Open science (OS) awareness and skills are increasingly becoming an essential part of everyday scientific work as e.g., many journals require authors to share data. However, following an OS workflow can seem challenging at first. Thus, instructions by journals and other guidelines are important. But how comprehensive are they in the field of ecology and evolutionary biology (Ecol Evol)? To find this out, we reviewed 20 published OS guideline articles aimed for ecologists or evolutionary biologists, together with the data policies of 17 Ecol Evol journals to chart the current landscape of OS guidelines in the field, find potential gaps, identify field-specific barriers for OS and discuss solutions to overcome these challenges. We found that many of the guideline articles covered similar topics, despite being written for a narrow field or specific target audience. Likewise, many of the guideline articles mentioned similar obstacles that could hinder or postpone a transition to open data sharing. Thus, there could be a need for a more widely known, general OS guideline for Ecol Evol. Following the same guideline could also enhance the uniformity of the OS practices carried on in the field. However, some topics, like long-term experiments and physical samples, were mentioned surprisingly seldom, although they are typical issues in Ecol Evol. Of the journals, 15 out of 17 expected or at least encouraged data sharing either for all articles or under specific conditions, e.g. for registered reports and 10 of those required data sharing at the submission phase. The coverage of journal data policies varied greatly between journals, from practically non-existing to very extensive. As journals can contribute greatly by leading the way and making open data useful, we recommend that the publishers and journals would invest in clear and comprehensive data policies and instructions for authors. Avoimen tieteen ymm & auml;rrys ja taitojen hallinta on yh & auml; t & auml;rke & auml;mpi osa tutkijan arkea, sill & auml; esimerkiksi monet tieteelliset lehdet odottavat aineiston avointa jakamista. Avoimen tieteen ty & ouml;tapojen noudattaminen voi kuitenkin tuntua alkuun haastavalta, mink & auml; vuoksi esimerkiksi tieteellisten lehtien ja muiden tahojen laatimat ohjeet ovat t & auml;rkeit & auml;. Mutta kuinka kattavia ne ovat ekologian ja evoluutiobiologian alalla? K & auml;vimme l & auml;pi 20 julkaistua ekologeille tai evoluutiobiologeille suunnattua avoimen tieteen ohjeistusta sek & auml; 17 ekologian ja evoluutiobiologian tieteellisen lehden datak & auml;yt & auml;nn & ouml;t, tarkoituksenamme kartoittaa alojen avoimen tieteen ohjeiden nykytilaa, l & ouml;yt & auml;& auml; mahdollisia puutteita, tunnistaa alakohtaisia esteit & auml; avoimen tieteen k & auml;yt & auml;nt & ouml;jen toteutumiselle sek & auml; keskustella ratkaisuista, joilla n & auml;m & auml; haasteet voitaisiin ratkaista. Havaitsimme, ett & auml; monet ohjeistukset k & auml;sitteliv & auml;t samankaltaisia aiheita, vaikka ne oli tarkoitettu kapealle erityisalalle tai suunnattu hyvin rajoitetulle kohderyhm & auml;lle. Samoin monissa ohjeistuksissa mainittiin samankaltaisia aineistojen avoimen jakamisen hidastamista tai est & auml;mist & auml; aiheuttavia haasteita. Toiset aiheet, kuten pitk & auml;aikaiskokeet ja fyysiset n & auml;ytteet, sen sijaan mainittiin yll & auml;tt & auml;v & auml;n harvoin, vaikka niiss & auml; on tyypillisi & auml; ekologian ja evoluutiobiologian alojen haasteita. Tieteellisist & auml; lehdist & auml; 15:ss & auml; 17:sta vaadittiin tai v & auml;hint & auml;& auml;n kannustettiin jakamaan aineisto avoimesti joko kaikkien artikkelien osalta tai tietyin edellytyksin, esim. rekister & ouml;ityjen tutkimusraporttien osalta. Lis & auml;ksi 10 n & auml;ist & auml; lehdist & auml; edellytti aineiston avointa jakamista jo submittointivaiheessa. Tieteellisten lehtien aineisto-ohjeiden kattavuus vaihteli suuresti lehtien v & auml;lill & auml;, k & auml;yt & auml;nn & ouml;ss & auml; olemattomasta hyvin laajaan. Koska tieteellisill & auml; lehdill & auml; on suuri vaikutusvalta avoimen tieteen k & auml;yt & auml;nt & ouml;jen edist & auml;miseen, suosittelemme kustantajia ja lehti & auml; panostamaan selkeisiin ja kattaviin aineistolinjauksiin ja ohjeistuksiin. Open Science (OS) awareness and skills are increasingly becoming an essential part of everyday scientific work. We reviewed OS guidelines given to ecologists and evolutionary biologists. We also reviewed data policies and instructions of the top journals of this field of science.image

Anticoagulant rodenticide (AR) resistance is widespread across Europe, but its prevalence has not been previously studied in Finland. We sampled 48 house mice (Mus musculus) and 48 brown rat (Rattus norvegicus) in farms and urban locations from southern Finland and here we report the identification of Vkorc1 polymorphisms linked to AR resistance.Two thirds of mice carried Vkorc1 polymorphisms. The most common was the type Y139C (resistance to first-generation ARs, bromadiolone and difenacoum), whereas the type L128S (resistance to FGARs and bromadiolone) was much less abundant. Only two rats tested positive, both with a rare type R33P (resistance probably to warfarin). No clear patterns based on subregion, sex and urban/rural division were seen.We suggest that the type Y139C polymorphism should be taken into account when planning control actions targeting house mice in Finland. The low number of AR resistant rats found suggests a low prevalence of resistance in rats. To fully understand the state of AR resistance in Finland, additional studies on yellow-necked mouse (Apodemus flavicollis) and bank voles (Myodes glareolus) are needed.

Most studies on intraspecific competition, i.e., competition among individuals of the same species, have been conducted during the breeding season. Yet, at northern latitudes, intraspecific competition is expected to be particularly strong under the harsh weather conditions of the non-breeding season with limited number of resources available per individual. We studied the food-hoarding behaviour of wintering Eurasian Pygmy Owls (Glaucidium passerinum) along with sex- and age-specific spatial distribution in relation to fluctuating main prey abundance (voles) and conspecific density using a 15-year dataset. In low vole abundance years, increasing conspecific density reduced the total prey number stored by an owl, suggesting high costs of exploitative competition. The distance between the stores of nearest neighbours was greater when both were females, suggesting that the spatial avoidance is driven by sex-specific competition. However, food stores of females had a larger amount of prey items, especially when the nearest neighbour was of the same sex. The number of stores hoarded by an owl increased with increasing conspecific densities. Distributing the prey items to multiple store-sites instead of one (shifting from larder-hoarding towards scatter-hoarding) can help to reduce the overall loss to potential pilfering when conspecific density is high. These results combined suggest that high conspecific density inflames sex-specific interference competition, rather than solely exploitative competition, and in turn drives the observed sex-specific spatial distribution. Adopting a sex-specific spatial distribution according to hoarding and aggressive behaviour can be a way to reduce the severity of intraspecific competition locally and could have cascading effects on the prey community.

Abstract Large areas of forests are annually damaged or destroyed by outbreaking insect pests. Understanding the factors that trigger and terminate such population eruptions has become crucially important, as plants, plant‐feeding insects, and their natural enemies may respond differentially to the ongoing changes in the global climate. In northernmost Europe, climate‐driven range expansions of the geometrid moths Epirrita autumnata and Operophtera brumata have resulted in overlapping and increasingly severe outbreaks. Delayed density‐dependent responses of parasitoids are a plausible explanation for the 10‐year population cycles of these moth species, but the impact of parasitoids on geometrid outbreak dynamics is unclear due to a lack of knowledge on the host ranges and prevalences of parasitoids attacking the moths in nature. To overcome these problems, we reviewed the literature on parasitism in the focal geometrid species in their outbreak range and then constructed a DNA barcode reference library for all relevant parasitoid species based on reared specimens and sequences obtained from public databases. The combined recorded parasitoid community of E. autumnata and O. brumata consists of 32 hymenopteran species, all of which can be reliably identified based on their barcode sequences. The curated barcode library presented here opens up new opportunities for estimating the abundance and community composition of parasitoids across populations and ecosystems based on mass barcoding and metabarcoding approaches. Such information can be used for elucidating the role of parasitoids in moth population control, possibly also for devising methods for reducing the extent, intensity, and duration of outbreaks.

European aspen (Populus tremula L.) is a keystone species in boreal forests that are dominated by coniferous tree species. Both living and dead aspen trees contribute significantly to the species diversity of forest landscapes. Thus, spatial and temporal continuity of aspen is a prerequisite for the long-term persistence of viable populations of numerous aspen-associated species. In this review, we collate existing knowledge on the ecological role of European aspen, assess the knowledge needs for aspen occurrence patterns and dynamics in boreal forests and discuss the potential of different remote sensing techniques in mapping aspen at various spatiotemporal scales. The role of aspen as a key ecological feature has received significant attention, and studies have recognised the negative effects of modern forest management methods and heavy browsing on aspen occurrence and regeneration. However, the spatial knowledge of occurrence, abundance and temporal dynamics of aspen is scarce and incomprehensive. The remote sensing studies reviewed here highlight particularly the potential of three-dimensional data derived from airborne laser scanning or photogrammetric point clouds and airborne imaging spectroscopy in mapping European aspen, quaking aspen (Populus tremuloides Michx.) and other Populus species. In addition to tree species discrimination, these methods can provide information on biophysical, biochemical properties and even genetic diversity of aspen trees. Major obstacles in aspen detection using remote sensing are the low proportion and scattered occurrence of European aspen in boreal forests and the overlap of spectral and/or structural properties of European aspen and quaking aspen with some other tree species. Furthermore, the suitability of remote sensing data for aspen mapping and monitoring depends on the geographical coverage of data, the availability of multitemporal data and the costs of data acquisition. Our review highlights that integration of ecological knowledge with spatiotemporal information acquired by remote sensing is key to understanding the current and future distribution patterns of aspen-related biodiversity.

Competing species and predators can alter the habitat use of animals, but both factors are rarely simultaneously controlled. We studied in experimental enclosures how closely related species, the sibling vole (Microtus levis Miller, 1908) and the field vole (Microtus agrestis (Linnaeus, 1761)), adjust their habitat use when facing either the competing species or simultaneously competition and predation risk. The species responded differently in their proportional use of two habitat types, a low cover (productive but riskier) and a high cover (safer but poorer). When alone, field voles used the low-cover habitat according to availability at low densities, but decreased its use with increasing density. Sibling voles, however, avoided the low-cover habitat in single-species populations. Under interspecific competition, the habitat-use patterns switched between species: sibling voles used the low-cover habitat according to availability, with decreasing use as densities increased. Sibling voles responded to predation risk by showing a stronger density-dependent decrease in the use of low-cover habitat. Field voles, initially using mostly high cover, did not change behaviour under risk of predation. Our results highlight the importance of considering both predation risk and interspecific competition when interpreting patterns of habitat selection among coexisting species.

The most common rodent control method worldwide is anticoagulant rodenticides (ARs), which cause death by internal bleeding. ARs can transfer to non-target predators via secondary exposure, i.e. by consuming contaminated rodents. Here we quantify the prevalence of seven AR substances in the liver tissues of altogether 17 mammalian or avian predator or scavenger species in Finland. In addition, we identify the environmental and biological factors potentially linked to secondary AR poisoning. No previous AR screenings have been conducted in the country, despite the widespread use of ARs and their potential impacts on the high levels of the ecosystem food chain. ARs were detected (≥0.3 μg/kg) in 82% of the 131 samples. The most prevalent and the AR with highest concentrations was bromadiolone (65% of samples). In 77% of the positive samples more than one (2-5) different ARs were detected. Of the environmental variables, we only found a weakly positive relationship between the coumatetralyl concentration and the livestock farm density. Conversely, overall AR concentration and number, as well as the concentration of three separate ARs (coumatetralyl, difenacoum and bromadiolone) differed among the three species groups tested, with the group "other mammals" (largely represented by red fox and raccoon dog) having higher values than the groups presented by mustelids or by birds. ARs are authorized only as biocides in Finland and a national strategy on risk management (e.g. for minimising secondary poisoning of non-target species) of ARs was adopted in 2011. Based on these results it appears that the risk mitigation measures (RMMs) either have not been followed or have not been effective in preventing wide scale secondary exposure. Continued monitoring of AR residues in non-target species is needed in order to evaluate the effectiveness of current RMMs and a need for new ones to reduce the risk of secondary poisoning.

Browsing effects of voles on plants can be mitigated by means of non-lethal predation, i.e. by subjecting voles to predator cues. Earlier studies largely focused on mammalian olfactory cues, whereas here we exposed Microtus voles to owl calls to examine whether the introduced predation risk reduces browsing on tree seedlings (silver birch, Scots pine, Norway spruce). We found that owl risk was associated with higher growth rate of birch seedlings, but only late in the growing season, while there was no obvious effect in case of pine and spruce. Early in the season, in the absence of predation risk, voles were able to move freely and search for their preferred food, grasses. Late in the season, when grasses were less palatable, voles likely targeted birch seedlings to a higher degree. Our results suggest that owl calls could alleviate vole herbivory on birches but, at least on a short temporal scale, will not protect seedlings of conifers.

Previous studies on intraguild predation have mainly focused on within-class assemblages, even though avian top predators may also influence mammalian mesopredator prey. By using nation-wide long-term data from Finland, northern Europe, we examined the impacts of golden eagles (Aquila chrysaetos) together with red foxes (Vulpes vulpes) and pine martens (Martes martes) on forest-dwelling herbivores, black grouse (Tetrao tetrix) and hazel grouse (Tetrastes bonasia). We hypothesized that eagles may alleviate the overall predation pressure on grouse by imposing intraguild predation risk on mesopredators. The predation impact of eagle was modelled using eagle density estimates and distance to eagle nest. Wildlife triangle counts were used as predation impact proxies of mammalian mesopredators and as measures of response in grouse. Our results show that eagle density correlated negatively with black grouse abundance indices while being positively associated with the proportion of juveniles in both grouse species, irrespective of the abundance of mesopredators. Yet, foxes and martens alone had a negative effect on the abundance indices and the proportion of young in the two grouse species. This suggests that the possible cascading effects of eagles are not mediated by decreased mesopredator numbers, but instead by fear effects. Alternatively, they may be mediated by other species than fox or marten studied here. In conclusion, we found support for the hypothesis that eagles provide protection for juvenile black and hazel grouse, whereas they are a threat for adult grouse. This important information helps us to better understand the role of avian top predators in terrestrial ecosystems.

Competition, habitat structure and predation risk may alter mobility and activity of animals. In particular, small predators need to weigh the positive and negative impacts of the presence of competing larger predator species. Other predators present a risk but may also enhance the predation efficiency of smaller predators if avoiding one predator species increases prey exposure to the other. We studied how habitat structure and the presence of avian predators affected the activity patterns of radio-collared least weasels Mustela nivalis nivalis, small-bodied predators highly specialized on hunting small rodents and subject to intraguild predation. Least weasels appeared to schedule their mobility according to the peak activity of the avian predator present in large outdoor enclosures. In the presence of diurnal European kestrels Falco tinnunculus, least weasels moved less during night compared with evening and day, whereas in the presence of nocturnal Tengmalm's owls Aegolius funereus, least weasels moved more during night than during other time periods. Least weasels almost exclusively used the tall grass habitat in all treatments, whereas owls used the short grass habitat most and kestrels used both habitats equally. Our results suggest that beneficial interactions play an important role in the interaction between least weasels, kestrels and owls, and that the avoidance of risky habitats by least weasels minimizes the risk of intraguild predation.

The phosphatase and actin regulator 1 (PHACTR1) locus is a very commonly identified hit in genome-wide association studies investigating coronary artery disease and myocardial infarction (MI). However, the function of PHACTR1 in the heart is still unknown. We characterized the mechanisms regulating Phactr1 expression in the heart, used adenoviral gene delivery to investigate the effects of Phactr1 on cardiac function, and analyzed the relationship between MI associated PHACTR1 allele and cardiac function in human subjects. Phactr1 mRNA and protein levels were markedly reduced (60%, P<0.01 and 90%, P<0.001, respectively) at 1 day after MI in rats. When the direct myocardial effects of Phactr1 were studied, the skeletal α-actin to cardiac α-actin isoform ratio was significantly higher (1.5-fold, P<0.05) at 3 days but 40% lower (P<0.05) at 2 weeks after adenovirus-mediated Phactr1 gene delivery into the anterior wall of the left ventricle. Similarly, the skeletal α-actin to cardiac α-actin ratio was lower at 2 weeks in infarcted hearts overexpressing Phactr1. In cultured neonatal cardiac myocytes, adenovirus-mediated Phactr1 overexpression for 48 hours markedly increased the skeletal α-actin to cardiac α-actin ratio, this being associated with an enhanced DNA binding activity of serum response factor. Phactr1 overexpression exerted no major effects on the expression of other cardiac genes or LV structure and function in normal and infarcted hearts during 2 weeks’ follow-up period. In human subjects, MI associated PHACTR1 allele was not associated significantly with cardiac function (n = 1550). Phactr1 seems to regulate the skeletal to cardiac α-actin isoform ratio.

Top predators may induce extensive cascading effects on lower trophic levels, for example, through intraguild predation (IGP). The impacts of both mammalian and avian top predators on species of the same class have been extensively studied, but the effects of the latter upon mammalian mesopredators are not yet as well known. We examined the impact of the predation risk imposed by a large avian predator, the golden eagle (Aquila chrysaetos, L.), on its potential mammalian mesopredator prey, the red fox (Vulpes vulpes, L.), and the pine marten (Martes martes, L.). The study combined 23years of countrywide data from nesting records of eagles and wildlife track counts of mesopredators in Finland, northern Europe. The predation risk of the golden eagle was modeled as a function of territory density, density of fledglings produced, and distance to nearest active eagle territory, with the expectation that a high predation risk would reduce the abundances of smaller sized pine martens in particular. Red foxes appeared not to suffer from eagle predation, being in fact most numerous close to eagle nests and in areas with more eagle territories. This is likely due to similar prey preferences of the two predators and the larger size of foxes enabling them to escape eagle predation risk. Somewhat contrary to our prediction, the abundance of pine martens increased from low to intermediate territory density and at close proximity to eagle nests, possibly because of similar habitat preferences of martens and eagles. We found a slightly decreasing trend of marten abundance at high territory density, which could indicate that the response in marten populations is dependent on eagle density. However, more research is needed to better establish whether mesopredators are intimidated or predated by golden eagles, and whether such effects could in turn cascade to lower trophic levels, benefitting herbivorous species.

The transforming growth factor (TGF)-β is one of the key mediators in cardiac remodelling occurring after myocardial infarction (MI) and in hypertensive heart disease. The TGF-β-stimulated clone 22 (TSC-22) is a leucine zipper protein expressed in many tissues and possessing various transcription-modulating activities. However, its function in the heart remains unknown.

The negative impacts of mammalian herbivores on plants have been studied quite extensively, but typically with only a single herbivore species at a time. We conducted a novel comparison of the browsing effects of voles, hares and cervids upon the growth and survival of boreal tree seedlings. This was done by excluding varying assemblages of these key mammalian herbivores from silver birch, Scots pine and Norway spruce seedlings for 3 years. We hypothesised that the pooled impacts of the herbivores would be greater than that of any individual group, while the cervids would be the group with the strongest impact. Growth of birch seedlings advanced when cervids were excluded whereas growth of seedlings accessible to cervids was hindered. Survival of all seedlings was lowest when they were accessible to voles and voles plus hares, whereas cervids seemed not to influence seedling survival. Our results show that the impact of herbivores upon woody plants can be potent in the boreal forests, but the mechanism and strength of this link depends on the tree and herbivore species in question. Risk of abated stand regeneration appears highest for the deciduous birch, though there is need for seedling protection also in coniferous stands. The clear cervid-mediated growth limitation of birch also indicates potential for a trophic cascade effect by mammalian top predators, currently returning to boreal ecosystems.

AIMS:Activating transcription factor 3 (ATF3) is a stress-activated immediate early gene suggested to have both detrimental and cardioprotective role in the heart. Here we studied the mechanisms of ATF3 activation by hypertrophic stimuli and ATF3 downstream targets in rat cardiomyocytes. METHODS AND RESULTS:When neonatal rat cardiomyocytes were exposed to endothelin-1 (ET-1, 100 nM) and mechanical stretching in vitro, maximal increase in ATF3 expression occurred at 1 hour. Inhibition of extracellular signal-regulated kinase (ERK) by PD98059 decreased ET-1- and stretch-induced increase of ATF3 protein but not ATF3 mRNA levels, whereas protein kinase A (PKA) inhibitor H89 attenuated both ATF3 mRNA transcription and protein expression in response to ET-1 and stretch. To characterize further the regulatory mechanisms upstream of ATF3, p38 mitogen-activated protein kinase (MAPK) signaling was investigated using a gain-of-function approach. Adenoviral overexpression of p38α, but not p38β, increased ATF3 mRNA and protein levels as well as DNA binding activity. To investigate the role of ATF3 in hypertrophic process, we overexpressed ATF3 by adenovirus-mediated gene transfer. In vitro, ATF3 gene delivery attenuated the mRNA transcription of interleukin-6 (IL-6) and plasminogen activator inhibitor-1 (PAI-1), and enhanced nuclear factor-κB (NF-κB) and Nkx-2.5 DNA binding activities. Reduced PAI-1 expression was also detected in vivo in adult rat heart by direct intramyocardial adenovirus-mediated ATF3 gene delivery. CONCLUSIONS:These data demonstrate that ATF3 activation by ET-1 and mechanical stretch is partly mediated through ERK and cAMP-PKA pathways, whereas p38 MAPK pathway is involved in ATF3 activation exclusively through p38α isoform. ATF3 activation caused induction of modulators of the inflammatory response NF-κB and Nkx-2.5, as well as attenuation of pro-fibrotic and pro-inflammatory proteins IL-6 and PAI-1, suggesting cardioprotective role for ATF3 in the heart.

INTRODUCTION:Food availability is an important environmental cue for animals for deciding how much to invest in reproduction, and it ultimately affects population size. The importance of food limitation has been extensively studied in terrestrial vertebrate populations, especially in birds, by experimentally manipulating food supply. However, the factors explaining variation in reproductive decisions in response to food supplementation remain unclear. By performing meta-analyses, we aim to quantify the extent to which supplementary feeding affects several reproductive parameters in birds, and identify the key factors (life-history traits, behavioural factors, environmental factors, and experimental design) that can induce variation in laying date, clutch size and breeding success (i.e., number of fledglings produced) in response to food supplementation.RESULTS:Food supplementation produced variable but mostly positive effects across reproductive parameters in a total of 201 experiments from 82 independent studies. The outcomes of the food effect were modulated by environmental factors, e.g., laying dates advanced more towards low latitudes, and food supplementation appeared not to produce any obvious effect on bird reproduction when the background level of food abundance in the environment was high. Moreover, the increase in clutch size following food addition was more pronounced in birds that cache food, as compared to birds that do not. Supplementation timing was identified as a major cause of variation in breeding success responses. We also document the absence of a detectable food effect on clutch size and breeding success when the target species had poor access to the feed due to competitive interactions with other animals.CONCLUSIONS:Our findings indicate that, from the pool of bird species and environments reviewed, extra food is allocated to immediate reproduction in most cases. Our results also support the view that bird species have evolved different life-history strategies to cope with environmental variability in food supply. However, we encourage more research at low latitudes to gain knowledge on how resource allocation in birds changes along a latitudinal gradient. Our results also emphasize the importance of developing experimental designs that minimise competition for the supplemented food and the risk of reproductive bottle-necks due to inappropriate supplementation timings.

cAMP-dependent protein kinase (PKA) regulates the L-type calcium channel, the ryanodine receptor, and phospholamban (PLB) thereby increasing inotropy. Cardiac contractility is also regulated by p38 MAPK, which is a negative regulator of cardiac contractile function. The aim of this study was to identify the mechanism mediating the positive inotropic effect of p38 inhibition. Isolated adult and neonatal cardiomyocytes and perfused rat hearts were utilized to investigate the molecular mechanisms regulated by p38. PLB phosphorylation was enhanced in cardiomyocytes by chemical p38 inhibition, by overexpression of dominant negative p38α and by p38α RNAi, but not with dominant negative p38β. Treatment of cardiomyocytes with dominant negative p38α significantly decreased Ca(2+)-transient decay time indicating enhanced sarco/endoplasmic reticulum Ca(2+)-ATPase function and increased cardiomyocyte contractility. Analysis of signaling mechanisms involved showed that inhibition of p38 decreased the activity of protein phosphatase 2A, which renders protein phosphatase inhibitor-1 phosphorylated and thereby inhibits PP1. In conclusion, inhibition of p38α enhances PLB phosphorylation and diastolic Ca(2+) uptake. Our findings provide evidence for novel mechanism regulating cardiac contractility upon p38 inhibition.

Objectives: Transforming-growth-factor β-stimulated clone 22 (TSC-22) is a leucine zipper protein expressed in many tissues and possessing various transcription-modulating activities. However, its function in the heart remains largely unknown. The aim of the present study was to characterize the cardiac TSC-22 expression. Methods: Acute pressure overload was accomplished in conscious Sprague-Dawley (SD) rats by intravenous infusion of arginine 8 -vasopressin (AVP, 0.05 μg/kg/min) for 4 hours and subcutaneous infusion of angiotensin II (Ang II, 33 μg/kg/h) with and without Ang II receptor type 1 blocker losartan (400 μg/kg/h) by using osmotic minipumps for 2 weeks. Adenovirus-mediated intramyocardial gene transfer of TSC-22 was performed into left ventricle (LV) of SD rats. Experimental myocardial infarction (MI) was produced by ligation of the left anterior descending coronary artery. Cultured neonatal rat ventricular myocytes (NRVM) were treated with endothelin-1 (ET-1, 100 nM). Results: A significant 1.6-fold increase ( P <0.05) in LV TSC-22 mRNA levels was noted already after 1 hour AVP infusion. Moreover, Ang II infusion markedly upregulated TSC-22 expression, LV mRNA levels being highest at 6 hours (11-fold, P <0.001). Simultaneous infusion of losartan completely abolished Ang II-induced increase in TSC-22 mRNA levels. Adenovirus-mediated gene transfer of TSC-22 into LV resulted a 1.9-fold ( P <0.001) increase in TSC-22 mRNA levels, accompanied by upregulated BNP mRNA levels (1.4-fold, P <0.01). In response to experimental MI, TSC-22 mRNA levels were elevated 4.1-fold ( P <0.001) at 1 day and 1.9-fold ( P <0.05) at 4 weeks. In cultured NRVM, ET-1 treatment increased TSC-22 mRNA levels from 1 h to 24 h, the greatest increase being observed at 12 h (2.7-fold, P <0.001). TSC-22 protein levels were upregulated from 4 h to 24 h with the highest increase at 24 h (4.7-fold, P <0.01). Conclusion: These results indicate that TSC-22 expression is rapidly activated in response to pressure overload, MI and in ET-1 treated cultured NRVM. Moreover, adenovirus-mediated overexpression of TSC-22 mRNA was associated with elevated left ventricular BNP mRNA levels.

Rationale: p38 MAPK is a negative regulator of cardiac contractile function, but the underlying molecular mechanism is not well understood. Objective: To determine the molecular mechanism mediating the positive inotropic effect of p38 inhibition. Methods and Results: The effect of p38 inhibition on calcium regulatory proteins and calcium cycling was investigated in isolated neonatal and adult rodent cardiomyocytes using chemical inhibitors, recombinant adenoviruses and RNAi. Chemical inhibition of p38 enhanced phospholamban (PLB) phosphorylation at serine 16 residue. This was recapitulated with dominant negative p38α adenovirus and by p38α RNAi, but not with dominant negative p38β adenovirus. Chemical inhibition of p38 and dominant negative p38α resulted in significant decrease in Ca 2+ -transient decay time in adult rat cardiomyocytes. Inhibition of p38α, but not p38β, induced an increase in phosphorylation of protein phosphatase inhibitor-1 at the protein kinase A site threonine 35. Analysis of underlying mechanisms revealed that p38 inhibition increased phosphorylation of G protein-coupled receptor kinase 2 (GRK2), but not GRK3 or GRK5. p38 inhibition- induced PLB phosphorylation was abolished in cardiomyocytes overexpressing wild type GRK2. In contrast, increased PLB phosphorylation upon p38 inhibition was not reduced in cardiomyocytes overexpressing GRK2 harboring a non-phosphorylatable mutation in serine 670 residue. Finally, p38 inhibition-induced shortening of Ca 2+ -transient decay time was absent in cardiomyocytes overexpressing wild type GRK2. Conclusions: Inhibition of p38α regulates protein phosphatase inhibitor-1 resulting in increased PLB phosphorylation and enhanced SERCA2a function.