outcomes were analysed with regard to embryological data, and the rate of biochemical pregnancy, clinical pregnancy and live birth, in addition maternal and neonatal outcomes were assessed. Fifty-six patients underwent 80 cycles of IVM treatment and 65 patients underwent 98 cycles of standard IVF treatment. MAIN RESULTS AND THE ROLE OF CHANCE: For fresh cycles, the differences in the biochemical pregnancy, clinical pregnancy or miscarriage rates between the two treatment groups were not statistically significant. The IVM group showed significantly lower live birth rates in fresh cycles in comparison to standard IVF treatment (18.8 versus 31.0%, P = 0.021). For frozen embryo transfer (FET) cycles the differences in biochemical pregnancy, clinical pregnancy, live birth or miscarriage rates between the two treatments groups were not statistically significant. The cumulative biochemical pregnancy (67.5 versus 83.7%, P = 0.018), clinical pregnancy (51.3 versus 65.3%, P = 0.021) and live birth rates (41.3 versus 55.1%, P = 0.005) were significantly lower in the IVM group in comparison to the standard IVF treatment group. There was no overall difference in the cumulative miscarriage rates between the two treatment groups. There was no difference between treatment methods with regard to the neonatal outcomes, and the IVM group had a significantly lower rate of ovarian hyperstimulation syndrome (0 versus 7.1%, P < 0.001). LIMITATIONS, REASONS FOR CAUTION: This was an observational study and further randomized clinical trials are required to clarify the difference in outcomes between standard IVF and IVM for patients with PCO/PCOS. WIDER IMPLICATIONS OF THE FINDINGS: This is the first study to compare IVM with standard IVF in PCO/PCOS patients using blastocyst development and single embryo transfer. Furthermore, it is the first study to show the results of fresh, frozen and cumulative treatment cycle outcomes between the two groups. Our results show similar success rates to those reported from other groups, particularly in relation to the incidence of miscarriage in fresh IVM cycles and improved success from FET cycles. Maternal and neonatal outcomes are consistent with the limited literature available. STUDY FUNDING/COMPETING INTERESTS: The study was supported by the Women's and Infant's Research Foundation of Western Australia. Professor Hart is Medical Director of Fertility Specialists of Western Australia (FSWA) and a shareholder Western IVF. He has received educational sponsorship from MSD, Merck-Serono and Ferring Pharmaceuticals. T.H. is a consultant with FSWA and a shareholder in Western IVF. She has received educational sponsorship from MSD, Merck-Serono and Ferring Pharmaceuticals. The other authors have no competing interests.

Shaped by Their Environment: Variation in Blue Whale Morphology across Three Productive Coastal Ecosystems.

Species ecology and life history patterns are often reflected in animal morphology. Blue whales are globally distributed, with distinct populations that feed in different productive coastal regions worldwide. Thus, they provide an opportunity to investigate how regional ecosystem characteristics may drive morphological differences within a species. Here, we compare physical and biological oceanography of three different blue whale foraging grounds: (1) Monterey Bay, California, USA; (2) the South Taranaki Bight (STB), Aotearoa New Zealand; and (3) the Corcovado Gulf, Chile. Additionally, we compare the morphology of blue whales from these regions using unoccupied aircraft imagery. Monterey Bay and the Corcovado Gulf are seasonally productive and support the migratory life history strategy of the Eastern North Pacific (ENP) and Chilean blue whale populations, respectively. In contrast, the New Zealand blue whale population remains in the less productive STB year-round. All three populations were indistinguishable in total body length. However, New Zealand blue whales were in significantly higher body condition despite lower regional productivity, potentially attributable to their non-migratory strategy that facilitates lower risk of spatiotemporal misalignment with more consistently available foraging opportunities. Alternatively, the migratory strategy of the ENP and Chilean populations may be successful when their presence on the foraging grounds temporally aligns with abundant prey availability. We document differences in skull and fluke morphology between populations, which may relate to different feeding behaviors adapted to region-specific prey and habitat characteristics. These morphological features may represent a trade-off between maneuverability for prey capture and efficient long-distance migration. As oceanographic patterns shift relative to long-term means under climate change, these blue whale populations may show different vulnerabilities due to differences in migratory phenology and feeding behavior between regions. Spanish abstract La ecología y patrones de historia de vida de las especies a menudo se reflejan en la morfología animal. Las ballenas azules están distribuidas globalmente, con poblaciones separadas que se alimentan en diferentes regiones costeras productivas de todo el mundo. Por lo tanto, brindan la oportunidad de investigar cómo las características regionales de los ecosistemas pueden impulsar diferencias morfológicas dentro de una especie. Aquí, comparamos la oceanografía física y biológica de tres zonas de alimentación diferentes de la ballena azul: (1) Bahía de Monterey, California, EE. UU., (2) Bahía del sur de Taranaki (BST), Nueva Zelanda, y (3) Golfo de Corcovado, Chile. Adicionalmente, comparamos la morfología de las ballenas azules de estas regiones utilizando imágenes de aeronaves no tripuladas. La Bahía de Monterey y el Golfo de Corcovado son estacionalmente productivos y apoyan la estrategia migratoria de la historia de vida de las poblaciones de ballena azul chilena y del Pacífico Norte Oriental (PNO), respectivamente. Por el contrario, la población de ballena azul de Nueva Zelanda permanece en la menos productiva BST durante todo el año. Las tres poblaciones eran indistinguibles en cuanto a la longitud corporal total. Sin embargo, las ballenas azules de Nueva Zelanda tenían una condición corporal significativamente mayor a pesar de una menor productividad regional, potencialmente atribuible a su estrategia no migratoria que facilita un menor riesgo de desalineación espaciotemporal con oportunidades de alimentación disponibles de manera más consistente. Alternativamente, la estrategia migratoria de las poblaciones de ballenas PNO y chilena puede tener éxito cuando su presencia en las zonas de alimentación se alinea temporalmente con la abundante disponibilidad de presas. Documentamos diferencias en la morfología del cráneo y la aleta caudal entre poblaciones, que pueden estar relacionadas con diferentes comportamientos de alimentación adaptados a las características de hábitat y presas específicas para cada región. Estas características morfológicas pueden representar una compensación entre la maniobrabilidad para la captura de presas y una migración eficiente a larga distancia. A medida que los patrones oceanográficos cambian en términos de mediano a largo plazo debido al cambio climático, estas poblaciones de ballenas azules pueden mostrar diferentes vulnerabilidades debido a diferencias en la fenología migratoria y el comportamiento de alimentación entre regiones.

IVF versus ICSI for the fertilization of in-vitro matured human oocytes.

Traditional dogma suggests that intracytoplasmic sperm injection (ICSI) should be performed to ensure successful oocyte fertilization in an in-vitro maturation (IVM) cycle. This study postulated that there would be no difference in the fertilization rate when ICSI was compared with IVF. This hypothesis was tested in a randomized trial of IVF versus ICSI in IVM. A total of 150 immature oocytes were collected in eight cycles of IVM for patients diagnosed with polycystic ovarian syndrome (PCOS). Patients were primed with minimal FSH before transvaginal oocyte aspiration. Sibling oocytes were inseminated by 50% IVF and 50% ICSI. There was no significant difference in fertilization, useable or total blastocyst development between the two insemination technique groups. Clinical pregnancy results for combined fresh and cryopreserved transfers were identical between the two insemination techniques with a total of two fresh and five cryopreserved IVF-inseminated embryos resulting in three clinical pregnancies (42.9%) and five fresh and two cryopreserved ICSI-derived embryos resulting in three clinical pregnancies (42.9%). This research has shown IVF to be a legitimate fertilization technique for IVM oocytes in PCOS patients and provides a greater awareness of the use of a fertilization method previously not utilized with IVM. In-vitro maturation (IVM) is an alternative treatment method to traditional IVF. Due to the minimal use of stimulating hormones in this treatment, IVM has a lower risk of ovarian hyperstimulation syndrome, it can be used for fertility preservation in cancer patients and it is more cost conservative. Early research into the effects of IVM showed a hardening effect on the membrane surrounding the egg (the zona pellucida). It was initially believed that, to overcome this hardening in order to allow the egg to be fertilized, spermatozoa would need to be injected into the egg using intracytoplasmic sperm injection. Due to recent advances in hormonal stimulation protocols (FSH priming) and culture conditions, we postulated that, for patients suffering from polycystic ovarian syndrome (PCOS), fertilization, embryo development and clinical pregnancy would not be superior in the injected oocytes compared with those inseminated by IVF. We found that by using the two insemination techniques on sibling oocytes from eight PCOS patients, there was no significant difference in fertilization, useable or total blastocyst development (day 5 or 6 embryos) and that clinical pregnancy results were identical. This research provides a greater awareness of a fertilization technique which is not normally utilized for IVM treatment, providing a less invasive, more cost-effective approach for the patient.

Epidemiology, clinical features and outcomes of incident tuberculosis in children in Canada in 2013-2016: results of a national surveillance study.

PURPOSE: Childhood tuberculosis disease is difficult to diagnose and manage and is an under-recognised cause of morbidity and mortality. Reported data from Canada do not focus on childhood tuberculosis or capture key epidemiologic, clinical and microbiologic details. The purpose of this study was to assess demographics, presentation and clinical features of childhood tuberculosis in Canada. METHODS: We conducted prospective surveillance from 2013 to 2016 of over 2700 paediatricians plus vertical tuberculosis programmes for incident tuberculosis disease in children younger than 15 years in Canada using the Canadian Paediatric Surveillance Program (CPSP). RESULTS: In total, 200 cases are included in this study. Tuberculosis was intrathoracic in 183 patients of whom 86% had exclusively intrathoracic involvement. Central nervous system tuberculosis occurred in 16 cases (8%). Fifty-one per cent of cases were hospitalised and 11 (5.5%) admitted to an intensive care unit. Adverse drug reactions were reported in 9% of cases. The source case, most often a first-degree relative, was known in 73% of cases. Fifty-eight per cent of reported cases were Canadian-born Indigenous children. Estimated study rates of reported cases (per 100 000 children per year) were 1.2 overall, 8.6 for all Indigenous children and 54.3 for Inuit children. CONCLUSION: Childhood tuberculosis may cause significant morbidity and resource utilisation. Key geographies and groups have very high incidence rates. Elimination of childhood tuberculosis in Canada will require well-resourced community-based efforts that focus on these highest risk groups.

In vitro developmental potential of macaque oocytes, derived from unstimulated ovaries, following maturation in the presence of glutathione ethyl ester.

BACKGROUND: The inadequacies of oocyte in vitro maturation (IVM) systems for both non-human primates and humans are evidenced by reduced fertilization and poor embryonic development, and may be partly explained by significantly lower glutathione (GSH) contents compared with in vivo matured (IVO) oocytes. As this influence has not been fully explored, this study investigated the effect of the GSH donor, glutathione ethyl ester (GSH-OEt), on the IVM and development of macaque oocytes as a model of human oocyte IVM. METHODS: Macaque oocytes derived from unstimulated ovaries were cultured in mCMRL-1066 alone or supplemented with 3 or 5 mM GSH-OEt. In vitro matured oocytes were subjected to the GSH assay, fixed for the assessment of spindle morphology or prepared ICSI. Embryo development of zygotes cultured in mHECM-9 was assessed up to Day 9 post-ICSI. RESULTS Supplementation of the maturation medium with GSH-OEt significantly increased oocyte maturation and normal fertilization rates compared with control oocytes, but only 5 mM GSH-OEt significantly increased the oocyte and cumulus cell GSH content. Confocal microscopy revealed significant differences in the spindle morphology between IVO and control in vitro matured metaphase II oocytes. Oocytes matured with 5 mM GSH-OEt exhibited spindle area and spindle pole width similar to that seen in the IVO oocyte. While no significant differences were observed in blastocyst rates, addition of 3 mM GSH-OEt during IVM significantly increased the proportion of embryos developing to the 5-8 cell stage while 5 mM GSH-OEt significantly increased the proportion of morula-stage embryos compared with controls. CONCLUSIONS: Supplementation of the IVM medium with GSH-OEt promotes better maturation and normal fertilization of macaque oocytes compared with non-supplemented medium. However, further improvement of the primate oocyte IVM culture system is required to support better blastocyst development of oocytes derived from unstimulated ovaries.

Developmental potential of bovine oocytes following IVM in the presence of glutathione ethyl ester.

Glutathione (GSH) is synthesised during oocyte maturation and represents the oocyte's main non-enzymatic defence against oxidative stress. Inadequate defence against oxidative stress may be related to poor embryo quality and viability. In the present study, bovine oocytes were matured in vitro in the presence of GSH ethyl ester (GSH-OEt), a cell permeable GSH donor, and its effects on subsequent fertilisation and embryo development were assessed. GSH-OEt significantly increased the GSH content of IVM oocytes without affecting fertilisation or Day 3 cleavage rates. Maturation in the presence of GSH-OEt did not significantly increase the blastocyst rate compared with control oocytes. However, 5 mM GSH-OEt treatment resulted in significantly higher blastocyst total cell number. The GSH level of IVM oocytes was significantly decreased in the absence of cumulus cells and when cumulus-oocyte complexes were cultured in the presence of buthionine sulfoximine (BSO), an inhibitor of GSH synthesis. The addition of GSH-OEt to cumulus-denuded or BSO-treated oocytes increased the GSH content of bovine oocytes and restored the rate of normal fertilisation, but not embryo development, to levels seen in control oocytes. Thus, GSH-OEt represents a novel approach for effective in vitro elevation of bovine oocyte GSH and improvement in blastocyst cell number.

Oocyte glutathione and fertilisation outcome of Macaca nemestrina and Macaca fascicularis in in vivo- and in vitro-matured oocytes.

Fertilisation and development of IVM non-human primate oocytes is limited compared with that of in vivo-matured (IVO) oocytes. The present study describes the IVM of macaque oocytes with reference to oocyte glutathione (GSH). Timing of maturation, comparison of IVM media and cysteamine (CYS) supplementation as a modulator of GSH were investigated. A significantly greater proportion of oocytes reached MII after 30 h compared with 24 h of IVM. Following insemination, IVM oocytes had a significantly lower incidence of normal fertilisation (i.e. 2PN = two pronuclei and at least one polar body) and a higher rate of abnormal fertilisation (1PN = one pronucleus and at least one polar body) compared with IVO oocytes. Immunofluorescence of 1PN zygotes identified incomplete sperm head decondensation and failure of male pronucleus formation as the principal cause of abnormal fertilisation in IVM oocytes. The IVO oocytes had significantly higher GSH content than IVM oocytes. Cumulus-denuded oocytes had significantly lower GSH following IVM compared with immature oocytes at collection. Cysteamine supplementation of the IVM medium significantly increased the GSH level of cumulus-intact oocytes and reduced the incidence of 1PN formation, but did not improve GSH levels of the denuded oocyte. Suboptimal GSH levels in macaque IVM oocytes may be related to reduced fertilisation outcomes.

Dedicated orthogeriatric service reduces hip fracture mortality.

BACKGROUND: Hip fracture is a common serious injury afflicting the geriatric population and is associated with poor clinical outcomes, functional and walking disabilities and high 1-year mortality rates. A multidisciplinary approach has been shown to improve outcomes of geriatric patients with fragility fracture. AIMS: We piloted a dedicated orthogeriatric service for hip fracture patients to determine if the service facilitated a change in major patient outcomes, such as mortality, length of stay and dependency. METHODS: A dedicated orthogeriatrics service for hip fracture was established as a collaborative project between the Department of Geriatric Medicine and Department of Orthopaedic Surgery at a university teaching hospital. Orthogeriatrics service data were collected prospectively on an orthogeriatric filemaker database from July 2011 to July 2012 (N = 206). Data were compared to previously recorded data (Irish Hip Fracture Database) on a cohort of hip fracture patients admitted to the same orthopaedic trauma unit from July 2009 to July 2010 (N = 248). RESULTS: Patients in the orthogeriatric service group experienced significant reductions in 1-year mortality (χ2 = 13.34, P < 0.001), length of acute hospital stay (U = -3.77, P < 0.001) and requirements for further rehabilitation (χ 2 = 26.59, P < 0.001). Patients in the pre-service establishment group were significantly more dependent following their fracture than the patients in the orthogeriatric service group (χ 2 = 5.34, P = 0.021). CONCLUSIONS: A multidisciplinary management approach to fragility fracture of the femoral neck that involves comprehensive geriatric assessment, daily medical involvement of a geriatric team and specialised follow-up assessment leads to a significant reduction in mortality and improved outcomes.

Effects of the selective serotonin reuptake inhibitor, fluoxetine, on regional gastric contractility.

UNLABELLED: Selective serotonin reuptake inhibitors (SSRIs) are increasingly used to treat a variety of disorders but have gastrointestinal side-effects. AIM: To determine the effects of the SSRI, fluoxetine, on gastric smooth muscle contractility. METHODS: Fundic, antral, and pyloric circular muscle contractility of guinea pig muscle strips were measured in vitro. Fluoxetine was added in concentrations from 0.1 nmol L(-1) to 100 mumol L(-1). Receptor antagonists were used to determine the neural pathways involved. RESULTS: Fluoxetine caused concentration dependent contractions, which were greatest in fundus compared with the antrum or pylorus. The contractile effects of fluoxetine in the antrum were reduced by tetrodotoxin, atropine, phentolamine, and the 5-HT(4) receptor antagonist GR 113808. The contractile effects of fluoxetine in the fundus were reduced by atropine, phentolamine, and GR 113808. CONCLUSIONS: Fluoxetine affects gastric contractility with regional variability - contracting the fundus more than the antrum or pylorus. The fluoxetine contractile effect is reduced by tetrodotoxin, atropine, phentolamine, and a 5-HT(4) receptor antagonist. These results suggest fluoxetine interacts with muscarinic, alpha-adrenergic, and serotoninergic receptors and/or ongoing reuptake/release of serotonin in the stomach.

Predicting mortality following hip fracture: an analysis of comorbidities and complications.

BACKGROUND: Hip fracture is common in the geriatric population. These patients have multiple comorbidities that complicate treatment and recovery such that poor functional outcomes often result. Since functional outcomes are associated with comorbidities and complications it is important to define the contributing factors. AIMS: To describe comorbidities common to geriatric hip fracture patients and determine predictability of complications and mortality based on comorbidities. METHODS: Data in this study were sourced from information prospectively collected for evaluation of a new orthogeriatric service established at a University Teaching Hospital over the period of 1 year. RESULTS: The median age was 82 years (range 54-100) and 73 % were female (N = 206). Common comorbidities included hypertension (51 %), dementia (28 %), osteoporosis (19 %), ischaemic heart disease (IHD) (15 %) and chronic obstructive pulmonary disease (15 %). In predicting 1-year mortality based on comorbidities, the final model included age, IHD, delay to surgery and explained 26 % of the variability in mortality. Predicting 1-year mortality based on complications, the final model included age and respiratory complications and explained 26 % of the variability in mortality. There was a significant association between having respiratory complications and chronic obstructive pulmonary disease (p < 0.001) with 63 % of those with respiratory complications having chronic obstructive pulmonary disease. CONCLUSIONS: This study highlights specific patient comorbidities and medical complications that could be used to guide clinical assessment, management and targeted interventions that improve outcomes in this patient group.

The Community Assessment of Risk Instrument: Investigation of Inter-Rater Reliability of an Instrument Measuring Risk of Adverse Outcomes.

BACKGROUND: Frailty is increasingly common in community dwelling older adults and increases their risk of adverse outcomes. Risk assessment is implicit in the Aged Care Assessment Teams process, but few studies have considered the factors that influence the assessor's decision making or explored the factors that may contribute to their interpretation of risk. OBJECTIVE: to examine the inter-rater reliability of the Community Assessment of Risk Instrument (CARI), which is a new risk assessment instrument. DESIGN: A cohort study was used. SETTING AND PARTICIPANTS: A sample of 50 community dwelling older adults underwent comprehensive geriatric assessment by two raters: a geriatrician and a registered nurse. Procedure and measurements: Each participant was scored for risk by the two raters using the CARI. This instrument ranks risk of three adverse outcomes, namely i) institutionalisation, ii) hospitalisation and iii) death within the next year from a score of 1, which is minimal risk to 5, which is extreme risk. Inter-rater reliability was assessed with Gamma, Spearman correlation and Kappa statistics. Internal consistency was assessed with Cronbach's alpha. RESULTS: There were 30 female (mean age 82.23 years) and 20 male (mean age 81.75 years) participants. Items within domains showed good-excellent agreement. The gamma statistic was >0.77 on 6/7 Mental State items, 14/15 items in the Activities of Daily Living domain. In the Medical domain, 6/9 items had Gamma scores >0.80. The global domain scores correlated well, 0.88, 0.72 and 0.87. Caregiver network scores were 0.71, 0.73 and 0.51 for the three domains. Inter-rater reliability scores for global risk scales were 0.86 (institutionalisation) and 0.78 (death). The gamma statistic for hospitalisation was 0.29, indicative of lower inter-rater reliability. Cronbach's alpha was 0.86 and 0.83 for the Activities of Daily Living domain, 0.51 and 0.42 for the Mental state domain and 0.23 and 0.10 for the Medical state domain. CONCLUSIONS: Overall, the instrument shows good inter-rater reliability. Poor correlations on some items relate to poor communication of clinical data and variable interpretation based on professional background. Lack of internal consistency in the medical condition domain confirms the discrete nature of these variables.

A comparison of preterm and adult airway smooth muscle mechanics.

To determine whether airway smooth muscle undergoes a maturational change regarding force generation, length-tension relationships were determined in isolated trachealis strips from adult and preterm sheep. At the length of maximum force generation, passive active and total tensions of the adult muscle were 2.5 times greater than preterm values (P less than 0.001). KCl stimulation yielded a greater peak tension in the adult strips than in the preterm strips (P less than 0.01). Preterm strips required higher concentrations of KCl to initiate contractions and higher concentrations to reach peak tension. Acetylcholine- (ACh) induced contraction resulted in greater force development at each dose in the adult strips compared with preterm strips (P less than 0.001). The dose of ACh required to reach a half-maximal response was significantly less for the adult strips than for the preterm strips (P less than 0.005). These data demonstrate that both force generation and receptor sensitivity increase with age. This inability of immature smooth muscle to generate as much force as adult smooth muscle may help explain why very preterm neonates requiring intermittent positive-pressure ventilation are at risk for developing structural airway problems.

Epithelial modulation of trachealis muscle tension is calcium and temperature dependent.

The tracheobronchial epithelium produces inhibitory substance(s) that alter the tracheal smooth muscle tension. This study examined the effect of changes in extracellular Ca2+ and temperature in vitro on the tension response of rabbit trachealis muscle to mechanical removal of the epithelium. Tension during acetylcholine- and KCl-induced contractions was examined at 0, 0.75, 1.5, 2.5, and 5 mM bath Ca2+ concentrations and at 37, 30, 23, and 41 degrees C bath temperature. At most extracellular Ca2+ concentrations (i.e., 0.75, 1.5, 2.5, and 5 mM), epithelial removal shifted the acetylcholine concentration response approximately one-half log to the left (P less than 0.001 for each condition) but had no effect on the responses to KCl (P = NS). Reductions in bath Ca2+ to 0 mM eliminated the epithelial inhibitory effect on the acetylcholine response. In contrast to the effects of reductions in Ca2+, cooling the airway to 30 and 23 degrees C progressively diminished the magnitude of the epithelial inhibitory effect. Our results indicate that the influence of the tracheal epithelium on tracheal smooth muscle responses to constrictor agonists is substance specific and can be diminished by reductions in tracheal temperature and extracellular Ca2+ concentration.

PACAP and VIP inhibit pyloric muscle through VIP/PACAP-preferring receptors.

UNLABELLED: Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide with structural homology to vasoactive intestinal polypeptide (VIP). Two receptor types for PACAP have been described: PACAP preferring receptors are selective for PACAP; whereas VIP/PACAP preferring receptors have similar affinity for both PACAP and VIP. Both VIP and PACAP are present in enteric nerves at the pylorus. VIP is known to exert inhibitory effects on pyloric muscle; the effect of PACAP is unknown. The aims of this study were to determine the effect of PACAP on pyloric muscle and to characterize the PACAP receptor. METHODS: Rabbit pyloric muscle strips were cut parallel to circular muscle fibres and placed in muscle baths. The effect of PACAP and VIP were quantitated as percent of basal motility index (MI). RESULTS: PACAP-27, PACAP-38, and VIP had dose dependent inhibitory effects on the spontaneous phasic contractions of the pylorus. The PACAP-27- induced relaxation was inhibited by the PACAP receptor antagonist PACAP6-27, but was not affected by tetrodotoxin. VIP also had dose dependent inhibitory effects on pyloric muscle. The VIP relaxation was inhibited by PACAP6-27, but not affected by tetrodotoxin. CONCLUSIONS: These studies indicate that, similar to VIP, PACAP inhibits pyloric muscle. The inhibitory effect of the PACAP receptor antagonist on both PACAP and VIP-induced relaxation suggest that PACAP and VIP act at the same receptor, a VIP/PACAP preferring receptor.

Effect of modulating voltage-dependent calcium channels on cholecystokinin and acetylcholine-induced contractions of the guinea pig gallbladder.

UNLABELLED: The aim of this study was to investigate the role of extracellular Ca2+ utilization in cholecystokinin (CCK) and acetylcholine-induced guinea pig gallbladder contractions by using agents that modulate influx of extracellular Ca2+ through voltage-dependent calcium channels. METHODS: Guinea pig gallbladder muscle strips were studied isometrically at Lmax in vitro. RESULTS: (1) Acetylcholine and CCK caused dose-dependent contractions, with EDmax of 10(-4) and 10(-6) M, respectively. (2) Preventing influx of extracellular Ca2+ by incubation in Ca(2+)-free/0.1 mM EGTA solution inhibited the acetylcholine (10(-4) M)-induced contraction by 60 +/- 3% compared to only 46 +/- 5% (P < 0.05) for CCK (10(-6) M)-induced contraction. (3) Nifedipine (3 microM) inhibited the response to acetylcholine (10(-4) M) by 54 +/- 3%, compared to only 34 +/- 3% (P < 0.01) for CCK (10(-6) M). (4) Bay K 8644 (10(-7) M) significantly increased (P < 0.05) the contractile responses to low doses of each agonist: acetylcholine (10(-6) M) by 121 +/- 44% and CCK (10(-9) M) by 94 +/- 31%, but had no effect on the contraction to the EDmax of each agonist. CONCLUSIONS: These studies demonstrate: (1) acetylcholine and CCK cause guinea pig gallbladder contraction by both intracellular Ca2+ release and influx of extracellular Ca2+ through voltage-dependent calcium channels; (2) the CCK-induced contraction is more dependent on intracellular Ca2+ than is acetylcholine; and (3) acetylcholine and CCK-induced contractions can by modulated by manipulating influx of extracellular Ca2+ through voltage-dependent calcium channels.

Effects of clonidine and tricyclic antidepressants on gastric smooth muscle contractility.

To determine if and how clonidine and tricyclic antidepressants affect gastric contractility. Guinea pig fundic and antral circular muscle strips were studied in vitro. The effects of clonidine or amitriptyline added in graded concentrations on contractions to electric field stimulation (EFS), acetylcholine (ACh), and SP in the presence of N(epsilon)-nitro-l-arginine methyl ester (l-NAME) were studied. EFS produced frequency dependent contractions of fundic and antral muscle that were abolished by atropine or tetrodotoxin (TTX). ACh contractions were abolished by atropine but not TTX. Clonidine reduced contractile response to EFS but had no effect on ACh contractions. The threshold concentration of clonidine to inhibit EFS contractions was lower in the fundus than in the antrum. Amitriptyline reduced contractions to both EFS and ACh but not to SP. The threshold concentration of amitriptyline to inhibit EFS contractions was lower in the antrum than in the fundus. Both clonidine and amitriptyline affect gastric contractility. At threshold concentrations, clonidine affects fundic contractility whereas amitriptyline affects antral contractility. Clonidine affects gastric contractility in response to EFS but not to ACh, suggesting alpha-2 receptors on cholinergic nerves that reduce ACh release. Amitriptyline inhibits gastric contractility to EFS and ACh suggesting an inhibitory muscle effect.

Morphine alters preproenkephalin gene expression.

Rats made tolerant/dependent to morphine by s.c. implantation of drug pellets displayed a significant decrease in striatal preproenkephalin mRNA that persisted during the period of withdrawal. Levels of Met-enkephalin were normal at the end of treatment, but reduced after withdrawal. The direction and time-course of these alterations are consistent with roles for altered neuronal gene expression in the phenomena of opiate tolerance and dependence.

Cross-tolerance between morphine- and bombesin-induced inhibition of intestinal transit in rats.

Intracerebroventricular (i.c.v.) injection of either morphine or bombesin to rats inhibits intestinal transit of an intraduodenally administered radiochromium marker. In this work, we show that tolerance develops to this effect of bombesin after i.c.v. infusion of the peptide (0.5 micrograms/h for 4 days via an s.c. implanted Alzet 2001 osmotic minipump). Tolerance also develops to the inhibition of intestinal transit associated with i.c.v. morphine after s.c. injections of morphine. Bombesin-induced delay of transit is not attenuated by naltrexone (10 mg/kg, s.c.), a standard narcotic antagonist. Nevertheless, two-way cross-tolerance develops between bombesin and morphine in this system. This is a surprising result since both bombesin and morphine are believed to act on different receptors and cause opposite effects on intestinal motility in rats.