How to write an umbrella review? A step-by-step tutorial with tips and tricks.

The number of meta-analyses (MA) and systematic reviews (SR) on various medical issues has increased during the last two decades. The MA and SR results may differ from one another due to a number of factors such as inaccurate or diverse searches through the databases, discrepancies in the extraction process or in statistical analysis, among others. Some results may even contradict one another, resulting in confusion among readers. Umbrella reviews (UR) have allowed the collection of all available data on a medical issue into one concise study, making it the source of evidence-based medical knowledge to the highest degree. Furthermore, UR can resolve those problems by collecting all data and taking into account both MA and SR, making it the superior tool for physicians. Although the pros of UR are clear and the overall popularity of these types of study has increased tremendously, there is no available step-by-step guide on how to conduct one. Therefore, the objective of the present study was to provide researchers with a detailed tutorial on how to conduct an UR. UR represent the next major step in the advancement of evidence-based medicine, with great practical potential for physicians looking for the most up-to-date data on their topic of interest. We hope that our step-by-step guide may be a useful tool for researchers conducting UR in the future.

Can the kisspeptin help us in the understanding of pathology of some neurodegenerative brain diseases?

It is already known that the discovery of kisspeptin was a revolutionary step in the understanding of neuroendocrine regulation of reproduction. Kisspeptin is one of the main moderators of the gonadotropic axis, but the kisspeptin gene is known to be expressed in various regions of the central nervous system. The activity of kisspeptin is not limited to hypothalamic pituitary gonadal axis; it participates in the regulation of multiple neuronal circuits in the limbic system. The limbic system is a part of the brain involved in behavioural and emotional reactions, and disturbances in its functioning may be the source of some psychiatric as well as degenerative disorders. In the present review, we summarise the current state of knowledge concerning the role of kisspeptin in the limbic system and a new hope for the treatment of disturbances in its functioning.

Pyruvate dehydrogenase deficiency: morphological and metabolic effects, creation of animal model to search for curative treatment.

The main source of energy for brain and other organs is glucose. To obtain energy for all tissue, glucose has to come through glycolysis; then as pyruvate it is converted to acetyl-CoA by pyruvate dehydrogenase complex (PDC) and finally enters citric acid cycle. What happens when one of these stages become disturb? Mutation in genes encoding subunits of PDC leads to pyruvate dehydrogenase deficiency. Abnormalities in PDC activity result in severe metabolic and brain malformations. For better understanding the development and mechanism of pyruvate dehydrogenase deficiency the murine model of this disease has been created. Studies on a murine model showed similar malformation in brain structures as in the patients suffered from pyruvate dehydrogenase deficiency such as reduced neuronal density, heterotopias of grey matter, reduced size of corpus callosum and pyramids. There is still no effective cure for PDC-deficiency. Promising therapy seemed to be ketogenic diet, which substitutes glucose to ketone bodies as a source of energy. Studies have shown that ketogenic diet decreases lactic acidosis and inhibits brain malformations, but not the mortality in early childhood. The newest reports say that phenylbutyrate increases the level of PDC in the brain, because it reduces the level of inactive form of PDH. Experiments on human fibroblast and zebra fish PDC-deficiency model showed that phenylbutyrate is promising cure to PDC-deficiency. This review summarizes the most important findings on the metabolic and morphological effects of PDC-deficiency and research for treatment therapy.

The influence of early postnatal chronic mild stress stimulation on the activation of amygdala in adult rat.

Amygdala is a limbic structure involved in the stress response. The immunohistochemical and morphometric methods were used to examine whether the chronic mild psychological stress during the early postnatal period would change activation of amygdaloid nuclei in response to the same stressor in adult. In the study we focused on the role of neurons containing calbindin (CB), calretinin (CR), parvalbumin (PV) and nitric oxide synthase (NOS). The rats were divided into three groups: control non-stressed animals and two experimental: EI consisted of animals that were exposed to acute stress in the high-light, open-field test (HL-OF) at P90 (P - postnatal day) and EII consisted of rats that were exposed to chronic stress in HL-OF, daily during the first 21 postnatal days and then once at P90. The scheme of activation of amygdaloid nuclei under stress in EI and EII group was similar. The highest density of c-Fos-ir cells (c-Fos - a marker of neuronal activation) was demonstrated by the medial nucleus (Me) and bed nucleus of the accessory olfactory tract (BAOT). The amygdaloid nuclei diversity after HL-OF was determined by the high activation of the NOS-ir cells in the Me and NOS- and CR-ir cells in the BAOT. These are probably projection neurons involved in modulation of defensive, reproductive and autonomic behavior in stress response and creation/storage of aversive memory. However, in comparison with EI group, significant decrease in density of c-Fos-ir cells, in almost all amygdaloid nuclei of EII group was revealed. Particularly in BAOT and Me the strong decrease of activity of NOS- and CR-ir neurons was observed. It probably results in attenuation of stress responses what, depending on the circumstances, can be adaptive or maladaptive.

Brain Organoids: Expanding Our Understanding of Human Development and Disease.

Stem cell-derived brain organoids replicate important stages of the prenatal human brain development and combined with the induced pluripotent stem cell (iPSC) technology offer an unprecedented model for investigating human neurological diseases including autism and microcephaly. We describe the history and birth of organoids and their application, focusing on cerebral organoids derived from embryonic stem cells and iPSCs. We discuss new insights into organoid-based model of schizophrenia and shed light on challenges and future applications of organoid-based disease model system. This review also suggests hitherto unrevealed potential applications of organoids in combining with new technologies such as nanophotonics/optogenomics for controlling brain development and atomic force microscopy for studying mechanical forces that shape the developing brain.

Organisation of the dopamine neuronal subsets within midbrain of the feathertail glider (Acrobates pygmaeus, Acrobatidae, Marsupialia).

The Marsupial feathertail glider has a unique set of morphological, anatomical and behavioural features that make it a promising model for study of primate evolution. Among them it has many locomotor adaptations to arboreal life, such as diagonal gait of movements, gliding, fast climbing and running along branches. These ecological and behavioural specialisations could result in differences in anatomy of the brain systems involved in their integration. It is well acknowledged that dopaminergic neurons are involved in motor control, motivation and cognition. Due to the fact that there are no data on morphological organisation of dopaminergic system in the midbrain of this species, we decided to investigate it using immunohistochemical and quantitative methods. Our study showed that the general distribution and characteristics of the dopaminergic cells within midbrain nuclei of the pygmy acrobat is similar to that in other species, but it lack the substantia nigra compact part - ventral tier and "tail" of the substantia nigra subnuclei. This study provides the first description of the dopaminergic cells and nuclei in the midbrain of the feathertail glider and we hope it will start interest in the neurobiology of this species.

Schizophrenia: a neurodevelopmental disorder--integrative genomic hypothesis and therapeutic implications from a transgenic mouse model.

Schizophrenia is a neurodevelopmental disorder featuring complex aberrations in the structure, wiring, and chemistry of multiple neuronal systems. The abnormal developmental trajectory of the brain appears to be established during gestation, long before clinical symptoms of the disease appear in early adult life. Many genes are associated with schizophrenia, however, altered expression of no one gene has been shown to be present in a majority of schizophrenia patients. How does altered expression of such a variety of genes lead to the complex set of abnormalities observed in the schizophrenic brain? We hypothesize that the protein products of these genes converge on common neurodevelopmental pathways that affect the development of multiple neural circuits and neurotransmitter systems. One such neurodevelopmental pathway is Integrative Nuclear FGFR1 Signaling (INFS). INFS integrates diverse neurogenic signals that direct the postmitotic development of embryonic stem cells, neural progenitors and immature neurons, by direct gene reprogramming. Additionally, FGFR1 and its partner proteins link multiple upstream pathways in which schizophrenia-linked genes are known to function and interact directly with those genes. A th-fgfr1(tk-) transgenic mouse with impaired FGF receptor signaling establishes a number of important characteristics that mimic human schizophrenia - a neurodevelopmental origin, anatomical abnormalities at birth, a delayed onset of behavioral symptoms, deficits across multiple domains of the disorder and symptom improvement with typical and atypical antipsychotics, 5-HT antagonists, and nicotinic receptor agonists. Our research suggests that altered FGF receptor signaling plays a central role in the developmental abnormalities underlying schizophrenia and that nicotinic agonists are an effective class of compounds for the treatment of schizophrenia.

Effect of light-dark changes on the locomotor activity in open field in adult rats and opossums.

There have been no reports on how the light-dark changes determine the locomotor activity of animals in the group of high reactivity (HR) and low reactivity (LR). In the present study we have compared selected parameters of the locomotor activity of the HR and the LR groups of the laboratory opossums and Wistar rats during consecutive, light and dark phases in the open field test. Sixty male Wistar adult rats, at an average weight of 350 g each, and 24 adult Monodelphis opossums of both sexes at an average weight of 120 g each were used. The animals' activity for 2 h daily between the hours of 17:30 and 19:30, in line with the natural light-dark cycle were recorded and then analysed using VideoTrack ver.2.0 (Vievpoint France). According to our results, we noted that a change of the experimental conditions from light to dark involves an increase in the locomotor activity in rats and opossums of the HR group, while there is no effect on the activity of the rats and opossums in the LR group. Locomotor activity in the HR rats, both in the light and dark conditions is characterised by a consistent pattern of change - higher activity in the first stage of the recording and a slowdown (habituation) in the second phase of the observation. The locomotor activity of the opossum, during both light and dark conditions, was observed to be at a consistently high level compared to the rats.

Nerve growth factor (NGF) immunoreactive neurons in the juvenile rat hippocampus: response to acute and long-term high-light open-field (HL-OF) or forced swim (FS) stress stimulation.

This study aimed at examining and comparing the influence of two different stress stimuli on the density (number of cells/mm²) of nerve growth factor (NGF) containing neurons in the hippocampal CA1 and CA3 pyramidal cell layers and the dentate gyrus (DG) granule cell layer in juvenile rats (P28; P-postnatal day). The high-light open-field (HL-OF) test and forced swim (FS) test were employed to investigate the effects of a single, 15-min acute exposure and repeated (15 min daily for 21 days) long-term exposure to stress. In order to detect NGF-ir neurons, immunohistochemical (-ir) techniques were used. In comparison with nonstressed animals, acute and long-term HL-OF or FS stimulation resulted in a marked increase (P<0.001) in the density of NGF-ir containing cells in all the hippocampal structures. The frequency of stress application (acute vs. long-term), however, did not have a substantial impact on the studied parameter, with the exception of the CA3 sector, where a decreased density (P<0.001) of NGF-ir neurons was observed after long-term exposure to FS. It may be concluded that a rise in the density of NGF-ir neurons in the juvenile rat hippocampus after exposure to HL-OF or FS stressors could have affected the activity of the hypothalamic-pituitary-adrenocortical (HPA) stress axis. Prolonged HL-OF or FS stress was probably aggravating enough not to trigger the habituation process. The type of stressor applied (HL-OF vs. FS) was not essentially a factor determining the density of NGF-ir cells in the hippocampus.

Stress-induced changes of interleukin-1beta within the limbic system in the rat.

The aim of this study was to investigate the influence of two periods of life, namely P28 and P360, on the changes in interleukin-1beta (IL-1beta) immunoreactivity (-ir) in the hippocampus (CA1, CA3, DG) and amygdala (central-CeA, medial-MeA) caused by acute and repeated open field (OF), or by forced swim (FS) exposition. Rats were divided into groups: non-stressed, exposed to acute (one-time for 15 min) and chronic stressors (21 days for 15 min daily). We found IL-1beta-ir in the control group to be higher in P360 than in P28. In P28, under OF and FS exposure, IL-1beta-ir in the CeA remained unaltered but increased in the MeA and in the hippocampus after acute and chronic stress. In P360 no changes were observed in the IL-1beta-ir level after acute and chronic stimulation. These data demonstrate that only the levels of IL-1beta-ir in juvenile rat brains are affected by FS and OF. Additionally, there was no significant difference between FS and OF stimulation in IL-1beta-ir.

Changes in NGF/c-Fos colocalization in specific limbic structures of juvenile and aged rats after open field stimulation.

Changes in NGF release during stressful events have been associated with the activation of neurons expressing NGF receptors. This study examined the influence of acute stress-induced stimulation on NGF/c-Fos colocalization in the following limbic regions: the paraventricular (PV) nucleus of the hypothalamus, medial (MeA) nucleus of the amygdala, and CA3 hippocampus. Juvenile (P21) and aged rats (P360) were exposed to a 15-minute acute open field (OF) test. Double immunofluorescence staining, used to detect NGF-ir and c-Fos-ir cells, revealed a higher percentage of NGF/c-Fos-ir neurons in the P21 control group than in the P360 control group. Under OF acute stimulation, a statistically significant (p < 0.05) increase of NGF/c-Fos level in CA3 of juvenile animals and in PV and CA3 of the aged rats was observed. These observations indicate that the investigated structures in both age groups show a different response to acute OF stimulation. Acute OF affects the levels of NGF/c-Fos more significantly in aged rats.

ORMOSIL nanoparticles as a non-viral gene delivery vector for modeling polyglutamine induced brain pathology.

Studies have shown the presence of expanded polyQ containing proteins in brain cells related to Huntington disease (HD) and other poly-glutamine disorders. We report the use of organically modified silica (ORMOSIL) nanoparticles as an efficient non-viral gene carrier in an effort to model brain pathology associated with those disorders induced by expanded polyQ peptides. In experiment 1, plasmids expressing Hemaglutinin-tagged polypeptides with 20 glutamine repeats (Q20) or with extended 127-glutamine repeats (Q127) were complexed with ORMOSIL nanoparticles and injected twice (2 weeks apart) into the lateral ventricle of the mouse brain. Fourteen days post-injection of Q127, immunocytochemistry revealed the presence of the characteristic nuclear and cytoplasmic Q127 aggregates in numerous striatal, septal and neocortical neuronal cells as well as ubiquitin-containing aggregates indicative of the neuronal pathology. The mice receiving Q127 showed a marked increase in the reactive GFAP (+) astrocytes in striatum, septum and brain cortex, further indicating the neurodegenerative changes, accompanied by motor impairments. In experiment 2, plasmids Q20 or Q127 were complexed with ORMOSIL and were injected into the brain lateral ventricle or directly into the striatum of adult rats. In both routes of transfection, Q127 induced the appearance of reactive GFAP (+) astrocytes and activated ED1 antigen expressing microglia. An increase in the size of the lateral ventricle was also observed in rats receiving Q127. In transgenic mouse polyQ models, extensive pathologies occur outside the nervous system and the observed brain pathologies could reflect developmental effects of the toxic polyQ proteins. Our experiments show that the nervous tissue restricted expression of poly Q-extended peptides in adult brain is sufficient to evoke neuropathologies associated with HD and other polyQ disorders. Thus, nanotechnology can be employed to model pathological and behavioral aspects of genetic brain diseases in mice as well as in other species, providing a novel research tool for in vivo testing of single or multi-gene therapies.

The influence of acute and chronic open-field exposure on the hippocampal formation: an immunohistochemical study.

The hippocampus plays a role in new learning, memory and emotion and is a component of the neuroanatomical stress circuit. The structure is involved in terminating hypothalamic-pituitary-adrenocortical (HPA) axis responses to stress and attenuates stress responses by shutting off this axis. The immunoreactivity (-ir) of c-Fos, NGF and its receptor TrkA following acute and chronic open-field stress were studied in CA1-CA3 and the DG of the hippocampus. The material consisted of 21 male adult rats divided into three groups: nonstressed (control) animals and rats exposed to acute (15 min once) and chronic (15 min daily for 21 days) aversive stimulation (open-field exposure). The brains were stained with use of immunohistochemical methods for c-Fos, NGF or TrkA. In the animals exposed to acute open-field stress the number of c-Fos-, TrkA and NGF-ir cells was higher in all the structures studied than in the control animals. However they were differentiated only in c-Fos immunoreactivity. In the rats exposed to chronic open-field stress the number of c-Fos-ir cells in the structures of the hippocampal formation studied was smaller than in rats exposed to acute stress and was comparable to that in the control group. No differences were observed between the groups exposed to acute and chronic stress in the number of TrkA-ir cells in the structures under investigation. The number of NGF-ir neurons in CA1 and CA2 was lower after exposure to chronic than after exposure to acute stress but was still higher than that in the control group. Our findings indicate that neurons of CA1-CA3 and the DG are engaged in the stress response after acute as well as chronic open-field exposure. This is probably related to the important role of the hippocampus in processing new spatial information as well as in the habituation processes, although these appear to have different mechanisms.

Immunoreactivity of c-Fos, NGF and its receptor TrkA in the periventricular zone of the rat hypothalamus after open field exposure.

The immunoreactivity (ir) for c-Fos, NGF and TrkA, following an acute and chronic open field stress, were studied in the periventricular zone of rat hypothalamus. Adult rats were divided into three groups: control, exposed to acute (single exposure--15 minutes) and chronic (multiple exposures--15 minutes daily for 21 days) open field stress. In the control rats neurons immunoreactive to c-Fos, TrkA and NGF were found. The number of TrkA- and NGF-ir cells was high, whereas this of c-Fos-ir ones was low. In animals exposed to acute open field stress the number of c-Fos-ir cells in the examined nuclei varied, however it was much higher than that in the control animals. The number of TrkA-ir neurons in all the studied nuclei was also higher than that in the control animals, but the increase of the number of NGF-ir neurons was not observed in supraoptic nucleus. In the animals exposed to chronic open field stress the number of c-Fos-ir cells was increased in comparison to that in the control rats. After chronic stress exposure the number of TrkA-ir neurons in supraoptic nucleus remained high in comparison to that in animals exposed to acute stress, whereas it was decreased in other studied nuclei. No significant differences in the number of NGF-ir cells were observed between the groups exposed to the acute and chronic stress. Observed decrease of c-Fos- and TrkA-ir in the studied nuclei in the animals suffering from chronic stress in comparison with the acute one may indicate the occurrence of habituation phenomenon. This phenomenon does not concern NGF-ir.

Influence of the "open field" exposure on calbindin D28K, calretinin, and parvalbumin containing cells in the rat midbrain - developmental study.

The aim of our study was to analyze the influence of the open field (OF) exposure on: 1. Distribution of c-Fos positive nuclei in: ventral tegmental area, substantia nigra, periaqueductal gray. 2. Appearance of calbindin-D28k, calretinin and parvalbumin in midbrain neurons that are engaged in the stress response. 3. Changes of c-Fos and calcium-binding proteins expression during maturation. The material consisted of Wistar rats of age between 0 and 90 days. The OF exposure was applied throughout 10 min and 90 min before the death of the animals. The brain sections were double stained using the antibodies against c-Fos, CB, CR or PV. Our results showed that in all studied nuclei age-related increase of c-Fos expression (without changing of its distribution properties) was found. PV didn't show any co-localization with c-Fos in neurons of studied regions at any ages, however some PV-immunoreactive (PV-ir) basket-like structures around c-Fos-immunoreactive (c-Fos-ir) neurons were observed. In the youngest group of rats c-Fos-ir cells and cells immunoreactive for CB and CR constituted separate neuronal populations. During maturation increases in the level of their co-localization with c-Fos was observed. We may conclude that in adult rat midbrain structures CB-immunoreactive (CB-ir) and CR-immunoreactive (CR-ir) cells (probably projection neurons) are mainly activated in the stress response following OF exposure. In the contrary PV-ir cells has only an indirect (modulatory) influence upon the c-Fos-ir cells.

Open field stress and neurons containing calcium-binding proteins in the piriform cortex of the rat.

In the present study we wanted to check whether the expression of the c-Fos protein (the marker of cellular activity) appears in cells containing calcium-binding proteins (CaBPs) in animals exposed to the open field test. Eight adult Wistar rats were examined. In the first step the open field test was applied throughout 10 minutes. After perfusional fixation brains were frozen and cut on the cryostat in the coronal plane and stained with the standard immunohistochemical method. Sections were double stained for c-Fos and CaBPs: parvalbumin (PV), calbindin (CB), calretinin (CR). c-Fos positive cells were localized predominantly in layers II and III of the piriform cortex (PC). The double labeling study showed that neurons containing CaBPs are rarely c-Fos-immunoreactive. Often PV-positive and CB-positive fibers surround c-Fos-positive neurons in layers II and III in a form of a basket. It seems that cells containing CaBPs are not directly involved in the response to aversive stimuli but cells containing those calcium-binding proteins might influence directly c-Fos positive neurons of PC.

The relationships between neurons containing dopamine and nitric oxide synthase in the ventral tegmental area.

Ventral tegmental area (VTA) is a heterogeneous group of dopaminergic cells which contains interfascicular (IF), parabrachial (PBP) and rostral linear (RLi) nuclei. Neurons of this area are involved in the regulation of motor and motivational aspects of behavior and reveal high neuronal plasticity. Among many various neurotransmitters and neuromodulators, nitric oxide (NO) is localized in this region. In the present study, we investigated morphology and distribution of nitric oxide synthase (NOS)-positive neurons in VTA and their colocalization with dopaminergic neurons. The study was performed on six adult Wistar rats. After perfusional fixation, the brains were cut, immunostained for tyrosine hydroxylase (TH) and NOS and studied by confocal laser microscopy. In each of the three studied nuclei of VTA we investigated three different neuronal populations. Numerous TH-immunoreactive (TH-ir) and NOS-immunoreactive (NOS-ir) neurons are present in the studied region. Among them, a considerable number showed coexistence of both neurotransmitters. The populations of TH-ir and NOS-ir neurons interact with each other as manifested by the presence of NOS-ir endings on TH-ir neurons and vice versa. Taking the above into account, it may be suspected that NO is involved in the modulation of dopaminergic transmission.

The developmental pattern of c-fos expression in the rat thalamus following open-field stress stimulation.

The thalamic nuclei with their defined set of input-output connections are the primary channel for information flow to the cerebral cortex. Several data suggest that neurons of that area are involved in the response to various aversive stimulations. However the pattern of activation seems to depend on the stress model as well as the stage of maturation. In the present study we would like to check which nuclei of the thalamus show expression of c-fos in the response to the "open field test", and how this response pattern changes during the maturation process. 30 rats of age ranged from P0 to P120 (P-postnatal day) were studied. The experimental group was exposed to the "open field test" for 10 minutes. After perfusion and fixation, brains were cut and stained for c-fos with immunohistochemical method. Our results showed that during development the pattern of c-fos activity in the thalamic nuclei after stress stimulation undergoes significant changes. Distinct c-fos expression was observed in the paraventricular nucleus, intergeniculate leaflet and ventral lateral geniculate nucleus. These findings suggest that these nuclei may play a direct role in the stress reaction involved in the response to the "open field test".

Facilitatory effect of unilateral lesion of the ventral tegmental area on locomotor response to stimulation of the contralateral ventral tegmental area: involvement of GABAergic transmission.

It was found previously that in the rat, unilateral electrolytic lesion of the ventral tegmental area (VTA) facilitated feeding induced by electrical stimulation of the homologous VTA tissue in the contralateral hemisphere. In the present work, VTA stimulation-induced locomotor response was tested in male Wistar rats using a latency to move/stimulation frequency curve shift paradigm in order to check for functional generality of the "contralateral facilitation effect" and also with the aim of elaborating an easy and reliable behavioral model to study this phenomenon. In a further step, the hypothesis was tested that enhancement of function of the intact VTA results from elimination of tonic GABAergic influence derived normally from the lesioned VTA. GABA(A) (bicuculline, doses 0, 0.5 and 5.0 ng) and GABA(B) (phaclofen, doses 0, 500 and 1000 ng) receptors antagonists, and for comparison, a GABA(A) receptor agonist (muscimol, doses 0, 12.5, 25. 0 and 50.0 ng), were injected unilaterally to VTA and their effect on locomotor response elicited by electrical stimulation of the contralateral VTA was tested in a latency/frequency paradigm. It was found that similar to feeding, locomotor response evoked by unilateral electrical stimulation of the VTA was facilitated after contralateral VTA lesion which manifested as a decrease of the locomotion threshold and a leftward shift of the function relating latency to move to stimulation frequency. The effect was immediate, long-lasting and specific to the VTA destruction; lesions outside the VTA area caused gradual impairment of the locomotor response to stimulation. The facilitatory effect of the electrolytic lesion could be replicated by bicuculline, which significantly facilitated stimulation-induced behavior. Phaclofen exerted slight facilitating influence only at a low dose. No effect of muscimol on the locomotion threshold was found. We conclude that "the contralateral facilitation effect" at the level of VTA reflects the interhemispheric regulation of activity of the dopaminergic (DA) cells in which GABA(A)-mediated interhemispheric communication plays a significant role.