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.

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.

Age-Related (Aged vs. Adult) Comparison of the Effect of Two Mild Stressors on the Nerve Growth Factor (NGF) in the Rat Hypothalamic Supraoptic Nucleus (SON) - Immunohistochemical Study.

The ontogenetic period of life and stress can have different effects on the nerve growth factor (NGF) in the hypothalamus. The aim of our study was to investigate the influence of two mild stressors, acute and chronic exposure to forced swim (FS) or high-light open field (HL-OF), on neurons containing NGF. Immunofluorescence staining was used to reveal the density of NGF-immunoreactive (ir) cells in the hypothalamic supraoptic nucleus (SON) in adult (postnatal day 90; P90) and aged (P720) rats. The P90 and P720 rats that were subjected to acute and chronic FS showed no differences in the density of NGF-ir neurons in the SON compared with nonstressed rats. However, a significant increase in NGF-ir cells was noted after acute but not after chronic HL-OF only in P90 rats. What is more, there were no age-related (P90 vs. P720) changes in the density of NGF-ir neurons in non-stressed and FS- or HL-OF-stressed rats. Our results indicate that acute HL-OF was the only factor inducing changes in the density of NGF-ir neurons in the SON of adult rats. This could be related to the neuroprotective role of NGF-ir cells in response to acute HL-OF. The absence of age-dependent changes in the density of NGF-ir neurons may indicate that the ageing processes in SON do not generate changes in the NGF immunoreactivity of its neurons.

The effect of mild stress stimulation on the nerve growth factor (NGF) and tyrosine kinase receptor A (TrkA) immunoreactivity in the paraventricular nucleus (PVN) of the hypothalamus and hippocampus in aged vs. adult rats.

Ontogenetic life and stress can have different effects on the nerve growth factor (NGF) and its tyrosine kinase receptor A (TrkA) in the structures of the limbic system. This study aimed to explore the influence of two different stressors, acute and chronic exposure to forced swim (FS) stress or high-light open-field (HL-OF) stress, on cells containing NGF and TrkA. Immunofluorescence staining was used to reveal the density of NGF and TrkA immunoreactive (ir) cells in the paraventricular nucleus (PVN) of the hypothalamus or hippocampal subfields CA1, CA3 and dentate gyrus (DG) in adult (postnatal day 90; P90) and aged (P720) rats. Data revealed that neither acute nor chronic FS caused any alteration in NGF-ir and TrkA-ir cells in any of the structures investigated in P90 and P720 rats. However, a significant increase in NGF-ir was detected in the CA1 and CA3 after acute but not after chronic HL-OF in both age groups. The TrkA-ir remained unchanged after exposure to HL-OF in the PVN and hippocampus. Despite lack of change in the density of NGF-ir and TrkA-ir cells between P90 and P720 non-stressed rats, a significant age-related decrease in NGF-ir and TrkA-ir cells in the PVN of FS- and HL-OF-stressed rats was noted. However, in the hippocampus, an age-related decrease in NGF-ir or TrkA-ir cells was observed in all rats except acute FS-stressed rats. The changes are possibly associated with involutional aging processes caused by insufficient control of hypothalamic-pituitary-adrenal (HPA) axis functioning in P720 rats and may contribute to disturbances in NGF signaling.

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.

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.

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.

The immunoreactivity of c-Fos, NGF and its receptor TrkA after open-field exposure in the central and medial nuclei of the rat amygdala.

The amygdala is a critical component of the neuroanatomical stress circuit. It plays a role in the generation of responses to emotional stimuli. The central (CeA) and medial (MeA) amygdaloid nuclei are implicated in activation of the hypothalamic-pituitary-adrenocortical (HPA) axis. The immunoreactivity (-ir) of c-Fos, NGF and its receptor, TrkA, following acute and chronic open-field stress were studied in the CeA and MeA nuclei of the amygdala. The material consisted of 21 male adult rats divided into three groups: non-stressed (control) animals, rats exposed to acute (once only lasting 15 min) and chronic (15 min daily over 21 days) aversive stimulation (open-field exposure). The brains were stained with the use of immunohistochemical methods for c-Fos, NGF or TrkA. In the control rats c-Fos-, TrkA- and NGF-ir cells were observed in the nuclei studied, but the quantity varied, being moderate or high (immunoreactive to TrkA and NGF) or low (immunoreactive to c-Fos). In the animals exposed to acute open-field stress the number of c-Fos-ir, NGF-ir and TrkA-ir cells in the nuclei under examination was differentiated but higher than that in the control animals. In the animals exposed to chronic open-field stress the number of c-Fos-ir cells in the nuclei studied was similar and was smaller than those in animals exposed to acute stress. The number of TrkA-ir neurons was also lower in comparison to that in animals exposed to acute stress. However, no significant differences in the number of NGF-ir cells were observed between the groups exposed to acute and chronic stress. Diverse expression of c-Fos protein following both acute and chronic stress stimulation may prove the functional heterogeneity of the amygdaloid nuclei investigated. The decrease observed in both c-Fos- and TrkA-ir in MeA (only TrkA in CeA) of animals exposed to chronic stress may indicate the phenomenon of habituation.

Quantitative analysis of influence of sevoflurane on the reactivity of microglial cells in the course of the experimental model of intracerebral haemorrhage.

BACKGROUND: Microglial cells play an important role in the pathophysiology of intracerebral haemorrhage. We have examined the possible influence of sevoflurane on the reactivity of microglial cells during intracranial haemorrhage. METHODS: Forty adult male rats were divided into two groups. All animals were anaesthetized with fentanyl, dehydrobenzperidol and midazolam. In the experimental group animals additionally received sevoflurane 2.2 vol% end-tidal concentration. Intracranial haemorrhage was produced through infusion of blood into the striatum. The microglial cell population (numerical density of immunoreactive cells and their distribution) was assessed on days 1, 3, 7, 14 and 21 after producing a haematoma using antibodies OX42 and OX6. RESULTS: In the control group significant differences in the density of OX42-ir cells between 3rd and 7th (81.86 vs. 129.99) (95% CI: -77.99 to -18.25, P = 0.0035) and between 14th and 21st (105.36 vs. 63.81) (95% CI: 13.21 to 69.89, P = 0.006) survival days were observed. However, significant increase of percentage of amoeboid OX42-ir cells between 3rd and 7th (0.98 vs. 48.71) (95% CI: -52.17 to -43.30, P = 0.0001) and between 7th and 14th (48.71 vs. 58.47) (95% CI: -13.96 to -5.55, P = 0.0002) and then their decrease - between 14th and 21st (58.47 vs. 31.74) (95% CI: 22.52 to 30.93, P = 0.0001) days of observation were noted. In the sevoflurane groups OX42-ir cells were not found. On the 3rd day the density of OX6-ir cells in the sevoflurane group was significantly lower than that in the control group (12.39 vs. 34.57) (95% CI: -49.78 to -2.96, P = 0.02). The percentage of an amoeboid form of OX6-ir cells was significantly lower in the sevoflurane group than that in the control group (27.31 vs. 82.03) (95% CI: -72.52 to -36.92, P = 0.0001) (58.76 vs. 82.37) (95% CI: -38.81 to -8.41, P = 0.003) (42.87 vs. 81.55) (95% CI: -53.23 to -24.10, P = 0.0001) respectively for 3rd, 7th and 14th days of survival. CONCLUSION: Administration of sevoflurane during anaesthesia in animals with intracerebral haemorrhage evoked a decrease of activation of the microglial cells.

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.

The influence of sevoflurane on the reactivity of astrocytes in the course of the experimental intracerebral haemorrhage in rat.

40 adult Wistar rats were divided into two groups depending on the applied anaesthesia. In both groups animals were generally anaesthetized with fentanyl, dehydrobenzperidol administered intraperitoneally and midazolam given intramuscularly. In the second group (SEVO) animals received sevoflurane of 2.2 vol% end-tidal concentration. Intracerebral haematoma was produced through infusion of 100 microl of autologous blood into the striatum. Each group was divided into five subgroups depending on the length of survival period: 1, 3, 7, 14, 21 days. The astrocytic population was studied by means of anti-GFAP staining. Stereological analysis was applied to estimate the numerical density of immunoreactive cells and the distribution of their types. On 7th day of observation the density of GFAP-immunoreactive astrocytes in SEVO was lower (p<0,05) than that in the control group. In the control group, the increase (p<0.05) of per cent of activated astrocytes between the 1st and 3rd survival day was noted, which remained at this level till the end of observation. In SEVO group, the increase (p<0.05) of per cent of activated astrocytes between the 3rd and 7th day and the decrease (p<0.05) between the 14th and 21st survival day were observed. During days of observation the per cent of activated astrocytes was lower (p<0.05) in the SEVO group than that in the control group. Administration of sevoflurane during anaesthesia to animals with intracerebral haemorrhage has evoked not only the delay of the activation of astrocytes but also decrease in its level.

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".

Apoptotic cell death in the porcine endometrium during the oestrous cycle.

It has been reported that apoptosis plays an essential role in controlling the physiological cell kinetics in the human and rodent endometrium but this type of death has never been studied in the porcine endometrium. The aim of this study was to investigate the apoptotic cell death in the porcine endometrium during the middle (Days 9-11) and late (Day 13) luteal phase, during the luteolysis (Day 15) and early follicular phase (Days 17-19) of the oestrous cycle. Apoptotic cells were identified by in situ DNA 3'-end labelling method. It was revealed that the greatest number of apoptotic cells in the luminal and glandular epithelium was found on Days 17-19 and on Day 15 of the oestrous cycle, respectively. In the stroma, the greatest number of these cells was found on Days 9-11. Our data have shown that in the porcine endometrium, both epithelial and stromal cells undergo apoptosis and that the number of apoptotic cells varies depending on the phase of the oestrous cycle.

Microglial and astroglial cells in the rat paraclaustral reservoir during postnatal development: an immunohistochemical study.

A immunohistochemical study of postnatal development of the paraclaustral reservoir of migrating cells in the rat brain was performed using anti-GFAP (for astroglia), ED1 and OX-42 (for microglia) antibodies. From birth to the 4th day of postnatal life most GFAP-positive cells in the paraclaustral reservoir are similar to transitional astroglia. From the end of the first postnatal week they have the morphology of mature astrocytes, although during the next week, their density was a slightly higher than in neighboring structures. On the 21st day, the morphology and density of astroglial cells in the ventral part of the external capsule did not differ from the surrounding regions. ED1/OX-42- positive microglial cells present in the paraclaustral reservoir during the first postnatal week represented ameboid microglia; their density was clearly higher than in the neighboring structures. During the second week they began to transform into ramified microglia and from the 21st day on, only OX-42 positive resting microglial cells were observed in the ventral part of the external capsule. We suggest that the paraclaustral reservoir is a place of accumulation of astroglia and microglia during brain development and may possibly serve as source of glial cells for neighboring structures. Alternatively, these glial populations may perform local developmental functions.

Cholinergic endings on various neurones containing calcium binding proteins and glutamic acid decarboxylase in the hippocampus of the rat.

Immunohistochemical study of the cholinergic innervation of the hippocampal cells containing glutamic acid decarboxylase (GAD) and calcium binding proteins: parvalbumin (PV), calbindin D28k (CB) and calretinin (CR) was conducted on 5 adult rat brains. Analysis of sections with double immunostaining for vesicular acetylcholine transporter (VAChT; the marker of cholinergic cells, fibres and terminals) and respectively either GAD or PV, CB, CR, using confocal laser-scanning microscope shows that the intensive cholinergic innervations receive GAD, PV and CB-positive hippocampal cells. Cholinergic afferentiations of the CR-positive neurones are considerably fewer.