"A perfect storm": Autistic experiences of menopause and midlife.

LAY ABSTRACT: Previous studies report that menopause can be a very difficult transition for some autistic people. This study focuses on how autistic people experience menopause and what support and information might help them. Autistic Community Research Associates played an important role in the research and co-authored this article. We held four focus groups and eight interviews online with 24 autistic participants who lived in either Canada (n = 13) or the United Kingdom (n = 11). We analysed participant conversations using a method called reflexive thematic analysis. Participants described many intense challenges during menopause. Four themes and eight subthemes were identified across participant groups: (1) Complexity, multiplicity and intensity of symptoms (0 subthemes); (2) Life experience and adversity converging at midlife (three subthemes); (3) The importance of knowledge and connection (two subthemes); and (4) Barriers to support and care (three subthemes). The experiences of our participants may not be the same as other autistic people, and the study could have been more inclusive of diverse autistic groups. However, hearing about the experiences of others may provide reassurance to autistic people who struggle with menopause and let them know they are not alone.

Advancements in Cortisol Detection: From Conventional Methods to Next-Generation Technologies for Enhanced Hormone Monitoring.

The hormone cortisol, released as the end-product of the hypothalamic-pituitary-adrenal (HPA) axis, has a well-characterized circadian rhythm that enables an allostatic response to external stressors. When the pattern of secretion is disrupted, cortisol levels are chronically elevated, contributing to diseases such as heart attacks, strokes, mental health disorders, and diabetes. The diagnosis of chronic stress and stress related disorders depends upon accurate measurement of cortisol levels; currently, it is quantified using mass spectroscopy or immunoassay, in specialized laboratories with trained personnel. However, these methods are time-consuming, expensive and are unable to capture the dynamic biorhythm of the hormone. This critical review traces the path of cortisol detection from traditional laboratory-based methods to decentralised cortisol monitoring biosensors. A complete picture of cortisol biology and pathophysiology is provided, and the importance of precision medicine style monitoring of cortisol is highlighted. Antibody-based immunoassays still dominate the pipeline of development of point-of-care biosensors; new capture molecules such as aptamers and molecularly imprinted polymers (MIPs) combined with technologies such as microfluidics, wearable electronics, and quantum dots offer improvements to limit of detection (LoD), specificity, and a shift toward rapid or continuous measurements. While a variety of different sensors and devices have been proposed, there still exists a need to produce quantitative tests for cortisol ─ using either rapid or continuous monitoring devices that can enable a personalized medicine approach to stress management. This can be addressed by synergistic combinations of technologies that can leverage low sample volumes, relevant limit of detection and rapid testing time, to better account for cortisol's shifting biorhythm. Trends in cortisol diagnostics toward rapid and continuous monitoring of hormones are highlighted, along with insights into choice of sample matrix.

Through the Eyes of a Young Carer: A Photo Elicitation Study of Protective Resilience.

Caregiving is recognised as a source of stress with potential for negative health impacts as well as positive outcomes and development of resilience. For young carers, children, and adolescents providing care for close family members, adaptation through resilience is crucial, yet work using a resilience approach is limited. This study explored protective factors and pathways to resilience in a sample of young carers, through application of the socioecological model in caring relationships. An in-depth qualitative approach was used, with in-person interviews facilitated by auto-driven photo elicitation. Deductive thematic analysis was applied, framed by three levels (individual, community, and society) of the socioecological model of resilience. Twelve participants (nine girls and three boys) aged 5-18 years, each providing care to a family member, were recruited using opportunity and volunteer sampling via carers' centres in the southwest of England. Ten key themes were identified, four at the individual level: pre-empting challenges and planning, cognitive strategies, emotional strategies, and seeking solitude; three at the community level: family support, friendships, and pets and inanimate objects; and three at the society level: professional support, access to caregiver activities and community, and being outdoors. The location of themes at each level indicated relevance of the socioecological model to identification of protective factors in a young carer population. These findings have important applications for guidance to charities and organisations supporting young carers. Identification of factors that promote resilience offers support for the development of well-informed interventions, which harness these protective factors to develop resilience and improve health for young carers.

Death comes for us all: relating movement-integrated habitat selection and social behavior to human-associated and disease-related mortality among gray wolves.

Avoiding death affects biological processes, including behavior. Habitat selection, movement, and sociality are highly flexible behaviors that influence the mortality risks and subsequent fitness of individuals. In the Anthropocene, animals are experiencing increased risks from direct human causes and increased spread of infectious diseases. Using integrated step selection analysis, we tested how the habitat selection, movement, and social behaviors of gray wolves vary in the two months prior to death due to humans (being shot or trapped) or canine distemper virus (CDV). We further tested how those behaviors vary as a prelude to death. We studied populations of wolves that occurred under two different management schemes: a national park managed for conservation and a provincially managed multi-use area. Behaviors that changed prior to death were strongly related to how an animal eventually died. Wolves killed by humans moved slower than wolves that survived and selected to be nearer roads closer in time to their death. Wolves that died due to CDV moved progressively slower as they neared death and reduced their avoidance of wet habitats. All animals, regardless of dying or living, maintained selection to be near packmates across time, which seemingly contributed to disease dynamics in the packs infected with CDV. There were no noticeable differences in behavior between the two management areas. Overall, habitat selection, movement, and sociality interact to put individuals and groups at greater risks, influencing their cause-specific mortality.

Erratum: The problem and promise of scale in multilayer animal social networks.

[This corrects the article DOI: 10.1093/cz/zoaa052.].

Bibliometric investigation of the integration of animal personality in conservation contexts.

Consistent individual differences in behavior, commonly termed animal personality, are a widespread phenomenon across taxa that have important consequences for fitness, natural selection, and trophic interactions. Animal personality research may prove useful in several conservation contexts, but which contexts remains to be determined. We conducted a structured literature review of 654 studies identified by combining search terms for animal personality and various conservation subfields. We scored the relevance of personality and conservation issues for each study to identify which studies meaningfully integrated the 2 fields as opposed to surface-level connections or vague allusions. We found a taxonomic bias toward mammals (29% of all studies). Very few amphibian or reptile studies applied personality research to conservation issues (6% each). Climate change (21%), invasive species (15%), and captive breeding and reintroduction (13%) were the most abundant conservation subfields that occurred in our search, though a substantial proportion of these papers weakly integrated conservation and animal personality (climate change 54%, invasive species 51%, captive breeding and reintroduction 40%). Based on our results, we recommend that researchers strive for consistent and broadly applicable terminology when describing consistent behavioral differences to minimize confusion and improve the searchability of research. We identify several gaps in the literature that appear to be promising and fruitful avenues for future research, such as disease transmission as a function of sociability or exploration as a driver of space use in protected areas. Practitioners can begin informing future conservation efforts with knowledge gained from animal personality research.

Investigación bibliométrica sobre la integración de la personalidad animal a los contextos de conservación Resumen Las diferencias individuales y constantes en el comportamiento, comúnmente llamadas personalidad animal, son un fenómeno generalizado en los taxones con consecuencias importantes para la aptitud, selección natural e interacciones tróficas. Las investigaciones sobre la personalidad animal pueden ser útiles en varios contextos de conservación, aunque falta determinar cuáles son estos contextos. Realizamos una revisión literaria estructurada de 654 estudios identificados mediante la combinación de los términos de búsqueda para la personalidad animal y varios subcampos de la conservación. Puntuamos la relevancia de la personalidad y los temas de conservación en cada estudio para identificar cuáles de estos integraron significativamente a ambos campos, contrario a las conexiones a nivel superficial o alusiones vagas. Descubrimos un sesgo taxonómico por los mamíferos (29% de todos los estudios). Pocos estudios enfocados en anfibios o reptiles aplicaron un estudio de personalidad a los temas de conservación (6% para cada uno). El cambio climático (21%), las especies invasoras (15%) y la reproducción en cautiverio y las reintroducciones (13%) fueron los subcampos de conservación más abundantes que aparecieron en nuestra búsqueda, aunque una proporción significativa de estos artículos integraron muy poco a la conservación y la personalidad animal (cambio climático 54%, especies invasoras 51%, reproducción en cautiverio y reintroducciones 40%). Con base en nuestros resultados, recomendamos que los investigadores procuren tener terminologías consistentes y de aplicación generalizada cuando describan las diferencias conductuales para así minimizar las confusiones y facilitar la búsqueda durante la investigación. Identificamos varios vacíos en la literatura que prometen ser vías fructíferas para las investigaciones en el futuro, como la transmisión de enfermedades como una función sociable o la exploración como un impulsor del uso del espacio en las áreas protegidas. Los practicantes pueden comenzar por guiar los siguientes esfuerzos de conservación con el conocimiento obtenido de las investigaciones sobre la personalidad animal.

The Influence of Geographical Origin, Age, Sex, and Animal Husbandry on the Spontaneous Histopathology of Laboratory Cynomolgus Macaques (Macaca Fascicularis): A Contemporary Global and Multisite Review of Historical Control Data.

To investigate the influence of geographical origin, age, and sex on toxicologically relevant spontaneous histopathology findings in cynomolgus macaques (Macaca fascicularis), we performed a comparative analysis of historical control data (HCD) from 13 test sites that included 3351 animals (1645 females and 1706 males) sourced from Mauritius, China, Vietnam, and Cambodia, aged from 2 to 9.5 years, and from 446 toxicology studies evaluated between 2016 and 2021. The most common findings were mononuclear infiltrates in the kidney, liver, brain, and lung, which showed highest incidences in Mauritian macaques, and heart, salivary glands, and gastrointestinal tract (GIT), which showed highest incidences of mononuclear infiltrates in mainland Asian macaques. Developmental and degenerative findings were more common in Mauritian macaques, while lymphoid hyperplasia and lung pigment showed higher incidences in Asian macaques. Various sex and age-related differences were also present. Despite origin-related differences, the similarities in the nature and distribution of background lesions indicate that macaques from all geographical regions are suitable for toxicity testing and show comparable lesion spectrum. However, in a toxicity study, it is strongly recommended to use animals from a single geographical origin and to follow published guidelines when using HCD to evaluate and interpretate commonly diagnosed spontaneous lesions.

Early-life social experience affects offspring DNA methylation and later life stress phenotype.

Studies in rodents and captive primates suggest that the early-life social environment affects future phenotype, potentially through alterations to DNA methylation. Little is known of these associations in wild animals. In a wild population of spotted hyenas, we test the hypothesis that maternal care during the first year of life and social connectedness during two periods of early development leads to differences in DNA methylation and fecal glucocorticoid metabolites (fGCMs) later in life. Here we report that although maternal care and social connectedness during the den-dependent life stage are not associated with fGCMs, greater social connectedness during the subadult den-independent life stage is associated with lower adult fGCMs. Additionally, more maternal care and social connectedness after den independence correspond with higher global (%CCGG) DNA methylation. We also note differential DNA methylation near 5 genes involved in inflammation, immune response, and aging that may link maternal care with stress phenotype.

Toxoplasma gondii infections are associated with costly boldness toward felids in a wild host.

Toxoplasma gondii is hypothesized to manipulate the behavior of warm-blooded hosts to promote trophic transmission into the parasite's definitive feline hosts. A key prediction of this hypothesis is that T. gondii infections of non-feline hosts are associated with costly behavior toward T. gondii's definitive hosts; however, this effect has not been documented in any of the parasite's diverse wild hosts during naturally occurring interactions with felines. Here, three decades of field observations reveal that T. gondii-infected hyena cubs approach lions more closely than uninfected peers and have higher rates of lion mortality. We discuss these results in light of 1) the possibility that hyena boldness represents an extended phenotype of the parasite, and 2) alternative scenarios in which T. gondii has not undergone selection to manipulate behavior in host hyenas. Both cases remain plausible and have important ramifications for T. gondii's impacts on host behavior and fitness in the wild.

The problem and promise of scale in multilayer animal social networks.

Scale remains a foundational concept in ecology. Spatial scale, for instance, has become a central consideration in the way we understand landscape ecology and animal space use. Meanwhile, scale-dependent social processes can range from fine-scale interactions to co-occurrence and overlapping home ranges. Furthermore, sociality can vary within and across seasons. Multilayer networks promise the explicit integration of the social, spatial, and temporal contexts. Given the complex interplay of sociality and animal space use in heterogeneous landscapes, there remains an important gap in our understanding of the influence of scale on animal social networks. Using an empirical case study, we discuss ways of considering social, spatial, and temporal scale in the context of multilayer caribou social networks. Effective integration of social and spatial processes, including biologically meaningful scales, within the context of animal social networks is an emerging area of research. We incorporate perspectives that link the social environment to spatial processes across scales in a multilayer context.

Early-life relationships matter: Social position during early life predicts fitness among female spotted hyenas.

How social development in early-life affects fitness remains poorly understood. Though there is growing evidence that early-life relationships can affect fitness, little research has investigated how social positions develop or whether there are particularly important periods for social position development in an animal's life history. In long-lived species in particular, understanding the lasting consequences of early-life social environments requires detailed, long-term datasets. Here we used a 25-year dataset to test whether social positions held during early development predicted adult fitness. Specifically, we quantified social position using three social network metrics: degree, strength and betweenness. We determined the social position of each individual in three types of networks during each of three stages of ontogeny to test whether they predict annual reproductive success (ARS) or longevity among adult female spotted hyenas Crocuta crocuta. The social positions occupied by juvenile hyenas did predict their fitness, but the effects of social position on fitness measures differed between stages of early development. Network metrics when individuals were young adults better predicted ARS, but network metrics for younger animals, particularly when youngsters were confined to the communal den, better predicted longevity than did metrics assessed during other stages of development. Our study shows how multiple types of social bonds formed during multiple stages of social development predict lifetime fitness outcomes. We suggest that social bonds formed during specific phases of development may be more important than others when considering fitness outcomes.

Small-molecule intramimics of formin autoinhibition: a new strategy to target the cytoskeletal remodeling machinery in cancer cells.

Although the cancer cell cytoskeleton is a clinically validated target, few new strategies have emerged for selectively targeting cell division by modulating the cytoskeletal structure, particularly ways that could avoid the cardiotoxic and neurotoxic effects of current agents such as taxanes. We address this gap by describing a novel class of small-molecule agonists of the mammalian Diaphanous (mDia)-related formins, which act downstream of Rho GTPases to assemble actin filaments, and their organization with microfilaments to establish and maintain cell polarity during migration and asymmetric division. GTP-bound Rho activates mDia family members by disrupting the interaction between the DID and DAD autoregulatory domains, which releases the FH2 domain to modulate actin and microtubule dynamics. In screening for DID-DAD disruptors that activate mDia, we identified two molecules called intramimics (IMM-01 and -02) that were sufficient to trigger actin assembly and microtubule stabilization, serum response factor-mediated gene expression, cell-cycle arrest, and apoptosis. In vivo analysis of IMM-01 and -02 established their ability to slow tumor growth in a mouse xenograft model of colon cancer. Taken together, our work establishes the use of intramimics and mDia-related formins as a new general strategy for therapeutic targeting of the cytoskeletal remodeling machinery of cancer cells.

Loss of heterozygosity and SOSTDC1 in adult and pediatric renal tumors.

BACKGROUND: Deletions within the short arm of chromosome 7 are observed in approximately 25% of adult and 10% of Wilms pediatric renal tumors. Within Wilms tumors, the region of interest has been delineated to a 2-Mb minimal region that includes ten known genes. Two of these ten candidate genes, SOSTDC1 and MEOX2, are particularly relevant to tumor development and maintenance. This finding, coupled with evidence that SOSTDC1 is frequently downregulated in adult renal cancer and regulates both Wingless-Int (Wnt)- and bone morphogenetic protein (BMP)-induced signaling, points to a role for SOSTDC1 as a potential tumor suppressor. METHODS: To investigate this hypothesis, we interrogated the Oncomine database to examine the SOSTDC1 levels in adult renal clear cell tumors and pediatric Wilms tumors. We then performed single nucleotide polymorphism (SNP) and sequencing analyses of SOSTDC1 in 25 pediatric and 36 adult renal tumors. Immunohistochemical staining of patient samples was utilized to examine the impact of SOSTDC1 genetic aberrations on SOSTDC1 protein levels and signaling. RESULTS: Within the Oncomine database, we found that SOSTDC1 levels were reduced in adult renal clear cell tumors and pediatric Wilms tumors. Through SNP and sequencing analyses of 25 Wilms tumors, we identified four with loss of heterozygosity (LOH) at 7p and three that affected SOSTDC1. Of 36 adult renal cancers, we found five with LOH at 7p, two of which affected SOSTDC1. Immunohistochemical analysis of SOSTDC1 protein levels within these tumors did not reveal a relationship between these instances of SOSTDC1 LOH and SOSTDC1 protein levels. Moreover, we could not discern any impact of these genetic alterations on Wnt signaling as measured by altered beta-catenin levels or localization. CONCLUSIONS: This study shows that genetic aberrations near SOSTDC1 are not uncommon in renal cancer, and occur in adult as well as pediatric renal tumors. These observations of SOSTDC1 LOH, however, did not correspond with changes in SOSTDC1 protein levels or signaling regulation. Although our conclusions are limited by sample size, we suggest that an alternative mechanism such as epigenetic silencing of SOSTDC1 may be a key contributor to the reduced SOSTDC1 mRNA and protein levels observed in renal cancer.

Methionine-dependence phenotype in the de novo pathway in BRCA1 and BRCA2 mutation carriers with and without breast cancer.

Methionine-dependence phenotype (MDP) refers to the reduced ability of cells to proliferate when methionine is restricted and/or replaced by its immediate precursor homocysteine. MDP is a characteristic of human tumors in vivo, human tumor cell lines, and normal somatic tissue in some individuals. It was hypothesized that MDP is a risk factor for developing breast cancer in BRCA (BRCA1 and BRCA2) germline mutation carriers. To test the hypothesis, human peripheral blood lymphocytes of BRCA carriers with and without breast cancer and healthy non-carrier relatives (controls) were cultured for 9 days in medium containing either 0.1 mmol/L L-methionine or 0.2 mmol/L D,L-homocysteine, with the ratio of viable cell growth in both types of medium after 9 days used to calculate the methionine-dependence index (MDI), a measure of MDP. We also tested whether MDP was associated with common polymorphisms in methionine metabolism. Viable cell growth, MDI, and polymorphism frequency in MTRR (A66G and C524T) and MTHFR (A1298C and A1793G) did not differ among the study groups; however, MDI tended to be higher in BRCA carriers with breast cancer than those without and was significantly increased in MTHFR 677T allele carriers relative to wild-type carriers (P=0.017). The presence of MTR A2756G mutant allele and MTHFR C677T mutant allele in carriers was associated with increased breast cancer risk [odds ration, 3.2 (P=0.16; 95% confidence interval, 0.76-13.9) and 3.9 (P=0.09; 95% confidence interval, 0.93-16.3), respectively]. The results of this study support the hypothesis that defects in methionine metabolism may be associated with breast cancer risk in BRCA carriers.

Lymphocytes of BRCA1 and BRCA2 germ-line mutation carriers, with or without breast cancer, are not abnormally sensitive to the chromosome damaging effect of moderate folate deficiency.

Mutations in BRCA1 and BRCA2 genes may cause defective DNA repair and increase the risk for breast cancer. Folate deficiency is associated with increased breast cancer risk and induces chromosome abnormalities. We hypothesized that BRCA1 and BRCA2 germline mutation carriers are more sensitive to the genome damaging effect of folate deficiency compared with healthy non-carrier controls and that this sensitivity is further increased in those carriers who develop breast cancer. We tested these hypotheses in lymphocytes cultured in a medium containing 12 or 120 nM folic acid (FA) for 9 days and measured proliferative capacity and chromosomal instability using the cytokinesis-block micronucleus assay. BRCA1 and BRCA2 mutation carriers with or without breast cancer were not abnormally sensitive to FA deficiency-induced chromosome instability; however, BRCA2 mutation carriers had significantly reduced cell proliferation. FA deficiency reduced cell proliferation and increased micronucleus formation significantly, accounting for 45-59% and 70-75% of the variance in these parameters compared with 0.3-8.5% and 0.2-0.3% contributed by BRCA1 or BRCA2 mutation carrier status, respectively. The results of this study suggest that moderate folate deficiency has a stronger effect on chromosomal instability than BRCA1 or BRCA2 mutations found in breast cancer families.

Folic acid deficiency increases chromosomal instability, chromosome 21 aneuploidy and sensitivity to radiation-induced micronuclei.

Folic acid deficiency can lead to uracil incorporation into DNA, hypomethylation of DNA, inefficient DNA repair and increase chromosome malsegregation and breakage. Because ionising radiation increases demand for efficient DNA repair and also causes chromosome breaks we hypothesised that folic acid deficiency may increase sensitivity to radiation-induced chromosome breakage. We tested this hypothesis by using the cytokinesis-block micronucleus assay in 10 day WIL2-NS cell cultures at four different folic acid concentrations (0.2, 2, 20, and 200 nM) that span the "normal" physiological range in humans. The study showed a significant dose-dependent increase in frequency of binucleated cells with micronuclei and/or nucleoplasmic bridges with decreasing folic acid concentration (P<0.0001, P=0.028, respectively). These biomarkers of chromosomal instability were also increased in cells irradiated (1.5 Gy gamma-rays) on day 9 relative to un-irradiated controls (P<0.05). Folic acid deficiency and gamma-irradiation were shown to have a significant interactive effect on frequency of cells containing micronuclei (two-way ANOVA, interaction P=0.0039) such that the frequency of radiation-induced micronucleated cells (i.e. after subtracting base-line frequency of un-irradiated controls) increased with decreasing folic acid concentration (P-trend<0.0001). Aneuploidy of chromosome 21, apoptosis and necrosis were increased by folic acid deficiency but not by ionising radiation. The results of this study show that folate status has an important impact on chromosomal stability and is an important modifying factor of cellular sensitivity to radiation-induced genome damage.

Low intake of calcium, folate, nicotinic acid, vitamin E, retinol, beta-carotene and high intake of pantothenic acid, biotin and riboflavin are significantly associated with increased genome instability--results from a dietary intake and micronucleus index survey in South Australia.

The aim of this study was to determine the association between dietary intake, determined using a food frequency questionnaire, and genome damage in lymphocytes measured using the micronucleus (MN) assay. The study, performed on 190 healthy individuals (mean age 47.8 years, 46% males), also examined whether a supplementation with beta-carotene, vitamins C and E along with zinc (ACEZn), in a randomized trial for 6 months, improves genome stability. Multivariate analysis of baseline data showed that (1) the highest tertile of intake of vitamin E, retinol, folic acid, nicotinic acid (preformed) and calcium is associated with significant reductions in MN frequency, i.e. -28, -31, -33, -46 and -49%, respectively (P < 0.005) relative to the lowest tertile of intake and (2) the highest tertile of intake of riboflavin, pantothenic acid and biotin was associated with significant increases in MN frequency, i.e. +36% (P = 0.054), +51% (P = 0.021), and +65% (P = 0.001), respectively, relative to the lowest tertile of intake. Mid-tertile beta-carotene intake was associated with an 18% reduction in MN frequency (P = 0.038); however, the highest tertile of intake (>6400 microg/day) resulted in an 18% increment in MN frequency. Supplementation with ACEZn significantly reduced the MN index by 13% (P = 0.038). The study also showed interactive additive effects such as the protective effect of increased calcium intake (-46%) and the exacerbating effect of riboflavin (+42%) on increased genome damage caused by low folate intake. The results from this study illustrate the strong impact of a wide variety of micronutrients and their interactions on genome health, depending on the level of intake.

Intra- and inter-laboratory variation in the scoring of micronuclei and nucleoplasmic bridges in binucleated human lymphocytes. Results of an international slide-scoring exercise by the HUMN project.

One of the objectives of the HUman MicroNucleus (HUMN) project is to identify the methodological variables that have an important impact on micronucleus (MN) or micronucleated (MNed) cell frequencies measured in human lymphocytes using the cytokinesis-block micronucleus assay. In a previous study we had shown that the scoring criteria used were likely to be an important variable. To determine the extent of residual variation when laboratories scored cells from the same cultures using the same set of standard scoring criteria, an inter-laboratory slide-scoring exercise was performed among 34 laboratories from 21 countries with a total of 51 slide scorers involved. The results of this study show that even under these optimized conditions there is a great variation in the MN frequency or MNed cell frequency obtained by individual laboratories and scorers. All laboratories ranked correctly the MNed cell frequency in cells from cultures that were unirradiated, or exposed to 1 or 2Gy of gamma rays. The study also estimated that the intra-scorer median coefficient of variation for duplicate MNed cell frequency scores is 29% for unexposed cultures and 14 and 11% for cells exposed to 1 and 2Gy, respectively. These values can be used as a standard for quality or acceptability of data in future studies. Using a Poisson regression model it was estimated that radiation dose explained 67% of the variance, while staining method, cell sample, laboratory, and covariance explained 0.6, 0.3, 6.5, and 25.6% of the variance, respectively, leaving only 3.1% of the variance unexplained. As part of this exercise, nucleoplasmic bridges were also estimated by the laboratories; however, inexperience in the use of this biomarker of chromosome rearrangement was reflected in the much greater heterogeneity in the data and the unexplained variation estimated by the Poisson model. The results of these studies indicate clearly that even after standardizing culture and scoring conditions it will be necessary to calibrate scorers and laboratories if MN, MNed cell and nucleoplasmic bridge frequencies are to be reliably compared among laboratories and among populations.

Effectiveness of an antagonist to gonadotrophin releasing hormone on the FSH and LH response to GnRH in perifused equine pituitary cells, and in seasonally acyclic mares.

We wish to use a gonadotrophin-releasing hormone (GnRH) antagonist in the mare as a tool for investigating the control of the oestrous cycle. The aim of this study was to test the effectiveness of the antagonist cetrorelix by testing both in vitro, using perifused equine anterior pituitary cells, and in vivo in seasonally acyclic mares. Pituitary cells were prepared and after 3-4 days incubation, loaded onto columns and given four pulses of GnRH (at 0, 30, 60 and 90 min; dose-response study). After the second GnRH pulse, infusion of cetrorelix began (0, 100, 1000 and 2000 pmol/l) and continued until the end of the experiment. To mimic luteal phase conditions, cells were pre-incubated and perifused with progesterone (25 nmol/l) and GnRH pulses given at 0, 90, 180 and 270 min. Cetrorelix (0 or 1000 pmol/l) began after the second GnRH pulse. Follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations were measured in 5 min fractions. Both FSH and LH response areas (above baseline) after GnRH were inhibited by 1000 pmol/l cetrorelix (P < 0.01, P < 0.01, respectively) but not by 100 pmol/l cetrorelix. Similarly, in the presence of progesterone, cetrorelix inhibited the FSH (P < 0.001) and LH (P = 0.0002) response area. Seasonally acyclic mares, pre-treated for 3 days with progesterone (150 mg i.m. per day) were given cetrorelix as (i) a loading dose of 1 microg/kg then infusion at 2.2 ng/(kg min) for 90 min, (ii) a s.c. injection at 20 microg/kg, (iii) infusion at 2.2 ng/(kg min) for 48 h, and (iv) no cetrorelix (control mares). At 90 min, 6, 24 and 48 h after cetrorelix was first administered, mares were given a bolus injection of GnRH (22.2 ng/kg i.v.) and the FSH and LH responses measured. All doses of cetrorelix inhibited the FSH response at 90 min. The response was no longer suppressed at 6 h in the 90 min infusion group, showing a rapid recovery from inhibition. At 24 h, the FSH responses in the injected and 48 h infusion group were suppressed. The LH concentrations were low and showed no significant changes. This study has defined the time course and dose of cetrorelix with respect to its effect on FSH in the horse. It is concluded that cetrorelix could be used to elucidate the role of FSH in follicular development in cyclic mares.